February 2012 5-1 Draft – Do Not Cite or Quote

CHAPTER 5 INTEGRATED HEALTH EFFECTS 

OF LEAD EXPOSURE

5.1 Introduction

1 This chapter reviews, summarizes, and integrates the evidence for the broad spectrum of 

2 health effects associated with exposure to Pb. The chapter begins (Section 5.2) with a 

3 discussion of the evidence for the modes of action that mediate the health effects of Pb, 

4 including those modes of action that are shared by all of the health effects evaluated in 

5 this ISA and those modes of action that are specific for particular endpoints. Subsequent 

6 sections comprise evaluations of the epidemiologic and toxicological evidence for the 

7 health effects of Pb exposure on major outcome categories such as nervous system effects 

8 (Section 5.3), cardiovascular effects (Section 5.4), renal effects (Section 5.5), immune 

9 effects (Section 5.6), effects on heme synthesis and red blood cell function (Section 5.7), 

10 and reproductive and developmental effects (Section 5.8). Section 5.9 provides reviews 

11 of the evidence for the effects of Pb on other noncancer health outcomes, for which the 

12 cumulative bodies of evidence are smaller, including those related to the hepatic system 

13 (Section 5.9.1), gastrointestinal system (Section 5.9.2), endocrine system (Section 5.9.3), 

14 bone and teeth (Section 5.9.4), ocular health (Section 5.9.5), and respiratory system 

15 (Section 5.9.6). Chapter 5 concludes with a discussion of the evidence for Pb effects on 

16 cancer (Section 5.10).

17 Individual sections for major outcome categories (e.g., nervous system, cardiovascular, 

18 renal) begin with a brief summary of conclusions from the 2006 Pb AQCD followed by 

19 an evaluation of recent evidence that is intended to build upon evidence from previous 

20 reviews. Within each of these sections, results are organized by endpoint (e.g., cognitive 

21 function, behavior, neurodegenerative diseases) then by specific scientific discipline 

22 (i.e., epidemiology, toxicology). Sections for each of the major outcome categories 

23 (e.g., nervous system, cardiovascular, renal effects) conclude with an integrated summary 

24 of the evaluation of evidence and a conclusion regarding causality. Based upon the 

25 framework (described in the Preamble), a determination of causality was made for a 

26 broad outcome category (i.e., nervous system effects) by evaluating the coherence of 

27 evidence across disciplines and across a spectrum of related endpoints. Each discussion 

28 leading up to the causal determination characterizes the evidence on which the causal 

29 judgment was based, including the strength of evidence for the individual endpoints 

30 within the major outcome category.
February 2012 5-2 Draft – Do Not Cite or Quote

5.2 Modes of Action

5.2.1 Introduction

1 The diverse health effects of Pb are dependent on multiple factors, including the 

2 concentration and duration of exposure, the particular Pb compounds constituting the 

3 exposure, and which tissues are affected. A mode of action (MOA) is the common set of 

4 biochemical, physiological, or behavioral responses (i.e., empirically observable 

5 precursor steps) that can cumulatively result in the formation of negative health 

6 outcomes. Although the effects of Pb appear to be mediated through multiple modes of 

7 action, alteration of cellular ion status (including disruption of calcium homeostasis, 

8 altered ion transport mechanisms, and perturbed protein function through displacement of 

9 metal cofactors) seems to be the major unifying mode of action underlying all subsequent 

10 modes of action (Figure 5-1). This section draws information from all of the subsequent 

11 health effects sections in Chapter 5, and identifies the major modes of action operating at 

12 the molecular, cellular, and tissue/organ level. In turn, the individual health effect 

13 sections bridge these effects to those observed on the organismal level. Each of the 

14 individual health effect sections includes a more detailed description of the mechanisms 

15 specific to the individual health effect. Accordingly, this section differs in structure and 

16 content from other health effects sections as it does not primarily focus on the literature 

17 published since the 2006 Pb AQCD, but rather incorporates recent information with older 

18 studies (which represent the current state of the science) on the possible modes of action 

19 of Pb. 
February 2012 5-3 Draft – Do Not Cite or Quote

Note: The subsections where these MOAs are discussed are indicated in parentheses.

Figure 5-1 Schematic representation of the relationships between the 

various MOAs by which Pb exerts its health effects.

5.2.2 Altered Ion Status

1 Physiologically-relevant metal ions (e.g., Ca, Mg, Zn, Fe) are known to have a multitude 

2 of functions in biological systems, including roles as charge carriers, intermediates in 

3 enzymatically-catalyzed reactions, and as structural elements in the proper maintenance 

4 of tertiary protein conformations (Garza et al., 2006). It is through disruption of these 

5 biological functions that Pb effects its negative actions, ultimately interfering with such 

6 tightly regulated processes as cell signaling, intracellular ion homeostasis, ion transport, 

7 energy metabolism, and enzymatic function.

5.2.2.1 Disruption of Ca2+ Homeostasis

Calcium (Ca2+ 8 ) is one of the most important carriers of cell signals and regulates virtually 

9 all aspects of cell function, including energy metabolism, signal transduction, hormonal 

regulation, cellular motility, and apoptosis (Carafoli, 2005). Ca2+ 10 homeostasis is 

11 maintained through a tightly regulated balance of cellular transport and intracellular 

storage (Pentyala et al., 2010). Disruption of Ca2+ 12 homeostasis by Pb has been observed 

13 in a number of different cell types and cell-free environments, indicating that this is a 

14 major mode of action for Pb-induced toxicity on a cellular level.
February 2012 5-4 Draft – Do Not Cite or Quote

Ca2+ 1 homeostasis is particularly important in bone cells, as the skeletal system serves as 

the major dynamic reservoir of Ca2+ 2 in the body (Wiemann et al., 1999; Long et al., 

3 1992). Bone cells also are unique in that they can exist in a microenvironment that is high 

in both Ca2+ 4 and Pb concentrations, potentially increasing their relative susceptibility to 

5 Pb-induced toxicity (Long et al., 1992). A series of studies from the laboratory of Long, 

6 Dowd, and Rosen have indicated that exposure of cultured osteoblastic bone cells to Pb 

disrupts intracellular Ca2+ levels ([Ca2+ 7 ]i). Exposure of osteoblasts to 1, 5, or 25 μM Pb 

for 40-300 minutes resulted in prolonged increases in [Ca2+ 8 ]i of 36, 50 and 120% over 

9 baseline, respectively (Schanne et al., 1997; Schanne et al., 1989). Long et al. (1992)

10 observed that exposure of osteoblasts to either 400 ng parathyroid hormone (PTH)/mL

culture for 1 hour or 25 μM Pb for 20 hours increased [Ca2+]i 11 . Pretreatment of Pbexposed cells with PTH increased [Ca2+ 12 ]i above concentrations observed in either single 

13 exposure, indicating that Pb may disrupt the ability of bone cells to respond to normal 

hormonal control. A similar additive increase in [Ca2+ 14 ]i was also observed when bone 

15 cells were co-treated with epidermal growth factor (EGF) plus Pb, versus Pb alone (Long 

and Rosen, 1992). Pb-induced increases in [Ca2+ 16 ]i were blocked by a protein kinase C 

17 (PKC) inhibitor, indicating that PKC activation may serve as the mechanism by which Pb 

perturbs [Ca2+ 18 ]i (Schanne et al., 1997). Schirrmacher et al. (1998) also observed 

alterations in Ca2+ 19 homeostasis in osteoblasts exposed to 5 μM Pb for 50 minutes due to 

potential disruption of Ca2+ 20 -ATPases. However, Wiemann et al. (1999) demonstrated that 

exposure to 5 or 12.5 μM Pb inhibited the Ca2+-release-activated calcium influx of Ca2+ 21

independently of any inhibitory effect on Ca2+ 22 -ATPases.

Ca2+ 23 homeostasis has also been shown to be disturbed in erythrocytes exposed to Pb

24 (Quintanar-Escorza et al., 2010; Quintanar-Escorza et al., 2007; Shin et al., 2007). In 

25 blood samples taken from Pb-exposed workers (mean [SD] blood Pb level: 74.4 

[21.9] μg/dL), the [Ca2+ 26 ]i was approximately 2.5-fold higher than that seen in nonexposed 

27 workers (mean [SD] blood Pb level: 9.9 [2] μg/dL) (Quintanar-Escorza et al., 2007). The 

increase in [Ca2+ 28 ]i was associated with higher osmotic fragility and modifications in 

29 erythrocyte shape. In a separate investigation, erythrocytes from 10 healthy volunteers 

30 were exposed to Pb at concentrations of 0.2 to 6.0 μM for 24 or 120 hours, concentrationrelated increases in [Ca2+ 31 ]i were observed across all concentrations for both durations of 

32 exposure (Quintanar-Escorza et al., 2010). Subsequent exposures of erythrocytes to either 

33 0.4 or 4.0 μM Pb [corresponding to 10 or 80 μg/dL in exposed workers (Quintanar34 Escorza et al., 2007)] for 12-120 hours resulted in duration-related increases with 

35 durations >12 hours. Osmotic fragility (measured as percent hemolysis) was increased in 

36 erythrocytes exposed to 0.4 μM Pb for 24 hours. Co-incubation with a vitamin E analog 

mitigated these effects, indicating that the increase in [Ca2+ 37 ]i is dependent on the 

38 oxidative state of the erythrocytes. Shin et al. (2007) observed that incubation of human 

erythrocytes with 5 μM Pb for 1 hour resulted in a 30-fold increase in [Ca2+ 39 ]i in vitro, 
February 2012 5-5 Draft – Do Not Cite or Quote

1 inducing the pro-coagulant activity of exposed erythrocytes. Induction of pro-coagulant 

2 activity in erythrocytes could lead to thrombus formation and negatively contribute to 

3 overall cardiovascular health, whereas increased osmotic fragility could substantially 

4 reduce erythrocyte life span and ultimately lead to anemic conditions.

Similar to effects seen in erythrocytes, Pb has been observed to interfere with Ca2+ 5

6 homeostasis in platelets and white blood cells. Dowd and Gupta (1991) observed that 

7 1 μM Pb (for 3.5 hours) was the lowest exposure concentration to result in increases in 

[Ca2+]i in human platelets. The observed increase in Ca2+ 8 levels was attributed to the 

increased influx of external Ca2+, possibly through receptor-operated Ca2+ 9 channels. In 

10 mouse splenic lymphocytes, 1 μM Pb was the lowest exposure concentration found to 

increase [Ca2+ 11 ]i, with incubation periods of 10 minutes or greater (Li et al., 2008c). These 

increases in Ca2+ appeared to be reversible as [Ca2+ 12 ]i returned to baseline after one hour. 

13 Pretreatment with a calmodulin antagonist slightly mitigated the effects of Pb exposure, 

indicating a role for calmodulin in disruption of Ca2+ 14 homeostasis in lymphocytes. In rat 

tail arteries exposed to 1.2 μM Pb-acetate for 1 hour, intracellular stores of Ca2+ 15 increased 

over controls, possibly through increased transmembrane influx of Ca2+ 16 (Piccinini et al., 

17 1977).

18 Exposure of the microsomal fraction of rat brain cells to as little as 0.25 μM Pb for 

2 minutes resulted in increased release of Ca2+ 19 into the media (Pentyala et al., 2010).

Further, Pb exposure also decreased the activity of the microsomal Ca2+ 20 -ATPase, thus 

decreasing the sequestration of Ca2+ 21 into microsomes. The results of this study suggest 

that disruption of microsomal release and re-uptake of Ca2+ may alter Ca2+ 22 homeostasis, 

23 ultimately leading to altered signal transduction and neuronal dysfunction. However, 

Ferguson et al. (2000) observed that [Ca2+ 24 ]i was decreased in rat hippocampal neurons in 

25 response to exposure to 0.1 μM Pb for 1-48 hours, although the observed decreases were 

not time-dependent. The decrease in [Ca2+ 26 ]i was shown to be due to increased efflux of 

Ca2+ 27 out of the neuron via a calmodulin-regulated mechanism, possibly through 

stimulated Ca2+ efflux via Ca2+ 28 -ATPase.

5.2.2.2 Disruption of Ion Transport Mechanisms

As described above, deregulation of Ca2+ 29 homeostasis results in negative effects in 

30 multiple organ systems. Under normal conditions in the life cycle of most cells, cytosolic 

concentrations of free Ca2+ fluctuate around 100-200 nM and Ca2+ 31 that has entered the 

32 cell must be removed in order to maintain normal homeostatic concentrations (Carafoli, 

2005) . An important component of the maintenance of Ca2+ 33 homeostasis is

transmembrane transport of Ca ions via Ca2+ 34 -ATPase and voltage-sensitive gates
February 2012 5-6 Draft – Do Not Cite or Quote

(Carafoli, 2005). Pb has been shown to disrupt the normal movement of Ca2+ 1 ions, as 

2 well as other physiologically important ions through interactions with these transport 

3 mechanisms. 

Multiple studies have reported the effects of Pb exposure on Na+-K+-ATPase, Ca2+ 4 -

ATPase, and Mg2+ 5 -ATPases in animal models. Decreases in the activity of all three 

6 ATPases were observed in the kidneys and livers of rats exposed to 750 ppm Pb in 

7 drinking water for 11 weeks (mean [SD] blood Pb level: 55.6 [6.3] μg/dL) (Kharoubi et 

8 al., 2008a) and in erythrocytes of rats exposed to 0.2% Pb in drinking water for 5 weeks 

9 (mean [SD] blood Pb level: 97.56 [11.8] μg/dL) (Sivaprasad et al., 2003). Increases in 

10 lipid peroxidation were seen in both studies, and the decrements in ATPase activities may 

11 be explained by generation of free radicals in Pb-exposed animals. A decrease in the 

activity of Na+-K+ 12 -ATPase was observed in rabbit kidney membranes exposed to 0.01 to 

13 10 μM Pb, possibly due to Pb inhibiting the hydrolytic cleavage of phosphorylated 

14 intermediates in the K-related branch of the pump (Gramigni et al., 2009). Similar 

decreases in Na+-K+ 15 -ATPase activity were observed in synaptosomes isolated from rats 

16 exposed to 200 ppm Pb in drinking water for 3 months (blood Pb level: 37.8 μg/dL)

17 (Rafalowska et al., 1996) or 15 mg/kg Pb injected (i.p.) for 7 days (blood Pb level: 

112.5 μg/dL) (Struzynska et al., 1997a). Inhibition of Na+-K+ 18 -ATPase activity was also 

19 observed in primary cerebellar granule neuronal cultures obtained from rats pre- and 

20 postnatally exposed to Pb (0.1 % Pb-acetate in dams drinking water, resulting in blood Pb 

level of 4 μg/dL) (Baranowska-Bosiacka et al., 2011a). The activity of Ca2+ 21 -ATPase in 

22 the sarcoplasmic reticulum of rabbits exposed to 0.01 μM Pb was similarly decreased

23 (Hechtenberg and Beyersmann, 1991). The inhibitory effect of Pb was diminished in the 

24 presence of high MgATP concentrations. The activity of generic ATPase was reported to 

25 be altered in the testes of rats exposed to 300 ppm Pb-acetate gestationally, and in 

26 drinking water after weaning to the age of 6, 8, 10, or 12 weeks (Liu et al., 2008). In 

27 pregnant rats fed a Pb-depleted (20 ± 5 μg/kg) or control (1 mg/kg) diet during gestation 

and lactation, no difference was observed in the activity of Na+-K+-ATPase and Ca2+ 28 -

Mg2+ 29 -ATPase in the parental generation (Eder et al., 1990). However, the offspring 

30 (exposed via placental and lactational transfer of Pb) of Pb-depleted rats displayed

31 decreased activities in both enzymes compared with offspring of rats with higher Pb 

exposures. A similar increase in the Na+-K+ 32 -ATPase activity was observed in rats 

33 exposed (i.p.) to 20 mg/kg Pb for 14 consecutive days (Jehan and Motlag, 1995). Co34 exposure of Pb with zinc and copper greatly attenuated the increase in ATPase activity. 

35 Although the precise mechanism was not investigated, Navarro-Moreno et al. (2009)

reported that Ca2+ 36 uptake was diminished in proximal renal tubule cells in rats chronically 

37 exposed to 500 ppm Pb in drinking water for 7 months (mean [SD] blood Pb level: 43.0 

38 [7.6] μg/dL).
February 2012 5-7 Draft – Do Not Cite or Quote

1 In vitro studies of ATPase activities in human erythrocyte ghosts have also shown that Pb 

2 affects the transport of metal ions across membranes. Calderon-Salinas et al. (1999b)

observed that 1-5x104μM Pb and Ca2+ 3 were capable of inhibiting the passive transport of 

4 each other in human erythrocyte ghosts incubated with both cations. Subsequent 

5 inhibition experiments indicated that both cations share the same electrogenic transport 

6 pathway (Sakuma et al., 1984). Further study by this group (Calderon-Salinas et al., 

1999a) demonstrated that Pb can noncompetitively block the transport of Ca2+ 7 by 

inhibiting the activity of Ca2+-Mg2+-ATPase at concentrations of 1-5x103 8 μM. Mas-Oliva

(1989) demonstrated that the activity of Ca2+-Mg2+ 9 -ATPase in human erythrocyte ghosts 

10 was inhibited by incubation with 0.1-100 μM Pb. The inhibitory action was most likely 

11 due to direct reaction with sulfhydryl groups on the ATPase at Pb concentrations greater 

12 than 1 μM, but due to the action of Pb on calmodulin at lower concentrations. Grabowska 

13 and Guminska (1996) observed that 10 μg/dL Pb was the lowest concentration to 

decrease the activity of Na+-K+-ATPase in erythrocyte ghosts; activity of Ca2+- Mg2+ 14 -

ATPase was less sensitive to Pb exposure, and Mg2+ 15 -ATPase activity was not affected.

16 In a study investigating ATPase activities in occupationally-exposed workers in Nigeria, 

Abam et al. (2008) observed that the activity of erythrocyte membrane-bound Ca2+-Mg2+ 17 -

18 ATPase was decreased by roughly 50% in all occupational groups (range of mean [SD] 

19 blood Pb level across nine occupational groups: 28.75 [11.31] - 42.07 [12.01] μg/dL) 

20 compared to nonexposed controls (mean [SD] blood Pb level: 12.34 [2.44] in males and 

16.85 [6.01] μg/dL in females). Higher membrane concentrations of Ca2+ 21 and magnesium 

22 were also observed, indicating that Pb prevented the efflux of those cations from the cell, 

23 most likely by substituting for those metals in the active site of the ATPase. In a study of 

24 247 mother-newborn pairs, Campagna et al. (2000) observed that newborn (cord) blood 

25 Pb (geometric mean [5th, 95th percentile]: 4.8 [2.8-9.2] μg/dL) was negatively and 

significantly associated with maternal blood Ca2+ 26 pump activities; however, newborn 

(cord) blood Pb was not significantly associated with cord blood Ca2+ 27 pump activities.

28 Newborn hair Pb (geometric mean [5th, 95th percentile]: 1.1 [0.1-8.0] μg/g) was 

negatively and significantly associated with both maternal and cord blood Ca2+ 29 pump 

30 activities. In a population of 81 newborns, Huel et al. (2008) found that newborn hair and 

31 cord blood Pb levels (mean [SD] newborn hair Pb and blood Pb levels: 1.22 [1.41] μg/g 

and 3.54 [1.72] μg/dL) were negatively associated with Ca2+ 32 -ATPase activity in plasma 

33 membranes of erythrocytes isolated from cord blood; newborn hair Pb levels were more 

strongly associated with cord Ca2+ 34 pump activity than were cord blood Pb levels. 

35 Pb has also been shown to disrupt cation transport mechanisms through direct action on 

36 voltage-sensitive cation channels. Audesirk and Audesirk (1993, 1991) demonstrated that 

extracellular free Pb inhibits the action of multiple voltage-sensitive Ca2+ 37 channels, with 

38 free Pb IC50 (half maximal inhibitory concentration) values of 0.7 μM for L-type 
February 2012 5-8 Draft – Do Not Cite or Quote

1 channels and 1.3 μM for T-type channels in neuroblastoma cells, and IC50 values as low 

2 as 0.03 μM for L-type channels in cultured hippocampal neurons. Sun and Suszkiw

(1995) corroborated the inhibitory action of extracellular Pb on Ca2+ 3 channels, 

4 demonstrating an IC50 value of 0.3 μM in adrenal chromaffin cells. The observed 

disruption of the Ca2+ channels most likely reflects competition between Pb and Ca2+ 5 for 

the extracellular Ca2+ 6 binding domain of the channel. Research by other laboratories 

supported these findings: Pb inhibited the action of multiple Ca2+ 7 channels in human 

8 embryonic kidney cells transfected with L-, N-, and R-type channels (IC50 values of 

9 0.38 μM, 1.31 μM, and 0.10 μM, respectively) (Peng et al., 2002) and P-type channels in 

10 cultured hippocampal neurons at concentrations up to 3 μM (Ujihara et al., 1995).

However, intracellular Pb was observed to enhance Ca2+ 11 currents through attenuation of

the Ca2+ dependent deactivation of Ca2+ 12 channels at an EC50 value of 200 μM, possibly 

through blocking the intracellular Ca2+ binding domain, or through Ca2+ 13 dependent 

14 dephosphorylation of the channel (Sun and Suszkiw, 1995). Recently, Pb has also been 

shown to enter cells (HEK293, HeLa, and PC12 cell lines) through store-operated Ca2+ 15

16 channels (Chiu et al., 2009; Chang et al., 2008b). In particular, the Orai1-STIM1 complex 

17 was shown to be critical in the entry of Pb ions into cells, and increased Pb permeation 

was directly related to decreased [Ca2+ 18 ]i concentrations at exposure concentrations as low 

19 as 0.1 μM.

Pb also disrupts the action of Ca2+ 20 -dependent potassium channels. Alvarez et al. (1986)

21 observed that Pb promoted the efflux of potassium from inside-out erythrocyte vesicles in 

22 a concentration-dependent manner at concentrations of 1-300 μM, either through action 

on a Mg modulatory site or through direct interaction with the Ca2+ 23 binding site. Fehlau 

24 et al. (1989) also demonstrated Pb-induced activation of the potassium channel in 

25 erythrocytes. However, Pb only activated the potassium channels at concentrations below 

26 10 μM; higher concentrations of Pb completely inhibited channel activity, indicating the 

27 modulation of potassium permeability is due to alterations in channel gating. Silken et al.

28 (2001) observed that Pb activated potassium channels in erythrocytes from the marine 

29 teleost Scorpaena porcus in a concentration-dependent manner after a 20-minute 

30 incubation; minor loss of potassium was seen at Pb concentrations of 1-2 μM, whereas 

31 exposure to 20-50 μM Pb resulted in approximately 70% potassium loss. Competitive 

32 and inhibitory binding assays suggest that Pb directly activates potassium channels in 

33 S. porcus.

Disruption of Neurotransmitter Release

Pb has been shown to inhibit the evoked release of neurotransmitters by inhibiting Ca2+ 34

35 transport through voltage-sensitive channels in in vitro experiments (Cooper and Manalis, 

36 1984; Suszkiw et al., 1984). However, in these same experiments, concentrations of Pb as 
February 2012 5-9 Draft – Do Not Cite or Quote

1 5 μM and higher were also observed to actually increase the spontaneous release of 

2 neurotransmitters. Subsequent research by other groups affirmed that Pb demonstrates 

Ca2+ 3 -mimetic properties in enhancing neurotransmitter release from cells in the absence 

of Ca2+ and Ca2+ 4 -induced depolarization. Tomsig and Suszkiw (1995, 1993) reported that 

5 Pb exposure induced the release of norepinephrine (NE) from bovine adrenal chromaffin 

cells, and was considerably more potent at doing so than was Ca2+ (K0.5 of 4.6x103 6 μM for 

Pb versus 2.4 μM for Ca2+ 7 ). Activation of PKC was observed to enhance the Pb-induced 

8 release of NE. Westerrink and Vijverberg (2002) observed that Pb acted as a high affinity 

substitute for Ca2+ 9 , and triggered enhanced catecholamine release from PC12 cells at 

10 μM in intact cells and 3x104μM in permeabilized cells. The suppression of Ca2+ 10 -

11 induced evoked release of neurotransmitters combined with the ability of Pb to enhance 

12 spontaneous releases could result in higher noise in the synaptic transmission of nerve 

13 impulses in Pb-exposed animals. In rats exposed to Pb at concentrations of 0.1-1.0% in 

14 drinking water beginning at gestational day (GD)15-16 and continuing to 120 days 

15 postnatal, decreases in total potassium-stimulated hippocampal GABA release were seen 

16 at exposure levels of 0.1-0.5% (range of mean [SD] blood Pb levels: 26.8 [1.3] - 61.8 

17 [2.9] μg/dL) (Lasley and Gilbert, 2002). Maximal effects were observed at 0.2% Pb in 

18 drinking water, but effects were less evident at 0.5%, and were absent at 1.0%. In the 

absence of Ca2+ 19 , potassium-induced GABA release was increased with the two highest 

20 exposure concentrations, suggesting a Pb-induced enhancement of evoked release of 

21 GABA. The authors suggest that this pattern of response indicates that Pb is a potent 

suppressor of evoked release at low concentrations, but a Ca2+ 22 mimic in regard to 

23 independently evoking exocytosis and release at higher concentrations (Lasley and 

24 Gilbert, 2002). Suszkiw (2004) reports that augmentation of spontaneous release of 

25 neurotransmitters may involve Pb-induced activation of CaMKII-dependent 

26 phosphorylation of synapsin I or direct activation of synaptotagmin I. Further, Suszkiw

27 (2004) suggests that unlike the intracellularly mediated effects of Pb on spontaneous 

28 release of neurotransmitters, Pb-induced inhibition of evoked transmitter releases is 

largely due to extracellular blockage of the voltage-sensitive Ca2+ 29 channels. 

5.2.2.3 Displacement of Metal Ions and Perturbed Protein 

Function

30 The binding of metal ions to proteins causes specific changes in protein shape, and the 

31 specific cellular function of many proteins may be altered by conformational changes

32 (Kirberger and Yang, 2008). Metal binding sites on proteins are generally ion-specific 

33 and are influenced by multiple factors, including binding geometries, ligand preferences, 

34 ionic radius, and metal coordination numbers (Kirberger and Yang, 2008; Garza et al., 

35 2006). The coordination chemistry that normally regulates metal-protein binding makes 
February 2012 5-10 Draft – Do Not Cite or Quote

1 many proteins particularly susceptible to perturbation from Pb, as it is able to function 

2 with flexible coordination numbers and can bind multiple ligands (Kirberger and Yang, 

3 2008; Garza et al., 2006). However, due to differences in its physical properties, Pb 

4 induces abnormal conformational changes when it binds to proteins (Kirberger and Yang, 

5 2008; Bitto et al., 2006; Garza et al., 2006; Magyar et al., 2005), and these structural 

changes elicit altered protein function. It is known that [Ca2+ 6 ]i is an important second 

7 messenger in cell signaling pathways, and operates by binding directly to and activating 

8 proteins such as calmodulin and PKC (Goldstein, 1993). Alterations in the functions of 

9 both of these proteins due to direct interaction with Pb have been well documented in the 

10 literature.

11 PKC is a family of serine/threonine protein kinases critical for cell signaling and 

12 important for cellular processes, including growth and differentiation (Goldstein, 1993).

PKC contains a C2 Ca2+ 13 -binding domain and requires the cation, as well as 

14 diacylglycerol and phospholipids, for proper cellular activity (Garza et al., 2006).

Markovac and Goldstein (1988b) observed that, in the absence of Ca2+ 15 , exposure to 

16 picomolar concentrations of Pb for 5 minutes directly activated PKC purified from rat 

brains. The activation of PKC by Pb was more potent than was Ca2+ 17 -dependent activation 

18 by five orders of magnitude. Long et al. (1994) affirmed these findings, reporting that Pb 

had a Kact 4,800 times smaller than that of Ca2+ (5.5x10-5 19 μM versus 25 μM, following a 

3 minute exposure). However, Ca2+ 20 had a higher maximal activation of PKC than did Pb. 

This possibly indicates the presence of multiple Ca2+ 21 -binding sites on the protein, and 

that Pb may bind the first site more efficiently than does Ca2+ 22 , but not subsequent sites. 

23 Tomsig and Suszkiw (1995) further demonstrated the ability of Pb to activate PKC at 

24 picomolar concentrations in adrenal chromaffin cells incubated with Pb for 10 minutes 

25 but also reported that activation of PKC by Pb was only partial (approximately 40% of 

the maximum activity induced by Ca2+ 26 ) and tended to decrease at concentrations greater 

27 than one nanomolar. 

28 Contrary to the above findings, Markovac and Goldstein (1988a) observed that Pb and 

Ca2+ 29 activated PKC at equivalent concentrations and efficacies when broken cell 

30 preparations of rat brain microvessels were incubated with either cation for 45 minutes. 

31 However, when PKC activation was investigated in whole vessel preparations, no 

32 activation was observed, but PKC did become redistributed from the cytosolic to the 

33 particulate fraction. This suggests that Pb redistributes PKC at micromolar 

34 concentrations, but does not activate the protein in brain microvessels. In human 

35 erythrocytes exposed to Pb-acetate for 60 minutes, the amount of PKC found in 

36 erythrocyte membranes and total PKC activity was increased at concentrations greater 

than 0.1 μM (Belloni-Olivi et al., 1996). The observation that neither Ca2+ 37 nor 

38 diacylglycerol was increased due to exposure indicates that Pb-induced activation of PKC 
February 2012 5-11 Draft – Do Not Cite or Quote

1 is due to direct interaction with the protein. Pb-induced alterations in PKC have also been 

2 observed in other tissues, including increased activity in rabbit mesenteric arteries at 

3 picomolar concentrations of Pb (Watts et al., 1995; Chai and Webb, 1988) and human 

4 erythrocytes from Pb-exposed workers (range of blood Pb levels: 5.4 to 69.3 μg/dL)

5 (Hwang et al., 2002), and decreased activity in mouse macrophages and the rat brain 

6 cortex at micromolar concentrations (Murakami et al., 1993; Lison et al., 1990).

Calmodulin is another important protein essential for proper Ca2+ 7 -dependent cell 

signaling. Calmodulin contains an “EF-hand” Ca2+ 8 binding domain, and is dependent on 

9 the cation for proper activity (Garza et al., 2006). Calmodulin regulates events as diverse 

10 as cellular structural integrity, gene expression, and maintenance of membrane potential

11 (Vetter and Leclerc, 2003; Saimi and Kung, 2002). Habermann et al. (1983) observed 

12 that exposure to Pb altered numerous cellular functions of calmodulin, including 

13 activation of calmodulin-dependent phosphodiesterase activity after 10 minutes 

14 incubation (minimal activation at 0.1 μM, EC50 = 0.5-1.0 μM), stimulation of brain 

15 membrane phosphorylation at Pb concentrations greater than 0.4 μM after 1 minute

16 incubation, and increased binding of calmodulin to brain membranes at Pb concentrations 

17 greater than 1 μM after 10 minutes incubation. Habermann et al. (1983) reported that the 

affinity of Pb for Ca2+-binding sites on calmodulin was approximate to that of Ca2+ 18 itself 

19 (Kd ~ 20 μM), whereas Richardt et al. (1986) observed that Pb was slightly more potent 

than was Ca2+ 20 at binding calmodulin (IC50 = 11 and 26 μM, respectively). Both studies 

21 indicated that Pb was much more effective at binding calmodulin than was any other 

22 metal cation investigated (e.g., mercury, cadmium, iron). Kern et al. (2000) observed that 

23 Pb was more potent in binding to, and affecting conformational changes in, calmodulin 

compared to Ca2+ (EC50 values of 4-5.5x10-4μM (threshold = 1x10-4 24 μM) and 0.45-

0.5 μM (threshold = 0.1 μM), respectively). Pb, in the absence of Ca2+ 25 , was also observed 

26 to activate calmodulin-dependent cyclic nucleotide phosphodiesterase activity at much 

lower concentrations compared to Ca2+ (EC50 value 4.3x10-4μM [threshold = 3x10-4 27 μM 

versus EC50 1.2x10-3 28 μM (threshold = 0.2 μM; 50 minute incubation]). When incubated 

with physiological concentrations of Ca2+ 29 , Pb induced phosphodiesterase activity at 

concentrations as low as 5x10-5 30 μM. Pb activated calcineurin, a phosphatase with 

31 widespread distribution in the brain and immune system, at threshold concentrations as 

low as 2x10-5 μM in the presence of Ca2+ 32 (incubation time = 30 minutes), but inhibited its 

activity at concentrations greater than 2x10-4 33 μM (Kern and Audesirk, 2000). Thus, 

34 picomolar concentrations of intracellular Pb appear to amplify the activity of calmodulin 

and thus can be expected to alter intracellular Ca2+ 35 signaling in exposed cells (Kern et al., 

36 2000). Mas-Oliva (1989) observed that low-exposure (<1 μM, 20 minute incubation) 

stimulatory effects of Pb exposure on the activity of Ca2+-Mg2+ 37 -ATPase was due to Pb 

38 binding to calmodulin and subsequent activation of the ion pore. Ferguson et al. (2000)
February 2012 5-12 Draft – Do Not Cite or Quote

1 observed that exposure of rat hippocampal neurons to Pb for 1 to 48 hours resulted in 

increased activation of a calmodulin-dependent Ca2+ 2 extrusion mechanism.

Pb has also been observed to alter the activity of other proteins that rely on Ca2+ 3 binding 

4 for normal cellular function. Osteocalcin is a matrix protein important in bone resorption, 

osteoclast differentiation, and bone growth and has three Ca2+ 5 -binding sites (Dowd et al., 

2001). Incubation of osteocalcin in solution with Ca2+ 6 and Pb resulted in the competitive 

displacement of Ca2+ 7 by Pb (Dowd et al., 1994). Pb was found to bind to osteocalcin 

more than 1000-times more tightly than was Ca2+ (Kd = 1.6x10-2 8 μM versus 7 μM, 

9 respectively), and analysis with nuclear magnetic resonance (NMR) indicated that Pb 

10 induced similar, though slightly different, secondary structures in osteocalcin, compared 

to Ca2+ 11 . The authors hypothesized that the observed difference in Pb-bound osteocalcin 

12 structure may explain previous findings in the literature that Pb exposure reduced 

13 osteocalcin adsorption to hydroxyapatite (Dowd et al., 1994). Further research by this 

14 group also found that Pb bound osteocalcin approximately 10,000-times more tightly than 

did Ca2+ (Kd = 0.085 μM versus 1.25 × 103 15 μM, respectively) (Dowd et al., 2001).

16 However, the authors reported that Pb exposure actually caused increased hydroxyapatite 

adsorption at concentrations 2-3 orders of magnitude lower than that seen with Ca2+ 17 . 

Additionally, Pb can displace Ca2+ in numerous other Ca2+ 18 -binding proteins important in 

muscle contractions, renal Ca2+ 19 transport and neurotransmission, including troponin C, 

20 parvalbumin, CaBP I and II, phospholipase A2, and synaptotagmin I, at concentrations as 

21 low as the nanomolar range (Bouton et al., 2001; Osterode and Ulberth, 2000; Richardt et 

22 al., 1986).

Pb can displace metal cations other than Ca2+ 23 that are requisite for protein function. One 

24 of the most researched targets for molecular toxicity of Pb is the second enzyme in the 

25 heme synthetic pathway, aminolevulinic acid dehydratase (ALAD). ALAD contains four 

26 zinc-binding sites and all four need to be occupied to confer full enzymatic activity

27 (Simons, 1995). ALAD has been identified as the major protein binding target for Pb in 

28 human erythrocytes (Bergdahl et al., 1997a), and exposure to Pb results in inhibition of 

29 the enzyme in the erythrocytes of Pb-exposed workers and adolescents (blood Pb levels 

30 >10 μg/dL) (Ahamed et al., 2006; Ademuyiwa et al., 2005b), in human erythrocytes 

exposed to Pb for 60 minutes (Ki = 7x104 31 μM) (Simons, 1995), and in rats exposed to 

32 25 mg/kg Pb once a week for 4 weeks (mean [SD] blood Pb level: 6.56 [0.98] μg/dL)

33 (Lee et al., 2005). Additional experiments indicated that lower concentrations of zinc 

34 result in greater inhibition of enzyme activity by Pb, suggesting a competitive inhibition 

35 between zinc and Pb at a single site (Simons, 1995).

36 Zinc-binding domains are also found in transcription factors and proteins necessary for 

37 gene expression, including GATA proteins and transcription factors TFIIIA, Sp1, and 
February 2012 5-13 Draft – Do Not Cite or Quote

1 Erg-1 (Ghering et al., 2005; Huang et al., 2004; Reddy and Zawia, 2000; Hanas et al., 

2 1999; Zawia et al., 1998). Pb was found to form tight complexes with the cysteine 

residues in GATA proteins (Pb stability constant (βPb) = 6.4 × 109 M-1 3 for single zinc 

fingers and β2

Pb2 = 6.4 × 1019 M-2 4 ), and was able to displace bound zinc from the protein 

5 under physiologically relevant conditions (Ghering et al., 2005). Once Pb was bound to 

6 GATA proteins, they displayed decreased ability to bind to DNA (Pb concentrations ≥ 

1.25 μM) and activate transcription (Pb concentration = 1x106 7 μM). Pb at a minimum 

8 concentration of 10 μM also binds to the zinc domain of TFIIIA, inhibiting its ability to 

9 bind DNA at concentrations (Huang et al., 2004; Hanas et al., 1999). Huang et al. (2004)

10 also reported that exposure to Pb caused the dissociation of TFIIIA-DNA adducts and 

11 using NMR spectroscopy, found that altered TFIIIA activity was the result of a Pb12 induced abnormal protein conformation. 

13 Pb exposure modulated the DNA-binding profiles of the transcription factors Sp1 and 

14 Erg-1 in rat pups exposed to 0.2% Pb-acetate via lactation, resulting in a shift in DNA15 binding toward early development (i.e., the first week following birth) (Reddy and Zawia, 

16 2000; Zawia et al., 1998). The shifts in Sp1 DNA-binding profiles were shown to be 

17 associated with abnormal expression of genes related to myelin formation 

18 (Section 5.2.7.5). Further mechanistic research utilizing a synthetic peptide containing a 

19 zinc finger motif demonstrated that Pb can bind the histidine and cysteine residues of the 

20 zinc finger motif, thus displacing zinc and resulting in an increase in the DNA-binding 

21 efficiency of the synthetic peptide (Razmiafshari et al., 2001; Razmiafshari and Zawia, 

22 2000). However, in DNA-binding assays utilizing recombinant Sp1 (which has three zinc 

23 finger motifs, opposed to only one in the synthetic peptide), 37 μM Pb was the lowest 

24 concentration observed to abolish the DNA-binding capabilities of Sp1 (Razmiafshari 

25 and Zawia, 2000).

26 Pb has also been reported to competitively inhibit Mg binding and thus inhibit the 

27 activities of adenine and hypoxanthine/guanine phosphoribosyltransferase in erythrocyte 

28 lysates of rats exposed to 0.1% Pb in drinking water for 9 months (mean [SD] blood Pb 

29 level: 7.01 [1.64] μg/dL) and in human erythrocyte lysates exposed to 0.1 μM Pb for as 

30 little as 5 minutes (Baranowska-Bosiacka et al., 2009), and cGMP phosphodiesterase at 

31 picomolar concentrations in homogenized bovine retinas (Srivastava et al., 1995). Pb was 

32 also reported to inhibit pyrimidine 5’-nucleotidase through competitive inhibition of 

33 magnesium binding, resulting in conformational changes and improper amino acid 

34 positioning in the active site (Bitto et al., 2006).

35 In summary, Pb has the ability to displace metal cations from the active sites of multiple 

36 enzymes and proteins, and thus to alter the functions of those proteins in occupationally 

37 exposed humans with blood Pb levels of 5.4-69.3 μg/dL, in adult rodents with blood Pb 
February 2012 5-14 Draft – Do Not Cite or Quote

1 levels of 6.5 μg/dL (exposure 4 weeks), in suckling rats exposed to 0.2% Pb via lactation, 

2 and in cell-free and cellular in vitro experiments conducted at exposure concentrations 

3 ranging from micromoles to picomoles. These alterations in protein function have 

4 implications for numerous cellular and physiological processes, including cell signaling, 

5 growth and differentiation, gene expression, energy metabolism, and biosynthetic 

6 pathways. Table 5-1 provides a list of enzymes and proteins whose function may be 

7 perturbed by Pb exposure.

Table 5-1 Enzymes and proteins potentially affected by exposure to Pb and the 

metal cation cofactors necessary for their proper physiological 

activity

Metalloprotein/Enzyme Direction of Actiona Metal Cation; Reference

Enzymes

Aminolevulinic acid dehydratase ↓ Zn; Simons (1995)

Ferrochelatase ↓ Fe (2Fe-2S Cluster); Crooks et al. (2010)

Superoxide dismutase ↓↑ Mn, Cu, Zn, Fe; Antonyuk et al. (2009),

Borgstahl et al. (1992)

Catalase ↓↑ Fe (Heme); Putnam et al. (2000)

Glutathione peroxidase ↓↑ Se; Rotruck et al. (1973)

Guanylate cyclase ↓ Fe (Heme); Boerrigter and Burnett (2009)

cGMP phosphodiesterase ↓ Mg, Zn; Ke (2004)

NAD synthase ↓ Mg; Hara et al. (2003)

NAD(P)H oxidase ↑ Ca; Leseney et al. (1999)

Pyrimidine 5’-nucleotidase ↓ Mg, Ca; Bitto et al. (2006), Amici et al. (1997),

Paglia and Valentine (1975)

Erythrocyte 

phosphoribosyltransferase 

↓ Mg (Mn, Ca, Co, Ni, Zn); Deng et al. (2010),

Arnold and Kelley (1978)

Ion Channels/ 

Transport

ATPase ↓↑ Ca, Mg, Na-K; Technische Universitat 

Braunschweig (2011)

Mitochondrial transmembrane pore ↑ Ca; He et al. (2000)

Calcium-dependent potassium 

channel

↑ Ca; Silkin et al. (2001), Alvarez et al. (1986)

Signal Transduction

Protein kinase C ↓↑ Ca; Garza et al. (2006)

Calmodulin ↑ Ca; Garza et al. (2006)

Pb Binding Metallothionein ↑ Zn, Cu; Yu et al. (2009)

DNA Binding GATA transcriptional factors ↓ Zn; Hanas et al. (1999), Huang et al. (2004)

a

↑ indicates increased activity; ↓ indicates decreased activity; ↓↑ indicates activity can be alternatively increased or decreased.
February 2012 5-15 Draft – Do Not Cite or Quote

5.2.2.4 Mitochondrial Abnormality

1 Alterations in mitochondrial function, including disruptions in ion transport, 

2 ultrastructural changes, altered energy metabolism, and perturbed enzyme activities due 

3 to Pb intoxication are well documented in the scientific literature. Exposure of rats to Pb 

4 in feed (1% Pb for 4, 6, 8, 10, 12, or 20 weeks) or drinking water (300 ppm for 8 weeks, 

5 500 ppm for 7 months, or 1% for 9 months) resulted in gross ultrastructural changes in 

6 renal tubules and epididymal mitochondria characterized as a general swollen appearance 

7 with frequent rupture of the outer membrane, distorted cristae, loss of cristae, frequent 

8 inner compartment vacuolization, observation of small inclusion bodies, and fusion with 

9 adjacent mitochondria (Wang et al., 2010d; Marchlewicz et al., 2009; Navarro-Moreno et 

10 al., 2009; Goyer, 1968; Goyer et al., 1968).

11 Transmembrane mitochondrial ion transport mechanisms are perturbed by exposure to 

Pb. Pb inhibits the uptake of Ca2+ 12 into mitochondria (Parr and Harris, 1976), while 

simultaneously stimulating the efflux of Ca2+ 13 out of the organelle (Simons, 1993a), thus 

disrupting intracellular/mitochondrial Ca2+ 14 homeostasis. Pb exposure has also been 

15 shown to decrease the mitochondrial transmembrane potential in primary cerebellar 

16 granule neuronal cultures from rats exposed to 0.1% Pb in drinking water throughout 

17 gestation and lactation (Baranowska-Bosiacka et al., 2011a), astroglia incubated with 0.1 

18 or 1.0 μM Pb for 14 days (Legare et al., 1993), proximal tubule cells exposed to 0.25, 0.5, 

19 and 1.0 μM for 12 hours (Wang et al., 2009c), and retinal rod photoreceptor cells 

20 incubated with 0.01 to 10 μM for 15 minutes (He et al., 2000). Further research indicated 

21 that Pb-induced mitochondrial swelling and decreased membrane potential is the result of 

22 the opening of a mitochondrial transmembrane pore (MTP), possibly by directly binding 

to the metal (Ca2+ 23 )-binding site on the matrix side of the pore (Bragadin et al., 2007; He 

24 et al., 2000). Opening of the MTP is the first step of the mitochondrial-regulated 

25 apoptotic cascade pathway in many cells (Rana, 2008; Lidsky and Schneider, 2003). He 

26 et al. (2000) additionally observed cytochrome c release from mitochondria, and caspase27 9 and -3 activation following exposure of rod cells to Pb. Induction of mitochondrially28 regulated apoptosis via stimulation of the caspase cascade following exposure to Pb has 

29 also been observed in rat oval cells (Agarwal et al., 2009).

Altered Energy Metabolism

30 Pb has been reported to alter normal cellular bioenergetics. In mitochondria isolated from 

31 the kidneys of rats exposed to 1% Pb in feed for 6 weeks, the rate of oxygen uptake 

32 during ADP-activated (state 3) respiration was lower compared to controls (Goyer et al., 

33 1968). The rate of ATP formation in exposed mitochondria was observed to be 

34 approximately 50% that of control mitochondria. A decrease in state 3 respiration and 
February 2012 5-16 Draft – Do Not Cite or Quote

1 respiratory control ratios (state 3/state 4 [succinate or pyruvate/malate-activated]) was 

2 also observed in kidney mitochondria from rats exposed continuously from conception to 

3 six or nine months of age (i.e., gestationally, lactationally, and via drinking water after 

4 weaning) to 50 or 250 ppm Pb (Fowler et al., 1980). Pb-induced decreases in ATP and 

5 adenylate energy charge (AEC) were observed concurrently with increases in ADP, 

6 AMP, and adenosine in rats exposed to 1% Pb in drinking water for 9 months

7 (Marchlewicz et al., 2009). Similarly, ATP and AEC were decreased, and AMP 

8 increased, in primary cerebellar granule neuronal cultures from rats exposed to 0.1% Pb 

9 in drinking water throughout gestation and lactation (Baranowska-Bosiacka et al., 

10 2011a). One μM Pb (48 hours) was the lowest concentration observed to decrease 

11 cellular ATP levels in differentiated PC-12 cells, and these changes were correlated with 

12 a Pb-induced decrease in the expression of the voltage-dependent anion channel, which 

13 maintains cellular ATP levels in neurons (Prins et al., 2010). Dowd et al. (1990) reported 

14 that oxidative phosphorylation was decreased up to 74% after exposure of osteoblasts to 

15 10 μM Pb. Parr and Harris (1976) reported that Pb inhibited coupled and uncoupled 

16 respiratory oxygen use in mitochondria, and that Pb prevented pyruvate, but not malate, 

17 uptake. Mitochondrial levels of ATP were diminished after exposure, and the authors 

18 compared the effects of Pb on the energy supply to the actions of classic respiratory 

19 inhibitors, low temperature, and chemical uncouplers. Bragadin et al. (1998) supported 

20 this view by demonstrating that alkylated Pb compounds acted as a chemical uncoupler 

21 of respiration by abolishing the proton gradient necessary for oxidative phosphorylation. 

22 Further, the enzymatic activities of complex I and IV of the respiratory chain have been 

23 shown to be decreased in the peroneous longus muscle of rats exposed to 250 ppm Pb or 

24 5 ppm thallium in drinking water for 90 days (Méndez-Armenta et al., 2011). Contrary to 

25 the above findings, Rafalowska et al. (1996) reported that, although ATP levels did 

26 decrease, chronic exposure to Pb did not inhibit oxidative phosphorylation in the 

27 synaptosomes of rats exposed to 200 ppm Pb in water for 3 months. Similar effects with 

28 regard to the activity of the mitochondrial oxidative chain were observed in rats injected 

29 with 15 mg/kg Pb i.p. daily for seven days, as reported by Struzynksa et al. (1997a),

30 although ATP levels were reported to increase after exposure to Pb. 

31 Pb has also been shown to decrease glycolysis in osteoblasts exposed to 10 μM Pb and in 

32 human erythrocytes exposed to 30 μg/dL Pb (Grabowska and Guminska, 1996; Dowd et 

33 al., 1990). Contrary to these findings, Antonowicz et al. (1990) observed higher levels of 

34 glycolytic enzymes in erythrocytes obtained from Pb workers directly exposed to Pb, 

35 compared to controls exposed to lower concentrations of Pb (blood Pb levels: 82.1 versus 

36 39.9 μg/dL), and suggested that Pb activated anaerobic glycolysis. In vitro exposure of 

37 human umbilical cord erythrocytes to 100-200 μg/dL Pb for 20 hours was observed to 

38 lower the cellular pools of adenine and guanine nucleotide pools, including NAD and 

39 NADPH (Baranowska-Bosiacka and Hlynczak, 2003). These decreases in nucleotide 
February 2012 5-17 Draft – Do Not Cite or Quote

1 pools were accompanied by an increase in purine degradation products (adenosine, etc.). 

2 Similar decreases in cellular nucleotide pools were observed when rats were exposed to 

3 1% Pb in drinking water for four weeks (Baranowska-Bosiacka and Hlynczak, 2004). In 

4 erythrocytes, nucleotides are synthesized via salvage pathways such as the adenine 

5 pathway, which requires adenine phosphoribosyltransferase. The activity of this enzyme 

6 is inhibited by exposure to Pb in human and rat erythrocytes (see above for concentration 

7 and duration) (Baranowska-Bosiacka et al., 2009).

8 Disruptions in erythrocyte energy metabolism have been observed in workers 

9 occupationally exposed to Pb. Nikolova and Kavaldzhieva (1991) reported higher ratios 

10 of ATP/ADP in Pb-exposed workers with an average duration of exposure of 8.4 years 

11 (blood Pb not reported). Morita et al. (1997) evaluated the effect of Pb on NAD 

12 synthetase in the erythrocytes of Pb-exposed workers (mean [SD] blood Pb level: 34.6 

13 [20.7] μg/dL) and observed an apparent concentration-dependent decrease in NAD 

14 synthetase activity with increased blood Pb level. The blood Pb level associated with 

15 50% inhibition of NAD synthetase, which requires a magnesium cation for activity (Hara 

16 et al., 2003), was 43 μg/dL.

Altered Heme Synthesis

17 Exposure to Pb is known to inhibit two key steps in the synthesis of heme: 

18 porphobilinogen synthase (i.e., δ-aminolevulinic acid dehydratase), a cytoplasmic 

19 enzyme requiring zinc for enzymatic activity that condenses two molecules of 

20 aminolevulinic acid into porphobilinogen, and ferrochelatase, a mitochondrial iron-sulfur 

containing enzyme that incorporates Fe2+ 21 into protoporphyrin IX to create heme. Farant 

22 and Wigfield (1990, 1987) observed that Pb inhibits the activity of porphobilinogen 

23 synthase in rabbit and human erythrocytes, and that the effect on the enzyme was 

24 dependent on the affinity for thiol groups at its active site. Taketani et al. (1985)

25 examined the activity of Pb on ferrochelatase in rat liver mitochondria and observed that 

26 10 μM Pb (30 minute incubation) reduced NAD(P)H-dependent heme synthesis by half 

when ferric, but not ferrous, iron was used. Pb inhibits the insertion of Fe2+ 27 into the 

28 protoporphyrin ring and instead, Zn is inserted into the ring creating zinc protoporphyrin 

29 (ZPP). While not directly measuring the activity of ferrochelatase, numerous studies have 

30 shown that blood Pb levels are associated with increased erythrocyte ZPP levels in 

31 humans (average blood Pb levels ranging from 21.92 to 53.63 μg/dL) (Mohammad et al., 

32 2008; Counter et al., 2007; Patil et al., 2006b; Ademuyiwa et al., 2005b) and animals 

33 (blood Pb level: 24.7 μg/dL) (Rendón-Ramirez et al., 2007).
February 2012 5-18 Draft – Do Not Cite or Quote

5.2.3 Protein Binding

1 Evidence indicates that Pb binds to proteins within cells through interactions with side 

2 group moieties (e.g., thiol residues) and can potentially disrupt cellular function 

3 (Sections 5.2.2.3 and 5.2.2.4). However, some proteins are also able to bind Pb and 

4 protect against its negative effects through sequestration. The ability of Pb to bind 

5 proteins was first reported by Blackman (1936): Pb intoxication in children autopsy cases 

6 was observed to induce the formation of intranuclear inclusion bodies in the liver and 

7 kidney. Since that time, further research has been conducted into characterizing the 

8 composition of intranuclear inclusion bodies and identifying specific Pb-binding proteins.

5.2.3.1 Intranuclear and Cytoplasmic Inclusion Bodies

9 Goyer (1968) and Goyer et al. (1968) observed the formation of intranuclear inclusion 

10 bodies in the renal tubules of rats fed 1% Pb in food for up to 20 weeks. The observation 

11 of inclusion bodies was accompanied by altered mitochondrial structure and reduced 

12 rates of oxidative phosphorylation. Pb has further been observed to form cytoplasmic 

13 inclusion bodies preceding the formation of the intranuclear bodies, and to be 

14 concentrated within the subsequently induced intranuclear inclusion bodies following i.p. 

15 injection, drinking water, and dietary exposures (Navarro-Moreno et al., 2009; Oskarsson 

16 and Fowler, 1985; Fowler et al., 1980; McLachlin et al., 1980; Choie and Richter, 1972; 

17 Goyer et al., 1970a; Goyer et al., 1970b). Inclusion bodies have also been observed in the 

18 mitochondria of kidneys and the perinuclear space in the neurons of rats exposed to 

19 500 ppm Pb-acetate in drinking water for 60 days or 7 months (Navarro-Moreno et al., 

20 2009; Deveci, 2006). Intranuclear and cytoplasmic inclusions have also been found in 

21 organs other than the kidney, including liver, lung, and glial cells (Singh et al., 1999; 

22 Goyer and Rhyne, 1973). Pb found within nuclei has also been shown to bind to the 

23 nuclear membrane and histone fractions (Sabbioni and Marafante, 1976).

24 Upon denaturing intranuclear inclusion bodies with strong denaturing agents, Moore et 

25 al. (1973) observed that proteins included in the bodies were rich in aspartic and glutamic 

26 acid, glycine, and cysteine. Further work by Moore and Goyer (1974) characterized the 

27 protein as a 27.5 kDa protein that migrates as a single band on acrylamide gel 

28 electrophoresis. In contrast with the findings of Moore and Goyer, Shelton and Egle

29 (1982) identified a 32 kDa protein with an isoelectric point of 6.3 from the kidneys of rats 

30 exposed to 1% Pb-acetate in feed or 0.75% in drinking water. This protein, dubbed 

31 p32/6.3, was not found in control rats, indicating that the protein was induced by Pb 

32 exposure. This finding was in agreement with studies that indicated formation of 

33 intranuclear inclusion bodies required protein synthesis (McLachlin et al., 1980; Choie et 
February 2012 5-19 Draft – Do Not Cite or Quote

1 al., 1975). In addition to its presence in kidneys of Pb-exposed animals, p32/6.3 has been 

2 observed to be present and highly conserved in the brains of rats, mice, dogs, chickens, 

3 and humans (Egle and Shelton, 1986). Exposure of neuroblastoma cells to 50 or 100 μM 

4 Pb glutamate for 1 or 3 days increased the abundance of p32/6.3 (Klann and Shelton, 

5 1989). Shelton et al. (1990) determined that p32/6.3 was enriched in the basal ganglia, 

6 diencephalon, hippocampus, cerebellum, brainstem, spinal cord, and cerebral cortex, and 

7 that it contained a high percentage of glycine, aspartic, and glutamic acid residues.

8 Selvin-Testa et al. (1991) and Harry et al. (1996) reported that pre- and post-natal 

9 exposure of rats to 0.2-1.0% Pb in drinking water increased the levels of another brain 

10 protein, glial fibrillary acidic protein, in developing astrocytes and that this increase may 

11 be indicative of a demand for astrocytes to sequester Pb.

5.2.3.2 Cytosolic Lead Binding Proteins

12 Numerous studies have also identified cytosolic Pb-binding proteins. Two binding 

13 proteins, with molecular weights of 11.5 and 63 kDa, were identified by (Oskarsson et 

14 al., 1982) in the kidney postmitochondrial cytosolic fraction after injection with 50 mg

15 Pb. The two proteins were also found in the brain, but not the liver or lung. Mistry et al.

16 (1985) identified three proteins (MW = 11.5, 63, and >200 kDa) in rat kidney cytosol, 

17 two of which, the 11.5 and 63 kDa proteins, were able to translocate into the nucleus. The 

18 11.5 kDa kidney protein was also able to reverse Pb binding to ALAD through chelation 

19 of Pb and donation of a zinc cation to ALAD (Goering and Fowler, 1985, 1984).

Cadmium and zinc, but not Ca2+ 20 or Fe, prevented the binding of Pb to the 63 and 11.5 

21 kDa cytosolic proteins, which agrees with previous observations that cadmium is able to 

22 reduce total kidney Pb and prevent the formation of intranuclear inclusion bodies (Mistry 

23 et al., 1986; Mahaffey et al., 1981; Mahaffey and Fowler, 1977). Additional cytosolic Pb24 binding proteins have been identified in the kidneys of Pb-exposed rats and humans, 

25 including the cleavage product of α2-microglobulin, acyl-CoA binding protein (MW = 9 

26 kDa), and thymosin β4 (MW = 5 kDa) (Smith et al., 1998; Fowler and DuVal, 1991).

27 Cytosolic Pb-binding proteins distinct from kidney proteins have also been identified in 

28 the brain of exposed rats and human brain homogenates exposed in vitro (Quintanilla29 Vega et al., 1995; DuVal and Fowler, 1989; Goering et al., 1986). One protein (MW = 

30 12 kDa) was shown to alleviate hepatic ALAD inhibition due to Pb exposure through 

31 competitive binding with Pb and donation of zinc to ALAD. Cytosolic Pb-binding

32 proteins have been shown to be high in glutamic acid, aspartic acid, and cysteine residues

33 (Fowler et al., 1993; DuVal and Fowler, 1989). Some evidence exists that cytosolic Pb34 binding proteins directly target Pb and compartmentalize intracellular Pb as a protective 

35 measure against toxicity (Qian et al., 2005; Qian et al., 2000).
February 2012 5-20 Draft – Do Not Cite or Quote

5.2.3.3 Erythrocytic Lead Binding Proteins

1 The majority (94%) of Pb in whole blood is found in erythrocytes (Ong and Lee, 1980a).

2 Originally, the major Pb-binding protein in erythrocytes was identified as hemoglobin

3 (Cohen et al., 2000; Lolin and O'Gorman, 1988; Ong and Lee, 1980a, b; Raghavan and 

4 Gonick, 1977). However, Bergdahl et al. (1997b) observed the principal Pb-binding 

5 protein to be 240 kDa and identified it as ALAD. Two smaller Pb-binding proteins were 

6 observed, but not identified (MW = 45 and <10 kDa). ALAD levels are inducible by Pb 

7 exposure; the total concentration of the enzyme, but not the activity, increases after 

8 exposure in both exposed humans (blood Pb = 30-75 μg/dL) and rats (Pb exposure = 25 

9 mM in drinking water) (Boudene et al., 1984; Fujita et al., 1982; Fujita et al., 1981).

10 ALAD is a polymorphic gene with three isoforms: ALAD 1-1, ALAD 1-2, or ALAD 2-2. 

11 Carriers of the ALAD-2 allele have been shown to have higher blood Pb levels than 

12 carriers of the homozygous ALAD-1 allele (Scinicariello et al., 2007; Zhao et al., 2007; 

13 Kim et al., 2004; Pérez-Bravo et al., 2004; Smith et al., 1995a; Wetmur, 1994; Wetmur et 

14 al., 1991a; Astrin et al., 1987). Some newer studies, however, either observed lower 

15 blood Pb levels in carriers of the ALAD-2 allele or no difference in Pb levels among the 

16 different allele carriers (Scinicariello et al., 2010; Krieg et al., 2009; Chen et al., 2008c; 

17 Chia et al., 2007; Chia et al., 2006; Wananukul et al., 2006).

18 The ALAD-2 protein binds Pb more tightly than the ALAD-1form: in workers carrying 

19 the ALAD-2 gene, 84% of blood Pb was bound to ALAD versus 81% in carriers of the 

20 ALAD-1 gene (p = 0.03) (Bergdahl et al., 1997a). This higher affinity for Pb in ALAD-2 

21 carriers may sequester Pb and prevent its bioavailability for reaction with other enzymes 

22 or cellular components. This is supported by the observation that carriers of the ALAD-2 

23 gene have higher levels of hemoglobin (Scinicariello et al., 2007), decreased plasma 

24 levulinic acid (Schwartz et al., 1997b), decreased levels of zinc protoporphyrin

25 (Scinicariello et al., 2007; Kim et al., 2004), lower cortical bone Pb (Smith et al., 1995b),

26 and lower amounts of DMSA-chelatable Pb (Scinicariello et al., 2007; Schwartz et al., 

27 2000a; Schwartz et al., 1997a). However, the findings that ALAD-2 polymorphisms 

28 reduced the bioavailability of Pb are somewhat equivocal. Wu et al. (2003a) observed 

29 that ALAD-2 carriers had lower blood Pb level (5.8 ± 4.2 μg/dL) than carriers of the 

30 ALAD-1 gene (blood Pb level = 6.2 ± 4.1 μg/dL), and that ALAD-2 carriers 

31 demonstrated decreased renal function at lower patellar Pb concentrations than those 

32 observed to decrease renal function in ALAD-1 carriers. This potentially indicates that 

33 ALAD-2 carriers have enhanced Pb bioavailability. Weaver et al. (2003b) observed that 

34 ALAD-2 polymorphisms were associated with higher DMSA-chelatable Pb 

35 concentrations, when normalized to creatinine levels. Further, Montenegro et al. (2006)

36 observed among individuals with ALAD 1-1 or ALAD 1-2/2-2 genotypes a significant 
February 2012 5-21 Draft – Do Not Cite or Quote

1 increase in the amount of Pb found in the plasma (0.44 μg/L versus 0.89 μg/L, 

2 respectively) and in the % plasma/blood ratio (0.48% versus 1.45%, respectively). This 

3 potentially suggests that individuals with the ALAD 1-2/2-2 genotype are at increased 

4 risk of Pb-induced health effects due to lower amounts of Pb sequestration by erythrocyte 

5 ALAD, although this study did not specifically investigate the clinical implications of 

6 ALAD polymorphisms.

7 ALAD has the estimated capacity to bind Pb at 85 μg/dL in erythrocytes and 40 μg/dL in 

8 whole blood (Bergdahl et al., 1998). The 45 kiloDalton (kDa) and <10 kDa Pb-binding 

9 proteins bound approximately 12-26% and <1% of the blood Pb, respectively. At blood 

10 Pb concentrations greater than 40 μg/dL, greater binding to these components likely 

11 would be observed. Bergdahl et al. (1998) tentatively identified the 45 kDa protein as 

12 pyrimidine-5’-nucleotidase and the <10 kDa protein as acyl-CoA binding protein. Smith 

13 et al. (1998) previously identified acyl-CoA binding protein as a Pb-binding protein 

14 found in the kidney.

15 Studies also observed the presence of an inducible, low-molecular weight (approximately 

16 10 kDa) Pb-binding protein in workers occupationally exposed to Pb (Gonick et al., 

17 1985; Raghavan et al., 1981, 1980; Raghavan and Gonick, 1977). The presence of this 

18 low molecular weight protein seemed to have a protective effect as workers that exhibited 

19 toxicity at low blood Pb concentrations were observed to have lowered expression of this 

20 protein or low levels of Pb bound to it (Raghavan et al., 1981, 1980). The presence of low 

21 molecular weight Pb-binding proteins in exposed workers was confirmed by Lolin and 

22 O’Gorman (1988) and Church et al. (1993a, b). Further Lolin and O’Gorman (1988)

23 reported that the observed protein was only present when blood Pb levels were greater 

24 than 39 μg/dL, in agreement with the Pb-binding capacity of ALAD, identified by 

25 Bergdahl et al. (1998). Xie et al. (1998) confirmed this, observing the presence of a 

26 second low molecular weight protein with greater affinity than ALAD only at higher 

27 blood Pb levels. Church et al. (1993a, b) observed the presence of a 6-7 kDa protein in 

28 the blood of 2 Pb workers (blood Pb >160 μg/dL); approximately 67% of Pb was bound 

29 to the protein in the blood of the asymptomatic worker, whereas only 22% of the Pb was 

30 bound to it in the symptomatic worker. The reported protein was rich in cysteine residues 

31 and tentatively identified as metallothionein. 

5.2.3.4 Metallothionein

32 Metallothionein is a low-molecular weight metal-binding protein, most often zinc or

33 copper, which is rich in cysteine residues and plays an important role in the protection 

34 against heavy metal toxicity, trace element homeostasis, and scavenging free radicals (Yu 
February 2012 5-22 Draft – Do Not Cite or Quote

1 et al., 2009). Exposure to Pb-acetate induces the production of Pb- and Zn2 metallothionein in mice exposed via i.p. or intravenous (i.v.) injection at 30 mg/kg

3 (Maitani et al., 1986), in mice exposed via i.p. injection at 300 μmol/kg (Yu et al., 2009),

4 or in rats exposed via i.p. injection at 24 μmol/100g (Ikebuchi et al., 1986). The induced 

5 Pb-metallothionein consisted of 28% half-cysteine and reacted with an antibody for Zn6 metallothionein II (Ikebuchi et al., 1986). In contrast, exposure of rats to Pb via drinking 

7 water (200 or 300 ppm) failed to induce metallothionein in the kidneys or intestines

8 (Wang et al., 2009b; Jamieson et al., 2007). Goering and Fowler (1987a, b) observed that 

9 pretreatment of rats with zinc before injection with Pb resulted in Pb and zinc co-eluting 

10 with zinc-thionein, and that zinc-thionein I and II were able to bind Pb in vitro (Goering 

11 and Fowler, 1987a, b). Further, Goering and Fowler (1987a) found that kidney and liver 

12 zinc-thionein decreased binding of Pb to liver ALAD and was able to donate zinc to 

13 ALAD, thus attenuating the inhibition of ALAD due to Pb exposure. These findings are 

14 in agreement with Goering et al. (1986) and DuVal and Fowler (1989) who demonstrated 

15 that rat brain Pb-binding proteins attenuated Pb-induced inhibition of ALAD. 

16 Metallothionein has been reported to be important in the amelioration of Pb-induced 

17 toxicity effects. Liu et al. (1991) reported that zinc-metallothionein reduced Pb-induced 

18 membrane leakage and loss of potassium in cultured hepatocytes incubated with 600-

19 3,600 μM Pb. Metallothionein-null mice exposed to 1,000, 2,000, or 4,000 ppm Pb for 20 

20 weeks suffered renal toxicity described as nephromegaly and decreased renal function 

21 compared to Pb-treated wild-type mice (Qu et al., 2002). Interestingly, metallothionein22 null mice were unable to form intranuclear inclusion bodies and accumulated less renal 

23 Pb than did the wild-type mice (Qu et al., 2002). Metallothionein levels were induced by 

24 Pb exposure in non-null mice. Exposure to Pb (1,000, 2,000, or 4,000 ppm), both for 

25 104 weeks as adults and from GD8 to early adulthood, resulted in increased preneoplastic 

26 lesions and carcinogenicity in the testes, bladder, and kidneys of metallothionein-null rats 

27 compared to wild type mice (Tokar et al., 2010; Waalkes et al., 2004). Inclusion bodies 

28 were not observed in null mice. The authors concluded that metallothionein is important 

29 in the formation of inclusion bodies and mitigation of Pb-induced toxic effects, and that 

30 those with polymorphisms in metallothionein coding genes may be at greater 

31 susceptibility to Pb. In support of this theory, Chen et al. (2010a) observed that Pb32 exposed workers with a mutant metallothionein allele had higher blood Pb levels than did 

33 carriers of the normal allele (24.17 and 21.27 versus 17.03 μg/dL), and were more 

34 susceptible to the effects of Pb on systolic BP and serum renal function parameters.
February 2012 5-23 Draft – Do Not Cite or Quote

5.2.4 Oxidative Stress

1 Oxidative stress occurs when free radicals or reactive oxygen species (ROS) exceed the 

2 capacity of antioxidant defense mechanisms. This oxidative imbalance results in 

uncontained ROS, such as superoxide (O2

-

), hydroxyl radical (OH• 3 ), and hydrogen 

4 peroxide (H2O2), which can attack and denature functional/structural molecules and, 

5 thereby, promote tissue damage, cytotoxicity, and dysfunction. Pb has been shown to 

6 cause oxidative damage to the heart, liver, kidney, reproductive organs, brain, and 

7 erythrocytes, which may be responsible for a number of Pb-induced health effects

8 (Salawu et al., 2009; Shan et al., 2009; Vaziri, 2008b; Gonick et al., 1997; Sandhir and 

9 Gill, 1995; Khalil-Manesh et al., 1994; Khalil-Manesh et al., 1992b). The origin of ROS 

10 produced after Pb exposure is likely a multipathway process, resulting from oxidation of 

11 δ-aminolevulinic acid (ALA), membrane and lipid oxidation, nicotinamide adenine 

12 dinucleotide phosphate (NAD(P)H) oxidase activation, and antioxidant enzyme 

13 depletion, as discussed below. Some of these processes result from the disruption of 

14 functional metal ions within oxidative stress proteins, such as superoxide dismutase 

15 (SOD), catalase (CAT), and glutathione peroxidase (GPx). Interestingly, Pb exposure in 

16 many species of plants, invertebrates, and vertebrates discussed in the Ecological Effects 

17 of Pb results in upregulation of antioxidant enzymes and increased lipid peroxidation 

18 (Chapter 7). Oxidative stress is a common mode of action for a number of other metals 

19 (e.g., Cd, Mn, As, Co, Cr) that are often found with Pb and by which possible interactions 

20 with Pb have been suggested to occur (Jomova and Valko, 2011; Jomova et al., 2011; 

21 Matović et al., 2011; HaMai and Bondy, 2004). Not all of these co-occurring metals 

22 directly produce ROS or redox cycle, but instead may suppress the free radical 

23 scavenging ability of the organism thus leading to oxidative stress.

5.2.4.1 δ-ALA Oxidation

24 The majority of Pb present in the blood accumulates in erythrocytes where it enters 

through passive carrier-mediated mechanisms including a vanadate-sensitive Ca2+ 25 pump. 

26 Once Pb enters erythrocytes, it is predominantly found in the protein-bound form, with 

27 hemoglobin and δ-ALAD both identified as targets (Bergdahl et al., 1997a). Through its 

28 sulfhydryl and metal ion disrupting properties, Pb incorporates with and inhibits a 

29 number of enzymes in the heme biosynthetic process, including δ-ALA synthetase, δ30 ALAD, and ferrochelatase. Pb has been shown to be able to disrupt the zinc ions requisite 

31 for the activity of δ-ALAD, the rate limiting step in heme synthesis, leading to enzyme 

32 inhibition at picomolar concentrations (Simons, 1995). Additionally, low blood Pb levels 

33 (mean: 7 μg/dL) have been found to inhibit the activity of δ-ALAD in humans, and the 

34 lowest blood Pb level observed to be associated with lower δ-ALAD activity in these 
February 2012 5-24 Draft – Do Not Cite or Quote

1 studies was 5 μg/dL (Ahamed et al., 2005; Sakai and Morita, 1996). A significant 

2 negative correlation (r = -0.6) was found between blood Pb levels in adolescents (range 

3 of blood Pb levels: 4-20 μg/dL) and blood δ-ALAD activity in adolescents (Ahamed et 

4 al., 2006). This inhibition of δ-ALAD results in the accumulation of δ-ALA in blood and 

5 urine, where δ-ALA undergoes tautomerization and autoxidation. Oxidized δ-ALA 

6 generates ROS through reduction of ferricytochrome c and electron transfer from oxyHb, 

7 metHb, and other ferric and ferrous complexes (Hermes-Lima et al., 1991; Monteiro et 

al., 1991). The autoxidation of δ-ALA produces O2

- 8 , OH•, H2O2, and an ALA radical

9 (Monteiro et al., 1989; Monteiro et al., 1986).

5.2.4.2 Membrane and Lipid Peroxidation

10 A large number of studies in humans and experimental animals have found that exposure 

11 to Pb can lead to membrane and lipid peroxidation. It is possible that ROS produced from 

12 δ-ALA oxidation, as described above, interacts with and disrupts membrane lipids

13 (Oteiza et al., 1995; Bechara et al., 1993). Additionally, Pb has the capacity to stimulate 

14 ferrous ion initiated membrane lipid peroxidation serving as a catalyst for these events

15 (Adonaylo and Oteiza, 1999; Quinlan et al., 1988). The extent of peroxidation of lipids 

16 varies based on the number of double bonds present in unsaturated fatty acids, since 

17 double bonds weaken the C-H bonds on the adjacent carbon, making H removal easier

18 (Yiin and Lin, 1995). After Pb exposure (4-12 μg/dL in vitro, 24 hours), the production 

19 of malondialdehyde (MDA), a marker of oxidative stress and lipid oxidation end product, 

increased relative to the number of double bonds of the fatty acid. In the absence of Fe2+ 20 , 

21 Pb has not been shown to promote lipid peroxidation; however, it may accelerate 

22 peroxidation by H2O2 (Quinlan et al., 1988). This could be due to altering membrane structure, 

23 restricting phospholipid movement, and facilitating the propagation of peroxidation. 

24 Pb induces changes in the fatty acid composition of a membrane, which could lead to 

25 oxidative damage. Exposure to Pb (>62.5 ppm in drinking water, 3 weeks) in chicks 

26 promoted an increase in arachidonic acid (AA, 20:4) as a percentage of total fatty acids, 

27 and decreased the relative proportion of shorter chain fatty acids (linoleic acid, 18:2)

28 (Lawton and Donaldson, 1991). It is possible that Pb depressed the desaturation of 

29 saturated fatty acids to the corresponding monoenoic fatty acids, while stimulating 

30 elongation and desaturation of linoleic acid to AA. Since fatty acid chain length and 

31 unsaturation are related to the oxidative potential, changes in fatty acid membrane 

32 composition may result in enhanced lipid peroxidation. In addition, changes in fatty 

33 acids, thus membrane composition, can result in altered membrane fluidity (Donaldson 

34 and Knowles, 1993). Changes in membrane fluidity will disturb the conformation of the 
February 2012 5-25 Draft – Do Not Cite or Quote

1 active sites of membrane associated enzymes, disrupt metabolic regulation, and alter 

2 membrane permeability and function.

3 A number of recent studies report increased measures of lipid peroxidation in various 

4 organs, tissues, and species. Occupational Pb exposure resulting in elevated blood Pb 

5 levels (means >8 μg/dL) in various countries provides evidence of lipid peroxidation, 

6 including increased plasma MDA levels (Ergurhan-Ilhan et al., 2008; Khan et al., 2008; 

7 Mohammad et al., 2008; Quintanar-Escorza et al., 2007; Patil et al., 2006a; Patil et al., 

8 2006b). One study found a correlation between the MDA levels and blood Pb levels even 

9 in the unexposed workers, although they had blood Pb levels higher than the mean blood 

10 Pb level of the current U.S. population (i.e., <12 μg/dL) (Quintanar-Escorza et al., 2007).

11 Other studies found evidence of increased lipid peroxidation among the general 

12 population, including children, with elevated blood Pb levels (means >10 μg/dL)

13 (Ahamed et al., 2008; Ahamed et al., 2006; Jin et al., 2006). In adolescents, Ahamed et 

14 al. (2006) found a blood MDA levels to be positively correlated (r = 0.7) with blood Pb 

15 levels ranging between 4-20 μg/dL. Similar results have been shown after Pb exposure in 

16 animal studies (Abdel Moneim et al., 2011b; Pandya et al., 2010; Dogru et al., 2008; Yu 

17 et al., 2008; Adegbesan and Adenuga, 2007; Lee et al., 2005). Enhanced lipid 

18 peroxidation has been found in Pb treated (50 μg, 1-4 hours) rat brain homogenates

19 (Rehman et al., 1995), rat proximal tubular cells (0.5-1 μM, 12 hours) (Wang et al., 

20 2011b), and in specific brain regions, hippocampus and cerebellum, after Pb exposure 

21 (500 ppm, 8 weeks) to rats (Bennet et al., 2007). Overall, there was a correlation between 

22 the blood Pb level and measures of lipid peroxidation often measured by MDA levels.

23 Interestingly, many species of plants, invertebrates, and vertebrates exhibit increased 

24 lipid peroxidation with Pb exposure (Sections 7.2.4, 7.3.5, and 7.3.12). The increase in 

25 lipid peroxidation following Pb exposure observed across species and kingdoms 

26 demonstrate an evolutionarily conserved oxidative response following Pb exposure.

5.2.4.3 NAD(P)H Oxidase Activation

NAD(P)H oxidase is a membrane bound enzyme that requires Ca2+ 27 in order to catalyze 

the production of O2

- 28 from NAD(P)H and molecular oxygen (Leseney et al., 1999). Two 

29 studies provide evidence for increased activation of NAD(P)H oxidase that may 

30 contribute to the production of ROS after Pb exposure (Ni et al., 2004; Vaziri et al., 

31 2003). Vaziri et al. (2003) found increased protein expression of the NAD(P)H subunit 

gp91 32 phox in the brain, heart, and renal cortex of Pb treated rats (100 ppm in drinking 

33 water, 12 weeks). This upregulation was present in Pb-treated (1-10 ppm) human 

34 coronary artery endothelial cells, but not vascular smooth muscle cells (VSMC), which 
February 2012 5-26 Draft – Do Not Cite or Quote

1 do not express the protein (Ni et al., 2004). It is possible that NAD(P)H oxidase serves as 

2 a potential source of ROS in cells that express this protein.

5.2.4.4 Antioxidant Enzyme Disruption

3 Oxidative stress can result not only from the increased production of ROS, but also from 

4 the decreased activity of antioxidant defense enzymes. Pb has been shown to alter the 

5 function of several antioxidant enzymes, including SOD, CAT, glucose-6-phosphate 

6 dehydrogenase (G6PD), and the enzymes involved in glutathione metabolism, GPx, 

7 glutathione-S-transferase (GST), and glutathione reductase (GR). These changes in the 

8 antioxidant defense system could be due to the high affinity of Pb for sulfhydryl groups 

9 contained within proteins and its metal ion mimicry. However changes could also be a 

10 consequence of increased oxidative damage by Pb.

11 Studies of the effects of Pb on the activities of SOD and CAT give divergent results. 

12 These metalloprotein enzymes require various essential trace elements for proper 

13 structure and function, making them a target for Pb toxicity. CAT is a heme containing 

14 protein that requires iron ions to function (Putnam et al., 2000). SOD exists in multiple 

15 isoforms that require copper and zinc (SOD1 and SOD3) (Antonyuk et al., 2009) or 

16 manganese (SOD2) (Borgstahl et al., 1992). A number of studies have found decreased 

17 activity of these enzymes (Pandya et al., 2010; Ergurhan-Ilhan et al., 2008; Mohammad 

18 et al., 2008; Yu et al., 2008; Patil et al., 2006a; Patil et al., 2006b; Conterato et al., In 

19 Press), whereas others have observed increased activity following Pb exposure (Ahamed 

20 et al., 2008; Lee et al., 2005). Pb exposure (500 ppm, 1, 4, and 8 weeks) in rats showed 

21 that organ SOD and CAT responded differently depending on the dose and tissue 

22 investigated. Activity of SOD and CAT varied based on the brain region analyzed and 

23 time of exposure (Bennet et al., 2007). Another study found that the brain had 

24 consistently decreased SOD activity, irrespective of dose in prenatally exposed animals 

25 (0.3 and 3.0 ppm, blood Pb level 20.4 and 24.5 μg/dL); however hepatic SOD activity 

26 increased at low level Pb administration and decreased after high level exposure

27 (Uzbekov et al., 2007). It is possible that the increased SOD and CAT protein is due to 

28 activation by ROS, while decreased enzyme activity is the result of metal ion substitution 

29 by Pb causing enzyme inactivation.

30 Glutathione is a tripeptide antioxidant containing a cysteine with a reactive thiol group 

31 that can act nonenzymatically as a direct antioxidant or as a cofactor in enzymatic 

32 detoxification reactions by GST. Glutathione will donate an electron while in its reduced 

33 state (GSH), which leads to conversion to the oxidized form, glutathione disulfide 

34 (GSSG). Pb binds to the thiol and can both interfere with the antioxidant capacity of and 
February 2012 5-27 Draft – Do Not Cite or Quote

1 decrease levels of GSH. Short-term administration of Pb in vitro (0.1 μM) and 

2 biomarkers of Pb exposure in humans (18 μg/dL mean blood Pb level) have been 

3 associated with decreased blood and organ GSH and cysteine content, which may be due 

4 to increased GSH efflux from tissues (Pandya et al., 2010; Pillai et al., 2010; Ahamed et 

5 al., 2009; Ahamed et al., 2008; Flora et al., 2007; Ahamed et al., 2005; Chetty et al., 

6 2005; Nakagawa, 1991, 1989). Long-term Pb exposure elicits a compensatory 

7 upregulation in the biosynthesis of GSH in the attempt to overcome Pb toxicity, thus 

8 often manifesting as an increase in Pb-induced GSH (Daggett et al., 1998; Corongiu and 

9 Milia, 1982; Hsu, 1981; Conterato et al., In Press). However, other studies have found 

10 that long-term Pb exposure resulting in mean blood Pb levels between 6.6 and 22 μg/dL, 

11 causes the depletion of GSH (Mohammad et al., 2008; Lee et al., 2005; Ercal et al., 

12 1996). Thus, the time of exposure is important to consider when measuring GSH levels.

13 Glutathione reductase is able to reduce GSSG back to GSH. Therefore, an increased 

14 GSSG/GSH ratio is considered to be indicative of oxidative stress. Epidemiologic studies 

15 have found higher blood Pb levels to be associated with increases in the GSSG/GSH ratio

16 (Mohammad et al., 2008; Ercal et al., 1996; Sandhir and Gill, 1995). In one study, this 

17 association was observed in a population of children with a mean blood Pb level below 

18 10 μg/dL (Diouf et al., 2006). Studies have found mixed effects on GR activation. GR 

19 possesses a disulfide at its active site that is a target for inhibition by Pb. Studies in 

20 animals and cells have reported decreased (Bokara et al., 2009; Sandhir and Gill, 1995; 

21 Sandhir et al., 1994), increased (Sobekova et al., 2009; Howard, 1974), and no change

22 (Hsu, 1981) in GR activity after Pb exposure. This could be because the effect of Pb on 

23 GR varies depending on sex (Sobekova et al., 2009) and organ or organ region (Bokara et 

24 al., 2009).

25 GSH is used as a cofactor for peroxide reduction and detoxification of xenobiotics by the 

26 enzymes GPx and GST. GPx requires selenium for peroxide decomposition (Rotruck et 

27 al., 1973), whereas GST functions via a sulfhydryl group. By reducing the uptake of 

28 selenium, depleting cellular GSH, and disrupting protein thiols, evidence indicates that 

29 Pb decreases the activity of GPx and GST (Pillai et al., 2010; Yu et al., 2008; Lee et al., 

30 2005; Nakagawa, 1991; Schrauzer, 1987). Similar to other antioxidant enzymes, 

31 compensatory upregulation of these enzymes is described after Pb exposure in animals 

32 and in Pb-exposed workers (painters with a mean blood Pb level of 5.4 μg/dL) (Bokara et 

33 al., 2009; Ergurhan-Ilhan et al., 2008; Conterato et al., 2007; Daggett et al., 1998; 

34 Conterato et al., In Press). However, in another study, these enzymes were not able to 

35 compensate for the increased Pb-induced ROS, further contributing to the oxidative 

36 environment (Farmand et al., 2005).
February 2012 5-28 Draft – Do Not Cite or Quote

1 Recently, γ-glutamyltransferase (GGT) within its normal range has been regarded as an 

2 early and sensitive marker of oxidative stress. This may be because cellular GGT 

3 metabolizes extracellular GSH to be used in intracellular GSH synthesis. Thus, cellular 

4 GGT acts as an antioxidant enzyme by increasing the intracellular GSH pool. However, 

5 the reasons for the association between GGT and oxidative stress have not been fully 

6 realized (Lee et al., 2004). In one study, occupational Pb exposure (mean blood Pb level 

7 of 29.1 μg/dL) was associated with increased serum GGT levels (Khan et al., 2008).

8 Interestingly, higher blood Pb level was similarly associated with higher serum GGT 

9 levels in a sample of the U.S. adult population (NHANES III) with lower blood Pb levels

10 (Lee et al., 2006a). In this study of nonoccupationally-exposed individuals, a 

11 concentration-dependent relationship was observed at blood Pb levels <7 μg/dL.

5.2.4.5 Nitric Oxide Signaling

• 12 NO, also known as endothelium-derived relaxing factor, is a potent endogenous 

13 signaling molecule involved in vasodilation. Short- and long-term Pb exposure in animals 

decreases the biologically active •NO, not through reduction in • 14 NO-production capacity

15 (Vaziri and Ding, 2001; Vaziri et al., 1999b), but as a result of inactivation and 

sequestration of • 16 NO by ROS (Malvezzi et al., 2001; Vaziri et al., 1999a). Endogenous 

•

NO can interact with ROS, specifically O2

- 17 , produced following exposure to Pb to form 

the highly cytotoxic reactive nitrogen species, peroxynitrite (ONOO- 18 ). This reactive 

19 compound can damage cellular DNA and proteins, resulting in the formation of 

20 nitrotyrosine among other products. Overabundance of nitrotyrosine in plasma and 

tissues is present after exposure to Pb (Vaziri et al., 1999a). • 21 NO is also produced by 

22 macrophages in the defense against certain infectious agents, including bacteria. Studies 

have indicated that Pb exposure can significantly reduce production of • 23 NO in immune 

24 cells (Pineda-Zavaleta et al., 2004; Lee et al., 2001b; Tian and Lawrence, 1995), possibly 

25 leading to reduced host resistance (Tian and Lawrence, 1996).

Production of • 26 NO is catalyzed by a family of enzymes called nitric oxide synthases 

27 (NOS), including endothelial NOS (eNOS), neuronal NOS (nNOS), and inducible NOS 

28 (iNOS), which require a heme prosthetic group and a zinc cation for enzymatic activity

(Messerschmidt et al., 2001). Paradoxically, the reduction in • 29 NO availability in vascular 

30 tissue following Pb exposure is accompanied by statistically significant upregulation in 

31 NOS isotypes (Vaziri and Ding, 2001; Vaziri et al., 1999b; Gonick et al., 1997). A direct 

32 inhibitory action of Pb on NOS enzymatic activity has been rejected (Vaziri et al., 

1999b). Instead, the upregulation of NOS occurs as compensation for the decreased • 33 NO 

34 resulting from ROS inactivation (Vaziri et al., 2005; Vaziri and Ding, 2001; Vaziri and 

35 Wang, 1999).
February 2012 5-29 Draft – Do Not Cite or Quote

Soluble Guanylate Synthase

Many biological actions of • 1 NO, such as vasorelaxation, are mediated by cyclic guanosine 

2 monophosphate (cGMP), which is produced by soluble guanylate cyclase (sGC) from the 

3 substrate guanosine triphosphate. Soluble guanylate cyclase is a heterodimer requiring 

4 one molecule of heme for enzymatic activity (Boerrigter and Burnett, 2009). In VSMC, 

sGC serves as the • 5 NO receptor. Marked reduction in plasma concentrations and urinary 

6 excretion of cGMP is observed after Pb exposure to rats (5 ppm for 30 days and 100 ppm

7 for 12 months [resulting in a mean blood Pb level of 29.4 μg/dL]) (Marques et al., 2001; 

8 Khalil-Manesh et al., 1993b). In addition, Pb exposure downregulated the protein 

9 abundance of sGC in vascular tissue (Farmand et al., 2005; Courtois et al., 2003; 

10 Marques et al., 2001). This downregulation in sGC was prevented by antioxidant therapy 

11 (ascorbic acid) suggesting that oxidative stress also plays a role in Pb-induced 

12 downregulation of sGC (no change in blood Pb level was observed after ascorbic acid 

13 treatment) (Marques et al., 2001).

5.2.5 Inflammation

14 Misregulated inflammation represents one of the major hallmarks of Pb-induced immune 

15 effects. It is important to note that this can manifest in any tissue where immune cell 

16 mobilization and tissue insult occurs. Enhanced inflammation and tissue damage occurs 

17 through the modulation of inflammatory cell function and production of pro18 inflammatory cytokines and metabolites. Overproduction of ROS and an apparent 

19 depletion of antioxidant protective enzymes and factors (e.g., selenium) accompany this 

20 immunomodulation (Chetty et al., 2005).

21 Traditional immune-mediated inflammation can be seen with bronchial 

22 hyperresponsiveness, asthma, and respiratory infections associated with exposure to Pb. 

23 But it is important to recognize that any tissue or organ can be affected by immune24 mediated inflammatory dysfunction given the distribution of immune cells as both 

25 permanent residents and infiltrating cell populations (Mudipalli, 2007; Carmignani et al., 

26 2000). Pb spheres implanted in the brains of rats produced neutrophil-driven 

27 inflammation with apoptosis and indications of neurodegeneration (Kibayashi et al., 

28 2010). Pb also induces renal tubulointerstitial inflammation (mean blood Pb level of 

29 18 μg/dL or 100 ppm exposure for 14 weeks) (Rodriguez-Iturbe et al., 2005; Ramesh et 

30 al., 2001), which has been coupled with activation of the redox sensitive nuclear 

31 transcription factor kappa B (NFκB) and lymphocyte and macrophage infiltration in rats 

32 (100 ppm for 14 weeks resulting in mean blood Pb levels ranging 23-27 μg/dL,) (Bravo 

33 et al., 2007). These events could be in response to the oxidative environment arising from 
February 2012 5-30 Draft – Do Not Cite or Quote

1 Pb exposure, since Pb-induced inflammation and NFκB activation can be ameliorated by 

2 antioxidant therapy (Rodriguez-Iturbe et al., 2004).

3 Inflammation can be mediated by the production of chemical messengers such as 

4 prostaglandins (PG). Pb exposure has been reported to increase arachidonic acid (AA) 

5 metabolism, thus elevating the production of PGE2, PGF2, and thromboxane in 

6 occupationally-exposed humans (mean blood Pb level 48 μg/dL) (Cardenas et al., 1993)

7 and animal and cell models (e.g., 0.01 μM, 48 h) (Chetty et al., 2005; Flohe et al., 2002; 

8 Knowles and Donaldson, 1997; Lee and Battles, 1994). Dietary Pb supplementation of 

9 animals (500 ppm, 19 days) can increase the percentage of cell membrane AA, the 

10 precursor of cyclooxygenase and lipoxygenase metabolism to PGs and leukotrienes

11 (Knowles and Donaldson, 1990). Additionally, Pb (1 μM) may promote the release of 

12 AA via activation of phospholipase A2, as shown in isolated VSMC (Dorman and 

13 Freeman, 2002).

14 Inflammation may be the result of increased pro-inflammatory signaling or may stimulate 

15 these signaling pathways. Pb can elevate the expression of the pro-inflammatory 

16 transcription factors NFκB and activator protein-1 (AP-1), as well as the AP-1 

17 component c-Jun (Korashy and El-Kadi, 2008; Korashy and Ei-Kadi, 2008; Bravo et al., 

18 2007; Ramesh et al., 1999; Pyatt et al., 1996). Pb exposure (25 μM) to dendritic cells 

19 stimulated phosphorylation of the Erk/MAPK pathway, but not p38, STAT3 or 5, or 

20 CREB (Gao et al., 2007)

5.2.5.1 Cytokine Production

21 There are three modes by which Pb affects immune cytokine production. First, Pb can act 

22 on macrophages to elevate the production of pro-inflammatory cytokines such as TNF-α 

23 and interleukin (IL)-6 (Cheng et al., 2006; Chen et al., 1999; Dentener et al., 1989). This 

24 can result in local tissue damage during the course of immune responses affecting such 

25 targets as the liver. Second, when Pb acts on dendritic cells, it skews the ratio of IL26 12/IL-10 such that T-derived lymphocyte helper (Th)1 responses are suppressed and Th2 

27 responses are promoted (Chen et al., 2004; Miller et al., 1998). Third, when acquired 

28 immune responses occur following exposure to Pb, Th1 lymphocyte production of 

29 cytokines is suppressed (e.g., IFN-γ) (Lynes et al., 2006; Heo et al., 1996). In contrast, 

30 Th2 cytokines such as IL-4, IL-5, and IL-6 are elevated (Gao et al., 2007; Kim and 

31 Lawrence, 2000). The combination of these three modes of cytokine changes induced by 

32 Pb creates a hyperinflammatory state among innate immune cells and acquired immunity 

33 is skewed toward Th2 responses.
February 2012 5-31 Draft – Do Not Cite or Quote

1 Iavicoli et al. (2006a) reported that low blood Pb concentrations produced significant 

2 changes in cytokine levels in mice. At a low dietary Pb concentration (0.11 ppm, blood 

3 Pb level of 1.6 μg/dL), IL-2 and IFN-γ were decreased compared to the controls 

4 (0.02 ppm, 0.8 μg/dL), indicating a suppressed Th1 response. As the dietary and blood Pb 

5 concentrations increased (resulting in blood Pb levels 12-61 μg/dL), a Th2 phenotype 

6 was observed with suppressed IFN-γ and IL-2 and elevated IL-4 production. These 

7 findings support the notion that the immune system is remarkably sensitive to Pb-induced 

8 functional alterations and that nonlinear effects may occur at low Pb exposures. TGF-β 

9 production is also altered by Pb exposure to cells (1 μM, 3 days) (Zuscik et al., 2007). IL10 2 is one of the more variable cytokines with respect to Pb-induced changes. Depending 

11 upon the protocol it can be slightly elevated in production or unchanged. Recently, Gao et 

12 al. (2007) found that Pb-treated dendritic cells (25 μM) promoted a slight but statistically 

13 significant increase in IL-2 production among lymphocytes. Proinflammatory cytokines 

14 have been measured in other organs and cell types after Pb exposure. Elevation of IL-1β 

15 and TNF-α were observed in the hippocampus after Pb exposure (15 ppm, i.p., daily for 2 

16 weeks, blood Pb level of 30.8 μg/dL) and increased IL-6 was found in the forebrain

17 (Struzynska et al., 2007).

18 Consistent with animal studies, epidemiologic studies also found higher concurrent blood 

19 Pb levels in children and occupationally-exposed adults to be associated with a shift 

20 toward production of Th2 cytokines relative to Th1 cytokines. The evidence in children 

21 was based on comparisons of serum cytokine levels among groups with different blood 

22 Pb levels without consideration of potential confounding factors. Among children ages 9 

23 months to 6 years in Missouri, Lutz et al. (1999) found that children with concurrent 

24 blood Pb levels 15-19 μg/dL had higher serum levels of IL-4 and IgE (Section 5.6.3) than 

25 did children with lower blood Pb levels. These results were consistent with the mode of 

26 action for IL-4 to activate B cells to induce B cell class switching to IgE. In another study 

27 of children in grades 5 and 6 in Taiwan, investigators did not group children by blood Pb 

28 levels but by potential for Pb exposures due to age of home and location of residence

29 (Hsiao et al., 2011). Concurrent blood Pb levels did not differ by residence in old versus 

30 new homes or by urban versus rural residence (means: 3.2-3.8 μg/dL) but were higher 

31 among children living near an oil refinery, in particular, among children with known 

32 respiratory allergies (mean: 8.8 μg/dL). This latter group of children also had the lowest 

33 serum levels of IFN-γ and highest levels of IL-4. There was no direct comparison of 

34 cytokine levels between blood Pb level groups in the population overall; however, 

35 cytokine levels were similar between healthy and allergy groups in the other Pb source 

36 groups that had similar blood Pb levels. Thus, the differences in cytokine levels between 

37 healthy and allergic children living near the oil refinery may have been influenced by 

38 differences in their blood Pb levels. 
February 2012 5-32 Draft – Do Not Cite or Quote

1 Evidence of association between blood Pb levels and cytokine levels in 

2 nonoccupationally-exposed adults was equivocal. Among adult university students in 

3 Incheon, Korea, Kim et al. (2007) found associations of concurrent blood Pb level with 

4 serum levels of TNF-α and IL-6 that were larger among male students with blood Pb 

5 levels 2.51-10.47 μg/dL. Notably, the relative contributions of lower recent versus higher 

6 past Pb exposures to these effects is not known. In models that adjusted for age, sex, 

7 BMI, and smoking status, a 1 μg/dL increase in blood Pb level was associated with a 

8 23% increase (95% CI: 4, 55%) in log of TNF-α and a 26% increase in log of IL-6 (95% 

9 CI: 0, 55%). The association between levels of blood Pb and plasma TNF-α was greater 

10 among men who were GSTM1 null or had the TNF-α GG genotype. For the association 

11 between blood Pb level and plasma IL-6, the effect estimate was slightly elevated in 

12 TNF-α GG genotype but not elevated in the GSTM1 positive group. The effects of Pb on 

13 several physiological systems have been hypothesized to be mediated by the generation 

14 of ROS (Daggett et al., 1998). Thus, the null variant of GSTM1, which is associated with 

15 reduced elimination of ROS, may increase the risk of Pb-associated immune effects. The 

16 results for the TNF-α polymorphism were difficult to interpret. The GG genotype is 

17 associated with lower expression of TNF-α, and the literature is mixed with respect to 

18 which variant increases risk of inflammation-related conditions. Among adults in Italy, 

19 blood Pb levels were not correlated with either Th2 or Th1 cytokine levels in men

20 (Boscolo et al., 1999) or women (Boscolo et al., 2000)

21 Results from studies of occupationally-exposed adults also suggested that Pb exposure 

22 may be associated with decreases in Th1 cytokines and increases in Th2 cytokines; 

23 however, analyses were mostly limited to comparisons of levels among different 

24 occupational groups with different mean blood Pb levels (Di Lorenzo et al., 2007; 

25 Valentino et al., 2007; Yucesoy et al., 1997a). The exception was a study of male foundry 

26 workers, pottery workers, and unexposed workers by Valentino et al. (2007). Multiple 

27 regression analyses were performed with age, BMI, smoking, and alcohol consumption 

28 included as covariates. Although information on concentration-response relationships 

29 was not provided, higher blood Pb level was associated with higher IL-10 and TNF-α. 

30 Levels of IL-2, IL-6, and IL-10 also increased from the lowest to highest blood Pb group. 

31 In contrast with most other studies, both exposed worker groups had lower IL-4 levels 

32 compared with controls. In a similar analysis, DiLorenzo et al. (2007) separated exposed 

33 workers into intermediate (9.1-29.4 μg/dL) and high (29.4-81.1 μg/dL) blood Pb level 

34 groups, with unexposed workers comprising the low exposure group (blood Pb levels 1-

35 11 μg/dL). Mean TNF-α levels showed a monotonic increase from the low to high blood 

36 Pb group. Levels of granulocyte colony-stimulating factor (G-CSF) did not differ 

37 between the intermediate and high blood Pb groups among the Pb recyclers; however, G38 CSF levels were higher in the Pb recyclers than in the unexposed controls. Furthermore, 

39 among all subjects, blood Pb showed a strong, positive correlation with G-CSF. Yucesoy 
February 2012 5-33 Draft – Do Not Cite or Quote

1 et al. (1997a) found lower serum levels of the Th1 cytokines, IL-1β and IFN-γ, in 

2 workers (mean blood Pb level of 59.4 μg/dL) compared with controls (mean blood Pb 

3 level of 4.8 μg/dL); however levels of the Th2 cytokines, IL-2 and TNF-α levels, were 

4 similar between groups. As most occupationally-exposed cohorts represent populations 

5 highly exposed to Pb, with mean blood Pb levels >22 μg/dL, effects observed within 

6 these cohorts may not be generalizable to the population as a whole. However, animal, 

7 general population, and occupational studies suggest that exposure to Pb increases the 

8 production of pro-inflammatory cytokines, skews the ratio of Th1 and Th2 cytokines to 

9 favor Th2 responses, and suppresses lymphocyte cytokine production.

5.2.6 Endocrine Disruption

5.2.6.1 Hypothalamic-Pituitary-Gonadal Axis

10 Pb is a potent endocrine disrupting chemical found to be associated with reproductive and 

11 developmental effects in both male and female animal models. Pb may act both at 

12 multiple points along the hypothalamic-pituitary-gonadal (HPG) axis and directly at 

13 gonadal sites. The HPG axis functions in a closely regulated manner to produce 

14 circulating sex steroids and growth factors required for normal growth and development. 

15 Long-term Pb exposure in animals has been shown to reduce serum levels of follicle16 stimulating hormone (FSH), luteinizing hormone (LH), testosterone, and estradiol

17 (Biswas and Ghosh, 2006; Rubio et al., 2006). Similar changes in serum HPG hormones 

18 have been observed after high-level Pb exposure in animals, resulting in blood Pb levels 

19 > 20 μg/dL (Dearth et al., 2002; Ronis et al., 1998b; Foster, 1992; Sokol and Berman, 

20 1991). Increases in LH and FSH have been associated with increasing concurrent blood 

21 Pb levels in adult women from the NHANES cohort (Krieg, 2007). The change in HPG 

22 hormones likely occurs through the inhibition of LH secretion and the reduction in the 

23 expression of the steroidogenic acute regulatory protein (StAR) (Huang and Liu, 2004; 

24 Srivastava et al., 2004; Huang et al., 2002; Ronis et al., 1996). StAR expression is the 

25 rate-limiting step essential in maintaining gonadotropin-stimulated steroidogenesis, 

26 which results in the formation of testosterone and estradiol. Prenatal Pb exposure 

27 (resulting in 3 μg/dL blood Pb) was found to decrease basal StAR synthesis, but not 

28 gonadotropin-stimulated StAR synthesis, suggesting that Pb may not directly affect 

29 ovarian responsiveness to gonadotropin stimulation (Srivastava et al., 2004). Instead, Pb 

30 may act at the hypothalamic-pituitary level to alter LH secretion, which is necessary to 

31 drive StAR production and subsequent sex hormone synthesis. Release of LH and FSH 

32 from the pituitary is controlled by gonadotropin-releasing hormone (GnRH). Pb exposure 

33 (10 μM, 90 min) in rat brain median eminence cells can block GnRH release (Bratton et 
February 2012 5-34 Draft – Do Not Cite or Quote

1 al., 1994). Pb may also interfere with release of pituitary hormones through interference 

2 with cation-dependent secondary messenger systems, which mediate hormone release and 

3 storage.

4 Endocrine disruption may also be a result of altered hormone binding to endocrine 

5 receptors. Prenatal and postnatal Pb exposure (20 ppm) to rats was able to decrease the 

6 number of estrogen, LH, and FSH receptors found in the uterus or ovaries and receptor 

7 binding affinity (Wiebe et al., 1988; Wiebe and Barr, 1988). Altered hormone binding 

8 ability may be due to the ion binding properties of Pb, resulting in changes in receptor 

9 tertiary structure that will disrupt ligand binding. In addition, Pb-induced changes in 

10 hormone levels that act as inducing agents for receptor synthesis may affect the number 

11 of hormone receptors produced. 

12 Some of these endocrine disrupting effects of Pb have been related to the generation of 

13 ROS. Treatment with antioxidants is able to counteract a number of the endocrine 

14 disrupting effects of Pb, including apoptosis and decreased sperm motility and production

15 (Salawu et al., 2009; Shan et al., 2009; Madhavi et al., 2007; Rubio et al., 2006; Wang et 

16 al., 2006a; Hsu et al., 1998a). Direct generation of ROS in epididymal spermatozoa was 

17 observed after Pb exposure in rats (i.p. 20 or 50 ppm, 6 weeks) (Hsu et al., 1998b). In 

18 addition, lipid peroxidation has been observed in Pb-exposed rats (i.p. 0.025 ppm, 15 

19 days) (Pandya et al., In Press). Lipid peroxidation in the seminal plasma was significantly 

20 increased in a group of Pb-exposed workers with high blood Pb levels (>40 μg/dL)

21 (Kasperczyk et al., 2008).

22 The liver is often associated with the HPG axis due in part to its production of insulin23 like growth factor 1 (IGF-1). Children with increased blood Pb levels (>4 μg/dL)

24 (Huseman et al., 1992) and Pb-exposed animals (blood Pb level of 14 μg/dL) (Pine et al., 

25 2006; Dearth et al., 2002) and gonadal cells (50 ppm Pb exposure) (Kolesarova et al., 

26 2010) show a decrease in plasma IGF-1, which may be the result of decreased translation 

27 or secretion of IGF-1 (Dearth et al., 2002). IGF-1 also induces LH-releasing hormone 

28 release, such that IGF-1 decrements may explain decreased LH and estradiol levels. IGF29 1 production is stimulated by growth hormone (GH) secreted from the pituitary gland and 

30 could be the result of GH depletion.

31 A number of studies have revealed that Pb exposure affects the dynamics of growth. 

32 Decreased growth after Pb exposure could be the result of Pb-induced decreased GH 

33 levels (Berry et al., 2002; Camoratto et al., 1993; Huseman et al., 1992; Huseman et al., 

34 1987). This decrease in GH could be a result of decreased release of GH releasing 

35 hormone (GHRH) from the hypothalamus or disrupted GHRH binding to its receptor, 

which has been reported in vitro after Pb treatment (IC50 free Pb in solution 5.2x10-5 36 μM, 
February 2012 5-35 Draft – Do Not Cite or Quote

1 30 minutes) (Lau et al., 1991). GH secretion may also be altered from decreased 

2 testosterone, a result of Pb exposure. 

5.2.6.2 Hypothalamic-Pituitary-Thyroid Axis

3 The effects of Pb on the hypothalamic-pituitary-thyroid (HPT) axis are mixed. Pb 

4 exposure impacts a variety of players in the thyroid hormone system. A number of human 

5 studies (blood Pb levels >7.3 μg/dL) have shown that elevated blood Pb are associated 

6 with lower thyroxine (T4) and free T4 levels without alteration in triiodothyronine (T3), 

7 suggesting that long-term Pb exposure may depress thyroid function in workers (Dundar 

8 et al., 2006; Tuppurainen et al., 1988; Robins et al., 1983). However, animal studies on 

9 thyroid hormones have shown mixed results. Pb-exposed cows (blood Pb levels 

10 >51 μg/dL) were reported to have an increase in plasma T3 and T4 levels (Swarup et al., 

11 2007), whereas mice and chickens manifested decreased serum T3 concentrations after Pb 

12 exposure accompanied by increased lipid peroxidation (Chaurasia et al., 1998; Chaurasia 

13 and Kar, 1997). Decreased serum T3 and increased lipid peroxidation were both restored 

14 by vitamin E treatment, suggesting the disruption of thyroid hormone homeostasis could 

15 be a result of altered membrane architecture and oxidative stress; however, no data were 

16 provided to exclude changes in Pb kinetics as the mechanism of protection (Chaurasia 

17 and Kar, 1997).

18 Decreased T4 and T3 may be the result of altered pituitary release of thyroid stimulating 

19 hormone (TSH). However, several studies have reported higher TSH levels in high-level 

20 Pb-exposed workers (blood Pb levels >39 μg/dL) (Lopez et al., 2000; Singh et al., 2000; 

21 Gustafson et al., 1989), which would result in increased T4 levels. Overall, results on the 

22 effects of Pb on the HPT axis are inconclusive.

5.2.7 Cell Death and Genotoxicity

23 A number of studies have attempted to characterize the genotoxicity of inorganic Pb in 

24 human populations, laboratory animals, and cell cultures. Endpoints investigated include 

25 DNA damage (single- and double-strand breaks, DNA-adduct formation), mutagenicity, 

26 clastogenicity (sister chromatid exchange, micronucleus formation, chromosomal 

27 aberrations), and epigenetic changes (changes in gene expression, mitogenesis). It is 

28 important to note that numerous studies have utilized exposure to Pb chromate to 

29 investigate genotoxicity endpoints; some studies have specifically attributed the observed 

30 increases in DNA damage and clastogenicity to the chromate ion while others have not. 

31 Due to the uncertainty regarding whether observed genotoxic effects are due to chromate 
February 2012 5-36 Draft – Do Not Cite or Quote

1 or Pb in studies using this form of inorganic Pb, only studies utilizing other forms of 

2 inorganic Pb (e.g., Pb nitrate, acetate) are discussed below. 

5.2.7.1 DNA Damage

3 A number of studies in human populations have observed associations between Pb 

4 exposure and increased DNA damage, as measured as DNA strand breaks. Most of these 

5 associations have been observed in occupationally-exposed populations (Grover et al., 

6 2010; Minozzo et al., 2010; Shaik and Jamil, 2009; Danadevi et al., 2003; Hengstler et 

7 al., 2003; Palus et al., 2003; Fracasso et al., 2002; de Restrepo et al., 2000). It is 

8 important to note that occupationally-exposed adults have very high blood Pb levels, and 

9 in one study (de Restrepo et al., 2000) the association between blood Pb level and DNA 

10 damage was observed only in workers with blood Pb levels greater than 120 μg/dL. Also, 

11 the studies were equivocal in regard to how blood Pb levels correlated with DNA 

12 damage: Fracasso et al. (2002) observed that DNA damage increased with increasing 

13 blood Pb levels (blood Pb levels, <25, 25-35, and >35 μg/dL), whereas Palus et al. (2003)

14 (mean blood Pb level: 50.4 μg/dL [range: 28.2-65.5 μg/dL]) and Minozzo et al. (2010)

15 (mean [SD]: 59.43 μg/dL [28.34]) observed no correlation. Lastly, Pb-exposed workers 

16 are also potentially exposed to other genotoxic materials, making it difficult to rule out 

17 confounding co-exposures. However, Hengstler et al. (2003) examined workers exposed 

18 to Pb, cadmium, and cobalt and observed that neither blood (mean: 4.4 [IQR: 2.84-

13.6] μg/dL) nor air Pb levels (mean: 3.0 [IQR: 1.6-50.0] μg/m3 19 ) were associated with 

20 DNA damage when examined alone, but that blood Pb influenced the occurrence of 

21 single strand DNA breaks when included in a multiple regression model along with 

22 cadmium in air and blood and cobalt in air indicating lack of confounding. Two studies 

23 were found that investigated Pb-induced DNA damage resulting from nonoccupational 

24 exposures. Mendez-Gomez (2008) observed that children living at close and intermediate 

25 distances to a Pb smelter had mean (range) blood Pb levels of 19.5 (11.3-49.2) and 28.6 

26 (11.4-47.5) μg/dL, respectively, compared to blood Pb level of 4.6 (0.1-8.7) μg/dL for 

27 children living distant to the smelter. DNA damage was increased in children living 

28 nearest to the smelter, compared to the children at the intermediate distance, but was not 

29 different from children living farthest away from the smelter. Multivariate analysis 

30 (which considered children urinary As levels, highest in children farthest from the 

31 smelter), revealed no statistically significant associations between DNA damage and 

32 blood Pb level. Further, DNA repair ability was also observed to be unrelated to blood Pb 

33 levels. Alternatively, Yanez et al. (2003) observed that children living close to a mining 

34 complex (mean [range] blood Pb level: 11.6 [3.0 to 19.5] μg/dL) did have higher levels 
February 2012 5-37 Draft – Do Not Cite or Quote

1 of DNA damage compared to control children who lived further away from the mining 

2 facility (mean [range] blood Pb level: 8.3 [3.0-25.0] μg/dL).

3 In mice given 0.7 to 89.6 mg/kg Pb nitrate by gavage for 24, 48, or 72 hours, or 1 or 2 

4 weeks, single strand DNA breaks in white blood cells were observed but did not increase 

5 with increasing concentration (Devi et al., 2000). The three highest concentrations had 

6 responses which were similar in magnitude and were actually lower than the responses to 

7 lower concentrations tested. In mice exposed to Pb (blood Pb level of 0.68 μg/dL) via 

8 inhalation for up to 4 weeks, differential levels of DNA damage were observed in 

9 different organ systems, with only the lung and the liver demonstrating statistically 

10 greater DNA damage compared to the respective organ controls after acute exposure

11 (Valverde et al., 2002). Statistically elevated levels of DNA damage were observed in the 

12 kidneys, lungs, liver, brain, nasal cavity, bone marrow, and leukocytes of mice exposed 

13 over a period of 4 weeks, although variability was high in all groups. The magnitude of 

14 the DNA damage was characterized as weak and did not increase with increasing 

15 durations of exposure. Xu et al. (2008) exposed mice to 10-100 mg/kg Pb-acetate via 

16 gavage for four weeks and observed a concentration-dependent increase in DNA single 

17 strand breaks in white blood cells that was statistically significant at 50 and 100 mg/kg. 

18 The authors characterized the observed DNA damage as severe. Pb nitrate induced DNA 

19 damage in primary spermatozoa in Pb-exposed rats (blood Pb levels of 19.5 and 

20 21.9 μg/dL) compared to control rats (Nava-Hernandez et al., 2009). The level of DNA 

21 damage was not concentration dependent and was comparable in both exposure groups. 

22 Narayana and Al-Bader (2011) observed no increase in DNA damage in the livers of rats 

23 exposed to 0.5 or 1% Pb nitrate in drinking water for 60 days. Interestingly, although the 

24 results were not statistically significant and highly variable within exposure groups, DNA 

25 fragmentation appeared to be lower in the exposed animals.

26 Studies investigating Pb-induced DNA damage in human cell cultures were 

27 contradictory. Pb-acetate did not induce DNA strand breaks in human HeLa cells when 

28 exposed to 500 μM Pb-acetate for 20-25 hours or 100 μM for 0.5-4 hours (Hartwig et al., 

29 1990; Snyder and Lachmann, 1989). Pb nitrate, administered to lymphoma cells at 1000-

30 10,000 μM for 6 hours, did not result in any DNA-protein crosslinks (Costa et al., 1996).

31 Pb-acetate was observed by Woźniak and Blasiak (2003) to result in DNA single and 

32 double strand breaks in primary human lymphocytes exposed to 1-100 μM for 1 hour, 

33 although the pattern of damage was peculiar. DNA damage was greater in cells exposed 

34 to 1 or 10 μM, compared to those exposed to 100 μM. DNA-protein crosslinks were only 

35 observed in the 100 μM exposure group, suggesting that the decreased strand breaks 

36 observed in the high exposure group may be a result of increased crosslinking in this 

37 group. Pasha Shaik et al. (2006) also observed DNA damage in human lymphocytes 

38 exposed to 2,100-3,300 μM Pb nitrate for 2 hours. Although there was a concentration-
February 2012 5-38 Draft – Do Not Cite or Quote

1 dependent increase in DNA damage from 2,100-3,300 μM, no statistics were reported 

2 and no unexposed control group was included making it difficult to interpret these results. 

3 Gastaldo et al. (2007) observed that exposure of human endothelial cells to 1-1,000 μM 

4 Pb nitrate for 24 hours resulted in a concentration-dependent increase in DNA double 

5 strand breaks.

6 Studies in animal cell lines were equally as ambiguous as those using human cell lines. 

7 Zelikoff et al. (1988) and Roy and Rossman (1992) reported that Pb-acetate 

8 (concentration not stated and 1,000 μM, respectively) did not induce single or double 

9 DNA strand breaks or DNA-protein or DNA-DNA crosslinks in CHV79 cells. However, 

10 both Xu et al. (2006) and Kermani et al. (2008) reported Pb-acetate-induced DNA 

11 damage in PC12 cells exposed to 0.1, 1, or 10 μM for 24 hours and in bone marrow 

12 mesenchymal stem cells exposed to 60 μM for 48 hours, respectively. Wedrychowski et 

13 al. (1986) reported that DNA-protein crosslinks were induced in a 

14 concentration-dependent manner in hepatoma cells exposed to 50-5,000 μM Pb nitrate 

15 for 4 hours. Pb-acetate and Pb nitrate increased the incidence of nick translation in 

16 CHV79 cells when a bacterial DNA polymerase was added.

17 Pb-acetate did not induce single strand DNA breaks in HeLa cells exposed to 500 μM for 

18 20-25 hours (Hartwig et al., 1990). However, exposure to both Pb-acetate and UV light 

19 resulted in increased persistence of UV-induced strand breaks, compared to exposure to 

20 UV light alone. Similar effects were seen in hamster V79 cells: UV-induced mutation 

21 rates and SCE frequency was exacerbated by co-incubation with Pb-acetate. Taken 

22 together, these data suggest that Pb-acetate interferes with normal DNA repair 

23 mechanisms triggered by UV exposure alone. Pb nitrate was observed to affect different 

24 DNA double strand break repair pathways in human endothelial cells exposed to 100 μM 

25 for 24 hours. Exposure to Pb inhibited nonhomologous end joining repair, but increased 

26 two other repair pathways, MRE11-dependent and Rad51-related repair (Gastaldo et al., 

27 2007). Interestingly, in contrast to the above studies, exposure of lung carcinoma cells to 

28 100, 300, or 500 μM Pb-acetate for 24 hours resulted in an increase in nucleotide excision 

29 repair efficiency (Li et al., 2008a). Roy and Rossman (1992) observed an increase in UV30 induced mutagenicity when CHV79 cells were co-exposed to 400 μM Pb-acetate (a 

31 nonmutagenic concentration of Pb-acetate), indicating an inhibition of DNA repair. 

32 Treatment of Chinese hamster ovary cells to 0.5-500 μM Pb-acetate resulted in a 

33 concentration-dependent accumulation of apurinic/apyrimidinic site incision activity, 

34 indicating that DNA repair was diminished (McNeill et al., 2007).
February 2012 5-39 Draft – Do Not Cite or Quote

5.2.7.2 Mutagenicity

1 Only one human study was found that investigated Pb-induced mutagenicity. Van 

2 Larebeke et al. (2004) investigated the frequency of mutations in the hypoxanthine 

3 phosphoribosyltransferase (HPRT) gene in Flemish women without occupational 

4 exposures to Pb or a number of other heavy metals and organic contaminants. Higher 

5 blood Pb level (range 1.6-5.2 μg/dL) was associated with greater HPRT mutation 

6 frequency in the total population. Also, women with high blood Pb levels (i.e., greater 

7 than the population median, not reported) demonstrated a greater mutation frequency 

8 compared to women with lower blood Pb levels.

9 Pb-induced mutagenicity was investigated in four studies using human cell cultures. Ye

10 (1993) exposed human keratinocytes to 0.1-100 μM/mL Pb-acetate for 2-24 hours. This 

11 study did not measure HPRT mutations directly, but rather measured the amount of 

12 tritium incorporated into DNA as an indicator of mutation. In the presence of 6-

13 thioguanine, tritium incorporation was increased in exposed cells, indicating weak 

14 mutagenicity. Hwua and Yang (1998) reported that Pb-acetate was not mutagenic in 

15 human foreskin fibroblasts exposed to 500-2,000 μM for 24 hours. Pb-acetate remained 

16 nonmutagenic in the presence of 3-aminotriazole, a catalase inhibitor, indicating that 

17 oxidative metabolism did not play a part in potential mutagenicity of Pb. Exposure to 

18 Pb-acetate alone did not induce mutagenicity in lung carcinoma cells (100-500 μM for 24 

19 hours) or fibroblasts (300-500 μM for 24 hours) (Li et al., 2008a; Wang et al., 2008c).

20 However, pretreatment with PKC inhibitors before Pb treatment did result in statistically 

21 significant increases in mutagenicity in both cell lines.

22 Results from investigations into Pb-induced mutagenicity using animal cell lines were as 

23 equivocal as were the findings from human cell line studies, although the mixed findings 

24 may be reflective of specific Pb compounds used. Pb-acetate was observed to be 

25 nonmutagenic (HPRT assay) in Chinese hamster V79 cells exposed to 1-25 μM of the 

26 compound for 24 hours (Hartwig et al., 1990), but elicited a mutagenic response in 

27 CHV79 cells (gpt assay) exposed to 1,700 μM for 5 days (Roy and Rossman, 1992).

28 Pb-acetate was observed to be nonmutagenic (HPRT assay) in Chinese hamster ovary 

29 cells exposed to 5 μM for 6 hours (McNeill et al., 2007). The observation of mutagenicity 

30 in the second study is complicated by the concurrent observation of severe cytotoxicity at 

31 the same concentration. Pb nitrate was alternatively found to be nonmutagenic in CHV79 

32 cells (gpt assay) exposed to 0.5-2,000 μM for 5 days (Roy and Rossman, 1992), but 

33 mutagenic in the same cell line (HPRT assay) exposed to 50-5,000 μM for 5 days

34 (Zelikoff et al., 1988). However, mutagenicity was only observed at 500 μM, and was 

35 higher than that observed at higher concentrations. Pb sulfate was also observed to be 

36 mutagenic in CHV79 cells (HPRT assay) exposed to 100-1,000 μM for 24 hours, but as 
February 2012 5-40 Draft – Do Not Cite or Quote

1 with Pb nitrate, it was not concentration-dependent (Zelikoff et al., 1988). Pb chloride 

2 was the only Pb compound tested in animal cell lines that was consistently mutagenic: 

3 three studies from the same laboratory observed concentration-dependent mutagenicity in 

4 the gpt assay in Chinese hamster ovary cells exposed to 0.1-1 μM Pb chloride for one 

5 hour (Ariza and Williams, 1999; Ariza et al., 1998; Ariza and Williams, 1996).

5.2.7.3 Clastogenicity

6 Clastogenicity is the ability of a compound to induce chromosomal damage, and is 

7 commonly observed as sister chromatid exchange, micronuclei formation, or incidence of 

8 chromosomal aberrations (i.e., breaks or gaps in chromosomes). The potential for Pb to 

9 be clastogenic has been investigated in numerous studies as described below.

Sister Chromatid Exchange

10 An association between blood Pb levels (means: 10.48 - 86.9 μg/dL) and sister chromatid 

11 exchange (SCE) was observed in a number of occupational studies (Wiwanitkit et al., 

12 2008; Duydu et al., 2005; Palus et al., 2003; Duydu et al., 2001; Pinto et al., 2000; 

13 Bilban, 1998; Anwar and Kamal, 1988; Huang et al., 1988). However, there are 

14 numerous methodological issues that limit firm conclusions from being drawn. Most 

15 notably, occupational co-exposures to other genotoxic materials were possible, although 

16 some studies excluded workers with exposures to known mutagens (Pinto et al., 2000; 

17 Huang et al., 1988). In most studies that attempted to investigate the concentration18 response relationship in workers, no association was observed between increasing blood 

19 Pb levels and the number of SCE (Palus et al., 2003; Duydu et al., 2001; Pinto et al., 

20 2000). However, Huang et al. (1988) did observe increased SCE in exposed workers in 

21 the two highest blood Pb groups (52.1 and 86.9 μg/dL), with a statistically significant 

22 association observed in the 86.9 μg/dL group. Pinto et al. (2000) did report an association 

23 with duration of exposure (range of years exposed: 1.6-40). Two studies reported no 

24 correlation between occupational exposure to Pb and number of SCE (Rajah and Ahuja, 

25 1996; Rajah and Ahuja, 1995). However, these two studies may have suffered from 

26 limited statistical power to observe an effect as they included very small numbers of Pb27 exposed workers. Mielzynska et al. (2006) found no association between blood Pb level 

28 and SCEs in children in Poland. Children had an average blood Pb level of 7.69 μg/dL 

29 and 7.87 SCEs/cell. 

30 Pb exposure has been observed to induce SCEs in multiple laboratory animal studies. In 

31 mice exposed to up to 100 mg/kg Pb-acetate i.p., Pb induced SCEs with 50 and 

32 100 mg/kg (Fahmy, 1999). Pb nitrate, also administered i.p. and induced the formation of 
February 2012 5-41 Draft – Do Not Cite or Quote

1 SCEs in a concentration-dependent manner (10-40 mg/kg) in the bone marrow of 

2 exposed mice (Dhir et al., 1993). Nayak et al. (1989b) exposed pregnant mice to 100-

3 200 mg/kg Pb nitrate via i.v. injection and observed an increase in SCEs in dams at 150 

4 and 200 mg/kg; no SCEs were observed in the fetuses. Tapisso et al. (2009) exposed rats 

5 to 21.5 mg/kg Pb-acetate (1/10th the LD50) via i.p. injection on alternating days for 11 or 

6 21 days, for a total of 5 or 10 exposures. Induction of SCEs in the bone marrow of 

7 exposed rats was increased over controls in a statistically significant duration-dependent 

8 manner. It is important to note that all three of these studies utilized an injection route of 

9 exposure that may not be relevant to routes of exposure in the general population 

10 (e.g., air, drinking water exposure). 

11 Few studies were found that investigated SCE formation in human cell lines due to Pb 

12 exposure. Statistically significant, concentration-dependent increases in SCEs were 

13 observed in human lymphocytes obtained from a single donor when incubated with 1, 5, 

14 10, or 50 μM Pb nitrate (Ustundag and Duydu, 2007). Melatonin and N-acetylcysteine 

15 were reported to ameliorate these effects, indicating Pb may induce SCEs through 

16 increased oxidative stress. Pb chloride was also observed to increase SCE levels in 

17 human lymphocytes exposed to 3 or 5 ppm (Turkez et al., In Press).

18 Studies investigating SCE in rodent cells were more equivocal than those in human cells. 

19 Pb sulfate, acetate, and nitrate were found not to induce SCE in Chinese hamster V79 

20 cells (Hartwig et al., 1990; Zelikoff et al., 1988). Both of these studies only examined 25-

21 30 cells per concentration, reducing their power to detect Pb-induced SCEs. Cai and 

22 Arenaz (1998), on the other hand, used 100 cells per treatment and observed that 

23 exposure to 0.05-1 μM Pb nitrate for 3-12 hours resulted in a weak, concentration24 dependent increase in SCEs in Chinese hamster ovary cells. Lin et al. (1994) also 

25 observed a concentration-dependent increase in SCEs in Chinese hamster cells exposed 

26 to 3-30 μM Pb nitrate for 2 hours.

Micronucleus Formation

27 Pb-induced micronucleus formation was observed in numerous occupational studies

28 (Grover et al., 2010; Khan et al., 2010b; Minozzo et al., 2010; Shaik and Jamil, 2009; 

29 Minozzo et al., 2004; Palus et al., 2003; Vaglenov et al., 2001; Pinto et al., 2000; Bilban, 

30 1998; Vaglenov et al., 1998). The workers in the occupational studies generally had high 

31 blood Pb levels (>20 μg/dL) making comparisons to the general population difficult, 

32 although Pinto et al. (2000) observed increased micronuclei in exposed workers with an 

33 average blood Pb level of 10.48 μg/dL compared with unexposed controls. In studies 

34 investigating the correlation between blood Pb levels and micronucleus formation, no 

35 association was observed (Minozzo et al., 2010; Minozzo et al., 2004; Palus et al., 2003; 
February 2012 5-42 Draft – Do Not Cite or Quote

1 Pinto et al., 2000), although Pinto et al. (2000), Grover et al. (2010), and Minozzo et al.

2 (2010) did report an association between micronuclei formation and duration of exposure. 

3 Only one study was found that investigated micronucleus formation in a nonworker 

4 population; Mielzynska et al. (2006) reported a statistically significant positive 

5 correlation between blood Pb levels and micronuclei frequency in children in Poland. 

6 Children, with an average blood Pb level of 7.69 μg/dL, were observed to have 4.44 

7 micronucleated cells per 1,000 cells analyzed. Children with blood Pb levels greater than 

8 10 μg/dL had significantly more micronucleated cells than did children with blood Pb 

9 levels less than 10 μg/dL.

10 Micronucleus formation in response to Pb exposure has been observed in rodent animal 

11 studies. Celik et al. (2005) observed that exposure of female rats to 140, 250, or 500 g/kg 

12 Pb-acetate once per week for 10 weeks resulted in statistically significantly increased 

13 numbers of micronucleated polychromatic erythrocytes (PCEs) compared to controls. 

14 Similarly, Alghazal et al. (2008b) exposed rats to 100 ppm Pb-acetate daily for 125 days 

15 and observed statistically significant increases in micronucleated PCEs in both sexes. 

16 Tapisso et al. (2009) exposed rats to 21.5 mg/kg Pb-acetate (1/10th the LD50) via i.p. 

17 injection on alternating days for 11 or 21 days, for a total of 5 or 10 exposures. Formation 

18 of micronuclei in the bone marrow of exposed rats was increased over that in controls in 

19 a significant duration-dependent manner. Two further studies investigated formation of 

20 micronuclei in the bone marrow of exposed mice: Roy et al. (1992) exposed mice to 10 

21 or 20 mg/kg Pb nitrate i.p. and observed a concentration-dependent increase in 

22 micronuclei, whereas Jagetia and Aruna (1998) observed an increase in micronuclei in 

23 mice exposed to 0.625-80 mg/kg Pb nitrate i.p., though the increase was not 

24 concentration-dependent. Mice exposed to 1 g/L Pb-acetate via drinking water, a more 

25 environmentally relevant route of exposure, for 90 days had statistically significant 

26 increases in micronucleated PCEs (Marques et al., 2006).

27 A few studies were found that reported increased micronucleus formation in human cell 

28 lines treated with Pb. Concentration-dependent micronucleus formation was observed in 

29 human lymphocytes when exposed to either 1, 5, 10, or 50 μM Pb nitrate or 3 or 5 ppm

30 Pb chloride (Ustundag and Duydu, 2007; Turkez et al., In Press). Gastaldo et al. (2007)

31 also observed a dose-dependent increase in micronuclei in human endothelial cells 

32 exposed to 1-1,000 μM Pb nitrate for 24 hours. Two animal cell culture studies 

33 investigating micronuclei formation produced contrasting results. One study observed 

34 that micronuclei were not induced in Chinese hamster cells exposed to 3-30 μM Pb 

35 nitrate for 2 hours (Lin et al., 1994), whereas the other observed that Pb-acetate induced a 

36 concentration-dependent increase in Chinese hamster cells when administered at 0.03-

37 10 μM for 18 hours (Bonacker et al., 2005).
February 2012 5-43 Draft – Do Not Cite or Quote

Chromosomal Aberrations

1 Chromosomal aberrations (e.g., chromosome breaks, nucleoplasmic bridges, di- and a2 centric chromosomes, and rings) were examined in a number of occupational studies

3 (Grover et al., 2010; Shaik and Jamil, 2009; Pinto et al., 2000; Bilban, 1998; De et al., 

4 1995; Huang et al., 1988). Methodological limitations outlined in previous sections, 

5 including potential for occupational co-exposure to genotoxic substances and generally 

6 high blood Pb levels (>20 μg/dL) that limit the relevance of findings to the general 

7 population, also pertain to the present findings. No correlation was observed between 

8 increasing blood Pb level and the number of chromosomal aberrations, although an 

9 association was observed between duration of exposure and chromosomal damage

10 (Grover et al., 2010; Pinto et al., 2000). Other studies reported no association between 

11 occupational exposure to Pb and chromosomal aberrations (Anwar and Kamal, 1988; 

12 Andreae, 1983). Smejkalova (1990) observed greater chromosomal damage and 

13 aberrations in children living in a heavily Pb-contaminated area of Czechoslovakia 

14 compared with children living in an area with less contamination, although the difference 

15 between the two areas was not statistically significant. Although blood Pb levels were 

16 statistically significantly higher in children living in the Pb-contaminated area than in 

17 children living in the less contaminated area, they were generally comparable (low 30s 

18 versus high 20s μg/dL, respectively), indicating there may not be enough of a dose 

19 contrast to detect a significant difference in aberration rates. 

20 The majority of animal studies investigating Pb-induced genotoxicity focused on the 

21 capacity of Pb to produce chromosomal damage. Fahmy (1999) exposed mice to 25-

22 400 mg/kg Pb-acetate i.p., either as a single exposure or repeatedly for 3, 5, or 7 days. 

23 Chromosomal damage was observed to increase in bone marrow cells (100-400 mg/kg) 

24 and spermatocytes (50-400 mg/kg) in a concentration-dependent manner after both 

25 exposure regimens. Pb nitrate was also observed to produce concentration-dependent 

26 chromosomal damage in mice exposed i.p. to a single exposure of 5, 10, or 20 mg/kg

27 (Dhir et al., 1992b). In a similar experiment, Dhir et al. (1990) exposed mice to 10, 20, or 

28 40 mg/kg Pb nitrate and saw an increase in chromosomal aberrations, although there was 

29 no concentration-dependent response as the response was similar in all concentrations 

30 tested. Nayak et al. (1989b) exposed pregnant mice to 100-200 mg/kg Pb nitrate via i.v. 

31 injection and observed no chromosomal gaps or breaks in dams or fetuses, but did report 

32 some karyotypic chromosomal damage and weak aneuploidy at the low exposure. In a 

33 similar experiment, low levels of chromosomal aberrations were observed in dams and 

34 fetuses injected with 12.5-75 mg/kg Pb nitrate, but there was no concentration-dependent 

35 response reported and few cells were analyzed (Nayak et al., 1989a). In rats given 

36 2.5 mg/100 g Pb-acetate i.p. daily for 5-15 days or 10-20 mg/100 g once and analyzed 

37 after 15 days, Pb-induced chromosomal aberrations were observed (Chakraborty et al., 
February 2012 5-44 Draft – Do Not Cite or Quote

1 1987). The above studies all suffer from the use of a route of exposure that may not be 

2 relevant to human environmental exposures. However, studies utilizing drinking water or 

3 dietary exposures also observed increases in chromosomal damage. Aboul-Ela (2002)

4 exposed mice to 200 or 400 mg/kg Pb-acetate by gavage for 5 days and reported that 

5 chromosomal damage was present in the bone marrow cells and spermatocytes of animals 

6 exposed to both exposure concentrations. Dhir et al. (1992a) also observed a 

7 concentration-dependent increase in chromosomal damage in mice exposed via gavage, 

8 albeit at much lower concentrations: either 5 or 10 mg/kg. Nehez et al. (2000) observed a 

9 Pb-induced increase in aneuploidy and percent of cells with damage after exposure to 

10 10 mg/kg administered by gavage 5 days a week for 4 weeks. In the only study that 

11 investigated dietary exposure, El-Ashmawy et al. (2006) exposed mice to 0.5%

12 Pb-acetate in feed, and observed an increase in abnormal cells and frequency of 

13 chromosomal damage.

14 In the few studies that investigated the capacity of Pb to induce chromosomal damage in 

15 human cell lines, Pb exposure did not induce chromosomal damage. Wise et al. (2005; 

16 2004) observed that Pb glutamate was not mutagenic in human lung cells exposed to 250-

17 2,000 μM for 24 hours. Pasha Shaik et al. (2006) observed that Pb nitrate did not increase 

18 chromosomal aberrations in primary lymphocytes (obtained from healthy volunteers) 

19 when incubated with 1,200 or 2,000 μM for 2 hours. Studies utilizing animal cell lines 

20 generally supported the finding of no Pb-induced chromosomal damage in human cell 

21 lines. Pb nitrate was found to induce no chromosomal damage in Chinese hamster ovary 

22 cells exposed to 500-2,000 μM for 24 hours (Wise et al., 1994), 3-30 μM for 2 hours (Lin 

23 et al., 1994), or 0.05-1 μM for 3-12 hours (Cai and Arenaz, 1998). Wise et al. (1994) did 

24 observe increased chromosomal damage in Chinese hamster ovary cells exposed to

25 1,000 μM Pb glutamate for 24 hours, but did not see any damage in cells exposed to 

26 higher concentrations (up to 2,000 μM).

5.2.7.4 Epigenetic Effects 

27 Epigenetic effects are heritable changes in gene expression resulting without changes in 

28 the underlying DNA sequence. A prime example of an epigenetic effect is the abnormal 

29 methylation of DNA, which could lead to altered gene expression and cell proliferation 

30 and differentiation. 

DNA Methylation

31 A single i.v. injection of 75 μmol/kg Pb nitrate resulted in global hypomethylation of 

32 hepatic DNA in rats (Kanduc et al., 1991). The observed hypomethylation in the liver 
February 2012 5-45 Draft – Do Not Cite or Quote

1 was associated with an increase in cell proliferation. A few additional studies in humans 

2 observed that higher bone Pb levels were associated with lower DNA methylation 

3 patterns in adults and cord blood of newborns (Wright et al., 2010; Pilsner et al., 2009).

4 Changes in DNA methylation patterns could potentially lead to dysregulation of gene 

5 expression and altered tissue differentiation.

Mitogenesis

6 Conflicting results have been reported regarding Pb-induced effects on mitogenesis, with 

7 both increased and decreased cell growth and mitogenesis. A discernable pattern of 

8 effects is difficult to detect when analyzing effects across human, in vivo animal, and in 

9 vitro studies. Only a few studies have investigated the potential epigenetic effects of Pb 

10 exposure in human populations indirectly by examining mitogenesis or the induction of 

11 cell proliferation, which can be a consequence of epigenetic changes. These studies

12 (Minozzo et al., 2010; Minozzo et al., 2004; Rajah and Ahuja, 1995) reported that Pb 

13 reduced mitogenesis in Pb-exposed workers (mean blood Pb levels: 35.4 μg/dL, 

14 59.4 μg/dL, and not reported, respectively). The observation of decreased cell division in 

15 exposed workers may indicate that cells suffered DNA damage and died during division, 

16 or that division was delayed to allow for DNA repair to occur. It is also possible that Pb 

17 exerts an aneugenic effect and arrests the cell cycle. 

18 Many studies have investigated the ability of Pb to induce mitogenesis in animal models, 

19 and have consistently shown that Pb nitrate can stimulate DNA synthesis and cell 

20 proliferation in the liver of animals exposed to 100 μM/kg via i.v. injection (Nakajima et 

21 al., 1995; Coni et al., 1992; Ledda-Columbano et al., 1992; Columbano et al., 1990; 

22 Columbano et al., 1987). Shinozuka et al. (1996) observed that Pb-induced hepatocellular 

23 proliferation was similar in magnitude to that induced by TNF-α at 100 μM/kg, and Pb 

24 was observed to induce TNF-α in glial and nerve cells and NF-κB, TNF-α, and iNOS in 

25 liver cells in exposed animals at 12.5 mg/kg and 100 μM/kg, respectively (Cheng et al., 

26 2002; Menegazzi et al., 1997). In the only study that examined Pb exposure via 

27 inhalation, exposure to 10,000 μM Pb-acetate for 4 weeks resulted in increased cellular 

28 proliferation in the lungs (Fortoul et al., 2005).

29 A great amount of research has been conducted investigating the potential effects of Pb 

30 on mitogenesis in human and animal cell cultures. In human cell cultures, Pb-acetate

31 inhibited cell growth in hepatoma cells (0.1-100 μM for 2-6 days) (Heiman and Tonner, 

32 1995) and primary oligodendrocyte progenitor cells (1 μM for 24 hours) (Deng and 

33 Poretz, 2002) but had no observable effects on growth in glioma cells (0.01-10 μM for 

34 12-72 hours) (Liu et al., 2000). Pb glutamate had no effect on cell growth in human lung 

35 cells, but did increase the mitotic index (250-1,000 μM for 24 hours) (Wise et al., 2005).
February 2012 5-46 Draft – Do Not Cite or Quote

1 The increase in the mitotic index was attributed to an arrest of the cell cycle at M-phase, 

2 and was not attributed to an actual increase of cell growth and proliferation. Gastaldo et 

3 al. (2007) also reported S and G2 cell cycle arrests in human endothelial cells following 

4 exposure to 100 μM Pb nitrate for 24 hours. Conflicting results with regard to DNA 

5 synthesis were reported, with a concentration-dependent inhibition of DNA synthesis 

6 reported in hepatoma cells (1-100 μM for 72 hours) (Heiman and Tonner, 1995), but an 

7 induction of synthesis observed in astrocytoma cells (1-50 μM for 24 hours) (Lu et al., 

8 2002).

9 In rat fibroblasts and epithelial cells, Pb-acetate, chloride, oxide, and sulfate were all 

10 observed to inhibit cell growth (10-1,000 μM for 1-7 days and 0.078-320 μM for 48 

11 hours, respectively) (Iavicoli et al., 2001; Apostoli et al., 2000). Iavicoli et al. (2001)

12 observed that in addition to inhibiting cell growth in rat fibroblasts, Pb-acetate caused 

13 GS/M and S-phase arrest. Pb-acetate decreased cell proliferation in mouse mesenchymal 

14 stem cells when administered at 20-100 μM for 48 hours (Kermani et al., 2008). Pb 

15 nitrate was alternatively reported to increase (Lin et al., 1994) and decrease (Cai and 

16 Arenaz, 1998) the mitotic index in Chinese hamster ovary cells exposed to 1 μM Pb 

17 nitrate. Lin et al. (1994) did not consider cell cycle arrest when measuring the mitotic 

18 index and did not observe a decrease at higher concentrations; in fact, the highest 

19 concentration tested, 30 μM, had a mitotic index equal to that in the untreated control 

20 cells. 

5.2.7.5 Gene Expression

21 Two animal studies have investigated the ability of Pb to alter gene expression in regard 

22 to phase I and II metabolizing enzymes. Suzuki et al. (1996) exposed rats to 100 μg/kg 

23 Pb-acetate or nitrate via i.p. injection and observed an induction of GST-P with both Pb 

24 compounds. The induction of GST-P by Pb was observed to occur on the transcriptional 

25 level and to be dependent on the direct activation of the cis-element GPEI enhancer. 

26 Degawa et al. (1993) reported that i.v. exposure to 20, 50, or 100 μmol/kg Pb nitrate 

27 selectively inhibited CYP1A2 levels. Pb was shown not to inhibit CYP1A2 by direct 

28 enzyme inhibition, but rather to decrease the amount of CYP1A2 mRNA. In contrast, 

29 Korashy and El Kadi (2004) observed that exposure of murine hepatoma cells to 10-100 

30 μM Pb nitrate for 24 hours increased the amount of CYP1A1 mRNA while not 

31 influencing the activity of the enzyme. NAD(P)H:quinone oxidoreductase and GST Ya 

32 activities and mRNA levels were increased after exposure to Pb. Incubation of primary 

33 human bronchial epithelial cells with 500 μg/L Pb-acetate for 72 hours resulted in the 

34 up-regulation of multiple genes associated with cytochrome P450 activity, glutathione 
February 2012 5-47 Draft – Do Not Cite or Quote

1 metabolism, the pentose phosphate pathway, and amino acid metabolism (Glahn et al., 

2 2008).

3 Additional animal studies provide further evidence that exposure to Pb compounds can 

4 perturb gene expression. Zawia and Harry (1995) investigated whether the observed Pb5 induced disruption of myelin formation in rat pups exposed postnatally was due to altered 

6 gene expression. In pups exposed to 0.2% Pb-acetate via lactation from postnatal day 

7 (PND)1-20, the expression of proteolipid protein, a major structural constituent of 

8 myelin, was statistically significantly elevated at PND20, compared to controls. The 

9 expression of another structural element of myelin, myelin basic protein, was similarly 

10 elevated in exposed animals, although not significantly so. The expression of both genes 

11 returned to control levels 5 days following the termination of exposure. These data 

12 suggest that altered gene expression in structural myelin proteins due to Pb exposure may 

13 be responsible for observed alterations in abnormal conduction of nerve impulses. Long 

14 et al. (2011) investigated the Pb-induced induction of ABCC5, an ATP-binding cassette 

15 transporter, in embryonic and adult zebrafish. In the initial in vitro portion of the study, 

16 exposure of zebrafish fibroblasts to 20 μM Pb nitrate for 24 hours significantly increased 

17 the induction of ABBC5 mRNA 2.68-fold over controls. Similar levels of induction were 

18 observed when embryonic zebrafish were exposed to 5 μM 24 to 96 hours; specifically, 

19 induction of ABCC5 was seen in the livers of developing embryos. In adult fish, 

20 induction of ABCC5 was observed in the brains, intestines, and kidneys of exposed fish, 

21 but decreased in their livers. Induction of ABCC5 was observed to attenuate the toxicity 

22 of Cd, but not Hg or As, in developing embryos, the attenuation of Pb-induced toxicity 

23 was not investigated. However, these findings indicate that increased expression of 

24 ABCC5 due to heavy metal exposure may play a part in cellular defense mechanisms.

5.2.7.6 Apoptosis

25 Occupational exposure to Pb and induction of apoptosis was investigated in a few studies. 

26 One study directly reported that exposure to Pb increased apoptosis compared to 

27 nonexposed controls (Minozzo et al., 2010), whereas the others reported that two early 

28 indicators of apoptosis, karyorrhexis and karyolysis, were elevated in exposed workers

29 (Grover et al., 2010; Khan et al., 2010b). Pb nitrate was also observed to induce apoptosis 

30 in the liver of exposed animals (Columbano et al., 1996; Nakajima et al., 1995).

31 Apoptosis was observed in rat fibroblasts exposed to Pb-acetate and rat alveolar 

32 macrophages exposed to Pb nitrate (Iavicoli et al., 2001; Shabani and Rabbani, 2000).

33 Observation of Pb-induced apoptosis may represent the dysregulation of genetically34 controlled cell processes and tissue homeostasis.
February 2012 5-48 Draft – Do Not Cite or Quote

5.2.8 Summary

1 The diverse health effects of Pb are mediated through multiple, interconnected modes of 

2 action. Each of the modes of action discussed has the potential to contribute to the 

3 development of a number of Pb-induced health effects (Table 5-2). While this section 

4 draws from older literature as well as newer lines of evidence, the inclusion of new 

5 evidence does not qualitatively change the conclusions regarding individual modes of 

6 action. However, the new evidence strengthens the conclusions. Evidence for the 

7 majority of these modes of action is observed at low blood Pb levels in humans, between 

8 2 and 17 μg/dL, with supporting evidence from in vitro assays. As many of these studies 

9 are examining adults with likely higher past than current Pb exposures, uncertainty exists 

10 as to the Pb exposure level, duration, and timing leading to these blood Pb levels 

11 associated with these modes of action. These observable effect levels are reflective of the 

12 data and methods available and do not imply that these modes of action are not acting at 

13 lower Pb exposure or blood Pb levels or that these concentrations represent the threshold 

14 of the effect. The observable effect levels in humans, reported in Table 5-2, are drawn 

15 from the available data; and, do not imply that theses modes of action are not acting at 

16 lower exposure levels or that these doses represent the threshold of the effect. Also, the 

17 data presented in this table does not inform regarding the exposure frequency and 

18 duration required to elicit a particular MOA.
February 2012 5-49 Draft – Do Not Cite or Quote

Table 5-2 MOAs, their related health effects, and information on concentrations 

eliciting the MOAs

Mode of Action

[Related Health Effects 

(ISA section)]

Concentrations or doses (Conditions)a

Blood Pb Dose

Altered Ion Status

[All Health Effects of Pb]

3.5 μg/dL

(mean in cord blood; association with cord blood Ca2+- ATPase pump activity)

Huel et al. (2008)

0.00005 μM free Pb2+

(In vitro; 30 minutes; calmodulin activation assay)

Kern et al. (2000)

Protein Binding

[Renal (5.5), Heme Synthesis and RBC 

Function (5.7)]

17.0 μg/dL 

(concurrent mean in adult workers with wildtype 

metallothionein expression; increased BP susceptibility) 

Chen et al. (2010a)

50 μM Pb glutamate

(In vitro; 24 hours; increased nuclear protein in 

neurological cell)

Klann and Shelton (1989)

Oxidative Stress

[All Heath Effects of Pb]

5.4 μg/dL

(concurrent mean in adult male workers; decreased CAT 

activity in blood) 

Conterato et al. (In Press)

0.1 μM Pb-acetate

(In vitro; 48 hours; decreased cellular GSH in 

neuroblastoma cells)

Chetty et al. (2005)

Inflammation

[Nervous System (5.3), Cardiovascular 

(5.4), Renal (5.5), Immune (5.6), 

Respiratory (5.6.4), Hepatic (5.9.1)]

2.5 μg/dL

(concurrent minimum in adult males; increased serum 

TNF-α and blood WBC count)

Kim et al. (2007)

0.01 μM Pb-acetate

(In vitro; 48 hours; increased cellular PGE2 in 

neuroblastoma cells)

Chetty et al. (2005)

Endocrine Disruption

[Reproductive and Developmental Effects

(5.8), Endocrine System 5.9.3), Bone and 

Teeth (5.9.4)]

1.7 μg/dL

(concurrent minimum in women with both ovaries 

removed; increased serum FSH)

Krieg (2007)

10 μM Pb nitrate

(In vitro; 30 minutes; displaced GHRH binding to rat 

pituitary receptors) 

Lau et al. (1991)

Cell Death/Genotoxicity

[Cancer (5.10), Reproductive and 

Developmental Effects (5.8), Bone and 

Teeth (5.9.4)]

3.3 μg/dL

(concurrent median in adult women; increased rate of 

HPRT mutation frequency)

Van Larebeke et al. (2004)

0.03 μM Pb-acetate

(In vitro; 18 hours; increased formation of micronuclei)

Bonacker et al. (2005)

a

This table provides examples of studies that report effects with low doses or concentration; they are not the full body of evidence used to characterize the weight 

of the evidence. In addition, the levels cited are reflective of the data and methods available and do not imply that these modes of action are not acting at lower 

Pb exposure or blood Pb levels or that these doses represent the threshold of the effect. Additionally, the blood concentrations and doses (indicating Pb exposure 

concentrations from in vitro systems) refer to the concentrations and doses at which these modes of action were observed. While the individual modes of action 

are related back to specific health effects sections (e.g., Nervous System, Cardiovascular), the concentrations and doses given should not be interpreted as 

levels at which those specific health effects occur.

The alteration of cellular ion status (including disruption of Ca2+ 1 homeostasis, altered ion 

2 transport mechanisms, and perturbed protein function through displacement of metal 

3 cofactors) appears to be the major unifying mode of action underlying all subsequent 

modes of action (Figure 5-1). Pb will interfere with endogenous Ca2+ 4 homeostasis, 

necessary as a cell signal carrier mediating normal cellular functions. [Ca2+ 5 ]i has been 

6 shown to increase after Pb exposure in a number of cell types including bone, 

7 erythrocytes, brain cells, and white blood cells, due to the increased flux of extracellular 

Ca2+ 8 into the cell. This disruption of ion transport is due in part to the alteration of the 

activity of transport channels and proteins, such as Na+-K+ 9 ATPase and voltage-sensitive 

Ca2+ 10 channels. Pb can interfere with these proteins through direct competition between Pb 

11 and the native metals present in the protein metal binding domain or through disruption 

of proteins important in Ca2+ 12 -dependent cell signaling, such as PKC or calmodulin. 
February 2012 5-50 Draft – Do Not Cite or Quote

Disruption of ion transport not only leads to altered Ca2+ 1 homeostasis, it can also result in 

2 perturbed neurotransmitter function. Pb has been shown to be able to displace metal ions, 

such as Zn, Mg, and Ca2+ 3 , from proteins due to the flexible coordination number of Pb 

4 and multiple ligand binding ability, leading to abnormal conformational changes to 

5 proteins and altered protein function. Evidence for this metal ion displacement and 

6 protein perturbation has been shown at picomolar concentrations of Pb. Additional effects 

7 of altered cellular ion status are the inhibition of heme synthesis and decreased cellular 

8 energy production due to perturbation of mitochondrial function.

9 Although Pb will bind to proteins within cells through interactions with side group 

10 moieties, thus potentially disrupting cellular function, protein binding of Pb may 

11 represent a mechanism by which cells protect themselves against the toxic effects of Pb. 

12 Intranuclear and intracytosolic inclusion body formation has been observed in the kidney, 

13 liver, lung, and brain following Pb exposure. A number of unique Pb binding proteins 

14 have been detected, constituting the observed inclusion bodies. The major Pb binding 

15 protein in blood is ALAD with carriers of the ALAD-2 allele potentially exhibiting 

16 higher Pb binding affinity. Additionally, metallothionein is an important protein in the 

17 formation of inclusion bodies and mitigation of the toxic effects of Pb.

18 A second major mode of action of Pb is the development of oxidative stress, due in many 

19 instances to the antagonism of normal metal ion functions. The origin of oxidative stress 

20 produced after Pb exposure is likely a multipathway process, resulting from oxidation of 

21 δ-ALA, NAD(P)H oxidase activation, membrane and lipid peroxidation, and antioxidant 

22 enzyme depletion. Through the inhibition of δ-ALAD due to displacement of Zn, 

23 accumulated δ-ALA goes through an auto-oxidation process to produce ROS. 

24 Additionally, Pb can induce the production of ROS through the activation of NAD(P)H 

25 oxidase. Pb-induced ROS can interact with membrane lipids to cause a membrane and 

26 lipid peroxidation cascade. Enhanced lipid peroxidation can also result from Pb 

potentiation of Fe2+ 27 initiated lipid peroxidation and alteration of membrane composition 

28 after Pb exposure. Increased Pb-induced ROS will also sequester and inactivate 

29 biologically active ·NO, leading to the increased production of the toxic product 

30 nitrotyrosine, increased compensatory NOS, and decreased sGC protein. Pb-induced 

31 oxidative stress not only results from increased ROS production but also through the 

32 alteration and reduction in activity of the antioxidant defense enzymes. The biological 

33 actions of a number of these enzymes are antagonized due to the displacement of the 

34 protein functional metal ions by Pb. 

35 In a number of organ systems Pb-induced oxidative stress is accompanied by 

36 misregulated inflammation. Pb exposure will modulate inflammatory cell function, 

37 production of pro-inflammatory cytokines and metabolites, inflammatory chemical 
February 2012 5-51 Draft – Do Not Cite or Quote

1 messengers, and pro-inflammatory signaling cascades. Cytokine production is skewed 

2 toward the production of pro-inflammatory cytokines like TNF-α and IL-6 as well as 

3 toward the promotion of a Th2 response and suppression of a Th1 response accompanied 

4 by decreased production of related cytokines.

5 Evidence indicates that Pb is a potent endocrine disrupting chemical. Pb will disrupt the 

6 HPG axis evidenced by a decrease in serum hormone levels, such as FSH, LH, 

7 testosterone, and estradiol. Pb interacts with the hypothalamic-pituitary level hormone 

8 control causing a decrease in pituitary hormones, altered growth dynamics, inhibition of 

9 LH secretion, and reduction in StAR protein. Pb has also been shown to alter hormone 

10 receptor binding likely due to interference of metal cations in secondary messenger 

11 systems and receptor ligand binding and through generation of ROS. Pb also may disrupt 

12 the HPT axis by alteration of a number of thyroid hormones, possibly due to oxidative 

13 stress. However, the results of these studies investigating HPT are mixed and require 

14 further investigation.

15 The association of Pb with increased genotoxicity and cell death has been investigated in 

16 humans, animals, and cell models. Occupational Pb exposure in humans has been 

17 associated with increased DNA damage, however lower blood Pb and exposure levels 

18 have been associated with these effects in experimental animals and cells. Results vary on 

19 the effect of Pb on DNA repair activity, however a number of studies reported decreased 

20 repair processes following Pb exposure. There is evidence of mutagenesis and 

21 clastogenicity in highly-exposed humans, however weak evidence has been shown in 

22 animals and cell based systems. Human occupational studies provide limited evidence for 

23 micronucleus formation (blood Pb levels >10 μg/dL), supported by Pb-induced effects in 

24 both animal and cell studies at higher exposure levels. Animal studies have also provided 

25 evidence for Pb-induced chromosomal aberrations. The observed increases in 

26 clastogenicity may be the result of increased oxidative damage to DNA due to Pb 

27 exposure, as co-exposures with antioxidants ameliorate the observed toxicities. Limited 

28 evidence of epigenetic effects is available, including DNA methylation, mitogenesis, and 

29 gene expression. Pb may alter gene expression by displacing Zn from multiple 

30 transcriptional factors, thus perturbing their normal cellular activities. Consistently 

31 positive results have provided evidence of increased apoptosis following Pb exposure. 

32 Similar to Pb, other polyvalent metal ions (e.g., Cd, Cr, Be, Ba, Se, Sr, As, Al, Cu) have 

33 demonstrated molecular mimicry and displacement of biological cation (Garza et al., 

34 2006). In this manner, these metal ions share with Pb a common central mode of action of 

35 disruption of ion status. Specifically, these metals have been shown to disrupt cellular 

processes as diverse as Ca2+ 36 homeostasis, cell signaling, neurotransmitter release, cation 

37 membrane channel function, protein-DNA binding, and cellular membrane structure
February 2012 5-52 Draft – Do Not Cite or Quote

1 (Pentyala et al., 2010; Huang et al., 2004; Atchison, 2003; Jehan and Motlag, 1995; 

2 Richardt et al., 1986; Cooper and Manalis, 1984; Habermann et al., 1983). Additionally,

3 presumably through their shared central mode of action, some of these metal ions also 

4 display corresponding downstream modes of actions such as oxidative stress, apoptosis, 

5 and genotoxicity (Jomova and Valko, 2011; Jomova et al., 2011; Matović et al., 2011; 

6 Agarwal et al., 2009; Méndez-Gómez et al., 2008; Rana, 2008; Hengstler et al., 2003).

7 Overall, Pb-induced health effects can occur through a number of interconnected modes 

8 of action that generally originate with the alteration of ion status.

5.3 Nervous System Effects

5.3.1 Introduction

9 The 2006 Pb AQCD concluded that the “overall weight of the available evidence 

10 provides clear substantiation of neurocognitive decrements being associated in young 

11 children with blood-Pb concentrations...” (U.S. EPA, 2006b). This conclusion was based 

12 on evidence from several prospective and cross-sectional studies conducted in diverse 

13 populations and after adjusting for potential confounding by socioeconomic status (SES), 

14 parental intelligence, and caregiving environment. This association was substantiated in a 

15 pooled analysis of children, 5 to 10 years of age, participating in seven prospective 

16 studies (Boston, MA; Cincinnati, OH; Rochester, NY; Cleveland, OH; Mexico City, 

17 Mexico; Port Pirie, Australia; and Kosovo, Yugoslavia) (Lanphear et al., 2005).

18 Associations between blood Pb levels and decrements in intelligence quotient (IQ), 

19 mental development, memory, and other specific indices of cognitive function in children 

20 ages 6 months to 17 years were most strongly indicated in children with population mean 

21 blood Pb levels (measured at various lifestages) in the range of 5-10 μg/dL; however, 

22 several results indicated associations in groups of children (ages 2-10 years) with mean 

23 blood Pb levels in the range of 3-5 μg/dL (Bellinger, 2008; Canfield, 2008; Hornung, 

24 2008; Rojo-Tellez, 2008). Based on fewer available studies, the 2006 Pb AQCD 

25 described consistent associations of blood Pb levels with behavioral outcomes, including 

26 inattention, and antisocial and delinquent behavior assessed in children ages 6 to 13 years

27 (U.S. EPA, 2006b).

28 Toxicological studies provided coherence with similarly consistent findings for Pb29 induced impairments in learning and behavior in rodents and monkeys (U.S. EPA, 

30 2006b). In contrast with studies in children, Pb exposure was not found consistently to 

31 affect memory of animals. Effects on learning were largely demonstrated as poorer 

32 performance on Morris water maze and discrimination reversal tasks, and effects on 
February 2012 5-53 Draft – Do Not Cite or Quote

1 behavior were largely demonstrated as distractibility, impulsivity, and insensitivity to 

2 changes in reinforcement. These effects in animals were predominantly found with Pb 

3 exposures that resulted in blood Pb levels between 20-50 μg/dL; however, some studies 

4 observed learning and behavioral impairments in animals with steady-state blood Pb 

5 levels of 10-15 μg/dL (Cory-Slechta, 1994; Altmann et al., 1993; Rice and Karpinski, 

6 1988; Gilbert and Rice, 1987). Toxicological studies also provided biological plausibility 

7 by characterizing modes of action for Pb-induced nervous system effects. In particular, 

8 toxicological evidence for Pb exposure interfering with neuronal metabolism at the 

9 cellular and histological level (e.g., synaptic architecture during development, 

10 neurotransmitter release, glia, neurite outgrowth, the blood brain barrier, and oxidative 

11 stress), provided biological plausibility for blood Pb levels in children being causally 

12 associated with deficits in multiple functional domains such as cognitive function, motor 

13 function, memory, mood, and behavior. Additional biological plausibility was provided 

14 by associations observed of childhood blood Pb levels with changes indicative of 

15 neuronal damage and altered brain physiology assessed in young adults using magnetic 

16 resonance imaging techniques (Yuan et al., 2006; Cecil et al., 2005; Meng et al., 2005; 

17 Trope et al., 2001).

18 A common finding across several epidemiologic studies of children was a supralinear 

19 concentration-response relationship between blood Pb level and neurocognitive deficits, 

20 i.e., a larger decrement in neurocognitive function per unit increase in blood Pb levels in 

21 children in the lower range of the population blood Pb level distribution (Kordas et al., 

22 2006; Schnaas et al., 2006; Tellez-Rojo et al., 2006; Lanphear et al., 2005; Rothenberg 

23 and Rothenberg, 2005; Bellinger and Needleman, 2003; Canfield et al., 2003a). Most of 

24 these epidemiologic results were based on the analysis of concurrent blood Pb levels and 

25 a cut-point of 10 μg/dL to define lower and higher blood Pb levels. These findings were 

26 corroborated by findings from analyses of the pooled cohort data indicating that a 

27 nonlinear relationship fit the data better than a linear relationship did (Lanphear et al., 

28 2005; Rothenberg and Rothenberg, 2005). Consistent with epidemiologic findings, 

29 toxicological studies observed nonlinear Pb concentration-response relationships for 

30 outcomes such as neuronal activation (Lewis and Pitts, 2004), neurogenesis (Gilbert et 

31 al., 2005), and retinal responses (Fox et al., 1991; Fox and Farber, 1988; Fox and Chu, 

32 1988).

33 Another area of focus included the comparison of various lifestages of Pb exposure in 

34 terms of risk of neurodevelopmental deficits. Toxicological studies clearly demonstrated 

35 that in utero with or without early postnatal exposure to Pb was an especially sensitive 

36 window for Pb-induced neurodevelopmental effects. Nonetheless, not all endpoints in 

37 animal toxicology studies had a single defined window of sensitivity but instead were 

38 shown to be affected by exposures at multiple periods during the lifespan of the 
February 2012 5-54 Draft – Do Not Cite or Quote

1 organism. Epidemiologic studies observed cognitive deficits in children ranging from 2 to 

2 10 years of age in association with prenatal, peak childhood, cumulative childhood, and 

3 concurrent blood Pb levels. Although examined in few studies, tooth or bone Pb levels 

4 were associated with cognitive and behavioral deficits in children and adolescents

5 (Wasserman et al., 2003; Greene and Ernhart, 1993; Bellinger et al., 1991; Needleman et 

6 al., 1979), also pointing to an effect of cumulative childhood Pb exposure. Among studies 

7 of children (ages 2-10 years) that examined blood Pb levels measured at multiple 

8 lifestages, several found that concurrent blood Pb was associated with an equal or larger 

9 decrement in IQ (Chen et al., 2005; Lanphear et al., 2005; Wasserman et al., 1994; 

10 Dietrich et al., 1993b; Bellinger et al., 1992). A common limitation of epidemiologic 

11 studies of children was the high correlation among blood Pb levels at different ages, 

12 making it difficult to ascertain which developmental periods of Pb exposure were 

13 associated with the greatest risk of neurodevelopmental decrements (Lanphear et al., 

14 2005). The issue of persistence of the neurodevelopmental effects of Pb exposure also 

15 was considered, with some evidence suggesting that the associations of biomarkers of 

16 early childhood Pb exposure (e.g., deciduous tooth, blood at age 2 or 6 years) with 

17 neurodevelopmental outcomes persisted into adolescence and young adulthood (Ris et al., 

18 2004; Tong et al., 1996; Needleman et al., 1990). Some studies in rats and monkeys also 

19 demonstrated that the effects of in utero and early postnatal Pb exposures on 

20 neurodevelopmental outcomes persisted into adulthood (Cory-Slechta, 1994; Altmann et 

21 al., 1993; Rice and Karpinski, 1988; Gilbert and Rice, 1987).

22 In epidemiologic studies of adults, a range of nervous system effects (e.g., impaired 

23 memory, attention, reaction time, visuomotor tasks and reasoning, alterations in visual or 

24 brainstem evoked potentials, postural sway) were mostly clearly indicated in Pb-exposed 

25 workers with blood Pb levels in the range of 14 to 40 μg/dL (Iwata et al., 2005; Bleecker 

26 et al., 1997; Baker et al., 1979; Cantarow and Trumper, 1944). In the limited literature 

27 examining nonoccupationally-exposed adults, the weight of evidence supported 

28 associations of bone Pb levels with cognitive function (Weisskopf et al., 2004; Wright et 

29 al., 2003b) but not concurrent blood Pb levels (Krieg et al., 2005; Nordberg et al., 2000; 

30 Payton et al., 1998; Muldoon et al., 1996). These findings suggested that rather than 

31 recent exposures, past or cumulative Pb exposures contributed to cognitive deficits in 

32 nonoccupationally-exposed adults. With regards to neurodegenerative diseases, whereas a 

33 few toxicological studies demonstrated Pb-induced amyloid plaques commonly 

34 associated with Alzheimer’s disease pathophysiology (Basha et al., 2005; Zawia and 

35 Basha, 2005), epidemiologic studies did not indicate that Pb exposure was associated 

36 with Alzheimer's Disease in adults. Pb biomarker (blood or bone) levels were 

37 inconsistently associated with amyotrophic lateral sclerosis (ALS) in adults in the general 

38 population; however, some case-control studies found that history of occupational Pb 

39 exposure was more prevalent among ALS cases than controls (Kamel et al., 2002; 
February 2012 5-55 Draft – Do Not Cite or Quote

1 Chancellor et al., 1993). Associations were reported for essential tremor and symptoms of 

2 anxiety and depression, but each was examined in only a few studies.

3 As discussed throughout this section, recent epidemiologic and toxicological studies 

4 continued to demonstrate associations of Pb exposure and biomarkers of Pb exposure 

5 with nervous system effects. The weight of evidence continued to be derived from 

6 associations observed of Pb exposure and blood Pb levels in young animals and children, 

7 respectively, with cognitive function decrements, inattention, and impulsivity. Expanding 

8 upon the previous body of evidence, several recent studies in children found similar 

9 associations with lower population mean (or quantile) blood Pb levels (1-5 μg/dL). 

10 Whereas previous evidence was inconsistent, several new studies in children reported 

11 associations between concurrent blood Pb levels and attention deficit hyperactivity 

12 disorder (ADHD). Recent studies in adults focused primarily on cognitive function 

13 decrements but also provided additional evidence for Pb-associated mood disorders, 

14 ALS, Parkinson's Disease, and essential tremor. New toxicological studies expanded 

15 evidence for the effects of prenatal and postnatal Pb exposure on learning, memory, and 

16 attention and provided insight into the contribution of social stress to this paradigm. New 

17 or expanded areas of toxicological research related to Pb exposure included mood 

18 disorders, neurofibrillary tangle formation, and adult dementia after early life Pb 

19 exposures. Historically important areas of toxicological research were further expanded 

20 with recent findings for Pb-induced effects on neurotransmitters, synapses, glia, neurite 

21 outgrowth, the blood brain barrier, and oxidative stress. The data detailed in the 

22 subsequent sections continue to enhance the understanding of nervous system effects 

23 associated with Pb exposure.

5.3.2 Cognitive Function and Learning

5.3.2.1 Epidemiologic Studies of Cognitive Function in Children

24 Epidemiologic studies have assessed global cognitive function most frequently by full25 scale IQ (FSIQ) and its verbal and performance subscale components in children ages 3 

26 to 17 years and by the Bayley Scales of Infant Development in children ages 6 months to 

27 3 years. These indices have strong psychometric properties and are among the most 

28 rigorously standardized measures. A large body of evidence also comprises associations 

29 of blood Pb levels with specific cognitive abilities, including memory and learning, 

30 executive function, language, and visuospatial processing. These specific indices of 

31 cognitive function are reflected in global measures of intelligence and also are more 

32 comparable to tests of learning and memory in animals. Fewer studies have examined 
February 2012 5-56 Draft – Do Not Cite or Quote

1 academic performance and achievement; however, these outcomes may provide 

2 information on the impact of Pb exposure on life success. The epidemiologic evidence for 

3 each of these categories of outcomes was evaluated separately in order of increasing 

4 weight of evidence. Emphasis was placed on prospective studies with repeated 

5 assessments of blood Pb levels and cognitive function and on studies examining blood Pb 

6 levels similar to those measured in contemporary U.S children (i.e., less than 5 μg/dL), 

7 and younger children whose blood Pb levels are less influenced by higher past Pb 

8 exposures.

9 Many factors have been shown to influence the intellectual abilities of children, including 

10 parental SES, parental education, parental IQ, quality and stability of the caregiving 

11 environment, nutritional status, and birth weight (Nation and Gleaves, 2001; Wasserman 

12 and Factor-Litvak, 2001). These and other influences on neurodevelopment often are 

13 correlated with blood Pb levels. Thus, due to their association with both blood Pb level 

14 and outcome, these other risk factors may potentially confound (i.e., bias due to an

15 association with blood Pb level and causal association with the outcome) the associations 

16 observed between blood Pb level and indices of cognitive function. In the evaluation of 

17 the effect of Pb independent from the effects of the other variables, greater weight was 

18 given to studies that accounted for potential confounding in the study design or in 

19 statistical analyses. A detailed discussion of the collective weight of evidence for the 

20 independent associations between Pb exposure and nervous system effects in relation to 

21 the adequacy of control for confounding by other risk factors is located in Section 5.3.11.

Full-scale IQ in Children

22 Several longitudinal cohort studies were initiated in the 1980s in order to address 

23 limitations of cross-sectional studies, including establishing a temporal association 

24 between blood Pb levels and cognitive outcomes, examining the persistence of cognitive 

25 deficits to older ages, and comparing risk estimates among blood Pb levels measured at 

26 different lifestages. Moreover, cooperation among investigators to adopt similar 

27 assessment protocols facilitated pooled and meta-analyses and comparison of results 

28 across populations that differed in the range of blood Pb levels, race/ethnicity, and SES. 

29 Individual cohort studies in diverse populations were consistent in demonstrating 

30 associations between higher blood Pb and lower FSIQ in populations of school-aged 

31 children with mean blood Pb levels in the range of 5 to 10 μg/dL (Schnaas et al., 2006; 

32 Bellinger and Needleman, 2003; Canfield et al., 2003a; Wasserman et al., 1997; Dietrich 

33 et al., 1993a; Baghurst et al., 1992; Bellinger et al., 1987) (Figure 5-2 and Table 5-3). In 

34 analyses restricted to children in the lower range of the blood Pb distribution (e.g., < 

35 10 μg/dL), associations were observed in groups of children with mean blood Pb levels 

36 3-4 μg/dL (Bellinger, 2008; Canfield, 2008; Hornung, 2008). Across cohort studies, 
February 2012 5-57 Draft – Do Not Cite or Quote

1 associations were observed with concurrent, prenatal, early childhood (e.g., age 2 years), 

2 and childhood average blood Pb levels (e.g., 0-8 year average) (Figure 5-2 and Table 

3 5-3). These findings were substantiated in a pooled analysis of seven prospective studies 

4 (Boston, MA; Cincinnati, OH; Rochester, NY; Cleveland, OH; Mexico City, Mexico; 

5 Port Pirie, Australia; and Kosovo, Yugoslavia) by Lanphear et al. (2005) as well as 

6 multiple meta-analyses that included both prospective and cross-sectional studies

7 (Pocock et al., 1994; Schwartz, 1994; Needleman and Gatsonis, 1990).

Note: Studies generally are presented in ascending order of mean blood Pb level. To facilitate comparisons among effect estimates 

across studies with different distributions of blood Pb levels and model structures (e.g., linear, log-linear), effect estimates are 

standardized to a 1 μg/dL increase in blood Pb level within the 10th to 90th percentile interval. The percentiles are estimated using 

various methods and are only approximate values. Effect estimates are assumed to be linear within the 10th to 90th percentile 

interval of blood Pb level. The various tests used to measure FSIQ are scored on a similar scale (approximately 40-160). Black 

diamonds, blue circles, orange triangle, and gray squares represent associations with concurrent, earlier childhood, prenatal, and 

lifetime average blood Pb levels, respectively. 

Figure 5-2 Associations of blood Pb levels with full-scale IQ (FSIQ) among 

children.

Study

Kim et al. (2009)

Chiodo et al. (2007)

Chiodo et al. (2004)

Jusko et al. (2011)

Canfield et al. (2003)

Bellinger et al. (1992)

Kordas et al. (2011)

Lanphear et al. (2005)

Min et al. (2009)

Min et al. (2009)

Min et al. (2009)

Schnaas et al. (2006)

Kordas et al. (2011)

Roy et al. (2011)

Dietrich et al. (1993)

Tong et al. (1996)

Wasserman et al. (1997)

Baghurst et al. (1992)

Blood Pb Timing

Concurrent

Concurrent

Concurrent

Concurrent

Concurrent

2 yr

Prenatal (cord)

Concurrent

Concurrent

4 yr

4 yr

Prenatal (maternal)

Concurrent

Concurrent

Concurrent

0 to 11-13 yr avg

0 to 7 yr avg

0 to 8 yr avg

Blood Pb Mean

(SD) (μg/dL)

1.73 (0.80)

5.0 (3.0)

5.4 (3.3)

5.0 (3.3)

5.8 (4.1)

6.5 (4.9)

6.6 (3.3)

6.9 (1.2)

7.0 (4.1)

7.0 (4.1)

7.0 (4.1)

7.8 (geometric)

8.7 (4.4)

11.4 (5.4)

11.8 (6.3) (Age 5)

14.0 (1.2) geometric

16.2 (geometric)

16.6 (mean of 25th-50th)

Blood Pb 10th-90th

percentile (μg/dL)

<1-2.8

2.1-8.7

2.3-9.5

1.9-9.0

2.1-10.7

2.2-12.3

1.2-2.4

3.4-25.4

3.0-12.1

3.0-12.1

3.0-12.1

3.2-19.1

1.4-2.7

5.8-18.4

5.5-19.7

12.7-18.1

6.0-44.2

10-27.5

FSIQ age (yr)

8-11

7

7.5

6

5

10

4

4.8-10

4

9

11

6-10

4

3-7

6.5

11-3

7

7-8

-1.5 -1 -0.5 0

Change in full scale IQ per 1 μg/dL increase in the 10th to 90th percentile

of blood Pb level (95% CI)
February 2012 5-58 Draft – Do Not Cite or Quote

Table 5-3 Additional characteristics and quantitative results for studies 

represented in Figure 5-2

Study

Population/

Location

Blood Pb Data 

(μg/dL) Statistical Analysis FSIQ Assessmenta Effect Estimate(95% CI)b,c

Kim et al.

(2009a)

279 children in Seoul, 

Seongnam, Ulsan, and 

Yeoncheon, Korea, ages 

8-11 yr in April-December 

2007

Concurrent

Mean (SD): 1.73 (0.80)

10th-90th: < 1-2.8

Log linear regression model 

adjusted for age, sex, maternal 

education, paternal 

education, yearly income, maternal 

smoking during pregnancy, indirect 

smoking after birth, birth weight,

maternal age

KEDI-WISC at ages 8-11 yr -0.62 (-1.05, -0.19)

Chiodo et al.

(2007)

506 African-American 

children in Detroit, MI area 

followed from birth 

(1989-1991) to age 7 yr.

Large proportions of 

children with prenatal 

exposure to cocaine or 

marijuana 

Concurrent

Mean (SD): 5.0 (3.0)

10th-90th: 2.1-8.7

Regression model adjusted for 

prenatal alcohol and drug use, SES, 

Symptom Checklist (component of 

HOME score), maternal IQ

WISC-III at age 7 yr -0.19 (-0.30, -0.08)b

Chiodo et al.

(2004)

246 African-American 

children in Detroit, MI area

followed from birth (not 

reported) to age 7.5 yr.

Large proportions of 

children with prenatal 

exposure to cocaine or 

marijuana

Concurrent

Mean (SD): 5.4 (3.3)

10th-90th: 2.3-9.5

Log linear regression model 

adjusted for SES, education, 

number of children <18 yr, HOME 

score, maternal vocabulary test 

score, sex, parity, family 

environment scale

WISC-III at age 7.5 yr -0.22 (-0.38, -0.05)b

Jusko et al.

(2011)

194 children in Rochester, 

NY followed from age 6 mo 

(1994-1995) to age 6 yr.

Concurrent

Mean (SD): 5.0 (3.3)

10th-90th: 1.9-9.0

Linear regression model adjusted 

for sex, birth weight, transferrin 

saturation, maternal race, maternal 

IQ, maternal education, HOME 

score, family income, and maternal 

prenatal smoking

WPPSI-R at age 6 yr -0.66 (-1.26, -0.06)

Canfield et al.

(2003a)

172 children in Rochester, 

NY born 1994-1995 

followed from age 6 mo to 

age 5 yr.

Concurrent

Mean (SD): 5.8 (4.1)

10th-90th: 2.1-10.7

Linear regression model adjusted 

for sex, maternal race, prenatal 

smoking, maternal education, child 

iron status, household income, 

maternal IQ, HOME score, birth 

weight

Stanford-Binet at age 5 yr -0.61 (-0.99, -0.24)

Bellinger et al.

(1992)

148 children in the Boston, 

MA area followed from birth 

(1979-1981) to age 15-17 

yr.

Early childhood (age 2 

yr)

Mean (SD): 6.5 (4.9)

10th-90th: 2.2-12.3

Linear regression model adjusted 

for HOME score (age 10 and 5 yr),

child stress, race, maternal IQ, 

SES, sex, birth order, marital status

WISC-R at age 10 yr -0.58 (-0.99, -0.18)

Lanphear et 

al. (2005)

1,333 children pooled from 

Boston, Cincinnati, 

Cleveland, Mexico City, Port 

Pirie, Rochester, and 

Yugoslavia cohorts

Concurrent

Mean (SD): 6.9 (1.2)

10th-90th: 3.4-25.4

Log linear regression model 

adjusted for HOME score, birth 

weight, maternal IQ, maternal 

education

FSIQ measured at ages 5-10 yr -0.25 (-0.34, -0.15)

Min et al.

(2009)

267 primarily AfricanAmerican children in the 

Cleveland, OH area 

followed from birth 

(1994-1996) to age 11 yr. 

Children were exposed 

prenatally to multiple drugs.

Age 4 yr

Mean (range): 7.0 

(1.3-23.8)

10th-90th: 3.0-12.1

Linear regression model adjusted 

for HOME score, caregiver's 

vocabulary test, sex, parity, 

maternal marital status, head 

circumference at birth

WISC-R at age 4 yr (concurrent)

WISC-R at age 9 yr

WISC-R at age 11 yr

-0.50 (-0.89, -0.11)

-0.41 (-0.78, -0.04)

-0.54 (-0.91, -0.17)

Schnaas et al.

(2006)

150 children in Mexico City, 

Mexico followed from birth 

(1987-1992) to age 10 yr.

Prenatal (maternal 

28-36 weeks)

Geometric mean (95% 

CI): 7.8 (2.5-24.5)

10th-90th: 3.2-19.1

Log linear mixed effects regression 

model adjusted for sex, SES, 

maternal IQ, HOME score, birth 

weight, postnatal blood Pb, random 

slope for subject

McCarthy GCI at ages 6-10 yr -0.44 (-0.73, -0.15)
February 2012 5-59 Draft – Do Not Cite or Quote

Study

Population/

Location

Blood Pb Data 

(μg/dL) Statistical Analysis FSIQ Assessmenta Effect Estimate(95% CI)b,c

Kordas et al.

(2011)

186 children in Mexico City, 

Mexico followed from birth 

(1994-1995) to age 4 yr

Prenatal (cord)

Mean (SD): 6.6 (3.3)

10th-90th: 1.2-2.4

Concurrent

Mean (SD): 8.7 (4.4)

10th-90th: 1.4-2.7

Linear regression model adjusted 

for sex, birth weight, gestational 

age, maternal age, years of 

schooling, IQ, smoking status, 

marital status, crowding in home, 

type of floor in home

McCarthy GCI at age 4 yr

Prenatal

Concurrent

-0.20 (-0.79, 0.39)

-0.60 (-0.99, -0.21)

Roy et al.

(2011)

717 children in Chennai, 

India ages 3-7 yr

Concurrent

Mean (SD): 11.4 (5.4)

10th-90th: 5.8-18.4

Log linear model adjusted for 

mid-arm circumference, age, sex, 

family income, parental education, 

parental IQ, family size

Binet-Kamat at ages 3-7 yr -0.39 (-0.65, -0.13)

Dietrich et al.

(1993a)

253 children in Cincinnati, 

OH followed from birth 

(1979-1985) to age 20-23 

yr.

Concurrent NR

Age 5 yr

Mean (SD): 11.8 (6.3)

10th-90th: 5.5-19.7

Linear regression model adjusted 

for HOME score, maternal IQ, birth 

weight, birth length, child sex, 

maternal cigarette consumption 

during pregnancy

WISC-R at age 6.5 yr -0.33 (-0.60, -0.06)

Tong et al.

(1996)

375 children in Port Pirie, 

Australia followed from birth 

(1979-1982) to age 11-13 

yr.

Lifetime avg

Geometric mean 

(GSD): 14.0 (1.2)

10th-90th: 12.7-18.1

Regression model adjusted for sex, 

age, school grade, parental 

occupational prestige, HOME score, 

maternal IQ, family functioning 

score, parental smoking, marital 

status, parental education, maternal 

age, birth weight, birth order, 

feeding method, breastfeeding 

duration, family size, life events, 

prolonged absences from school

WISC-R at age 11-13 yr -0.12 (-0.24, -

0.003)

Wasserman et 

al. (1997)

290 children in Kosovo, 

Yugoslavia followed from 

birth (1985-1986) to age 

10-12 yr.

Lifetime avg

Geometric mean: 16.2

10th-90th: 6.0-44.2

Generalized estimating equations 

with log-transformed blood Pb 

adjusted for age, sex, sibship size, 

birth weight, language spoken in 

home, HOME score, maternal age, 

maternal education, maternal 

Raven score

WISC-III at ages 10-12 yr -0.20 (-0.28, -0.11)

Baghurst et al.

(1992)

494 children in Port Pirie, 

Australia followed from birth 

(1979-1982) to age 11-13 

yr.

Lifetime avg

Mean of 25-50th: 16.6

10th-90th: 10-27.5

Log linear regression model 

adjusted for sex, birth weight, birth 

order, feeding method, 

breastfeeding duration, parental 

education, maternal age, parental 

smoking, SES, quality of home 

environment, maternal IQ, parents 

living together

WISC-R at age 7-8 yr -0.20 (-0.40, 0.10)

a

WISC = Wechsler Intelligence Scale for Children, WPPSI = Wechsler Preschool and Primary Scale of Intelligence, GCI = General Cognitive Index b

Effect estimates are standardized to a 1 μg/dL increase in blood Pb level within the 10th to 90th percentile interval. Effect estimates are assumed to be 

linear within the 10th to 90th percentile interval of blood Pb level. The percentiles are estimated using various methods and are only approximate values. c

95% CI was constructed using a standard error that was estimated for a p-value of 0.01. Authors specified a p-value of <0.01.

1 The analysis pooling data from seven prospective studies included 1,333 children 

2 ages 5-10 years of age with a median (5th-95th percentile) concurrent blood Pb level of 

3 9.7 μg/dL (2.5-33.2 μg/dL) (Lanphear et al., 2005). In multivariate models that adjusted 

4 for study site, maternal IQ, Home Observation for the Measurement of Environment 

5 (HOME) inventory (assessment of physical environment, parental responsivity, learning 

6 stimulation, emotional climate, and family interactions), birth weight, and maternal 

7 education, higher concurrent, peak, average lifetime, and early childhood blood Pb levels 

8 were associated with lower FSIQ measured at age 5-10 years, with the largest decrement 

9 in FSIQ estimated for concurrent blood Pb level (-0.25 points [95% CI: -0.34, -0.15] per 

10 1 μg/dL increase in blood Pb level in the 10th to 90th percentile interval [2.2-12.3 μg/dL] 
February 2012 5-60 Draft – Do Not Cite or Quote

of blood Pb level)1 1 . Various models were investigated to characterize the shape of the 

2 blood Pb concentration-response relationship. Consistent with findings from several 

3 individual cohort studies, Lanphear et al. (2005) found that a log-linear model best fit the 

4 data, with a greater decrease in FSIQ estimated for an increase in concurrent blood Pb 

5 from 2.4-10 μg/dL (3.9 points [95% CI: 5.3, 2.4]) than an increase from 10 to 20 μg/dL 

6 (1.9 points [95% CI: 2.6, 1.2]). Among children with concurrent blood Pb less than 

7 10 μg/dL, the median blood Pb level was 4.2 μg/dL (Hornung, 2008).

8 A key additional strength of the pooled analysis by Lanphear et al. (2005) was the 

9 extensive examination of the potential for confounding by several factors related to SES 

10 and the caregiving environment. Variables such as HOME score, birth weight, maternal 

11 IQ, and maternal education were statistically significantly associated with FSIQ and were 

12 included in the final model with blood Pb level. While a smaller decrement in FSIQ was 

13 estimated for concurrent blood Pb level in this adjusted model than in the unadjusted 

14 model (-0.43 points [95% CI: -0.53, -0.33] per 1 μg/dL increase in blood Pb level in the 

15 10th to 90th percentile interval of blood Pb level), the adjusted blood Pb effect estimate 

16 was nonetheless statistically significant. HOME score was not available in the Rochester 

17 study; however, exclusion of data from that cohort resulted in a 3% less negative effect 

18 estimate, indicating the lack of a strong influence of HOME score alone on blood Pb 

19 level-IQ associations. Other variables such as child sex, tobacco exposure during 

20 pregnancy, alcohol use during pregnancy, maternal age at delivery, marital status, and 

21 birth order were not statistically significantly associated with FSIQ and did not alter the 

22 effect estimate for concurrent blood Pb level. The individual study populations 

23 represented a wide range of SES, maternal education, and cultural backgrounds. 

24 Sensitivity analyses, in which one study was successively excluded, revealed that no 

25 single study was responsible for driving the results. Per 1 μg/dL increase in blood Pb 

26 level in the 10th to 90th percentile interval of blood Pb level, effect estimates excluding 

27 one study at a time ranged between -0.22 and -0.27, indicating the robustness of the 

28 concurrent blood Pb level effect estimate despite between-study variability in the 

29 distributions of potential confounding factors. 

30 The small number of studies published since the 2006 Pb AQCD continued to 

31 demonstrate associations between higher blood Pb level (primarily concurrent) and lower 

32 FSIQ in children between ages 3 and 11 years (Figure 5-2 and Table 5-3). Similar to 

33 studies reviewed in the 2006 Pb AQCD, most recent studies demonstrated associations 

34 between blood Pb level and lower FSIQ in populations with mean blood Pb level between 

35 5 to 10 μg/dL. New results from the prospective cohorts were limited. Mazumdar et al.

 1

To facilitate comparisons among effect estimates across studies with different distributions of blood Pb levels and 

model structures (e.g., linear, log-linear), effect estimates are standardized to a 1 μg/dL increase in blood Pb level 

within the estimated 10th to 90th percentile interval. Effect estimates are assumed to be linear within the 10th to 

90th percentile interval of blood Pb level.
February 2012 5-61 Draft – Do Not Cite or Quote

1 (2011) reported on the follow-up of the Boston cohort to age 28-30 years (55 out of the 

2 original 249 enrolled at birth). Blood Pb levels measured at age 6 months, 4 years, 

3 10 years, and levels averaged over childhood were associated with decrements in FSIQ in 

4 adults. The effect estimates were similar in magnitude for all childhood blood Pb 

5 measures, except for 6 month blood Pb level, which was associated with a smaller FSIQ 

6 decrement. These findings indicated that the effect of childhood Pb exposures may persist 

7 to adulthood.

8 Jusko et al. (2011; 2008) affirmed the findings in the Rochester cohort previously 

9 reported by Canfield et al. (2003a), who examined the cohort at age 5 years (Figure 5-2

10 and Table 5-3). Jusko et al. (2011) examined the Rochester cohort at age 6 years and, 

11 similar to Canfield et al. (2003a), found that a 1 μg/dL increase in concurrent blood Pb 

12 level was associated with a 0.66-point decrease (95% CI: -1.26, -0.06) in FSIQ. Model 

13 covariates were the same as those analyzed by Canfield et al. (2003a): sex, race, family 

14 income, maternal education, race, prenatal smoking, birth weight, transferrin saturation, 

15 maternal IQ, and HOME score. At both age 5 and 6 years, higher peak and lifetime 

16 average blood Pb levels also were associated with lower FSIQ, and incremental increases 

17 in blood Pb levels tended to be associated with larger FSIQ decrements in analyses 

18 restricted to children with blood Pb levels less than 10 μg/dL (Jusko et al., 2008; Canfield 

19 et al., 2003a). Canfield et al. (2003a) provided additional information on the extent of 

20 confounding by SES- and caregiving-related variables. The effect estimate in the 

21 covariate-adjusted model was 40% smaller than it was in the model with concurrent 

22 blood Pb level alone; however, the association with concurrent blood Pb level remained 

23 statistically significant. 

24 Additional evidence recently was provided for children in Mexico City, albeit in a 

25 separate cohort of children born later with lower blood Pb levels at corresponding ages. 

26 Among children born 1987-1992, Schnaas et al. (2006) previously reported larger Pb27 associated decrements in FSIQ for prenatal maternal (28-36 weeks) blood Pb levels than 

28 for concurrent blood Pb levels between ages 1 and 10 years. In contrast, Kordas et al.

29 (2011) found that concurrent blood Pb level was associated with a larger decrement in 

30 FSIQ at age 4 years than was cord blood Pb level. Children in the latter study were born 

31 between 1994 and 1995 and at age 4 years had a mean (SD) blood Pb level of 

32 8.7 (4.4) μg/dL. In Schnaas et al. (2006), the geometric mean (95% CI) blood Pb level at 

33 age 4 years was 10.3 (4.2, 20.5) μg/dL. It is not clear whether different temporal patterns 

34 of Pb exposure may have contributed to the contrasting associations for prenatal and 

35 concurrent blood Pb levels in the two studies.

36 Surkan et al. (2007) examined children from urban Boston, Massachusetts and rural 

37 Farmington, Maine participating in a trial designed to assess the effect of amalgam dental 
February 2012 5-62 Draft – Do Not Cite or Quote

1 fillings on child neurodevelopment. Prior to placement of amalgam fillings, blood Pb 

2 levels were measured, and children were administered an extensive battery of 

3 neuropsychological tests including tests of memory, learning, visual-motor ability, 

4 reading, reaction time. A strength of the study was that the range of concurrent blood Pb 

5 level in this study, 1-10 μg/dL, was lower than that examined in previous studies. Thus, it 

6 provided information on associations between blood Pb level and IQ decrements within a 

7 lower range of blood Pb levels. However, consistent with several previous studies, 

8 Surkan et al. (2007) found lower FSIQ in the group of children with concurrent blood Pb 

9 levels of 5 to 10 μg/dL. For example, adjusting for age, race/ethnicity, birth weight, SES, 

10 and primary caregiver IQ, children with blood Pb levels 5-10 μg/dL had lower FSIQ 

11 scores (-6.04 points [95% CI: -10.7, -1.36]) compared with children who had levels 

12 1-2 μg/dL (referent group). Children with blood Pb levels 2-5 μg/dL did not have FSIQ 

13 scores (-0.12 [95% CI: -3.30, 3.06]) that differed from children with blood Pb levels 

14 1-2 μg/dL. Another strength of this study was the analysis of potential confounding by a 

15 larger list of variables than that included in the final models. Although HOME score was 

16 not examined, several variables such as SES, caregiver education, parenting stress, 

17 marital status of caregiver, and maternal utilization of prenatal or annual health care were 

18 not significantly (p > 0.20) associated with child IQ either independently or in a model 

19 with blood Pb level (Surkan et al., 2007).

20 Other recent studies found blood Pb-associated lower FSIQ in populations of children in 

21 Asia with lower blood Pb levels (Kim et al., 2009b; Zailina et al., 2008) than were 

22 previously examined. Kim et al. (2009b) examined children ages 8 to 11 years in Korea 

23 (born mid- to late-1990s) with a mean (range) concurrent blood Pb level of 1.73 

24 (0.42-4.91) μg/dL. Children were tested using the Korean Educational Development 

25 Institute-WISC, which assesses vocabulary, arithmetic, picture arrangement, and block 

26 design. In a log-linear regression analysis adjusted for age, sex, maternal and paternal 

27 education, yearly income, prenatal smoking, postnatal environmental tobacco smoke 

28 exposure, birth weight, and maternal age at birth, a 1 μg/dL higher concurrent blood Pb 

29 level was associated with a 0.64-point lower (95% CI: -1.05, -0.19) FSIQ within the 10th30 90th percentile interval of blood Pb level (< 1 to 2.8 μg/dL). The potential confounders 

31 examined in this study were not found to have a strong influence on the findings as a 

32 similar magnitude of effect was estimated in a model that included only blood Pb level 

33 (-0.73 points [95% CI: -1.19, -0.27] per 1 μg/dL increase in blood Pb level in the 10th to 

34 90th percentile interval). Although several important SES-related confounders were 

35 considered, there was no direct assessment of the home environment and the primary 

36 caregiver IQ in this study, which are notable limitations.

37 Kim et al. (2009b) also examined effect modification of the blood Pb-FSIQ relationship 

38 by concurrent blood manganese (Mn) levels. The mean (range) blood Mn level was 
February 2012 5-63 Draft – Do Not Cite or Quote

1 14.3 (5.3-29.02) μg/dL. Blood Pb and Mn levels were not correlated (r = 0.03, p = 0.64). 

2 To examine effect modification, children were divided into two groups: blood Mn level 

3 above and below the median (14 μg/dL). Multivariate linear regression models predicting 

4 FSIQ and verbal IQ (VIQ) used concurrent blood Pb level as the predictor variable in the 

5 low and high Mn groups. As illustrated in Figure 5-3, the associations of concurrent 

6 blood Pb level with FSIQ and VIQ was larger in magnitude and statistically significant 

7 for children in the high Mn group compared with children in the low Mn group (-0.65 

8 points [95% CI: -1.26, -0.05] in high Mn group versus -0.50 points [95% CI: -1.23, 0.22] 

9 in low Mn group per 1 μg/dL increase in blood Pb level in the 10th to 90th percentile 

10 interval). However, higher blood Pb level was associated with lower FSIQ in children in 

11 the low Mn group.

Source: Reprinted with permission of Elsevier Science, Kim et al. (2009b).

Note: High and low Mn refer to levels above and below the median of 14 μg/dL, respectively.

Figure 5-3 Effect modification of the association between concurrent blood 

Pb level and FSIQ by blood Mn level.
February 2012 5-64 Draft – Do Not Cite or Quote

1 Higher concurrent blood Pb level also was associated with a lower IQ score (as assessed 

2 by the McCarthy General Cognitive Index) among children ages 6-8 years in Malaysia 

3 with a mean blood Pb level of 3.69 μg/dL (Zailina et al., 2008). The concentration4 response function was not reported; however, in a multivariate model, blood Pb level and 

5 not potential confounding factors such as paternal education, maternal education, 

6 household income, number of siblings, and birth order was reported to be significantly 

7 associated with FSIQ. As with Kim et al. (2009b), a notable limitation was the lack of 

8 consideration of confounding by the caregiving environment.

9 Pb has long been known to impact dopaminergic neurons, inhibit depolarization-evoked 

10 neurotransmitter release, and stimulate spontaneous neurotransmitter release

11 (Section 5.3.8.8). Further, dopaminergic activity is a key process in regulating cognitive 

12 function. Thus, variants in dopamine-related genes have the potential to act as effect 

13 modifiers of Pb-associated neurodevelopmental effects. Recent studies in children that 

14 examined effect modification by variants in dopamine genes produced contrasting results. 

15 In a study of children at ages 2 and 4 years, Kordas et al. (2011) did not find differences 

16 in association between concurrent blood Pb level and FSIQ by the Taq A1/A1 dopamine 

17 receptor 2 (DRD2) genotype, which is associated with reduced dopamine receptor 

18 density. Roy et al. (2011) examined children of similar age (3-7 years) in Chennai, India 

19 and found that among children with the Taq A1/A1 genotype, a 1 μg/dL higher blood Pb 

20 level was associated with a 0.84-point lower (95% CI: -1.66, -0.01) FSIQ within the 10th21 90th percentile interval of blood Pb level (5.8-18.3 μg/dL). The same increment in blood 

22 Pb level was associated with a 0.36-point lower (95% CI: -0.76, -0.04) FSIQ in children 

23 with the Taq A2/A2 higher receptor density genotype. Concurrent blood Pb level was 

24 associated with FSIQ in both studies in analyses including all subjects. However, there 

25 were many differences between study populations that may have contributed to 

26 differences in effect modification by the DRD2 variant. Compared with the group in 

27 India, the group in Mexico had a lower mean blood Pb level and lower mean FSIQ score. 

28 Additionally, children in the Mexico City group with the Taq A1/A1 genotype had a 

29 higher mean FSIQ score, whereas FSIQ scores were similar between Taq genotypes in 

30 the children in India.

31 Other recent studies examined children in the U.S.; however, in these studies, there were 

32 high proportions of children with prenatal exposure to alcohol, tobacco, marijuana, or 

33 cocaine that may limit the generalizability of findings. Chiodo et al. (2007) examined a 

34 population of African American children (age 7 years, born 1989-1991) in the Detroit, 

35 MI area with relatively low blood Pb level (mean [SD]: 5.0 [3.0]) but high prevalence of 

36 prenatal exposure to cocaine (38%) or marijuana (35%). In a detailed analysis of potential 

37 confounding factors, investigators found that cocaine exposure did not meet the criterion 

38 for inclusion in the model (i.e., p > 0.10 for association with FSIQ). Prenatal marijuana 
February 2012 5-65 Draft – Do Not Cite or Quote

1 exposure was associated with FSIQ (p < 0.10) and was included in a multivariate model 

2 along with caretaker education, SES, HOME score, symptom checklist, number of 

3 children in the home, maternal IQ, sex, and prenatal cigarette smoking. In this 

4 multivariate model, higher blood Pb level was associated with lower FSIQ (Figure 5-2

5 and Table 5-3). These findings supported an independent association of blood Pb level 

6 despite the high prevalence of prenatal drug exposure in the study population. 

7 Min and colleagues (2009) followed children in Cleveland, OH from age 4 to 11 years 

8 who were exposed prenatally to alcohol (77%), cigarette smoking (61%), marijuana 

9 (31%), or cocaine (51%). The study population was primarily African-American (86%) 

10 and of low SES (98%); 39% of mothers had not finished high school, and 86% were 

11 unmarried at the time of enrollment. The mean (range) blood Pb level, measured at age 

12 4 years, was 7.0 μg/dL (1.3-23.8). Several lines of evidence indicated that prenatal drug 

13 exposure did not heavily influence the association between blood Pb level and FSIQ. 

14 Prenatal alcohol exposure was correlated with blood Pb level but was not statistically 

15 significantly associated with IQ (p > 0.10) and did not change the blood Pb level effect 

16 estimate by more than 10%. Prenatal cocaine exposure was associated with FSIQ at age 

17 9 years; however, blood Pb level at age 4 years remained associated with a decrement in 

18 FSIQ at age 9 years after adjusting for cocaine exposure and several SES-related factors 

19 (Table 5-3). Based on the examination of interaction terms, associations between blood 

20 Pb level and FSIQ were not statistically significantly different between drug-exposed and 

21 -unexposed children. 

22 Similar to previous studies with repeated assessments of cognitive function over time, 

23 Min et al. (2009) found that the association between blood Pb level at age 4 years and 

24 FSIQ persisted to older ages. Higher blood Pb level at age 4 years was associated with 

25 similar magnitudes of decrements in FSIQ at ages 4, 9 and 11 years (Figure 5-2 and 

26 Table 5-3). Researchers also examined the shape of the concentration-response function 

27 using a restricted cubic spline function. Although the cubic spline term did not attain 

28 statistical significance (p = 0.19), qualitative analysis indicated that the association 

29 between blood Pb level and FSIQ decrements persisted and was greater at lower levels 

30 blood Pb (< 7 μg/dL). These findings were consistent with those from the pooled analysis

31 (Lanphear et al., 2005) and other individual studies (Tellez-Rojo et al., 2006).

Bayley Scales of Infant Development

32 The Bayley Scales of Infant Development are the most widely used tests of infant 

33 intelligence. The Mental Development Index (MDI) is statistically analogous to IQ, 

34 i.e., both scores have a population-standardized mean of 100 and standard deviation of 

35 15. While MDI assesses general cognitive function in infants, it is important to note that 
February 2012 5-66 Draft – Do Not Cite or Quote

1 MDI scores are not necessarily correlated with IQ scores measured later in childhood.

2 Associations between blood Pb levels and MDI scores were found in most of the 

3 prospective cohort studies in children between age 6 months and 3 years and mean blood 

4 Pb levels 5-10 μg/dL (Table 5-4). Recent studies examined children who were born in the 

5 mid-1990s to mid-2000s and continued to demonstrate associations between higher cord 

6 and concurrent blood Pb level and lower MDI scores (Table 5-4). Studies that found 

7 associations with concurrent blood Pb levels also tended to find associations with 

8 prenatal cord or maternal blood Pb levels. Recent studies provided new information on 

9 effect modification by nutritional status, maternal self-esteem, co-exposure to Mn, and 

10 genetic variants. While studies adjusted for multiple SES-related variables including 

11 maternal IQ and education, most did not consider confounding by the caregiving 

12 environment. Concurrent and cord blood Pb levels were associated with MDI, adjusting 

13 for HOME score in diverse cohorts (Solon et al., 2008; Wasserman et al., 1992; Bellinger 

14 et al., 1987). In the Cleveland cohort, associations of cord and postnatal blood Pb levels 

15 with MDI at ages 6 months to 2 years became null after adjusting for covariates including 

16 HOME score (Ernhart et al., 1988; Ernhart et al., 1987). In the analysis of the Cincinnati 

17 cohort, HOME score was not significantly associated with blood Pb level or MDI and 

18 thus, did not meet the criteria for model inclusion (Dietrich et al., 1987a). Collectively, 

19 evidence does not indicate that Pb-associated MDI decrements are driven by confounding 

20 by quality of the caregiving environment.
February 2012 5-67 Draft – Do Not Cite or Quote

Table 5-4 Associations of blood Pb level with Bayley MDI in children ages 

6 months to 3 yearsa

Study

Population/

Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Cognitive Index

Effect Estimate 

(95% CI)b

Jedrychowski et 

al. (2009b)

444 children born 

2001-2004 followed 

prenatally to age 36 

mo

Krakow, Poland

Prenatal (cord blood) 

Geometric mean 

(range): 1.29 (0.44-5)

10th-90th: 1.2-1.3

Log linear regression model adjusted for 

maternal education, birth order, prenatal 

smoking, sex, and within-subject MDI 

correlation

at age 12 mo

at age 24 mo

at age 36 mo

-1.9 (-3.8, 0.12)

-2.6 (-5.0, -0.21)

-2.3 (-4.3, -0.30)

Claus Henn et 

al.

455 children born 

1997-2000 followed 

prenatally to age 36 

mo

Mexico City

12 month

Mean (SD): 5.1 (2.6)

10th-90th: 2.5-8.4

Linear mixed effects regression adjusted 

for sex, hemoglobin, gestational age, 

maternal IQ, maternal education, blood 

Pb-blood Mn interaction

assessed ages 12 to 36 mo

12 mo blood Mn < 2.0 μg/dL

12 mo blood Mn 2.0-2.8 μg/dL

12 mo Mn > 2.8 μg/dL

-3.0 (-5.22, -0.78)

-0.07 (-0.39, 0.25)

-2.2 (0, 4.44)

Tellez-Rojo et 

al. (2006)

294 children born 

1997-1999 followed 

prenatally to age 24 

mo

Mexico City

Same cohort as above

Concurrent

12 month

Mean (SD): 4.7 (2.9)

10th-90th: 1.9-8.2

24 month

Mean (SD): 5.3 (4.1)

10th-90th: 2.1-10.7

Log linear regression model adjusted for

sex, age, birth weight, maternal IQ, 

cohort

at age 12 mo

at age 24 mo

-0.26 (-0.79, 0.26)

-0.89 (-1.32, -0.46)

Dietrich et al.

(1993a)

96 to 302 children born 

1979-1984 followed 

prenatally to age 6 mo

Cincinnati, OH

Prenatal (cord)

Mean (SD): 6.3 (4.5)

10th-90th: 2.3-11.7

Neonatal (10 day)

Mean (SD): 4.6 (2.8)

10th-90th: 1.9-8.1

3 month

Mean (SD): 5.9 (3.4)

10th-90th: 2.6-10.1

Linear regression model adjusted for 

birth weight, gestation, maternal age, 

race, sex, SES

assessed at 6 mo

Cord blood Pb

Neonatal blood Pb

3-month blood Pb

-0.66 (-1.4, 0.07)

-3.49 (-6.0, 0.96)

-0.48 (-1.0, 0.05)

Bellinger et al.

(1987)

249 children born 

1979-1981 followed 

prenatally to age 24 

mo

Prenatal (cord)

Low: < 3

Medium: 6-7

High: ≥ 10

Regression model adjusted for maternal 

age, race, maternal IQ, maternal 

education, number of years of smoking, 

number of alcohol drinks per week in 3rd 

trimester of pregnancy, SES, HOME, 

sex, birthweight, gestational age, birth 

order

assessed ages 6 to 24 mo

High vs. low cord blood

High vs. medium cord blood

-4.8 (-7.3, -2.3)

-3.8 (-6.3, -1.3)

Surkan et al.

(2008)

309 children ages 

12-36 mo during 

1996-2001 or 

2004-2005

Mexico City, Mexico 

Concurrent

Mean (SD): 6.4 (4.3)

10th-90th: 2.0-12.4

Linear mixed effects regression model 

adjusted for sex, maternal age, maternal 

IQ, maternal education, parity, alcohol 

consumption, smoking, cohort, maternal 

self-esteem

assessed ages 12 to 36 mo

All subjects

High maternal self-esteem

Low maternal self-esteem

-0.18 (-0.45, 0.09)

0.36 (-0.50, 1.2)

-0.31 (-0.60, -0.02)

Pilsner et al.

(2010)

255 children age 24 

mo born 1994-1995

Mexico City, Mexico

Prenatal (cord blood) 

Mean (SD): 6.7 (3.6)

10th-90th: 3.5-10.5

Linear regression model adjusted for 

maternal age, maternal IQ, marital 

status, parity, gestational age, 

inadequate folate intake, MTHFR 

genotype

at age 24 mo -0.73 (-1.2, -0.23)
February 2012 5-68 Draft – Do Not Cite or Quote

Study

Population/

Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Cognitive Index

Effect Estimate 

(95% CI)b

Hu et al. (2006) 146 children born 

1997-1999 followed 

prenatally to age 24 

mo

Mexico City, Mexico

Prenatal (maternal 

blood Pb) in 1st 

trimester

Mean (SD): 7.1 (5.1)

10th-90th: 2.5-13.1

3rd trimester

Mean (SD): 6.9 (4.2)

10th-90th: 2.8-12.1

Cord blood

Mean (SD): 6.2 (3.9)

10th-90th: 2.5-11.0

Concurrent

Mean (SD): 4.8 (3.7)

10th-90th: 1.6-9.1

Log linear regression model adjusted for

sex, maternal age, current weight, 

height-for-age Z score, maternal IQ, 

concurrent blood Pb (in models 

examining blood Pb at other lifestages)

assessed at age 24 mo

Prenatal 1st trimester

Prenatal 3rd trimester

Cord blood

Concurrent

-0.64 (-1.3, -0.03)

-0.38 (-1.0, 0.24)

-0.06 (-0.82, 0.70)

-0.23 (-0.92, 0.45)

Solon et al.

(2008)

502 children ages 6-35 

mo, born 1997-2004

Visayas, Philippines

Concurrent 

Mean (SD): 7.1 (7.7)

10th-90th: 1.6-14.9

Two-stage linear regression model to 

account for determinants of blood Pb 

(sex, roof materials, water source, 

breastfed for ≥ 4 months) and cognitive 

function (HOME score, maternal 

education, maternal smoking, born 

premature, region of residence)

assessed ages 6 to 35 mo -3.32 (-5.02, -1.60)

Vimpani et al.

(1985)

592 children followed 

prenatally to age 24

mo

Port Pirie, Australia

20% subjects had 

24-month blood Pb 

levels > 30

Linear regression model adjusted for 

maternal age, paternal education, 

maternal education, paternal workplace, 

maternal workplace, parental 

relationship, maternal prenatal marital 

status, child birth rank, mouthing activity, 

oxygen use at birth, apgar score, 

neonatal jaundice, size for gestational 

age, maternal IQ

at age 24 mo

Maternal avg prenatal blood Pb

Cord blood Pb

6 mo blood Pb

24 mo blood Pb

Lifetime avg

-0.64

0.03

-0.40, p < 0.05

-0.06

-0.31, p < 0.05

Wasserman et 

al. (1992)

392 children followed 

prenatally to age 24 

mo

Kosovska Mitrovica 

and Pristina, 

Yugoslavia

Concurrent Means:

35.5 (K. Mitrovica)

8.4 (Pristina)

Log linear regression model adjusted for 

sex, birth order, birth weight, ethnic 

group, HOME, maternal education, 

maternal age, maternal IQ

at age 24 mo

Cord blood Pb

6 mo blood Pb

12 mo blood Pb

18 mo blood Pb

24 mo blood Pb

Per tripling blood Pb

-1.7, p = 0.12

-1.1, p = 0.34

-1.7, p = 0.17

-1.8, p = 0.16

-2.5, p = 0.03

MDI = Mental Development Index, MTHFR = methylenetetrahydrofolate reductase

a

Studies are presented in order of increasing population mean blood Pb level.

b

Except where noted, effect estimates are standardized to a 1 μg/dL increase in blood Pb level within the 10th to 90th percentile interval of blood Pb level.

1 In a population of children in Krakow, Poland with lower cord blood Pb levels (median 

2 1.23 μg/dL, 95% CI: 1.24, 1.34 μg/dL) than those previously examined, increasing cord 

3 blood Pb level was associated with similar magnitudes of decrease in MDI at ages 12, 24, 

4 and 36 months (Jedrychowski et al., 2009b) (Table 5-4). Consistent with the hypothesis 

5 that the developing male central nervous system may be more vulnerable than that of 

6 females to environmental insults resulting in later behavioral problems (Moffitt et al., 

7 2001), investigators estimated a larger in MDI per unit increase in cord blood Pb level 

8 among the 233 males than among the 223 females (Figure 5-4). Although mean cord 

9 blood Pb levels were similar between males (1.35 μg/dL) and females (1.41 μg/dL), the 

10 mean 36-month MDI score was lower among males than among females (101 and 105, 

11 respectively, p = 0.0001). 
February 2012 5-69 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Elsevier Science, Jedrychowski et al. (2009a).

Figure 5-4 Regression of fitted MDI score at 36 months on log-transformed 

cord blood Pb level by sex.

1 Solon et al. (2008) examined children in the Philippines, ages 6 to 35 months. A key 

2 strength of this study was the adjustment for HOME score as well as maternal education, 

3 maternal smoking, premature birth, region of residence, and years of schooling of child. 

4 Not only did investigators find an association between higher concurrent blood Pb level 

5 and lower MDI score (Table 5-4), but they also found effect modification by red blood 

6 cell folate levels. Among children with folate levels less than or equal to 230 μg/mL, 

7 blood Pb level had a marginal association with lowering MDI scores in the range of 0.80 

8 to 2.44 points. Among children with higher folate levels, blood Pb level was not 

9 estimated to have a negative marginal impact. These findings from Solon et al. (2008)

10 indicated that children with folate deficiencies may be at increased risk of Pb-associated 

11 decreases in cognitive function. The results were consistent with observations that higher 

12 folate level is associated with lower blood Pb level since folate improves Pb excretion by 

13 inhibiting the binding of Pb to blood elements.

14 Multiple studies in different Mexico City mother-child pair cohorts recently reported 

15 associations between blood Pb levels (e.g., maternal, cord blood, or child postnatal) and 

16 decrements in Bayley MDI in children between age 12 and 36 months (Claus Henn et al.; 

17 Pilsner et al., 2010; Surkan et al., 2008; Hu et al., 2006; Tellez-Rojo et al., 2006). Recent 

18 studies extended findings from Tellez-Rojo et al. (2006) with follow-up of the same 
February 2012 5-70 Draft – Do Not Cite or Quote

1 cohort to age 3 years and observations of effect modification by maternal self-esteem

2 (Surkan et al., 2008) and blood Mn levels (Claus Henn et al.).

3 Claus Henn et al. examined interactions between blood Mn and Pb levels. Investigators 

4 selected mid-range (2.0-2.8 μg/dL) blood Mn levels as the reference group based on 

5 previous observations that MDI scores were least affected by increases in blood Mn level 

6 in this group. Among subjects with age 12-month blood Mn levels less than 2.0 μg/dL, a 

7 1 μg/dL higher age 12-month blood Pb level was associated with a 3.0-point (95% CI: -

8 5.2, -0.78) lower MDI. Among subjects with blood Mn levels greater than 2.8 μg/dL, a 

9 1 μg/dL higher age 12-month blood Pb level was associated with a 2.2-point (-4.4, 0) 

10 lower MDI. Interactions were not found using age 24-month blood Mn and Pb levels. 

11 Effects estimates were similar in magnitude before and after adjustment for sex, 

12 gestational age, hemoglobin, maternal IQ, maternal education, and visit. Findings in this 

13 Mexico City cohort added to those from Kim et al. (2009b) in older children that 

14 associations between blood Pb level and cognitive function may be influenced by Mn co15 exposure, and results from both studies indicated that blood Pb level was associated with 

16 decrements in cognitive function with both lower and higher levels of blood Mn. 

17 Biological plausibility for the interactive effects of Pb and Mn exposure is provided by 

18 observations that Mn has similar modes of action and cellular targets as does Pb, 

i.e., altering Ca+2 19 metabolism, inducing oxidative damage to neuronal cells, diminishing 

20 dopamine transmission. Claus Henn et al. also indicated that Pb-Mn co-exposure may 

21 affect cognitive function in children as young as 12 to 36 months of age.

22 Surkan et al. (2008) stratified data by the level of maternal self-esteem as reported by 

23 mothers. Higher age 24-month blood Pb level was associated with lower MDI score 

24 among children with mothers in the lowest three quartiles of self-esteem but not among 

25 children with mothers in the highest quartile of self-esteem (Table 5-4). Model covariates 

26 included cohort, sex, maternal IQ, maternal age, maternal education, parity, maternal 

27 smoking, maternal alcohol consumption, and maternal self-esteem. While HOME score 

28 was not examined, maternal self-esteem may incorporate many of the same aspects as 

29 HOME score, including mother-child interactions, educational interactions, and dietary 

30 practices. These findings indicated that higher maternal psychosocial functioning 

31 (e.g., lower stress, anxiety, and depression and higher self-esteem) may contribute to 

32 better caregiving, which in turn may improve neuropsychological functioning of the 

33 child. The biological plausibility of these findings in children is well-supported by 

34 observations in animals unrelated to Pb that environmental enrichment reverses the 

35 effects of early stress experiences on reactions such as depressed behavior, HPA 

36 activation, and immunosuppressant (Laviola et al., 2008; Laviola et al., 2004; Moreley37 Fletcher et al., 2003; Francis et al., 2002). With specific regards to Pb exposure, social 

38 isolation or enrichment has been found to exacerbate or protect against, respectively, Pb 
February 2012 5-71 Draft – Do Not Cite or Quote

1 exposure-induced learning impairments in animals (Section 5.3.2.2). It is worth 

2 mentioning in this context that the potential programming effects of stress on childhood 

3 health outcomes may occur at an even more fundamental level, i.e., through epigenetic 

4 programming (Dolinoy and Jirtle, 2008).

5 Hu et al. (2006) compared associations among prenatal blood Pb levels measured at 

6 different stages of gestation among 146 mother-child pairs meeting the following criteria: 

7 born at 37 weeks or greater gestational age, at least one valid Pb measurement during 

8 pregnancy, complete information on maternal age and IQ, and measurement of child 

9 blood Pb level at 24 months when the 24-month MDI was ascertained. Among blood Pb 

10 levels measured at various lifestages, 1st trimester maternal blood Pb level (whole or 

11 plasma Pb) was associated with larger decreases in subsequent 24-month MDI scores 

12 compared with maternal 3rd trimester, cord, and child concurrent blood Pb (Table 5-4). 

13 These results were adjusted for sex, 24-month blood Pb level, height-for-age Z score, 

14 weight, maternal age, and maternal IQ. HOME score was not included in the final model; 

15 however, investigators examined the potential for confounding by a larger list of 

16 unspecified variables. Prenatal Pb exposure effects also were indicated in a study 

17 conducted in Mexico City that examined FSIQ repeatedly during ages 6-10 years

18 (Schnaas et al., 2006). Third trimester (weeks 28-36) maternal blood Pb level, was 

19 associated with larger decreases in FSIQ than were maternal blood Pb level at 

20 12-20 weeks, maternal blood Pb level at delivery, or concurrent child blood Pb levels 

21 measured between ages 6 and 10 years.

22 In another recent study in Mexico City, Mexico, investigators found higher cord blood Pb 

23 level to be associated with a lower MDI score in children at age 24 months (-0.73 points 

24 [95% CI: -1.2, -0.23] in score per 1 μg/dL increase in cord blood Pb level in the 10th25 90th percentile interval [3.5-10.5 μg/dL]) (Pilsner et al., 2010). Investigators additionally 

26 examined effect modification by variants in the methylenetetrahydrofolate reductase 

27 (MTHFR) gene. The MTHFR enzyme is involved in folate metabolism, which, in turn, is 

28 involved in homocysteine methylation to the amino acid methionine. The transfer of 

29 methyl groups that results from folate metabolism is important for biological processes 

30 including Phase II detoxification reactions and epigenetic regulation of gene expression. 

31 The MTHFR gene has common functional variants, including the C677T SNP, which 

32 produces an enzyme with lower metabolic activity and is associated with lower serum 

33 folate levels (Kordas et al., 2009). Lower folate levels have been associated with higher 

34 blood Pb levels. Although Pilsner et al. (2010) found that both cord blood Pb levels and 

35 the MTHFR C677T allele were associated with lower child MDI score at age 24 months, 

36 they did not find a statistically significant interaction between blood Pb level and the 

37 MTHFR 677T allele. Results from stratified analyses were not reported, thus differences 

38 in the magnitude of association between genotypes could not be compared. 
February 2012 5-72 Draft – Do Not Cite or Quote

Specific Indices of Cognitive Function in Children

1 In addition to indices of global cognitive function, blood Pb levels also have been 

2 associated with specific cognitive abilities, including memory, learning, visuospatial 

3 processing, and other executive functions such as planning and problem solving in 

4 children and adolescents (Kordas et al., 2006; Tellez-Rojo et al., 2006; Chiodo et al., 

5 2004; Ris et al., 2004; Canfield et al., 2003b; Lanphear et al., 2000; Bellinger and Stiles, 

6 1993; Dietrich et al., 1992; Bellinger et al., 1991; Dietrich et al., 1991; Needleman et al., 

7 1979) (Figure 5-5 and Table 5-5). Studies often found associations with multiple indices 

8 of cognitive function, and because several tests are interrelated, it is difficult to attribute 

9 the effects of Pb exposure to a specific domain of cognitive function. Evidence that blood 

10 Pb levels are associated with a spectrum of cognitive indices provides coherence for the 

11 associations observed between blood Pb levels and FSIQ, a global measure of cognitive 

12 function. Furthermore, these tests of learning and memory in humans are homologous to 

13 tests in animals, and compared with that for FSIQ, evidence for these specific indices 

14 may improve understanding of the coherence between findings in humans and animals

15 (Rice, 1996). These complex cognitive functions have been described to be mediated by 

16 the actions of neurotransmitters dopamine and glutamate in the hippocampus, prefrontal 

17 cortex, and ventral striatum of the brain. In toxicological studies, Pb exposure has been 

18 shown to disrupt the function of these systems, which provides biological plausibility for 

19 associations observed between blood Pb levels and deficits in a range of executive 

20 functions in children.

21 Studies published since the 2006 Pb AQCD added to the evidence for associations 

22 between higher blood Pb level (concurrent most frequently examined) and lower 

23 performance in specific indices of cognitive function (Figure 5-5 and Table 5-5). The 

24 weight of evidence indicated that decrements in specific cognitive indices were 

25 associated with concurrent blood Pb level and in populations with mean blood Pb levels 

26 between 5-10 μg/dL. Most studies considered potential confounding by multiple SES27 related variables, and many found blood Pb-associated decrements in cognitive function 

28 in both unadjusted and covariate-adjusted models (Palaniappan et al., 2011; Chiodo et al., 

29 2007; Froehlich et al., 2007). Many studies did not examine potential confounding by 

30 HOME score. However, studies that did (Min et al., 2009; Solon et al., 2008; Froehlich et 

31 al., 2007; Canfield et al., 2004; Canfield et al., 2003b) and did not adjust for HOME score

32 (Krieg et al., 2010; Jedrychowski et al., 2008; Surkan et al., 2007; Lanphear et al., 2000)

33 both found associations between blood Pb level and decrements in specific cognitive 

34 indices.. Several studies found associations after adjusting for different potential 

35 confounding factors, including parental IQ or education and HOME score. The set of 

36 covariates varied among the specific cognitive endpoints, and HOME score was not 

37 associated with every cognitive index. For example, only household income remained 
February 2012 5-73 Draft – Do Not Cite or Quote

1 significantly associated with total errors in the spatial working memory task and was 

2 included in the final model (Froehlich et al., 2007). Solon et al. (2008) found that in a 

3 multivariable model with blood Pb level and several potential confounders, HOME score 

4 remained significantly associated with verbal IQ in 5 year-old children, whereas maternal 

5 education, maternal smoking, and premature birth did not. Combined, these findings 

6 support the independent association of blood Pb level with specific cognitive indices and 

7 indicate that the potential confounding variables vary among endpoints and study 

8 populations.

9 Several analyses of the Rochester cohort at ages 4 and 5 years indicated associations of 

10 blood Pb level with various indices of cognitive function, including visual and working 

11 memory, planning, and problem solving (Froehlich et al., 2007; Canfield et al., 2004; 

12 Canfield et al., 2003b). In particular, the associations observed with tests of memory and 

13 rule learning and reversal provided strong coherence with observations of Pb-induced 

14 impaired performance in animals in homologous tests of the Morris Water Maze, the 

15 delayed spatial alternation, and discrimination reversal learning (Section 5.3.2.2). 

16 Compared with other prospective cohort studies, findings from the Rochester cohort 

17 demonstrated associations between blood Pb level and decrements in executive functions 

18 in younger children with a lower mean blood Pb level (6.5 and 6.0 μg/dL at ages 4 and 

19 5 years, respectively). An additional strength of these studies was the detailed analysis of 

20 potential confounding by various demographic and SES-related variables (including 

21 HOME score) and other environmental exposures. Covariates were selected for specific 

22 models based on their association with a particular test (p ≤ 0.20), and several models 

23 adjusted for maternal IQ, maternal education, and HOME score. 

24 Canfield et al. (2003b) reported associations of concurrent blood Pb level with various 

25 parameters of the Shape School tasks, which test the ability of children to recall correctly 

26 shapes and colors that are attributed to specific cartoon characters. Concurrent blood Pb 

27 level was associated with color knowledge, shape knowledge, and number of correct 

28 responses at age 4 years (Figure 5-5 and Table 5-5). At age 5 years, higher lifetime 

29 average blood Pb level (area under the curve calculation using repeat measurements 

30 between age 6 months and 5 years) was associated with poorer performance on multiple 

31 tasks related to learning, working and spatial memory, and planning as assessed by the 

32 Cambridge Neuropsychological Testing Automated Battery (CANTAB) in both 

33 unadjusted and covariate-adjusted analyses (Canfield et al., 2004). These memory tests in 

34 children share homology with the Morris water maze test in animals in that both test the 

35 ability of subjects to use efficient search strategies to identify target locations, retain 

36 spatial information, and access remembered items in working memory. Both measure the 

37 time to complete the task and the number of errors made. At age 4 and 5 years, 

38 associations of concurrent blood Pb level with several indices of executive function were 
February 2012 5-74 Draft – Do Not Cite or Quote

1 robust to additional adjustment for attentiveness and child FSIQ, indicating that higher 

2 blood Pb level is associated with specific aspects of associative learning that are not 

3 represented by attentiveness and general knowledge base. 

a

Standard error was estimated from p-value. 

b

Sufficient data were not provided to calculate 95% CIs.

c

Direction of effect estimate was changed to indicate that a negative estimate represents poorer performance and a positive 

estimate represents improved performance. Black diamonds, gray squares, and blue circles represent associations with concurrent,

lifetime average, and earlier childhood blood Pb levels

Note: Regression coefficients were standardized to their standard deviation to facilitate comparisons among tests with different 

scales. Studies generally are presented in order of increasing mean blood Pb level. Effect estimates are standardized to a 1 μg/dL 

increase in blood Pb level within the 10th-90th percentile interval. The percentiles are estimated using various methods and are only 

approximate values. Effect estimates are assumed to be linear within the 10th to 90th percentile interval of blood Pb level. 

Figure 5-5 Standardized regression coefficients describing the associations 

of blood Pb levels with specific indices of cognitive function in 

children.

Study

Cho et al. (2010)

Lanphear et al. (2000)

Kreig et al. (2010)

Chiodo et al. (2007)

Chiodo et al. (2004)

Froehlich et al. (2007)

Canfield et al. (2004)

Canfield et al. (2003b)

Solon et al. (2008)

Min et al. (2009)

Palaniappan et al. (2011)

Blood Pb Mean

(SD) (μg/dL)

1.9 (0.67)

1.9 (geometric mean)

1.95 (0.16)

5.0 (3.0)

5.4 (3.3)

6.1 (4.9)

7.2 (3.6)

6.5

7.7 (7.7)

7.1 (5.1)

7.0 (range: 1.3-23.8)

11.5 (5.3)

10th-90th percentile

interval (μg/dL)

1.2-2.8

1.74-2.06

1.69-2.19

2.1-8.87

2.3-9.5

1.9-11.7

3.5-11.8

Data not available

1.6-14.9

3.0-12.1

6.0-18.3

Outcome

Color-word scorea

Digit Span

Math score

Digit Span

Reading score

Math score

Arithmeticb

Object assemblya

Spatial spanb

Perseverative errorsb

Spatial memorya

Rule learninga

Spatial spana

Nontarget errorsc

Total errorsc

Proportion correct

Color knowledge

Verbal IQ

Performance IQ

Verbal comprehension, age 9

Verbal comprehension, age 11

Math score, age 9

Math score, age 11

Reading score, age 9

Reading score, age 11

Visuomotor composite

◄

◄ ►

-0.6 -0.4 -0.2 0 0.2

Change in standardized test score per 1 μg/dL increase in blood Pb level 

within the 10th-90th percentile interval (95% CI)
February 2012 5-75 Draft – Do Not Cite or Quote

Table 5-5 Additional characteristics and quantitative results for studies 

represented in Figure 5-5

Study

Population/

Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Cognitive Index

Effect Estimate 

(95% CI)a

Cho et al.

(2010)

667 children ages 8-11 yr in 

2008

Five Korean cities

Concurrent

Mean (SD): 1.9 

(0.67)

10th-90th: 1.2-2.8

Log linear regression model 

adjusted for age, sex, parental 

education, maternal IQ, child IQ, 

birth weight, urinary cotinine

Color-word scorec

using Stroop test at ages 8-11 yr

0 (-0.09, 0.08)b

Lanphear et 

al. (2000)

4,852 children ages 

6-16 years

U.S. NHANES III 

(1991-1994)

Concurrent

Geometric mean: 1.9 

(95% CI: 1.70, 2.10)

10th-90th: 1.74, 2.06

Linear regression model adjusted 

for sex, race/ethnicity, poverty index 

ratio, reference adult education, 

serum ferritin levels, serum cotinine 

levels

Digit Span (WISC-R)

Math Score (WRAT-R)

at ages 6-16 yr

-0.02 (-0.03, -0.01)

-0.007 (-0.01, -0.001)

Krieg et al.

(2010)

842 children ages 12-16 yr

U.S. NHANES III 

(1991-1994)

Concurrent

Mean (95% CI): 1.95 

(1.63, 2.27)

10th-90th: 1.69-2.19

Log linear regression model 

adjusted for sex, caregiver 

education, family income, race- ethnicity, test language

Block design (WISC-R)

Digit span (WISC-R)

Reading score (WRAT-R)

Math score (WRAT-R)

assessed at ages 12-16 yr

-0.10 (-0.18, -0.02)

-0.08 (-0.13, -0.04)

-0.11 (-0.16, -0.05)

-0.07 (-0.12, -0.01)

Chiodo et al.

(2007)

506 African-American 

children in Detroit, MI area 

followed from birth 

(1989-1991) to age 7 yr.

Large proportions of 

children with prenatal 

exposure to cocaine or 

marijuana 

Concurrent

Mean (SD): 5.0 (3.0)

10th-90th: 2.1-8.7

Regression model adjusted for 

SES, caretaker education, maternal 

IQ (all outcomes)

Plus: age, HOME, Symptom 

Checklist (arithmetic and mazes)

Plus: sex, prenatal cigarettes/day, 

prenatal marijuana use (arithmetic)

Arithmetic

Mazes

Object assembly

WISC-III at age 7 yr

-0.07, p > 0.05c

-0.11 (-0.18, -0.04)

-0.13 (-0.21, -0.05)

Chiodo et al.

(2004)

246 African-American 

children in Detroit, MI area

followed from birth (not 

reported) to age 7.5 yr.

Large proportions of 

children with prenatal 

exposure to cocaine or 

marijuana

Concurrent

Mean (SD): 5.4 (3.3)

10th-90th: 2.3-9.5

Log linear regression model 

adjusted for SES, maternal 

education, sex (all outcomes)

Family Environmental Symptom 

(family functioning), number of 

children < 18 years, caregiver 

vocabulary, prenatal alcohol use 

(Perseverative errors)

Prenatal cocaine use (conceptual 

level responses)

HOME score, Symptom Checklist, 

caregiver age (Spatial span)

Corsi Spatial Span

WCST Perseverative errors

WCST Conceptual Level 

Responses

at age 7.5 yr

-0.10, p > 0.05c,d

-0.49, p > 0.05c,d

-0.77 (-1.5, 0)b

Froehlich et 

al. (2007)

174 children born 

1994-1995 followed from 

birth to age 5 yr

Rochester, NY

Concurrent

Mean (SD): 6.1 (4.9)

10th-90th: 1.9-11.7

Linear regression model adjusted 

for income (spatial memory); NICU, 

sex (rule learning); HOME score, 

maternal IQ, race (spatial span); or 

maternal IQ, transferrin saturation 

(problem solving) 

Spatial memory

Rule learning and reversal

Spatial span

Problem solving

using CANTAB at age 5 yr

-0.02 (-0.05. 0.01)b

-0.03 (-0.06, -0.001)b

-0.007 (-0.01, 0)b

-0.04 (-0.09, 0.01)b

Canfield et 

al. (2004)

174 children 174 children 

born 1994-1995 followed 

from birth to age 5 yr

Rochester, NY

Lifetime avg

Mean (SD): 7.2 (3.6)

10th-90th: 3.5-11.8

Linear regression model adjusted 

for maternal IQ, duration of 

breastfeeding, household income, 

maternal ethnicity, first prenatal visit 

(spatial span length), NICU 

admission, sex, spatial span length, 

spatial working memory problem 

(total nontarget errors, total errors), 

age at testing (nontarget errors), 

Maximum spatial span length

Total nontarget errors

Total errors

Using CANTAB at age 5 yr

-0.17 (-0.25, 0.59)d

-0.09 (-0.11, -0.06)d

0.04 (-0.07, 0.007)d
February 2012 5-76 Draft – Do Not Cite or Quote

Study

Population/

Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Cognitive Index

Effect Estimate 

(95% CI)a

Canfield et 

al. (2003b)

170 children born 

1994-1995 followed from 

age 6 mo to age 4 yr

Rochester, NY

Concurrent

Mean: 6.5

10th-90th: data not 

available

Linear mixed effects model adjusted 

for:

Sex, gestational age, birth order, 

maternal IQ, maternal education, 

prenatal smoking (correct 

responses)

Age, birth order, maternal IQ, 

maternal education, household 

income, marital status, HOME 

score, prenatal smoking (color 

knowledge)

Age, maternal IQ, household 

income, marital status, HOME 

score, race (shape knowledge)

Proportion correct responses

Color knowledge

Shape knowledge

using Shape School Task at 

ages 4 and 4.5 yr

-0.017 (-0.027, 0.007)

-0.074 (-0.101, -0.047)

-0.057 (-0.094, -0.020)

Solon et al.

(2008)

502 children ages 6-35 mo

377 children 3-5 yr,

2003-2004

Visayas, Philippines

Concurrent

Mean (SD): 7.1 (7.7)

10th-90th: 1.6-14.9

Two-stage linear regression model 

to account for determinants of blood 

Pb (sex, roof materials, water 

source, breastfed for ≥ 4 months) 

and cognitive function (HOME 

score, maternal education, maternal 

smoking, born premature, region of 

residence)

Verbal IQ

Performance IQ

using WPPSI-III at ages 3-5 yr

-0.34 (-0.63, -0.05)b

-0.45 (-2.37, 1.48)b

Min et al.

(2009)

267 primarily AfricanAmerican children in the 

Cleveland, OH area 

followed from birth 

(1994-1996) to age 11 yr. 

Children were exposed 

prenatally to multiple drugs.

Age 4 yr

Mean (range): 7.0 

(1.3-23.8)

10th-90th: 3.0-12.1

Linear regression model adjusted 

for HOME score, caregiver's 

vocabulary test, sex, parity, 

maternal marital status, head 

circumference at birth

Verbal comprehension age 9 yr

Verbal comprehension age 11 yr

Perceptual reasoning age 9 yr

Perceptual reasoning age 11 yr

Math at age 9 yr

Math at age 11 yr

Reading at age 9 yr

Reading at age 9 yr

Using WISC-R and WJTA

-0.028 (-0.055, 0)

-0.044 (-0.072, -0.015)

-0.033 (-0.064, -0.003)

-0.045 (-0.074, -0.016)

-0.028 (-0.057, 0.002)

-0.033 (-0.062, -0.004)

-0.039 (-0.069, -0.009)

-0.043 (-0.073, -0.014)

Palaniappan 

et al. (2011)

815 children ages 3-7 years 

in 2003-2006 in Chennai, 

India

Concurrent

Mean (SD): 11.5 (5.3)

10th-90th: 6.0-18.3

Generalized estimating equation 

adjusted for sex, hemoglobin level, 

maternal education, parental 

education, average monthly income, 

clustering at school and class level

Visuomotor composite

Using WRAVMA at ages 3-7 yr

-0.017 (-0.029, -0.005)

Studies not included in figure due to unavailability of blood Pb-cognitive function continuous effect estimates

Jedrychowski 

et al. (2008)

452 children followed from 

birth (2001-2003) to age 6 

mo

Prenatal (cord)

Mean (95% CI): 1.42 

(1.35, 1.48)

10th-90th: 1.36-1.46

Logistic regression model adjusted 

for maternal education, parity, sex

No maternal IQ

Verbal recognition memory 

score < 52.2

Using FTII at 6 mo

OR (95% CI)

1.47 (1.07, 2.01)

Surkan et al.

(2007)

534 children ages 6-10 yr

Boston, MA and 

Farmington, ME

Concurrent

Mean (SD): 2.2 (1.6)

Analysis of covariance adjusted for 

child IQ, caregiver IQ, age, SES, 

race, birth weight

WIAT at age 6-10 yr

Reading score

Math score

Visual memory score

Perseveration errors 

Stroop color-word interference

blood Pb level 5-10 

μg/dL vs. 1-2 μg/dLe

-5.20 (-9.45, -0.95) 

-4.02 (-7.6, -0.43) 

-6.47 (-11.9, -1.1)

-9.19 (-14.6, -3.7)d

0.75 (-1.6, 3.1)

Kordas et al.

(2006)

294 children with blood Pb 

levels < 10 μg/dL 

Concurrent

Range: 2.1-10.0

Linear regression model adjusted 

for sex, age, hemoglobin, family 

possessions, forgetting homework, 

house ownership, crowding, 

maternal education, birth order, 

family structure, arsenic exposure, 

tester, school

Visual memory span

< 10 correct vs. ≥ 10 correct: 

0.90

Stimulus discrimination 

< 20 correct vs. 20 correct

OR (95% CI)

0.90 (0.74, 1.10)

0.85 (0.63, 1.13)
February 2012 5-77 Draft – Do Not Cite or Quote

Study

Population/

Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Cognitive Index

Effect Estimate 

(95% CI)a

Bellinger and 

Stiles (1993)

148 children in the 

Boston, MA area followed 

from birth (1979-1981) to 

age 10 yr

57 month and 

concurrent blood Pb

Exact levels NR, 

mean reported to be 

< 8

Linear regression model adjusted 

for HOME score, family stress, race, 

marital status

Perseverative errors

57 month blood Pb

Concurrent blood Pb

Percent conceptual responses

57 month blood Pb

Concurrent blood Pb

Using WCST at age 10 yr

-0.05 (-0.09, -0.01)d

-0.05 (-0.11, 0.01)d

-0.93 (-2.0, 0.13)

NR

Bellinger et al.

(1994a)

79 children in the Boston, 

MA area followed from 

birth (1979-1981) to age 

19-20 yr

Deciduous tooth Pb

Q1: 2.9-5.9 μg/g

Q2: 6.0-8.7 μg/g

Q3: 8.8-19.8 μg/g

Q4: 19.9-51.8 μg/g

Regression model adjusted for 

parental IQ, sex, SES, current drug 

use, current alcohol use, current 

illicit drug use, maternal education, 

maternal age, birth order

Stroop Color-word test

WCST Number Correct

Mean (SE)

Q1: 103 (8.1), 

Q2: 116 (7.6), 

Q3: 127 (8.4), 

Q4: 125 (7.7)

Q1: 70.5 (2.9), 

Q2: 72.7 (2.8), 

Q3: 74.5 (3.1), 

Q4: 71.4 (2.9)

WISC = Wechsler Intelligence Scale for Children, WRAT = Wide Range Achievement Test, WCST = Wisconsin Card Sorting Test, CANTAB = Cambridge 

Neuropsychological Testing Automated Battery, WPPSI = Wechsler Preschool and Primary Scale of Intelligence, WRAVMA = Wide Range of Visual Motor 

Ability, FTII = Fagan Test of Infant Intelligence, WIAT = Wechsler Individual Achievement Test

a

Effect estimates are transformed to a z-score and standardized to a 1 μg/dL increase in blood Pb level in the 10th-90th percentile interval. b

95% CI was constructed using a standard error that was estimated from a p-value. c

Sufficient data were not provided to calculate 95% CI.

d

The direction of the effect estimate was changed such that a negative estimate represents poorer performance and a positive estimate represents better 

performance.

e

Effect estimates compare test performance of children in higher blood Pb groups to children in lowest blood Pb group.

1 A recent analysis of the Rochester cohort age 5 years extended previous findings with 

2 observations that higher concurrent blood Pb level also was associated with lower 

3 performance on many of the same tests executive function as assessed by CANTAB

4 (Froehlich et al., 2007) (Figure 5-5 and Table 5-5). In addition to spatial memory, 

5 Froehlich et al. (2007) found associations with rule learning and reversal tasks. Pb 

6 exposure of animals consistently has been found to induce impairments in homologous 

7 tests of cognitive flexibility, including delayed spatial alternation tests (Section 5.3.2.2). 

8 Both of these tests assess the ability of children and animals to complete a task according 

9 to a change in rules or reinforcement. Impaired performance in both children and animals 

10 is indicated by increased response errors, decreased percent of correct responses, and 

11 perseverance (i.e., animals repeatedly pressing the same lever without moving between 

12 the two locations or children repeatedly selecting the wrong target despite a change in 

13 rules or reinforcement. Some effect estimates describing the blood Pb-cognitive function 

14 association were attenuated after adjustment for covariates (e.g., spatial span and 

15 planning tasks), whereas others were magnified (e.g., total trials in rule learning, total 

16 errors in spatial working memory) (Froehlich et al., 2007). Further, despite attenuation, 

17 several associations did not change in statistical significance (Figure 5-5 and Table 5-5). 

18 Froehlich et al. (2007) also added to previous findings in the Rochester cohort by 

19 indicating that associations were modified by sex and dopamine receptor (DRD4) genetic 

20 variants. An increase in blood Pb level was associated with larger decrements in rule 
February 2012 5-78 Draft – Do Not Cite or Quote

1 learning and reversal and spatial memory in children with the DRD4 exon III 7-repeat 

2 microsatellite (assessed using a blood Pb-DRD4-7 interaction term, p = 0.042). The 

3 biological plausibility for these findings is provided by observations that the DRD4-7 

4 variant is associated with reduced dopamine-induced signaling in downstream pathways 

5 (e.g., cyclic AMP), that Pb exposure is found to impact dopaminergic activity 

6 (Section 5.3.8.8), and that dopamine is a key neurotransmitter that regulates cognitive 

7 processes. Associations of concurrent blood Pb level with rule learning and reversal also 

8 were greater in boys. Additional support for these observed interactions was provided by 

9 observations with children with the DRD4-7 variant and boys had poorer performance on 

10 rule learning and reversal tasks.

11 Several other studies found associations between higher blood Pb level and poorer 

12 performance on rule learning and reversal tasks in children and young adults (Surkan et 

13 al., 2007; Kordas et al., 2006; Chiodo et al., 2004; Bellinger et al., 1994a; Bellinger and 

14 Stiles, 1993). Across studies, the age range of subjects was 6 to 20 years, and most 

15 studies examined concurrent blood Pb levels. In the Boston cohort, age 4.5 year blood Pb 

16 levels were associated with poorer cognitive flexibility at age 10 years (Bellinger and 

17 Stiles, 1993), and deciduous tooth Pb (measured at ages 6-7 years) was associated with 

18 poorer cognitive flexibility at ages 19-20 years (Bellinger et al., 1994a).

19 Blood Pb-associated decrements in executive function in children with mean blood Pb 

20 levels < 5 were not as clearly demonstrated. Among children in five Korean studies, ages 

21 8-11 years, blood Pb level was not associated with performance a color-word association 

22 test. In the study of children (ages 6-10 years) participating in NECAT, Surkan et al.

23 (2007) provided information on the blood Pb-cognitive function concentration-response 

24 relationship. Children with concurrent blood Pb levels 3-4 μg/dL had lower scores 

25 compared with children with blood Pb levels 1-2 μg/dL; however, only a few 

26 associations were statistically significant (e.g., digit span, memory, motor speed). In 

27 analyses adjusted for caregiver IQ, age, SES, race, and birth weight, children with

28 concurrent blood Pb levels of 5 to 10 μg/dL had lower performance in a number of 

29 executive functioning domains such as working memory, visuospatial skills, cognitive 

30 flexibility, and ability to formulate, test, and adapt hypotheses (Figure 5-5 and Table 5-5). 

31 Compared with children with blood Pb levels of 1-2 μg/dL, children with blood Pb 

32 levels 5-10 μg/dL had lower scores on the reading (5.2 points [95% CI: 0.95, 9.45]) and 

33 math (4.0 points [95% CI: 0.43, 7.6]) composites of the Wechsler Individual 

34 Achievement Test. These decrements were robust to adjustment for child FSIQ. Thus, 

35 these findings combined with those from the Rochester cohort (Canfield et al., 2004; 

36 Canfield et al., 2003b) provide evidence that higher blood Pb level is associated with 

37 decrements in specific cognitive functions that may not be represented by FSIQ, a test of 

38 global function.
February 2012 5-79 Draft – Do Not Cite or Quote

1 In other studies of populations with mean concurrent or cord blood Pb level < 3 μg/dL, 

2 the contributions of likely higher past Pb exposures cannot be excluded. Also, these 

3 studies did not have as extensive consideration for potential confounding. In infants in 

4 Krakow, Poland with a mean cord blood Pb level of 1.42 μg/dL, higher cord blood Pb 

5 level was associated with a lower score on a visual recognition memory test

6 (Jedrychowski et al., 2008) (Table 5-5). The cord blood Pb levels may have been 

7 influenced by past Pb exposures of the mother mobilized from bone stores to the blood. 

8 While investigators adjusted for maternal education, parity, and sex, they did not adjust 

9 for other SES-related variables such as maternal IQ or HOME score. 

10 The influence of past Pb exposures also is uncertain in the NHANES analyses, which 

11 included adolescent subjects born in the 1970s. Krieg et al. (2010) examined an older 

12 subset of children (ages 12-16 years) from NHANES III (1988-1994) who were 

13 previously examined in Lanphear et al. (2000). Among all ages and older adolescents, 

14 higher concurrent blood Pb level was associated with lower scores on block design, digit 

15 span, reading score, and arithmetic tests (Figure 5-5 and Table 5-5) (Krieg et al., 2010; 

16 Lanphear et al., 2000). In analyses of quartiles of blood Pb level Lanphear et al. (2000)

17 additionally found a monotonic decrease in math and reading scores across increasing 

18 quartiles in covariate-adjusted models. Per unit increase in blood Pb level, the decrease in 

19 arithmetic and reading scores were larger among children with blood Pb levels less than 

20 2.5 μg/dL than among all subjects, with statistically significant associations found for 

21 children with blood Pb levels less than 5 μg/dL. While the NHANES results were 

22 adjusted for sex, race/ethnicity, test language, and multiple SES-related variables such as 

23 family income and caregiver education, information on HOME score was unavailable.

24 Krieg et al. (2010) provided additional information on effect modification by vitamin D 

25 receptor (VDR) variants. Although there were not differences in blood Pb levels among 

26 the various haplotypes of VDR, various polymorphisms and haplotypes modified the 

27 association between blood Pb level and a range of neurocognitive tests. The VDR 

28 regulates calcium absorption and metabolism, and effect modification by VDR variants is 

29 consistent with the well-established mode of action of Pb in mimicking calcium in shared 

30 transport and metabolic pathways. However, several inconsistencies were observed by 

31 Krieg et al. (2010) in that a particular variant was associated with a lower Pb-associated 

32 decrement in some indices but a greater Pb-associated decrement in other indices. For 

33 example, among children ages 12-16 years, the VDR rs2239185 CC genotype was 

34 associated with the largest blood Pb-associated decrease in digit span score and reading 

35 score. The decreases in digit span score (95% CI) per 1 μg/dL increase in concurrent 

36 blood Pb level were -1.5 points (-2.2, -0.71) for the CC genotype and -0.26 points (-0.99, 

37 0.46) for the TT genotype. Conversely, the TT genotype was associated with the greatest 

38 Pb-associated decrease in arithmetic score. The decreases (95% CIs) in math score per 
February 2012 5-80 Draft – Do Not Cite or Quote

1 1 μg/dL increase in concurrent blood Pb level were -8.4 points (-11.5, -5.2) for the TT 

2 genotype and -2.7 points (-10.1, 4.6) for the CC genotype. Effect modification by VDR 

3 rs731236 was more consistent across cognitive tests, with larger blood Pb-associated 

4 decrements in cognitive performance found in children with the CC genotype. It is 

5 important to recognize that not all of these VDR variants have been linked to functional 

6 changes.

7 Other recent studies conducted in diverse populations of children also found associations 

8 between higher blood Pb level and decrements in learning, memory, and visuospatial 

9 skills; however, they had limited or no consideration of potential confounding by SES10 related variables, which limits the implications of their findings (Nelson and Espy, 2009; 

11 Counter et al., 2008; Min et al., 2007; Vega-Dienstmaier et al., 2006).

Academic Performance in Children

12 As described in the preceding sections, a preponderance of evidence demonstrates Pb13 associated deficits in FSIQ and specific indices of cognitive function, including math, 

14 reading, and vocabulary skills. Deficits in global cognitive function and the 

15 aforementioned skills can lead to poorer academic achievement and school performance, 

16 which may be more objective measures of abilities and skills and have important 

17 implications for success later in life. Aptitude tests are used to predict future performance 

18 of an individual on a task or test. Achievement tests and school performance, in 

19 comparison, assess the actual knowledge of an individual in subject areas the individual 

20 has studied and measure the acquired knowledge of that subject. Studies reviewed in the 

21 2006 Pb AQCD consistently demonstrated associations of Pb biomarkers with measures 

22 of academic achievement and performance including scores on math or vocabulary tests, 

23 class rank, teacher assessment of academic functioning, and high school completion. 

24 Several studies found that blood or dentin Pb levels measured at an early age (ages 

25 2-8 years) were associated with academic performance at older ages (ages 8-18 years), 

26 suggesting the effect of early exposure to Pb may be persistent (Leviton et al., 1993; 

27 Bellinger et al., 1992; Needleman et al., 1990). Several studies found associations with 

28 concurrent blood Pb levels (Kordas et al., 2006; Wang et al., 2002a; Al-Saleh et al., 2001; 

29 Lanphear et al., 2000). Among recent studies, academic performance was examined less 

30 frequently; however, findings were consistent with the extant body of evidence.

31 The longitudinal study by Bellinger et al. (1992) was particularly noteworthy for 

32 examining associations of blood Pb level at several ages from the prenatal period to age 

33 10 years with Kaufman Test of Educational Achievement (KTEA) scores at age 10 years 

34 in the Boston cohort. While blood Pb levels at several ages were associated with lower 

35 academic achievement scores when in a model alone, only blood Pb level at age 2 years 
February 2012 5-81 Draft – Do Not Cite or Quote

1 showed a statistically significant association with lower predicted academic achievement 

2 after adjusting for HOME score, child stress, maternal age, race, maternal IQ, SES, sex, 

3 birth order, marital status, and number of residence changes. These covariates were 

4 selected because they were associated with blood Pb level and KTEA score, changed the 

5 blood Pb effect estimate by more than 10%, or because they were recognized to be 

6 important predictors of cognitive development (e.g., maternal IQ, birth order, sex). Age 

7 2 year blood Pb level was associated with a similar magnitude of decrease in the KTEA 

8 battery composite score in univariate and multivariate models. Additionally, the 

9 association was robust to adjustment for FSIQ, indicating that blood Pb levels may be 

10 associated with poorer performance on academic tasks not reflected in FSIQ. 

11 Miranda et al. (2007b) and (2009) linked blood Pb surveillance data collected between 

12 ages 0 and 5 years with end-of-grade (EOG) testing data for 8,600 fourth grade children 

13 in seven of the largest counties in North Carolina and 57,678 children in the entire state, 

14 respectively. A key strength of these analyses was the availability of individual-level data 

15 on blood Pb level and achievement score on a large number of children representative of 

16 the North Carolina fourth grade population, which provided large numbers of children 

17 with lower blood Pb levels (2-5 μg/dL). Thus, in this study, there was greater power to 

18 discern differences in achievement scores among children in the lower range of blood Pb 

19 levels. However, due to the records-based research design, investigators had a smaller set 

20 of available potential confounding variables than those typically considered in analyses 

21 of neurodevelopmental outcomes. In addition to adjusting for sex, race, school-type, 

22 school district, and age of blood Pb measurement, investigators adjusted for participation 

23 in a free or reduced lunch program as a measure of SES, and parental education as a 

24 measure of parental IQ. Miranda et al. (2007b) additionally adjusted for daily use of a 

25 computer as a measure of a stimulating home environment. Higher early childhood blood 

26 Pb levels were associated with lower reading and math EOG scores (Figure 5-6 and 5-7), 

27 and in both analyses, children with a blood Pb level of 2 μg/dL had lower EOG scores (p 

28 ≤ 0.05) compared with children with a blood Pb level of 1 μg/dL. Further, across deciles 

29 of blood Pb level, the decrease in EOG score generally was monotonic (Figure 5-6). In 

30 the statewide dataset, compared with children with an earlier (measured at some point 

31 between birth and age 5 years) blood Pb level of 1 μg/dL, children with an earlier blood 

32 Pb level of 2 μg/dL had a 0.30-point lower (95% CI: -0.58, -0.01) reading EOG score

33 (Miranda et al., 2009).
February 2012 5-82 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Elsevier Science, Miranda et al. (2007b).

Note: These results illustrate the decrease from a baseline score of 262.6 for a hypothetical referent white female, screened at 

2 years of age, living in Wake County, NC, with parents with a high school education, not enrolled in the school lunch program, and 

who does not use a computer every day (i.e., model covariates = zero).

Figure 5-6 Associations between childhood blood Pb levels and fourth grade 

End-of-Grade (EOG) math scores.

1 While the linear regression analyses indicated fractional-point decreases in EOG scores 

2 per 1 μg/dL increases in blood Pb level, Miranda et al. (2009) additionally used quantile 

3 regression to discern differential effects in various segments of the EOG distribution

4 (i.e., what is the 10th percentile of EOG scores conditioned on early childhood blood Pb 

5 levels). Compared with linear regression, quantile regression is more robust in response 

6 to outliers and predicts outcomes at the top and bottom tails of the outcome distribution 

7 rather than at the mean. With increasing blood Pb level, the lower tail of the EOG 

8 distribution was stretched out more so than were the middle or upper portions of the 

9 distribution. For example, an increase in blood Pb level from 1 to 5 μg/dL was associated 

10 with a greater decrease in EOG score among children in the 5th percentile of EOG than in 

11 children in the 95th percentile of EOG score (Figure 5-7). These findings indicated that 

12 children residing at the lowest performance regions of the EOG score distribution may be 

13 more affected by Pb exposure. Similarly, using quantile regression, Miranda et al. (2009)

14 showed that while cumulative social risk (lower parental education, being enrolled in a 

15 school lunch program) had a greater negative association with academic achievement in 

16 these children, blood Pb level was independently associated with EOG score decrements 

17 that were as large as 1 to 2 points.
February 2012 5-83 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Elsevier Science, Miranda et al. (2009)

Note: Baseline score calculated for a hypothetical referent individual with a blood Pb level of 1 μg/dL, parents completed college, 

and not enrolled in the school lunch program (i.e., model covariates = zero). 

Figure 5-7 Greater reduction in EOG achievement test scores with 

increasing blood Pb level in lower percentiles of the test score 

distribution.

1 Similar to Miranda et al. (2009), Chandramouli et al. (2009) observed associations 

2 between early childhood blood Pb levels (age 30 months) and later academic 

3 performance (Standard Assessment Tests [SAT] at age 7 years) among participants of the 

4 Avon Longitudinal Study of Parents and Children conducted in the U.K. However, in this 

5 study, decrements in SAT score were most clearly indicated in children with blood Pb 

6 levels of 5 to 10 μg/dL. Compared with children with blood Pb levels 0-2 μg/dL, children 

7 with blood Pb levels 5-10 μg/dL scored 0.51 points lower (95% CI: -0.82, -0.32) on the 

8 reading test and 0.49 points lower (95% CI: -0.78, -0.31) on the math test, adjusting for 

9 maternal education, home ownership, maternal smoking, home facilities score, parental 

10 SES, family adversity index, and parenting attitudes. Children with blood Pb levels 

11 2-5 μg/dL did not consistently have lower scores compared with children with blood Pb 

12 levels 0-2 μg/dL. While these aforementioned recent studies found associations for early 

13 childhood blood Pb levels, unlike the longitudinal assessment by Bellinger et al. (1992),

14 they did not have blood Pb measurements available at other lifestages. Therefore, recent 

15 studies could not provide information on associations with blood Pb levels at other 

16 lifestages. 
February 2012 5-84 Draft – Do Not Cite or Quote

5.3.2.2 Toxicological Studies of Cognition, Memory and Learning

1 As described in Section 5.3.2.1, epidemiologic studies in children have consistently 

2 demonstrated associations between blood Pb levels and decrements in a range of 

3 executive functions related to memory, reasoning, problem solving, and verbal skills. 

4 Studies in animals also demonstrate Pb-induced decrements in various indices of 

5 executive function, and several tests conducted in animals share homology with tests 

6 conducted in humans.

7 Data on neurocognition and learning as well as other nervous system endpoints with 

8 concentration-response data are shown in Figure 5-8 and accompanying Table 5-6. These 

9 data demonstrate that numerous studies in animal models have demonstrated that Pb 

10 exposure produces a spectrum of changes in the central nervous system, including 

11 deficits in CNS development and plasticity and altered homeostasis of mediators of 

12 cognitive function that manifest with deficits in memory, learning, and cognition. It is 

13 thought that early life Pb exposure can permanently alter CNS development and other 

14 pathways that contribute to these aforementioned memory and learning deficits, but the 

15 animal toxicology data show that multiple sensitive developmental windows for Pb 

16 exposure exist (Rice and Gilbert, 1990). As illustrated in Figure 5-8 and Table 5-6, Pb17 induced impairments in learning primarily previously were observed in animals with 

18 blood Pb levels > 30 μg/dL. Several new studies added to the evidence for impaired 

19 learning and memory in animals with lower blood Pb levels, 8-17 μg/dL (Cory-Slechta et 

20 al., 2010; Li et al., 2009c; Niu et al., 2009; Virgolini et al., 2008a; Stangle et al., 2007). In 

21 these studies of animals with lower blood Pb levels, Pb exposures began early, during the 

22 gestational or lactation period.

23
February 2012 5-85 Draft – Do Not Cite or Quote

Dosimetric representation reported by blood Pb level. (ID corresponds to Table 5-6.)

Figure 5-8 Nervous system summary array of toxicological outcomes after 

Pb exposure.
February 2012 5-86 Draft – Do Not Cite or Quote

Table 5-6 Summary of findings from neurotoxicological exposure-response 

array presented in Figure 5-8

Study ID

in Figure 

5-8 Reference

Blood Pb 

Level 

(μg/dL) Outcome

1 Beaudin et al. (2007) 13 & 31

Behavior, neonate: Lactational Pb exposure, offspring deficient in Reward Omission testing.

Physical development; Postnatal Pb exposure (birth to 4 weeks of age): Pb-dependent development of overreactivity to reward omission and errors is reversible with chelation treatment.

2 Grant et al. (Grant et al., 

1980) 57 Behavior, neonate: chronic Pb exposure (drinking water) to dams and pups, Changed behavior, males.

3 Kishi et al. (1983) 59 & 186

Behavior, neonate: Pb exposure (oral gavage of pups) during lactational period, Changed emotional behavior, 

males and females.

Motor function, neonate: Pb exposure (oral gavage of pups) during lactational period, motor function (rotarod 

performance) impaired, both sexes.

Physical development; Pb exposure during lactation (oral gavage): Delayed development of righting reflex in 

male rats.

4 Overmann (1977) 33, 174 & 

226

Behavior-Pb exposure (oral gavage of pups) during lactation: aversive conditioning affected by Pb exposure, 

male and females. 

Cognition-Pb exposure (oral gavage of pups) during lactation: Response inhibition impaired, both sexes.

Motor function- Pb exposure (oral gavage of pups) during lactation: Increased motor activity and impaired motor 

coordination (rotarod), male and females.

5 Stangle et al. (2007) 13 & 31

Cognition; Developmental Pb exposure (PND1-PND30): Impaired learning with visual discrimination task, 

heightened response to errors, both sexes.

Motor function; Developmental Pb exposure (PND1-PND30): Alcove latency and response latency significantly 

affected by Pb exposure, both sexes.

6 Gong & Evans (1997) 38 & 85

Cognition-Adult male 21 day Pb exposure: Hyperactivity with Habituation to new cage environment.

Morphology; 21 day Pb exposure to adult males: Marker of neuronal injury-elevated hippocampal glial fibrillary 

acidic protein (GFAP).

Physical development; Adult male rats (21 day Pb exposure): Neurotoxicity measured with brain glial fibrillary 

acidic protein (GFAP).

7 Rice (1990) 32 & 36 Cognition-Chronic Pb exposure from birth: Spatial discrimination reversal task impairment, both sexes.

8 Li et al. (2009c) 80 & 100

Cognition-Gestational & lactational Pb exposure: Morris water maze performance impaired.

Morphology; Gestational & lactational Pb exposure: Increased levels of inflammatory cytokines & exocytosis 

related proteins in brains of pups at weaning, both sexes.

Oxidative stress-gestational and lactational Pb exposure: Elevated hippocampal TNF levels in offspring, males 

and females.

9 Li et al. (2010b) 80 & 102

Cognition- Gestational & lactational Pb exposure: Morris water maze performance impaired.

Morphology: Increased levels of Alzheimer disease-associated proteins in mice with gestational and lactational 

Pb exposure, both sexes.

Proliferation/diff/survival, gestational & lactational Pb exposure: Increased hippocampal expression of P-tau and 

amyloid beta in male and female pups.

10 Cory-Slechta et al.

(2010) 10 & 13

Corticosterone: Lifetime Pb +/- stress: Correlation between 9-month old’s corticosterone level and frontal cortex 

dopamine levels in behaviorally tested female offspring. 

Stress: Corticosterone-Lifetime Pb plus stress: Affects FI performance, dopamine and serotonin levels in female 

offspring.

Stress: Corticosterone-neurotransmitter-Lifetime Pb exposure in female rats plus stress: Dopamine homeostasis 

affected.

11 Virgolini, Rossi-George, 

Weston et al. (2008b) 25

Corticosterone: Maternal Pb plus stress: Elevated corticosterone in male offspring with prenatal stress + offspring 

stress was further enhanced with Pb exposure. 

Stress: Corticosterone-Maternal Pb plus stress: Affects FI performance.

Neurotransmitter; Gestational and lactational Pb exposure: Induced DA and 5HT changes in rat offspring.
February 2012 5-87 Draft – Do Not Cite or Quote

Study ID

in Figure 

5-8 Reference

Blood Pb 

Level 

(μg/dL) Outcome

Stress induced neurotransmitter effects, Maternal Pb plus stress: 5-HT and 5-HIAA (5-HT metabolite), and 

dopamine turnover were significantly affected in males.

12 Virgolini et al. (2005) 15 & 27

Corticosterone: Chronic Pb exposure from weaning: Pb exposure alone decreased basal plasma corticosterone 

levels at 5 months of age, males.

Motor function: Chronic Pb exposure from weaning: Locomotor activity significantly decreased FI response rates 

& increased post-reinforcement pause period in a concentration-dependent manner, males. 

Stress: Corticosterone-Chronic Pb plus stress: Affects neurotransmitters & FI performance

13 Virgolini, Rossi-George, 

Lisek et al. (2008a)

31 Corticosterone: Maternal Pb exposure (gestation and lactation) +/- stress: Differential basal corticosterone levels 

between behavioral and non-behavioral tested rats, females.

11 &/or 31

Stress: Corticosterone-Maternal Pb plus stress: Affects FI performance, dopamine, serotonin, and NE levels.

Motor function: Maternal Pb +/- stress: Increased locomotor activity (run rate) with Pb and stress exposure.

31

Neurotransmitter; Gestational and lactational Pb exposure: Induced NE aberrations in adult rat offspring (both 

sexes).

Stress induced motor function: Maternal Pb +/- stress: Increased locomotor activity (run rate) with Pb and stress 

exposure.

Stress induced neurotransmitter; Gestational and lactational Pb exposure + stress: Induced HVA (monoamine 

neurotransmitter metabolite) and NE aberrations in female adult rat offspring.

14 Hu et al. (2008b) 4 & 12

Morphology; Gestational Pb exposure: Neurite outgrowth marker PSA-NCAM decreased in rat pups, both sexes.

Neurophysiology; gestation Pb exposure: decreased hippocampal sialyltransferase activity, both sexes.

Physical development; t-Gestational Pb exposure: Early brain synapse development impaired (hippocampal PSANCAM).

15 Wu et al. (2008a) 19 & 26

Morphology: Elevated expression of Alzheimer’s disease-related genes and Tc factors in aged brains of female 

monkeys (exposed to Pb as infants).

Oxidative stress: Elevated oxidative DNA damage in aged brains of female monkeys (exposed to Pb as infants).

16 Tavakoli-Nezhad et al.

(2001) 18, 29, & 54 Morphology; 3 to 6 weeks of Postnatal (starting at PND22) Pb exposure in males: Decreased number of 

spontaneously active midbrain dopamine neurons.

17 Leasure et al. (2008) 10 & 42

Motor function; Mouse maternal (dam) Pb exposure: Induced decreased rotarod performance in offspring (1 yearold male offspring).

Neurotransmitter; Mouse maternal (dam) Pb exposure: Affects 1 year old male offspring dopamine homeostasis, 

both sexes.

18 Fortune & Lurie (2009) 8 & 43 Neurotransmitter; Mouse maternal (dam) Pb exposure: Affects offspring superior olivary complex (auditory) 

neurotransmitters, both sexes.

Memory and Learning - Morris Water Maze

1 Blood Pb levels have been associated with decrements in both verbal (e.g., digit span) 

2 and spatial memory in children (e.g., spatial and visual memory span) (Figure 5-5 and 

3 Table 5-5). In the 2006 Pb AQCD, the results from different animal studies testing the 

4 effect of Pb on memory were mixed with impaired memory observed in animals with 

5 blood Pb levels 10-35 μg/dL but improved memory observed in other studies. Low-dose 

6 Pb was not found to affect short-term memory, i.e., recall of a learned task. Mixed results 

7 for memory also may be due to the fact that memory tests also test inattention. Memory 

8 tests may give incorrect results when opportunities exist for impaired attention to 

9 contribute to test results (U.S. EPA, 2006b). In animals, spatial memory is tested most 
February 2012 5-88 Draft – Do Not Cite or Quote

1 commonly using the Morris water maze. The 2006 Pb AQCD reported deficits in the 

2 Morris water maze performance with Pb exposure. New research continued to show Pb3 induced impaired Morris water maze performance. The Morris water maze tests spatial 

4 memory and learning by having a mouse swim and locate or remember the location of a 

5 platform submerged in opaque water. The Morris water maze is homologous to the 

6 spatial memory components of the CANTAB and WISC-R, which test the ability of 

7 subjects to recall correctly a sequence of locations (Froehlich et al., 2007; Canfield et al., 

8 2004; Chiodo et al., 2004).

9 Li et al. (2009c) examined Pb exposures from gestation through lactation. During this 

10 period, dams received Pb-acetate dissolved in drinking water (0.1%, 0.5%, and 1% with 

11 corresponding blood Pb levels of 4, 8 and 10 μg/dL at postnatal day [PND]21). 

12 Beginning at weaning, Pb-exposed pups were subjected to Morris water maze 

13 performance testing. Pb-exposed pups had statistically significant increases in escape 

14 latency and number of crossings of the platform area at 0.5% and 1% Pb-acetate exposure 

15 (blood Pb levels of 8 and 10 μg/dL, respectively), indicating impaired memory and 

16 learning (Li et al., 2009c). The pups in Li et al. (2009c) were not separated by sex. Cao et 

17 al. (2009) found that chronic administration of the supplement melatonin exacerbated Pb18 induced impairments in spatial memory and long-term potentiation. Adult male Wistar 

19 rats that had been exposed to Pb-acetate (0.2%) from birth received melatonin (3 mg/kg) 

20 from weaning via gastric gavage for 60 days. At this point (PND81-90), animals 

21 performed in the Morris water maze, and LTP of the hippocampal dentate gyrus was 

22 measured.

23 Another study found that various dietary supplements or methioninecholine concomitant 

24 with Pb exposure in weanling males shortened the escape latency of Pb-exposed pups to 

25 resemble more closely the escape latency of control pups (Fan et al., 2010; Fan et al., 

26 2009a). Zinc and methionine were effective dietary supplements in Fan et al. (2009a);

27 glycine, taurine, vitamin C, vitamin B1, tyrosine had no effect on the Pb-associated 

28 Morris water maze results. These data on the effect of early life Pb exposure on learning 

29 and memory in the Morris water maze affirm earlier findings including those by 

30 Kuhlmann et al. (1997) who used maternal Pb diet exposure (gestation and lactation), 

31 continuous Pb exposure (gestation through adulthood) or post-weaning Pb exposure and 

32 only found only significant impairments in the maternal and continuous exposure groups.

33 Another component of memory is working memory, which is the ability to temporarily 

34 keep information in mind while using the information to perform a related or unrelated 

35 task. The Morris water maze measures working memory in addition to learning. The 

36 2006 Pb AQCD reported that working memory as assessed by the Morris water maze was 
February 2012 5-89 Draft – Do Not Cite or Quote

1 significantly affected in chronic developmentally Pb-exposed (Pb-acetate in feed 10 days 

2 prior to mating through PND 21) female offspring at PND 21 (Jett et al., 1997).

3 Working memory also can be measured by testing delayed spatial alternation (DSA), 

4 which specifically tests spatial reversal learning. With DSA, an animal receives rewards 

5 based on alternating responses between two separate levers. In children, tests of spatial 

6 reversal learning includes the rule learning and reversal components of the CANTAB, 

7 Stroop Test, and WCST, and decrements in DSA have been associated with higher blood 

8 Pb levels in children (Figure 5-5 and Table 5-5). Studies detailed in earlier Pb AQCDs 

9 showed that Pb-exposed animals had deficits under DSA testing (Alber and Strupp, 1996; 

10 Rice and Gilbert, 1990; Rice and Karpinski, 1988; Levin et al., 1987; Levin and

11 Bowman, 1986). Studies in nonhuman primates showed Pb-induced behavioral 

12 impairment on DSA tasks (Rice and Gilbert, 1990) across many exposure periods, 

13 including early-life and chronic Pb exposures. Specifically, there were multiple lifestages 

14 during which Pb exposure induced deficits in this task. As in children, deficits in DSA 

15 included increased response errors, decreased percent of correct responses, and 

16 perseverance at a task (i.e., repeatedly pressing the same lever without moving between 

17 the two locations). These observations have been consistently made in nonhuman 

18 primates with continuous Pb exposure or juvenile to adult exposure but less consistently 

19 made in rats with juvenile only or juvenile to adult exposure. In fact, some rodent studies 

20 showed increased accuracy in the delayed alternation trials (Cory-Slechta et al., 1991).

21 Working memory is a subcategory of executive function or goal-oriented problem 

22 solving. Pb-induced deficits in working memory may be one of many factors that 

23 contribute to associations between blood Pb levels and inattention observed in humans

24 (Min et al., 2007; Surkan et al., 2007; Chiodo et al., 2004; Schweitzer et al., 2000; Stiles 

25 and Bellinger, 1993).

Learning - Y Maze

26 A recent study using the three-branch radial Y-maze showed Pb-induced effects on 

27 learning in rat offspring exposed during lactation and into adulthood (Niu et al., 2009).

28 The Y-maze has a light at the end of each branch. The branch with the illuminated light is 

29 a safe area whereas the other two branches are electrified and cause a mild electric shock 

30 when entered. The Y-maze test evaluates learning based on three criteria: 1) learning 

31 days or the number of days required to learn the maze (90% correctly); 2) the error 

32 number (EN) or the number of Y-maze runs required to learn the maze; and 3) the total 

33 reaction time (TRT) or the total amount of time spent in the maze per test day. Wistar 

34 Albino rat pups were exposed to Pb-acetate during lactation until the termination of the 

35 experiment at 12 weeks of age (dam drinking water 300 mg/L, blood Pb level: 17 μg/dL 

36 at 6 weeks of age. Rats were evaluated on the three Y-maze endpoints at 6, 8, 10 and 
February 2012 5-90 Draft – Do Not Cite or Quote

1 12 weeks of age.) Pb induced statistically significant impairments in learning, in 

2 particular, at 8, 10 and 12 weeks of age. EN and TRT were significantly affected only at 

3 8 and 10 weeks, respectively. Hippocampal glutamate levels were also significantly 

4 attenuated in Pb-exposed animals, and this is discussed in more detail in Section 5.3.8.4.

Learning - Response Inhibition and Schedule-Controlled Behavior Testing

5 Response inhibition is another measure of executive function and is measured with 

6 multiple tests of premature responses, decreased pause time between two scheduled 

7 events, and increased perseverance. These tests include Differential Reinforcement of 

8 Low Rates of Responding (DRL), Fixed Interval (FI) testing (Table 5-7), FI with 

9 Extinction (FI-Ext) or Fixed Ratio (FR)-FI, and Signal Detection with Distraction. 

10 Multiple studies from the 2006 Pb AQCD as well as earlier literature showed that early 

11 life Pb exposure contributed to response inhibition across the spectrum of these 

12 aforementioned tests. Monkeys with moderate blood Pb levels (11-13 μg/dL) learned the 

13 DRL task more slowly but eventually acquired reinforcement rates equal to that in 

14 controls. Newer data from female rats exposed to Pb (Stangle et al., 2007) continued to 

15 show animals with premature responses after Pb exposure or response inhibition 

16 decrements. These findings in animals are consistent with observations in children that 

17 blood Pb levels are associated with poorer impulse control (Figure 5-14 and Table 5-9). 

18 In children, impulsivity is commonly assessed by having parents or teachers rate problem 

19 behaviors such as trouble waiting, interrupting others, and responding at inappropriate 

20 times. However, studies also have specifically tested response inhibition assessed using 

21 the continuous performance test, which similar to animal tests, measures reactions to a 

22 stop signal (Section 5.3.3.1).

23 The 2006 Pb AQCD discussed learning or cognition as measured with schedule24 controlled behaviors including FI and FR operant conditioning and found that FI response 

25 rate was affected differentially with low-level (as low as 11 μg/dL) and high-level Pb 

26 (peak levels of 115 μg/dL) exposures increasing and decreasing FI response rate, 

27 respectively. This nonlinear response was since further explored in recent work, much of 

28 which also examined the interaction between stress and Pb exposure. 
February 2012 5-91 Draft – Do Not Cite or Quote

Table 5-7 Summary of effects of maternal and lifetime Pb exposure on 

FI performance

Pb (ppm)

Maternal Pbb Lifetime Pbc

Overall ratea PRPa Overall ratea PRPa

0 ppm:

0-PS No Significant Effectd No Significant Effect No Significant Effect No Significant Effect

0-OS No Significant Effect *↓ -23% No Significant Effect No Significant Effect

50 ppm:

50-NS No Significant Effect No Significant Effect *↑ 95% No Significant Effect

50-PS No Significant Effect No Significant Effect *↑ 79.2% *↓ -42%

50-OS *↑ 64.9% No Significant Effect *↑ 74.7% * ↓-39.3%

150 ppm:

150-NS *↑ 42.4% *↓ -30.3% No Significant Effect No Significant Effect

150-PS No Significant Effect *↓ -25.7% *↑ 90.7% *↓ -44.7%

150-OS *↑ 59.2% No Significant Effect *↑ 78.5% No Significant Effect

Note: *Dam blood Pb levels ranged from 5-13 μg/dL over gestation and lactation; offspring blood Pb ranged from 7-13 μg/dL from early life time points out 

to ten months of age. Thus, this study demonstrates that lifetime Pb exposure with or without prenatal stress induced learning deficits in female mice. 

Mechanistically, these authors propose that associations of Pb and stress with learning deficits may be related to aberrations in corticosterone and 

dopamine.

a

Based on calculation of group mean values across session block post-stress challenge for both maternal and lifetime Pb exposure studies. All calculations 

represent percent of 0-NS control values; ↑, represents increase; ↓, represents decrease. PRP = post-reinforcement pause. b

Data from Virgolini et al. (2005). *Denotes significant effect versus 0ppm control (p <0.05). c

Data from current study [Rossi-George et al. (2011)]

*Denotes significant effect vs. 0ppm control (p <0.05).

Source: Reprinted with permission of Elsevier Science, Rossi-George et al. (2011) (Table 1).

Learning Ability with Stress

1 The combined paradigm of Pb exposure and stress experienced by a laboratory animal is 

2 now being studied by multiple investigators who are focusing on the common pathway of 

3 HPA axis alteration and altered brain neurotransmitter levels. These studies indicated 

4 greater impairments in learning with Pb when combined with stress. These findings 

5 provide support for the association between higher blood Pb level and lower MDI score 

6 observed in children with mothers with low self-esteem but not children with mothers 

7 with high self-esteem (Surkan et al., 2008) (Table 5-4). In children, maternal self-esteem 

8 has been linked with the ability to cope with stress and improved maternal-child 

9 interactions. As indicated in Figure 5-8 and Table 5-6, Pb exposure has been shown to 

10 increase corticosterone levels and exacerbate Pb-induced dopamine release and learning 

11 ability. Cory-Slechta and colleagues have conducted multiple investigations in this area. 

12 Most recently, they showed enhanced learning deficits in female rat offspring following 

13 lifetime Pb exposure combined with maternal restraint or prenatal stress (Cory-Slechta et 

14 al., 2010). This exposure paradigm used dams who were exposed to Pb for 2 weeks prior 
February 2012 5-92 Draft – Do Not Cite or Quote

1 to mating through lactation and pups from a mixed sex litter from their dams during the 

2 aforementioned period and then via drinking water Pb (50 ppm) through the remainder of 

3 their lifetime. The resultant blood Pb levels of dams and pups ranged from 5 to 13 μg/dL.

4 In a separate investigation, Pb plus stress-related outcomes were examined in female 

5 offspring of dams who were exposed to Pb from 2 months prior to mating through 

6 lactation, i.e., developmental Pb exposure (2 exposure groups: 50 or 150 ppb Pb-acetate

7 in drinking water) (Virgolini et al., 2008a). Dams were subject to restraint stress at 

8 GD16-17. Compared with controls, marked increases in response rates on FI performance 

9 were found in the Pb plus stress female offspring, whose mean blood Pb level was 

10 11 μg/dL (50 ppb Pb-acetate). Because these animals did not show effects with maternal 

11 stress or Pb exposure alone, the results indicated a potentiation of effects with Pb and 

12 stress co-exposures. 

13 Similarly, lifetime Pb exposure (50 or 150 ppm, resulting in blood Pb levels of 11-16 and 

14 25-33 μg/dL, respectively) plus stress (maternal or offspring) also induced FI aberrations 

15 at the post-reinforcement pause (PRP) period in female offspring (Rossi-George et al., 

16 2011) (Table 5-7). Again, the results indicated a potentiation of effects. Within the FI 

17 schedule, the PRP represents timing capacity or proper temporal discrimination. Namely, 

18 the PRP is the period during which the animal must wait or pause before depressing the 

19 lever for a reward. In this case, Pb plus stress exposed animals started responding too 

20 early due to a decreased pause or PRP interval. Aberrant FI performance in infants and 

21 children has been used as a marker for impulsivity. Separately, overall FI response rate, a 

22 hyperactive behavior, was significantly increased with Pb exposure alone and with 

23 maternal or offspring stress at the 50 ppm exposure dose. At 150 ppm Pb, stress 

24 (maternal or offspring) increased FI response rate but Pb alone had no effect on FI. 

25 Biochemical analysis of possible mechanistic contributions to these aberrations revealed 

26 alterations in frontal cortex norepinephrine, reductions in dopamine homeostasis in the 

27 nucleus accumbens, and enhancement of the striatal monoamine system. This study on 

28 the effect of lifetime Pb exposure with or without stress on FI testing itself or during the 

29 PRP component of FI testing further affirms Pb-related learning deficits and provides 

30 possible mechanistic explanations.

31 Pb exposure over various developmental windows has been shown to affect 

32 corticosterone levels in rodents. These findings indicate that associations of Pb and stress 

33 with learning deficits (FI testing in females) may be related to aberrations in 

34 corticosterone and dopamine. Maternal Pb exposure (150 ppm drinking water from 

35 2 months prior to mating through lactation with restraint stress as detailed above) induced 

36 increased basal corticosterone in female and male offspring at 9 months of age; no 

37 interactions of Pb and stress were observed in this model (Cory-Slechta et al., 2004). By 
February 2012 5-93 Draft – Do Not Cite or Quote

1 14 months of age, these offspring had reduced corticosterone concentrations versus 

2 control animals, indicating a possible acceleration of age-related decreases in basal 

3 corticosterone levels (Cory-Slechta et al., 2008). Pb-induced decreases in corticosterone 

4 were enhanced with maternal stress. Postnatal exposure of male rodents to Pb (PND 

5 21-5 months of age) produced animals with significant decrements in baseline 

6 corticosterone; this effect produced a U-shaped concentration-response curve with 

7 significant decrements in basal corticosterone levels in the 50 ppm exposure group versus 

8 control (Virgolini et al., 2005). In summary, developmental (gestational and lactational) 

9 and post-weaning exposure to Pb induced changes in the HPA axis (corticosterone levels) 

10 in both sexes that were dynamic as the animal aged. 

11 Mechanistic understanding of the cognitive deficits observed with Pb and/or stress 

12 exposure was explored in a recent study. HPA hypofunction following dam Pb exposure 

13 (pup gestational and lactational Pb exposure) with or without maternal stress was 

14 reported (Rossi-George et al., 2011; Virgolini et al., 2008a). This study used the same 

15 model of developmental Pb exposure as is detailed in the preceding paragraph. Outcomes 

16 were examined in both male and female offspring. At 2 months of age without stress, 

17 basal corticosterone in females was significantly increased with 150 ppm Pb (resulting in

18 blood Pb level of 32 μg/dL at PND21) and 50 ppm Pb (resulting in blood Pb level of 

19 19 μg/dL) (Figure 5-9). Pb plus stress attenuated the Pb-induced elevations in 

20 corticosterone to baseline levels (Figure 5-9). At age 10 months, Pb and stress accelerated 

21 the age-dependent decrease in corticosterone levels in females. In males, basal 

22 corticosterone levels were not affected significantly by Pb and/or stress at 2 or 10 months 

23 of age (Figure 5-9). These authors also explored the function of the glucocorticoid 

24 negative feedback loop using the dexamethasone suppression test and found that Pb 

25 and/or maternal stress significantly impacted this negative feedback by increasing nuclear 

26 glucocorticoid receptor levels. This negative feedback loop was impacted more at the 

27 lower dose (50 ppm versus 150 ppm Pb-acetate). As summarized in Table 5-7, the results 

28 indicate that lifetime Pb exposure when combined with stress can exacerbate impairments 

29 in learning. The interaction between Pb and stress may be mediated via effects on 

30 corticosterone and dopamine. 
February 2012 5-94 Draft – Do Not Cite or Quote

Source: Rossi-George et al. (2011)

Note: *denotes significantly different from NS control; # denotes significantly different from corresponding Pb-NS value; + differs 

from 50-NS. (0, 50, 150 ppm) and/or stress (PS [dam stress] or OS [PS followed by offspring stress]).

Figure 5-9 Mean basal corticosterone levels of female and male offspring 

exposed to lifetime Pb.

1 Another study examined the effects of Pb on the HPA axis but examined the interaction 

2 with an outside stress administered using control vehicle injections (Rossi-George et al., 

3 2009). The corticosterone response to this vehicle injection stress was prolonged in a 

4 nonlinear concentration-dependent manner in both sexes with the most profound effects 

5 observed at the lower 50 ppm Pb dose. Maternal stress also prolonged the corticosterone 

6 stress response to vehicle injection and enhanced the Pb effect in males. To test the 

7 negative feedback of the HPA axis, exogenous dexamethasone (DEX) was administered 

8 to suppress endogenous corticosterone. The DEX test revealed HPA axis hypofunction. 

9 Specifically, Pb and Pb plus maternal stress initially reduced the ability of DEX to 

10 suppress corticosterone. With time, the effect of DEX in males induced prolonged 

11 corticosterone suppression or failure to return to baseline as was observed in control 

12 animals. In summary, dam Pb exposure induced HPA negative feedback hypofunction in 

13 both sexes of offspring with an inverse U concentration-response function. 

14 These studies of lifetime exposure (2011; Rossi-George et al., 2009) reported different 

15 basal stress hormone levels with Pb exposure. Males with lifetime Pb exposure had no 

16 statistically significant corticosterone response to Pb exposure; whereas males with dam 

17 Pb exposure had statistically significant decreases in corticosterone at 5 months of age in 
February 2012 5-95 Draft – Do Not Cite or Quote

1 the 50 ppm exposure group only (not in 150 ppm Pb exposure group). On the other hand, 

2 females had concentration-dependent corticosterone responses to Pb exposure in both 

3 exposure models (lifetime Pb exposure and dam Pb exposure). Maternal stress alone also 

4 led to HPA axis negative feedback hypofunction. Pb plus maternal stress enhanced 

5 negative feedback in males and attenuated this effect in females. Pb exposure with or 

6 without maternal stress prolonged the effect of DEX-dependent corticosterone 

7 suppression in males. These data together show that HPA axis alterations could provide a 

8 link between the contribution of Pb and stress to health effects. 

9 Schedule-controlled behavior is often measured using FI or FR testing. Because the FI 

10 animals are regularly handled by laboratory personnel and participate in tests of 

11 cognition, their baseline level of stress may be skewed from that of a laboratory animal 

12 that constantly remains in a cage without daily handling. Because effects on the HPA axis 

13 are of interest to Pb researchers, the baseline corticosterone levels of animals that have 

14 participated in behavior testing (FI) and those who have not (NFI) have been compared. 

15 Specifically, the corticosterone differences between FI and NFI animals after 

16 developmental Pb exposure (dam-only Pb exposure) have been measured. Virgolini et al.

17 (2008b) found that basal corticosterone levels were significantly different between FI and 

18 NFI animals. Also, the combination of dam Pb exposure with maternal stress was 

19 explored in FI and NFI animals. At the baseline age of 4-5 months, NFI animals 

20 displayed significant differences from FI animals. Pb exposure with or without stress did 

21 not induce differences in corticosterone levels in FI females. The corticosterone level of 

22 male FIs was affected by Pb and stress exposure (Virgolini et al., 2008b). In the FI males, 

23 the 50 ppb Pb exposure group (50Pb) had decreased corticosterone versus control (no Pb 

24 exposure) and the 150 ppb Pb exposure group (150Pb) had elevated corticosterone versus 

25 control. Male NFI animals showed a U shaped concentration-response curve with 50Pb 

26 animals having significantly less corticosterone than did control or 150Pb animals. In the 

27 NFI males, stress did not affect corticosterone levels or interact with the effect of Pb. NFI 

28 females exposed to 150Pb had significantly elevated corticosterone versus control (no Pb 

29 exposure). These data demonstrate that behaviorally trained animals have altered HPA 

30 axis and response to Pb exposure versus animals that are housed under conditions without 

31 daily handling by caregivers. 

32 Virgolini et al. (2008b) also expanded evidence for Pb exposure-stress interactions 

33 through the examination of the effects additional intermittent stress as an adult. The 

34 authors proposed that associations of Pb and stress with learning deficits (FI testing in 

35 females) may be related to aberrations in corticosterone and dopamine. Dam exposure to 

36 Pb (50 or 150 ppm Pb-acetate) followed by intermittent stress (cold, novelty or restraint) 

37 in offspring as adults induced statistically significant changes in FI response rate. 

38 Females were more sensitive to the adult intermittent stressors at the higher dose of Pb 
February 2012 5-96 Draft – Do Not Cite or Quote

1 (150 ppm) with statistically significant increases in FI response rate and decreased PRP, 

2 i.e., increased impulsivity (Figure 5-10). Males were more sensitive (decreased FI 

3 response rate due to decreased run rate) to the restraint stress at the lower Pb dose 

4 (50 ppm). At the higher dose of Pb, males were more sensitive to the cold stress 

5 (increased FI response rate and increased run rate) (Figure 5-11). Corticosterone levels 

6 were examined in this study and showed concentration-dependent correlations with FI 

7 outcomes in females but were independent of dose in males.

8 Another study examined female rats with lifetime Pb exposure combined with prenatal 

9 stress and found enhanced learning deficits (drinking water 50 ppm Pb-acetate, resulting 

10 in offspring blood Pb levels: 7-13 μg/dL) (Cory-Slechta et al., 2010). Learning was 

11 evaluated with multiple schedule of repeated learning (RL) and performance testing. RL 

12 was impaired, but performance was not affected with Pb exposure. The Pb-impaired RL 

13 was further enhanced with prenatal stress. There were statistically significant associations 

14 between Pb/stress and corticosterone concentration, dopamine from the frontal cortex, 

15 dopamine turnover in the nucleus accumbens, and total number of responses required to 

16 learn a sequence. Also, Pb-exposed offspring with and without maternal stress exposure 

17 had statistically significant decreases in hippocampal nerve growth factor (NGF) versus 

18 controls. Thus, this study demonstrated that lifetime Pb exposure with or without prenatal 

19 stress induced learning deficits in female mice. 
February 2012 5-97 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Elsevier Science, Virgolini, Rossi-George, Weston, et al. (2008b).

Figure 5-10 Changes in FI performance (FI overall performance, run rate, PRP) 

in female offspring with maternal Pb exposure plus various 

stressors (restraint, cold, novelty) in adulthood.
February 2012 5-98 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Elsevier Science, Virgolini, Rossi-George, Weston, et al. (2008b).

Figure 5-11 Changes in FI performance (FI overall performance, run rate, PRP) 

in male offspring with maternal Pb exposure plus various 

stressors (restraint, cold, novelty) in adulthood.

Cognitive Flexibility

1 Cognitive flexibility is a component of executive function that measures the ability to 

2 reallocate mental resources when situations change (Monsell, 2003). This flexibility is 

3 assessed as the ability to alter behavioral responses according to the context of the 

4 situation and incorporates the function of attention, working memory, and visual 

5 processing. Discrimination reversal learning and concurrent random interval (RI-RI) 

6 scheduling are used to measure cognitive flexibility. The 2006 Pb AQCD reported 

7 discrimination reversal learning deficits in monkeys with blood Pb levels of 11-20 μg/dL. 

8 Rats also showed similar deficits but the authors attributed the changes to learning-related 

9 problems instead of cognitive flexibility (Garavan et al., 2000; Hilson and Strupp, 1997).

10 Interestingly, recent work has shown that N-Methyl-D-aspartic acid or N-Methyl-D11 aspartate (NMDA) receptors and D2-like receptors, two well-characterized targets of Pb, 

12 are involved in discrimination reversal learning (Herold, 2010). Another test of cognitive 

13 flexibility is called concurrent random interval (RI-RI) scheduling in which depression on 

14 two response levers is reinforced at different frequencies. The 2006 Pb AQCD reported 

15 monkeys with Pb-induced cognitive flexibility impairment under RI-RI (Newland et al., 

16 1994). Coherence for these observations of Pb-induced deficits in cognitive flexibility is 

17 provided by consistent evidence in animals and children for associations of Pb with 
February 2012 5-99 Draft – Do Not Cite or Quote

1 individual tests of inattention, memory, and visual processing. Studies in children also 

2 demonstrate associations between blood Pb levels and decrements in specific tests of 

3 cognitive flexibility that are homologous to tests in animals such as discrimination 

4 reversal learning, the WCST, and Stoop test (Figure 5-5 and Table 5-5). While tests in 

5 animals measure the ability to complete a task according to changes in reinforcement, 

6 tests in humans assess the ability of subjects to complete a task according to changes in 

7 rules.

5.3.2.3 Toxicological Studies on the Effects of Chelation

8 Earlier work in the animal toxicological literature has shown that succimer or chelation 

9 treatment of Pb-exposed lab animals was able to normalize various aberrant Pb-induced 

10 behaviors including activity level, habituation (Gong and Evans, 1997) and forced-swim 

11 immobility (Stewart et al., 1996). A more recent study examined the effect of succimer 

12 treatment on various behavioral and cognitive outcomes in control and neonatally Pb13 exposed female animals (PND 1-30 Pb-acetate exposure, 300 ppm dam through lactation 

14 and either 30 or 300 ppm pup water) by drinking water, generating a moderate Pb (m-Pb) 

15 exposure and a high Pb (h-Pb) exposure group. Pb blood levels at PND52 in the control, 

16 m-Pb, h-Pb, m-Pb+succimer, and h-Pb+succimer were 1.5; 12.6; 31; 2.8; and 8.5 μg/dL, 

17 respectively. Brain Pb levels at the same time for the same groups were 41 (control),

18 1,040 (m-Pb), 3,690 (h-Pb); and 196 (m-Pb+succimer) and 1,370 (h-Pb+succimer) ng/g 

19 dry weight. Succimer treatment significantly attenuated the m-Pb induced impaired 

20 learning ability. Effects on arousal that were significantly affected in h-Pb rats were 

21 significantly attenuated with succimer treatment. Succimer treatment in the h-Pb animals 

22 only slightly improved learning ability but did not improve the impaired inhibitory 

23 control (Stangle et al., 2007). These are important findings because they provide evidence 

24 that certain neurobehavioral or cognitive impairments associated with Pb exposure appear 

25 to be reversible with chelation therapy.

26 In another study, a 3-week course of Pb-acetate (PND 1-17, dam drinking water) plus or 

27 minus succimer/chelator (PND 31-52) treatment was given to determine if succimer 

28 could alleviate behavioral deficits in rats exposed to Pb for the first 4 weeks of life. Pb29 exposed animals had altered reactivity and increased reward omission and errors. Pb30 exposed animals receiving chelation treatment had normalized reactivity to reward 

31 omission and errors (Beaudin et al., 2007). Pb-induced behavioral abnormalities were 

32 attenuated with chelation therapy.

33 The interaction between Pb and methionine choline also has been examined (Fan et al., 

34 2009a). Pb-exposed rats were supplemented with methionine choline to provide 
February 2012 5-100 Draft – Do Not Cite or Quote

1 information on its effect on Pb disposition in various tissues (blood, bone, brain) and its 

2 contributions to neurocognitive or neurobehavioral changes. As a sulfur source, 

3 methionine is a chelator and a free-radical scavenger. Choline is important for cell 

4 membranes and neurotransmitter synthesis (Zeisel and Blusztajn, 1994). In this model, 

5 methionine choline attenuated Pb-induced memory and learning deficits 

6 (Section 5.3.2.2). Exposure of weanling male rats to Pb-acetate in drinking water 

7 (300 mg/L) through PND60 produced a blood Pb level of 60 μg/dL, bone Pb level of 

8 165 μg/g, and brain Pb level of 0.63 μg/g. Methionine choline supplementation 

9 significantly attenuated blood and bone Pb levels but produced a nonsignificant 

10 attenuation of brain Pb (0.51 μg/g) in rats that had significant improvements in learning 

11 and memory (Section 5.3.2.2). Also, in another study, the metal chelators DP-109 and 

12 DP-460 were neuroprotective for Pb-related effects in the ALS mouse neurodegenerative 

13 model or Tg(SOD1-G93A) model (Petri et al., 2007).

14 In summary, succimer or chelation treatment appears to be able to restore Pb-dependent 

15 impairments of learning and arousal and be neuroprotective in a concentration-dependent 

16 fashion. In these studies, succimer use was more efficacious at lower doses of Pb 

17 exposure. Chelation did not restore Pb-impaired inhibitory control. Chelation with the 

18 antioxidant supplement affected the disposition of Pb in various tissues, significantly 

19 attenuating blood and bone Pb levels and nonsignificantly attenuating brain Pb. 

5.3.2.4 Integrated Summary of Cognitive Function in Children

20 Results from recent epidemiologic studies and animals studies expand the strong existing 

21 evidence base demonstrating that Pb exposure is associated with impaired cognitive 

22 function in children. A large epidemiologic evidence base demonstrates associations of 

23 higher blood Pb level with lower FSIQ in school-aged children, lower MDI scores in 

24 children ages 6 months to 3 years, and lower scores on tests of specific cognitive 

25 functions in children ages 6 months to age 16 years (Figure 5-2 and Figure 5-5; Table 

26 5-3, Table 5-4, and Table 5-5). Blood Pb level has been associated with a range of 

27 cognitive indices, including memory, verbal and math skills, and cognitive flexibility. 

28 There was no clear indication that blood Pb level was more strongly associated with 

29 performance in a particular domain of cognitive function. Studies in animals clearly 

30 demonstrated impaired performance on Morris Water Maze, the delayed spatial 

31 alternation, and discrimination reversal learning with Pb exposure. These animal 

32 observations provided strong coherence with associations in children with homologous 

33 tests of spatial memory and rule learning and reversal (Figure 5-5 and Table 5-5). Several 

34 studies found that blood or dentin Pb levels measured at an early age (ages 2-8 years) 

35 were associated with poorer academic performance at older ages (ages 8-18 years), 
February 2012 5-101 Draft – Do Not Cite or Quote

1 suggesting the effect of early exposure to Pb may be persistent and have important 

2 implications for success later in life.

3 The weight of epidemiologic evidence for Pb-associated cognitive function decrements 

4 was provided by several prospective studies conducted in diverse populations that found 

5 associations with blood Pb levels measured during various lifestages (concurrently, 

6 prenatally (cord or maternal), earlier in childhood, and averaged over multiple years) and 

7 had extensive evaluation of potential confounding variables. The association with FSIQ 

8 was substantiated in a pooled analysis of children, 5 to 10 years of age, participating in 

9 seven prospective studies (Boston, MA; Cincinnati, OH; Rochester, NY; Cleveland, OH; 

10 Mexico City, Mexico; Port Pirie, Australia; and Kosovo, Yugoslavia) (Lanphear et al., 

11 2005). Several new studies added evidence for associations of FSIQ with concurrent 

12 blood Pb level in populations with lower mean blood Pb levels. Previously, the weight of 

13 evidence supported these associations in populations with mean blood Pb levels in the 

14 range of 5-10 μg/dL. However, several new studies shifted the weight of evidence to 

15 lower blood Pb levels (primarily concurrent), with populations means in the range of 

16 2-7 μg/dL (2011; Kim et al., 2009b; Min et al., 2009; Zailina et al., 2008; Chiodo et al., 

17 2007). Studies of specific cognitive indices and academic performance did not 

18 consistently find decrements in performance in children with blood Pb levels < 5 μg/dL

19 (Cho et al., 2010; Miranda et al., 2010; Chandramouli et al., 2009; Surkan et al., 2007).

20 Several new toxicological studies added to the evidence for impaired learning and 

21 memory in animals with lower blood Pb levels, 8-17 μg/dL (Cory-Slechta et al., 2010; Li 

22 et al., 2009c; Niu et al., 2009; Virgolini et al., 2008a; Stangle et al., 2007). A large body 

23 of new evidence from Cory-Slechta and colleagues demonstrated that lifetime Pb 

24 exposure when combined with stress in animals exacerbated impairments in learning

25 (2011; Cory-Slechta et al., 2010; Rossi-George et al., 2009; Virgolini et al., 2008a).

26 Findings also indicated that these interactions potentially were mediated via effects on 

27 corticosterone and dopamine. 

28 With regards to important lifestages of Pb exposure, the weight of toxicological evidence 

29 demonstrates impaired learning and memory in animals exposed to Pb gestationally with 

30 or without early postnatal exposure. In particular, impairments in learning and memory 

31 observed with lower blood Pb levels (8-17 μg/dL), were found with Pb exposures that 

32 began during the gestational or lactation period. The prospective epidemiologic studies 

33 found decrements in cognitive function of children in association with concurrent, 

34 prenatal (cord and maternal), early childhood, and cumulative average blood Pb levels. 

35 However, collectively, based on the frequency of examination, the weight of evidence 

36 demonstrates associations between cognitive function decrements and concurrent blood 

37 Pb levels in children ages 4-10 years. Among studies that examined MDI in children ages 

38 6 months to 3 years, several found stronger associations of MDI with prenatal (maternal 
February 2012 5-102 Draft – Do Not Cite or Quote

1 or cord) blood Pb than with postnatal child blood Pb (Hu et al., 2006; Bellinger et al., 

2 1987; Dietrich et al., 1987a; Vimpani et al., 1985). Child concurrent or lifetime average 

3 blood Pb levels also were associated with MDI scores (Claus Henn et al.; Solon et al., 

4 2008; Surkan et al., 2007; Tellez-Rojo et al., 2006; Wasserman et al., 1992; Vimpani et 

5 al., 1985). Thus, both postnatal child and maternal Pb exposures may contribute to lower 

6 cognitive function in young children. It is important to note that in studies that examined 

7 maternal pregnancy or cord blood Pb levels, there is greater uncertainty regarding the 

8 relative contributions of maternal past and recent Pb exposures that contributed to 

9 associations. The influence of past or cumulative maternal Pb exposures was supported 

10 by observations that maternal patella Pb levels were associated with MDI among children 

11 at age 2 years in Mexico City (Gomaa et al., 2002). Nonetheless, results for prenatal 

12 blood Pb levels and concurrent child blood Pb levels measured during specific postnatal 

13 ages indicate that relatively short-duration Pb exposures (< 1 year) influence the 

14 cognitive development of children between birth and age 3 years. Short-duration Pb 

15 exposures may be especially important during early childhood because processes such as 

16 neurogenesis and synaptic pruning are highly active during the first few years of life

17 (Rice and Barone, 2000; Landrigan et al., 1999).

18 Epidemiologic studies found independent associations of blood Pb level after adjusting 

19 for multiple potential confounding factors, including parental IQ, SES, household 

20 income, and HOME score. In most studies that provided unadjusted and adjusted effect 

21 estimates, blood Pb level was found to be associated with a smaller but statistically 

22 significant decrement in FSIQ after adjusting for potential confounding factors

23 (Palaniappan et al., 2011; Kim et al., 2009b; Chiodo et al., 2007; Froehlich et al., 2007; 

24 Lanphear et al., 2005; Canfield et al., 2003a). Analyses of associations of covariates with 

25 blood Pb level and cognitive function indicated that the potential confounding variables 

26 may vary across populations and endpoints. HOME score was not associated with every 

27 cognitive index. Studies that adjusted for multiple SES-related factors but did not 

28 examine HOME score produced similar magnitudes of associations as did studies that

29 adjusted for HOME score (Table 5-3, Table 5-4, toTable 5-5). Thus, confounding by 

30 HOME score may be minimized when other correlated variables are accounted for in 

31 analyses. Further, it is uncertain the extent to which HOME score alone may confound 

32 blood Pb-IQ associations as studies have not reported the magnitude of change in the 

33 blood Pb effect estimate with just the addition of HOME score in the model. Thus, while 

34 the caregiving environment can be an important confounder, the overall weight of 

35 evidence indicates that it does not mitigate the strong findings linking higher blood Pb 

36 levels with lower FSIQ in children. Prenatal drug exposure also was not found to have a 

37 large influence on the relationship between blood Pb level and FSIQ (Min et al., 2009; 

38 Chiodo et al., 2007). That Pb exposure induces impairments in tests of learning and 

39 memory in animals further that are directly homologous to tests conducted in children 
February 2012 5-103 Draft – Do Not Cite or Quote

1 further demonstrates that confounding by SES and HOME does not fully account for the 

2 associations between blood Pb levels and cognitive function observed in children.

3 The evidence from epidemiologic and toxicological studies linking Pb exposure with 

4 decrements in multiple cognitive functional domains is strengthened by the well5 characterized toxicological evidence for Pb exposure interfering with development of the 

6 brain and activity of neurochemical processes that mediate cognitive function 

7 (Section 5.3.8). Pb has been shown to increase the permeability of the blood-brain barrier 

8 and deposit in the target central nervous system. Pb has been shown to impair 

9 neurogenesis, synaptic architecture, and neurite outgrowth. The high activity of these 

10 processes during fetal and infant development provides biological plausibility for young 

11 children being particularly at increased risk for Pb-associated impairments in cognitive 

12 function. Cognitive function is mediated by the cortical and subcortical structures of the 

13 brain that integrate function in the hippocampus, prefrontal cortex, and nucleus 

14 accumbens using dopamine and glutamate as primary neurotransmitters. Experimental 

15 studies have shown that Pb induces changes in dopamine release in these regions. 

16 Numerous studies also have shown Pb-induced changes in hippocampal function, 

17 including changes in glutamate release, receptor binding, and long-term potentiation. 

18 Thus, several lines of toxicological evidence establish a neuroanatomical and 

19 neurochemical basis for the effect of Pb on cognitive function

5.3.2.5 Epidemiologic Studies of Cognitive Function in Adults

Adults without Occupational Lead Exposures

20 As described in the preceding section, Pb exposure of animals that begins in gestation 

21 and lasts through the early postnatal period or for a lifetime has been shown to induce 

22 learning impairments in adult animals. Less well characterized are learning impairments 

23 in adult animals due to adult-only Pb exposures. In contrast, epidemiologic studies have 

24 examined cognitive performance in adults primarily in association with concurrently 

25 measured blood and bone Pb levels. For nonoccupationally-exposed adults, the 2006 Pb 

26 AQCD cited some evidence for associations of cognitive performance with bone Pb 

27 levels but not blood Pb levels (U.S. EPA, 2006b). Studies published since the 2006 Pb 

28 AQCD continued to indicate that bone Pb levels were associated more consistently with 

29 cognitive function in nonoccupationally-exposed adults (Table 5-8). Despite the large 

30 number of available publications, it is important to recognize that several studies are 

31 variants on analyses in the same population (e.g., NHANES or the Normative Aging 

32 Study [NAS]) and should not be considered as all independent assessments of the Pb33 cognitive function relationship. Another point to consider is that although cross-sectional 
February 2012 5-104 Draft – Do Not Cite or Quote

1 and longitudinal studies have been conducted, most examine the associations between 

2 one measurement of cognitive function and one concurrent measurement of blood Pb 

3 level. Such a design has the inherent weakness that temporality cannot be determined, 

4 which limits the causal inference regarding the effects of Pb exposure. When bone Pb 

5 level is used as the exposure measure, this issue is mitigated somewhat because bone Pb 

6 is a biomarker that reflects many cumulative years of exposure. Additionally, with single 

7 measurements of both blood and bone Pb level, it is difficult to characterize the specific 

8 timing, duration, and level of Pb exposure that contributed to the associations observed 

9 with cognitive function. This uncertainty may apply particularly for assessments of blood 

10 Pb levels, which are influenced both by current exposures and cumulative Pb stores in 

11 bone that are mobilized during bone remodeling compared with occupationally-exposed 

12 adults (Sections 4.3 and 4.7.3).

13 Several analyses of the large prospective Baltimore Memory Study and NAS have 

14 contributed to greater understanding of the relationship between biomarkers of Pb and 

15 cognitive effects in adults. In both cohorts, subjects were evaluated periodically with in16 person clinical assessments and self-administered questionnaires. Both studies were noted 

17 for comparisons of associations between bone and blood Pb levels within the same cohort 

18 and for their repeated assessments of Pb biomarker levels and cognitive function over 

19 time. In particular, the repeated assessments permitted the examination of associations of 

20 Pb biomarkers with changes in cognitive function over time, which allowed investigators 

21 to establish temporality between Pb biomarker levels and subsequent changes in 

22 cognitive function. The Baltimore Memory Study (BMS) included men and women, 

23 50-70 years of age, residing in Baltimore, MD. A total of 1,140 out of 2,351 (48.5%) 

24 subjects participated from neighborhoods that represented a diversity of race and SES. Of 

25 particular note, this study was unique in that it included a large proportion of African26 Americans (n=395). In comparison, the NAS involved only men (original n = 2,280) 

27 residing in the Greater Boston area. Subjects primarily were white and at enrollment were 

28 aged 21 to 80 years and had no current or past chronic medical conditions. Studies 

29 differed with respect to the potential confounding variables analyzed. The most notable 

30 difference was the inclusion of age, smoking and alcohol intake as covariates in the NAS 

31 analyses. Despite differences in study population characteristics and potential 

32 confounding variables considered, findings were similar between cohorts.

33 In the BMS, longitudinal analyses were conducted with repeat cognitive testing of study 

34 subjects at approximately 14-month intervals. Most subjects completed follow-up; 91% 

35 of the original cohort returned for a second round of testing and 83% for a third round 

36 each (Bandeen-Roche et al., 2009). An interquartile range higher tibia Pb level (12.7 

37 μg/g) was associated with a 0.019 units per year decrease in eye-hand coordination z38 score, adjusting for age, sex, interviewer, race and SES. Tibia Pb was associated with a 
February 2012 5-105 Draft – Do Not Cite or Quote

1 larger decrease among African Americans than among whites (Table 5-8). Tibia Pb levels 

2 were weakly associated with other time-related decreases in other indices of cognitive 

3 function (e.g., language, processing speed, executive function).

4 Cross-sectional analyses of the BMS included comparisons of the associations for blood 

5 and bone Pb level. The 991 adults in Shih et al. (2006) had a mean (SD) blood and tibia 

6 Pb levels of 3.46 (2.23) μg/dL and 18.7 (11.2) μg/g, respectively, Whereas higher 

7 concurrent blood and bone Pb level both were associated with poorer performance in the 

8 domains of language, processing speed, eye-hand coordination, executive function, 

9 verbal memory and learning, visual memory, and visuoconstruction, associations with 

10 tibia Pb level tended to be larger in magnitude (per SD increase) and statistically 

11 significant (Table 5-8). Tibia Pb levels were associated with worse performance on tests 

12 in all domains in models adjusted for age, sex, testing technician, and presence of the 

13 apolipoprotein (APO)E-ε4 allele (potential risk factor for Alzheimer's Disease). The 

14 magnitudes of associations were attenuated with additional adjustment for education, 

15 race, and SES. In these fully-adjusted models, higher tibia Pb levels were associated with 

16 poorer performance in domains except language and processing speed, with a borderline 

17 statistically significant association observed for visuoconstruction. In linear models, 

18 visuoconstruction scores were 0.0044 SDs (95% CI: -0.0091, 0.0003) lower per 1 μg/g 

19 bone higher tibia Pb level. Analysis of a quadratic term for tibia Pb indicated no evidence 

20 of nonlinearity. 

21 Other cross-sectional analyses indicated effect modification by race and neighborhood 

22 psychosocial hazards. In contrast with the longitudinal results, race-stratified analyses of 

23 persistent effects in cross-sectional analyses indicated that tibia Pb levels were associated 

24 with greater decreases in performance on tests of eye-hand coordination, executive 

25 functioning, and verbal memory and learning among whites than among African 

26 Americans (Bandeen-Roche et al., 2009) (Table 5-8). 
February 2012 5-106 Draft – Do Not Cite or Quote

Table 5-8 Associations of blood and bone Pb levels with cognitive function in 

adultsa

Study

Population/

Methods Cognitive Test

Subgroup/

Model

Blood Pb Effect Estimate

(95% CI)b Bone Pb Effect Estimate (95% CI)b

BandeenRoche et 

al. (2009)

1,140 adults, ages 50-70 yr

Baltimore Memory Study 

(BMS) cohort

Baltimore, MD

Marginal linear regression 

models adjusted for age, 

household wealth, 

education, race/ethnicity

Longitudinal associations

Eye/hand coordination

African-Americans

White

Cross-sectional associations

Verbal memory/learning

African-Americans

White

Executive Function

African-Americans

White

NOT EXAMINED Tibia Mean (SD): 18.8 (11.6) μg/g

Per 12.7 μg/g (IQR) increase 

-0.032 (-0.052, -0.012) per year

-0.009 (-0.024, 0.006) per year

0.006 (-0.09, 0.10)

-0.076 (-0.15, 0.001)

0.009 (-0.05, 0.07)

-0.07 (-0.12, -0.01)

Shih et al.

(2006)

985 adults, mean age: 59 yr

BMS, Baltimore, MD

Linear regression adjusted 

for:

Model A: age, sex, 

technician, presence of 

APOE-ε4 allele

Model B: Model I, years of 

education, race/ethnicity, 

wealth

Raven’s Colored Progressive 

Matrices

Language

Eye-hand coordination

Executive functioning

Visuoconstruction

Model A

Model B

Model A

Model B

Model A

Model B

Model A

Model B

Concurrent blood Pb Mean 

(SD): 3.5 (2.2) μg/dL

-0.013 (-0.064, 0.037)

-0.004 (-0.044, 0.036)

-0.024 (-0.0704, 0.022)

-0.017 (-0.057, 0.024)

-0.031 (-0.075, 0.012)

-0.022 (-0.059, 0.015)

-0.042 (-0.101, 0.017)

-0.031 (-0.084, 0.021)

Tibia Mean (SD): 18.7 (11.2) μg/g

-0.0896 (-0.146, -0.034)

0.007 (-0.034, 0.047)

-0.0896 (-0.134, -0.045)

-0.034 (-0.067, 0)

-0.0896 (-0.123, -0.056)

-0.016 (-0.056, 0.025)

-0.024 (-0.190, 0.141)

-0.049 (-0.101, 0.002)

TIBIA Pb STRONGER

Glass et 

al. (2009)

1,001 adults, 

mean age 59 yr

BMS, Baltimore, MD

Multilevel hierarchical 

regression model adjusted 

for age, sex, race/ethnicity, 

education, testing 

technician, time of day

Raven’s Colored Progressive 

Matrices

Language

Eye-hand coordination

Executive functioning

Visuoconstruction

Middle NPH

High NPH

Middle NPH

High NPH

Middle NPH

High NPH

Middle NPH

High NPH

NOT EXAMINED Tibia Pb Mean (SD): 18.8(11.1) 

μg/g

0.011 (-0.089, 0.111)c

-0.09 (-0.189, -0.011)c

-0.04 (-0.133, 0.044)c

-0.067 (-0.167, 0.033)c

-0.022 (-0.111, 0.067)c

-0.111 (-0.1998, -0.022)c

-0.003 (-0.155, 0.149)c

-0.007 (-0.189, 0.175)c

Weisskopf 

et al.

(2007a)

1,089 males, 

mean age 68.7 yr; 

Normative Aging Study 

(NAS), Boston, MA

Linear repeated measures 

analysis adjusted for age, 

age squared, education, 

smoking, alcohol intake, yr 

between bone Pb 

measurement and first 

cognitive test, yr between 

cognitive tests

CERAD, Neurobehavioral

Evaluation System, WIAS-R,

MMSE, VMI

Visuospatial, pattern

comparison (+ = poorer 

performance)

Executive function verbal 

fluency

Short-term memory, word list

NOT EXAMINED Mean (IQR): Tibia: 20 (15) μg/g

Patella: 25 (20) μg/g

Estimates per IQR increase:

Tibia: 0.79 (0.40, 1.2) over time

Patella: 0.73 (0.40, 1.2) over time

Tibia: -0.40 (-1.6, 0.80) over time

Patella: -0.86 (-2.00, 0.30) over 

time

Tibia: -0.28 (-1.2, 0.60) over time

Patella: -0.81 (-1.7, 0.05) over time
February 2012 5-107 Draft – Do Not Cite or Quote

Study

Population/

Methods Cognitive Test

Subgroup/

Model

Blood Pb Effect Estimate

(95% CI)b Bone Pb Effect Estimate (95% CI)b

Wang et 

al. (2007a)

358 males, median ages: 

67.2 yr (HFE wild-type)

67.7 yr (HFE variant)

Normative Aging Study, 

Boston, MA

Linear regression adjusted 

for age, years of education, 

nonsmoker, former smoker, 

pack-years, nondrinker, 

alcohol consumption, 

English as first language, 

computer experience, 

diabetes

MMSE HFE wildtype

One HFE variant

Two HFE variants

NOT EXAMINED Median: 19 μg/g (Tibia), 23 μg/g 

(Patella)

-0.20 (-1.0, 0.70) per year

-1.40 (-3.3, 0.40) per year

-6.3 (-10.4, -2.1) per year

Weuve et 

al. (2006)

720 males, ages ≥ 45 yr

NAS, Boston, MA

Linear mixed effects 

regression adjusted for 

smoking status, alcohol 

consumption, calorie 

adjusted calcium intake, 

regular energy expenditure 

on leisure time physical 

activity, diabetes

MMSE score ALAD wildtype

ALAD2 carrier

ALAD wildtype

ALAD2 carrier

Concurrent blood Pb Mean 

(IQR): 5.2 (3) μg/dL

-0.013 (-0.053, 0.027)

-0.087 (-0.180, 0.007)

Median (IQR): Tibia: 19 (15) μg/g; 

Patella: 27 (21) μg/g

Tibia

-0.03 (-0.14, 0.07)

-0.11 (-0.30, 0.06)

Patella

-0.03 (-0.11, 0.04)

-0.12 (-0.30, 0.06)

Rajan et 

al. (2008)

720 males, ages ≥ 45 yr

Normative Aging Study, 

Boston, MA

Linear regression adjusted 

for blood Pb main effect, 

ALAD genotype, age at 

cognitive test, education, 

alcohol consumption, 

cumulative smoking, English 

as first language

CERAD, Neurobehavioral 

Evaluation System, WIAS-R

Visuospatial, constructional

praxis

Executive function verbal 

fluency

Verbal memory, word recall

Perceptual speed, mean 

latency

Concurrent blood Pb Mean 

(SD): 5.3 (2.9) μg/dL (ALAD 

wildtype); 4.8 (2.7) μg/dL 

(ALAD2 carriers)

-0.048 (-0.216, 0.120)d

-0.028 (-0.2044, 0.148)d

0.003 (-0.168, 0.174)d

-0.168 (-0.392, 0.056)d

Mean (SD):

Tibia: 21.9 (13.8) μg/g (ALAD 

wildtype), 21.2 (11.6) μg/g (ALAD2 

carriers)

Patella: 29.3 (19.1) μg/g (ALAD

wildtype), 27.9 (17.3) μg/g (ALAD2 

carriers)

Tibia: -0.216 (-0.419, -0.013)d

Patella: 0.018 (-0.182, 0.218)d

Tibia: -0.089 (-0.292, 0.1143)d

Patella: -0.018 (-0.218 0.1082)d

Tibia: 0.064 (-0.127, 0.254)d

Patella: 0.127 (-0.728, 0.989)d

Tibia: -0.152 (-0.495, 0.191)d

Patella: -0.146 (-0.40, 0.109)d

TIBIA Pb STRONGER

Weuve et 

al. (2009)

587 females, ages 47-74 yr

Nurses’ Health Study, 

Boston, MA

Generalized estimating 

equations adjusted for age, 

age-squared at Pb 

assessment, age at 

cognitive assessment, 

education, husband’s 

education, alcohol 

consumption, smoking 

status, physical activity, 

aspirin use, ibuprofen use, 

use of Vitamin E 

supplements, menopausal 

status and postmenopausal 

hormone use

Telephone Interview for 

Cognitive Status and East 

Boston Memory Test

Composite cognitive score

Composite except letter 

fluency

Concurrent blood Pb Mean 

(SD): 2.9 (1.9) μg/dL

-0.015 (-0.068, 0.038)

0.015 (-0.070, 0.101)

Mean (SD)

Tibia Pb: 10.5 (9.7) μg/g

Patella Pb: 12.6 (11.6) μg/g

Tibia: -0.039 (-0.087, 0.0097)

Patella: -0.012 (-0.058, 0.035)

Tibia: -0.049 (-0.097, 0)

Patella: -0.035 (-0.081, 0.012)

TIBIA Pb STRONGER
February 2012 5-108 Draft – Do Not Cite or Quote

Study

Population/

Methods Cognitive Test

Subgroup/

Model

Blood Pb Effect Estimate

(95% CI)b Bone Pb Effect Estimate (95% CI)b

Krieg and 

Butler

(2009)

2,823 adults, ages 20-59 yr, 

U.S. NHANES III 

(1991-1994)

Log-linear regression model 

adjusted for age, sex, 

education, family income, 

race-ethnicity, computer or 

video-game familiarity, 

alcohol use within the last 

3 h, test language

Neurobehavioral Evaluation 

System 2

Symbol Digit Substitution

(mean total latency, sec)

Serial digit learning score

Ages 20-39 yr

Ages 40-59 yr

Ages 20-39 yr

Ages 40-59 yr

Concurrent Blood mean 

(SD): 2.88 (6.91) μg/dL

-0.097 (-0.422, 0.228)e

-0.290 (-0.601, 0.0207)e

-0.117 (-0.463, 0.228)e

0.401 (-0.193, 0.995)e

NOT EXAMINED

Krieg et al.

(2009)

2,090 adults, ages 20-59 yr

1976 adults, ages ≥ 60 yr

U.S. NHANES III 

(1991-1994)

Log linear regression model 

adjusted for sex, age, 

education, family income, 

race-ethnicity, computer or 

video game familiarity, 

alcohol use in the last 3 hrs, 

test language (20-59 yr) and 

sex, age, education, family 

income, race-ethnicity, test 

language (≥ 60 yr)

Neurobehavioral Evaluation 

System 2

Symbol Digit Substitution

(mean total latency, sec)

Serial digit learning score

Word recall

Story recall

Ages 20-59 yr

ALAD GG

ALAD CC/CG

Ages 20-59 yr

ALAD GG

ALAD CC/CG

Ages ≥ 60 yr

ALAD GG

ALAD CC/CG

Ages ≥ 60 yr

ALAD GG

ALAD CC/CG

Concurrent Blood Pb Mean 

(SD):

20-59 yr: 2.85 (7.31) μg/dL; 

≥ 60 yr: 4.02 (3.56) μg/dL

-0.132 (-0.358, 0.095)e

-0.526 (-1.118, 0.066)e

-0.022 (-0.526, 0.482)e

0.025 (-0.406, 0.456)e

-0.075 (-0.285, 0.135)e

0.025 (-0.406, 0.456)e

0.085 (-0.0997, 0.271)e

-0.466 (-1.072, 0.139)e

NOT EXAMINED
February 2012 5-109 Draft – Do Not Cite or Quote

Study

Population/

Methods Cognitive Test

Subgroup/

Model

Blood Pb Effect Estimate

(95% CI)b Bone Pb Effect Estimate (95% CI)b

Krieg et al.

(2010)

2,093 adults, ages 20-59 yr

1,799 adults, ages ≥ 60 yr

U.S. NHANES III 

(1991-1994)

Log linear regression model 

adjusted for sex, age, 

education, family income, 

race-ethnicity, computer or 

video game familiarity, 

alcohol use in the last three 

hours, test language (20-59 

yr) and sex, age, education, 

family income, raceethnicity, test language (≥ 60 

yr)

Neurobehavioral Evaluation 

System 2

Symbol Digit Substitution

(mean total latency, sec)

Serial digit learning score

(+ = poorer performance)

Word recall

Story recall

Ages 20-59 yr

VDR haplotype 

CC

VDR haplotype 

CT

VDR haplotype 

TC

VDR haplotype TT

Ages 20-59 yr

VDR haplotype

CC

VDR haplotype 

CT

VDR haplotype 

TC

VDR haplotype TT

Ages ≥ 60 yr

VDR haplotype 

CC

VDR haplotype 

CT

VDR haplotype 

TC

VDR haplotype TT

Ages ≥ 60 yr

VDR haplotype 

CC

VDR haplotype 

CT

VDR haplotype 

TC

VDR haplotype TT

Concurrent blood Pb Mean 

(SD): 20-59 yr: 2.85 (7.32) 

μg/dL; ≥ 60 yr: 4.02 (3.39) 

μg/dL

-3.916 (-8.638, 0.805)e

0.139 (-0.278, 0.556)e

-0.505 (-1.025, 0.015)e

-0.695 (-0.783, 0.871)e

-2.533 (-4.868, -0.198)e

-0.322 (-0.922, 0.278)e

0.447 (0.542, 0.351)e

0.044 (-0.783, 0.871)e

-0.766 (-1.817, 0.285)

-0.085 (-0.40, 0.21)

-0.034 (-0.471, 0.403)

-0.095 (-0.895, 0.705)

0.146 (-1.674, 1.966)

0.003 (-.193, 0.20)

0.034 (-0.322, 0.3899)

-0.166 (-0.434, 0.102)

NOT EXAMINED

Gao et al.

(2008)

188 adults, mean age 69.2 

yr

Sichuan and Shandong 

Provinces, China

ANCOVA adjusted for age, 

sex, education, BMI, 

APOE ε4

CERAD, CSID, IU story recall, 

Animal fluency test, IU token 

test

Composite cognitive score

Concurrent plasma Pb 

Mean (SD): 0.39 (0.63) 

μg/dL

-0.006 (-0.016, 0.004)

NOT EXAMINED
February 2012 5-110 Draft – Do Not Cite or Quote

Study

Population/

Methods Cognitive Test

Subgroup/

Model

Blood Pb Effect Estimate

(95% CI)b Bone Pb Effect Estimate (95% CI)b

Van 

Wijngaarden 

et al. (2009)

47 adults, mean age 61.5 

yr

Rochester, NY

Linear regression adjusted 

for age, gender, 

educational level, history 

of hypertension

CANTAB and Montreal 

Cognitive Assessment

Delayed matching, % correct

Total trials

NOT EXAMINED Mean (SD): Tibia: 2.0 (5.2) μg/g; 

Calcaneus: 6.1 (8.5) μg/g

Calcaneus

Lowest tertile: 87.56f

Medium tertile: 86.67

Highest Tertile: 80.67, p = 0.027

Tibia

Lowest tertile: 85.42f

Medium tertile: 87.08

Highest tertile: 82.44, p = 0.25

Calcaneus

Lowest tertile: 2.54f

Medium tertile: 2.61

Highest tertile: 2.72, p = 0.21

Tibia

Lowest tertile: 2.62f

Medium tertile: 2.59

Highest tertile: 2.66

a

Studies are presented by cohort then generally in the order of discussion in the text.

b

Effect estimates have been standardized to the standard deviation of the cognitive test scores and standardized to an SD or IQR increase in blood or bone Pb level. c

Effect estimates indicate interactions between Pb and category of neighborhood psychosocial hazard (NPH), with the lowest tertile of NPH serving as the reference 

group.

d

Effect estimates indicate interactions between Pb and ALAD genotype.

e

The directions of effect estimates were changed to indicate a negative slope as a decrease in cognitive performance. f

Results refer to mean cognitive function scores among tertiles of bone Pb.

1 Similar to the BMS, in the NAS cohort, higher tibia Pb levels were associated with 

2 decreases in cognitive performance over time (Weisskopf et al., 2007a). Weisskopf et al.

3 (2007a) expanded evidence by finding associations with patella Pb levels. Two 

4 measurements of cognitive function, collected approximately 3.5 years apart were 

5 available for 60-70% of participants. Longitudinal analyses were conducted with repeated 

6 measures plus a bone Pb-time interaction term in order to estimate the association 

7 between bone Pb level and decline in cognitive test score over time. Although bone Pb 

8 levels were associated with increased response latency on a pattern comparison test, they 

9 were associated with fewer errors on the same test. The authors proposed that this may be 

10 related to slowing reaction time to improve accuracy. When the nine men with the 

11 highest bone Pb levels were removed, the association with fewer errors was no longer 

12 statistically significant. However, the authors did not indicate whether the point estimate 

13 changed. In the analysis with patella Pb, Weisskopf et al. (2007a) found a nonlinear 

14 association, with latency times becoming worse over time (i.e., larger values or slower 

15 response time) up to approximately 60 μg/g patella Pb, but the change over time leveling 

16 off at higher levels (Figure 5-12). Below 60 μg/g, a 20 μg/g difference in patella Pb level 

17 was associated with an increase in latency of approximately 0.15 ms. 
February 2012 5-111 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Williams & Wilkins, Weisskopf et al. (2007a).

Note: Models are adjusted for age, age squared, education, smoking, alcohol intake, years between bone Pb measurement and first 

cognitive test, and years between the cognitive tests. The 9 subjects with the highest patella Pb concentrations (>89 μg/g bone 

mineral) were removed. The estimate is indicated by the solid line and the 95% confidence interval by the dashed lines. Patella Pb 

concentrations of all individual subjects are indicated by short vertical lines on the abscissa. (reference = 0 at mean of patella Pb 

concentration).

Figure 5-12 Nonlinear association between patella Pb level and the relative 

change over time in response latency on the pattern comparison 

test.

1 Longitudinal analysis of the NAS cohort also indicated effect modification by 

2 hemochromatosis (HFE) gene variants (Wang et al., 2007a). In models adjusted for 

3 age, years of education, nonsmoker, former smoker, pack-years, nondrinker, alcohol 

4 consumption, English as first language, computer experience, and diabetes, an 

5 interquartile range higher tibia Pb level (15μg/g) was associated with a 0.22 point steeper 

6 annual decline (95% CI: -0.39, -0.05) in MMSE score among men with either the H63D 

7 and C282Y variant. The association was found to be nonlinear, with larger Pb-associated 

8 declines observed at higher tibia Pb levels (Figure 5-13). This difference was comparable 

9 to the difference in MMSE score between men who were 4 years apart in age in their 

10 study sample. Tibia Pb level was associated with a smaller decline in MMSE score in 

11 men with only HFE wildtype alleles (Figure 5-13). HFE variants, H63D and C282Y, are 

12 associated with hemochromatosis, a disease characterized by higher iron body burden.

13 Higher iron body burden has been linked with lower Pb absorption, thus the apparent 

14 interaction could be related to altered Pb toxicokinetics rather than some other direct 

15 biological interaction. For example, the relation between tibia Pb level and Pb dose at the 

16 biologically relevant site(s) could be shifted to the left among for HFE variant allele 

17 carriers such that for the same tibia Pb level, Pb the dose at the relevant biological site(s) 
February 2012 5-112 Draft – Do Not Cite or Quote

1 may be higher leading to a greater annual decline in MMSE score as indicated in Figure 

2 5-13 (solid curve). This implies that similar decreases in MMSE score would still be 

3 found in wildtype HFE carriers only at higher tibia Pb levels.

4

Source: Wang et al. (2007a).

Note: The lines indicate curvilinear trends estimated from the penalized spline method. Among HFE wild-types, the association 

between tibia Pb and annual cognitive decline was nearly linear, but among variant allele carriers, the association tended to deviate 

from linearity (p = 0.08). The model was adjusted for age, years of education, nonsmoker, former smoker, pack-years, nondrinker, 

alcohol consumption, English as first language, computer experience, and diabetes. 

Figure 5-13 Exploration of nonlinear association of tibia Pb level with annual 

rate of cognitive decline, by hemochromatosis (HFE) gene variant.

5 The 2006 Pb AQCD described associations of blood and tibia Pb levels with poorer 

6 cognitive performance among 141 NAS men (Payton et al., 1998). Several new and 

7 larger cross-sectional NAS analyses corroborated previous findings for bone Pb but 

8 generally indicated weak associations with concurrent blood Pb levels and only in groups 

9 with specific genetic variants. In contrast with the longitudinal analyses, Weisskopf et al.

10 (2007a) found that repeat measures of bone Pb levels were not consistently associated 

11 with cognitive function in cross-sectional analyses. In a study of Mini-Mental State 

12 Examination (MMSE) tests scores (test of general cognitive function) among 720 men 

13 45 years of age and older, higher concurrent blood Pb levels were associated with lower 

14 MMSE scores among ALAD2 carriers (Weuve et al., 2006). A 3 μg/dL higher concurrent 

15 blood Pb level (the interquartile range) was associated with a 0.26 point lower mean 

16 MMSE score (95% CI: -0.54, -0.01) among ALAD2 carriers and a 0.04 point lower score 

17 (95% CI: -0.16, -0.07) among noncarriers. A subsequent study did not find a consistent 
February 2012 5-113 Draft – Do Not Cite or Quote

1 direction of modification of the association between blood or bone Pb levels and other 

2 tests of cognitive function by ALAD genotype (Rajan et al., 2008). Nonetheless, Rajan et 

3 al. (2008) found tibia Pb levels to be associated more strongly and consistently with 

4 poorer cognitive performance compared with concurrent blood or patella Pb levels (Table 

5 5-8).

6 Effect modification by stress was examined in both the NAS and BMS cohorts. In the 

7 NAS cohort, higher bone Pb levels were associated with poorer cognitive function among 

8 individuals with higher individual-level perceived stress (Peters et al., 2007). Higher tibia 

9 Pb levels also were associated with larger decrements in cognitive performance among 

10 BMS subjects living in neighborhoods with a greater number psychosocial hazards 

11 (e.g., number of violent crimes, emergency calls, off-site liquor licenses) (Glass et al., 

12 2009) (Table 5-8). These observation of effect modification by environmental stress in 

13 adult humans is supported by several observations of Pb-stress interactions in impaired 

14 learning and memory and adult animals with Pb exposures from gestation through post15 weaning and lifetime Pb exposures (Section 5.3.2.2).

16 Weuve et al. (2009) studied the association of blood and bone Pb levels with cognitive 

17 function in a subset of 587 healthy women in the Boston, MA area participating in the 

18 Nurses' Health Study. Blood and bone Pb levels were measured between the ages of 47 

19 and 74 years, and the mean (SD) blood Pb level in this group was 2.9 (1.9) μg/dL 

20 measured in samples collected an average of 5 years before cognitive testing. As in the 

21 aforementioned studies of adults, tibia and patella Pb levels were more consistently 

22 associated with cognitive performance than was blood Pb levels (Table 5-8). Contrary to 

23 expectation, higher patella and tibia Pb levels were associated with higher scores on the 

24 “f” naming test (naming words that begin with f). In separate models, the “f” naming test 

25 was omitted from a composite index of all cognitive tests, and a one SD (10 μg/g bone) 

26 higher tibia Pb level was associated with 0.051-point lower (95% CI: -0.010, -0.003) 

27 standardized composite score. A similar magnitude of decrease was estimated for an 

28 increase in age of 3 years in these women. The magnitude of association was slightly 

29 smaller for an SD unit increase in patella Pb level (-0.033 [95% CI: -0.080, 0.014).

30 Several studies analyzed data from the U.S.-representative NHANES III (1991-1994) 

31 population of men and women and investigated effect modification by age and genetic 

32 variants. Only blood Pb levels were available and were measured in samples collected 

33 concurrently with cognitive testing. Krieg and Butler (2009) did not find blood Pb level 

34 consistently to be associated with poorer performance on cognitive testing among 2,090 

35 adults 20-59 years of age or among 1,796 adults 60 years of age and older. Because 

36 different types and numbers of tests were conducted in the two age groups, it is difficult 

37 to compare findings between age groups. In the subset of the population with genetic 
February 2012 5-114 Draft – Do Not Cite or Quote

1 analysis, blood Pb-cognitive function associations were not found to be modified by 

2 ALAD genetic variants in the same direction (Krieg et al., 2009). Among adults ages 

3 20-59 years with the CC and CG ALAD genotype groups combined (i.e., ALAD2 

4 carriers), reaction time improved (i.e., faster reaction time) by 38 ms per 10-fold increase 

5 in concurrent blood Pb level. In contrast, among ALAD2 carriers, higher blood Pb levels 

6 were associated with poorer performance on a symbol-digit substitution task. The 

7 potential direction of effect modification by the ALAD2 genotype is not immediately 

8 clear as the greater affinity of the ALAD2 enzyme subunit for Pb may increase 

9 susceptibility to Pb-associated health effects by increasing blood Pb levels or it may 

10 diminish Pb-associated health effects by decreasing Pb bioavailability by maintaining it 

11 in a sequestered state in the bloodstream. Krieg et al. (2010) did find differences in the 

12 association between concurrent blood Pb level and scores on a symbol-digit substitution 

13 test by the VDR variants, rs731236 and VDR rs2239185, and by the VDR haplotype. 

14 Similar to observations in adolescent NHANES participants (Section 5.3.2.1), results 

15 were not uniform across the various tests. However, blood Pb level generally was 

16 associated with greater decrements in cognitive performance among adults with the CC 

17 genotypes of VDR variants.

18 Other studies with smaller numbers of subjects generally produced results consistent with 

19 those from the larger studies above. A cross-sectional study of 188 rural Chinese men and 

20 women found a weak association between higher plasma Pb levels and a lower composite 

21 cognitive score based on a battery of in-person administered tests (Gao et al., 2008). It 

22 should be noted, though, that Pb in plasma makes up a very small fraction of all Pb in 

23 blood and is a different, and much less used, biomarker than Pb in whole blood. The 

24 relevance of this Pb fraction is not entirely clear. Pb in plasma is not bound to 

25 erythrocytes, as is about 99% of blood Pb. Thus, it has been postulated that plasma Pb 

26 may be more toxicologically active (Chuang et al., 2001; Hernandez-Avila et al., 1998).

27 In another cross-sectional study of 47 men and women in Rochester, NY (55-67 years of 

28 age), subjects in the higher two tertiles of calcaneal bone (trabecular bone with higher 

29 turnover rate than tibia) Pb level performed worse on delayed matching-to-sample and 

30 paired associated learning tasks (Van Wijngaarden et al., 2009) (Table 5-8). In analyses 

31 of tibia Pb levels, subjects in the highest tertile of tibia Pb level did not consistently 

32 perform worse on cognitive tests (Table 5-8). The exact calcaneal and tibia Pb levels in 

33 tertiles were not reported.

Adults with Occupational Lead Exposures

34 The 2006 Pb AQCD concluded that in adults, blood Pb levels were associated with 

35 cognitive function most consistently among those with occupational Pb exposures. These 

36 findings were supported by results from a few recent studies of occupationally-exposed 
February 2012 5-115 Draft – Do Not Cite or Quote

1 adults. Dorsey et al. (2006) followed up on a cohort of Pb-exposed workers in Korea with 

2 a mean age of 43.4 years, on whom patella Pb measurements were made. This group 

3 represented a typically highly-exposed occupational group with an average blood Pb 

4 level of 30.9 μg/dL. In this cohort, both blood and tibia Pb levels previously were found 

5 to be associated with poorer performance on a battery of neurocognitive tests (Schwartz 

6 et al., 2005; Schwartz et al., 2001). Dorsey et al. (2006) found higher patella Pb levels to 

7 be associated with poorer manual dexterity, poorer sensory function, and greater 

8 depression symptoms. These associations for patella Pb levels were not as strong as the 

9 previously reported associations were with either blood or tibia Pb levels in this 

10 occupational cohort.

11 A follow-up study of the original 1982 Lead Occupational Study was conducted 

12 2001-2004 with 83 of the original 288 Pb-exposed workers and 51 of the original 181 

13 controls (Khalil et al., 2009b). Those originally in the exposed workers group had last 

14 worked in a job with Pb exposure from 0.02 to 16 years (median: 6) prior to follow-up 

15 testing. While the follow-up participation was somewhat low, participants did not differ 

16 from nonparticipants on most baseline cognitive tests except for performing slightly 

17 better on aspects of the grooved pegboard test and recall on a paired associates learning 

18 task. This suggests that the follow-up participation was not biased to poor performers. At 

19 follow-up, the former Pb-exposed workers performed worse than did the controls in total 

20 cognitive score and in the spatial and general intelligence domains (p <0.05). They also 

21 performed worse in all other domains (e.g., motor, executive, and memory) although the 

22 differences were not as large. A similar pattern was observed in analyses using tibia Pb 

23 levels measured at the follow-up visit to represent exposure. Weaker associations were 

24 observed with concurrent blood Pb levels (median among the exposed: 12 μg/dL). 

25 Among the former Pb workers, higher tibia Pb levels were associated with a greater 

26 decrease in total score and scores for spatial and executive domains between baseline and 

27 follow-up. Tibia Pb level were associated inversely with poorer performance in other 

28 domains as well. As in nonoccupationally-exposed adults, the stronger findings for tibia 

29 Pb levels in former Pb-exposed workers indicate stronger effects of higher past Pb 

30 exposures than lower current exposures on cognitive function.

31 Additional studies aimed to characterize factors that either mediate or modify the 

32 association between Pb biomarkers and cognitive function. A study of 61 current Pb 

33 smelter workers with a mean age of 40 years and blood Pb level of 29.1 μg/dL found that 

34 both a working lifetime time-weighted integrated blood Pb level (an index of cumulative 

35 exposure) (p = 0.09) and tibia Pb level (p = 0.08) were associated with longer times to 

36 complete the grooved pegboard test (Bleecker et al., 2007a). Among 112 Pb smelter 

37 workers, working lifetime time-weighted integrated blood Pb level was associated with 

38 poorer performance on attention and digit symbol tasks among those with low cognitive 
February 2012 5-116 Draft – Do Not Cite or Quote

1 reserve (assessed by performance on the Wide Range Achievement Test-R for reading)

2 (Bleecker et al., 2007b).

3 Apolipoprotein E is a transport protein for cholesterol and lipoproteins. The gene appears 

4 to regulate synapse formation (connections between neurons) and may be particularly 

5 critical in early childhood. A genetic variant, called the ApoE-ε4 allele is a haplotype 

6 between 2 exonic SNPs and is perhaps the most widely studied genetic variant with 

7 respect to increasing risk of neurologic disease. ApoE-ε4 carriers are at two-fold 

8 increased risk of developing Alzheimer’s disease, although the majority of such 

9 individuals still do not develop the disease. Thus, it is biologically plausible that ApoE-ε4 

10 carriers may be biologically susceptible to cognitive dysfunction. A study of 

11 occupationally-exposed adults found that among individuals with at least one ApoE-ε4 

12 allele, blood Pb level was associated with poorer performance on digit symbol, pegboard 

13 assembly, and complex reaction time tests (Stewart et al., 2002).

14 Studies also indicated that Pb-exposed workers may be at increased risk of motor 

15 dysfunction. Among Pb smelter workers, working lifetime time-weighted integrated 

16 blood Pb level was associated with poorer motor performance (p <0.05). Iwata et al.

17 (2005) examined the cross-sectional association between blood Pb level and aspects of 

18 postural sway among 121 Pb-exposed workers in Japan with blood Pb levels between 6 

19 and 89 μg/dL (mean: 40 μg/dL). In multiple regression analyses adjusted for age, height, 

20 and smoking and drinking status, higher blood Pb level was associated with greater 

21 sagittal sway with eyes open (p <0.05) and eyes closed (p <0.01) and transversal sway 

22 with eyes closed (p <0.05). The authors calculated a benchmark dose level (Budtz23 Jorgensen et al., 2001; NRC, 2000) of 14.3 μg/dL from a linear concentration-response 

24 model of their data. A supralinear concentration-response function was found to fit the 

25 data slightly better than was a linear function.

Summary of Cognitive Function in Adults

26 In summary, among nonoccupationally-exposed adults, there is weak evidence that 

27 cognitive function is associated with concurrently measured blood Pb levels. The 

28 strongest evidence was provided by NHANES analyses, in which concurrent blood Pb 

29 levels were associated with lower cognitive function in particular age and genetic variant 

30 subgroups (Krieg et al., 2010; Krieg and Butler, 2009; Krieg et al., 2009). These analyses 

31 did not have bone Pb measures for comparison. It is important to note that because bone 

32 Pb is a major contributor to blood Pb levels, blood Pb level also can reflect to a large 

33 extent longer term exposures, including higher past exposures, especially among adults 

34 without occupational exposures. Thus, in the NHANES analyses of adults, it is difficult 

35 to characterize the relative contributions of recent and past Pb exposures to the 
February 2012 5-117 Draft – Do Not Cite or Quote

1 associations observed between concurrent blood Pb level and cognitive function. 

2 Consistent with the conclusion of the 2006 Pb AQCD, several recent studies found 

3 associations between bone Pb levels and cognitive function in adults (Table 5-8). Much 

4 of the evidence was provided by analyses of two cohorts: BMS and NAS. Recent 

5 longitudinal analyses demonstrated that higher bone Pb levels measured at baseline were 

6 associated with subsequent declines in cognitive function over 2- to 4-year periods

7 (Bandeen-Roche et al., 2009; Weisskopf et al., 2007a). These findings suggested that 

8 long-term Pb exposure may contribute to ongoing declines in cognitive function in adults. 

9 Recent studies that analyzed both blood and bone Pb levels generally found stronger 

10 associations for bone Pb levels, in particular tibia Pb levels, across the various cognitive 

11 tests that were performed (Table 5-8). The discrepant findings for blood and bone Pb 

12 levels indicate that cumulative Pb exposure that likely included higher past exposures, 

13 may be a better predictor of cognitive function in adults than is blood Pb level. Patella 

14 and tibia Pb levels were examined in the NAS and Nurses' Health Study; however, 

15 evidence did not consistently indicate that tibia Pb levels were more strongly associated 

16 with decreases in cognitive performance (Weuve et al., 2009; Weisskopf et al., 2007a).

17 Additional support for the effects of cumulative or past Pb exposure is provided by 

18 analyses of the Boston prospective cohort as adults. Deciduous tooth Pb was associated 

19 with decrements in specific cognitive indices at ages 19-20 years (Bellinger et al., 1994a),

20 and blood Pb levels measured at age 6 months, 4 years, 10 years, and levels averaged 

21 over childhood were associated with decrements in FSIQ at ages 28-30 years (Mazumdar 

22 et al., 2011).

23 Although based on limited examination, there is some indication that certain variants in 

24 HFE, ALAD, or VDR genes modify the association between Pb and cognitive function in 

25 nonoccupationally-exposed adults; however, results were uniform across the various 

26 cognitive tests performed. Aside from identifying populations potentially at increased 

27 risk and elucidating underlying modes of action, such effect modification also serves to 

28 strengthen the basic inference about associations between Pb biomarkers and cognitive 

29 function. Specifically, when effect modification is identified, potential confounding 

30 factors would have to vary by levels of the modifying factor, which is usually unlikely, 

31 particularly when considering genotype. It also is important to bear in mind that it is not 

32 always clear whether the observed effect modification reflects a change in the 

33 toxicokinetics of Pb and therefore a change in dose at the biological site of action or a 

34 direct biological interaction that increases the toxicity of Pb at a particular target organ or 

35 tissue.

36 In contrast with nonoccupationally-exposed adults, in adults with current occupational Pb 

37 exposures, cognitive function was associated with both blood and bone Pb levels. These 

38 findings indicate that among adults with occupational Pb exposures, both current and 
February 2012 5-118 Draft – Do Not Cite or Quote

1 cumulative exposures affect cognitive function. In the study of former Pb workers, blood 

2 Pb levels (median: 12 μg/dL) and findings were more similar to those from 

3 nonoccupationally-exposed adults. Among former Pb workers, tibia Pb levels were 

4 associated more strongly with cognitive performance than were blood Pb levels (Khalil et 

5 al., 2009b). Thus, in the absence of higher current Pb exposures, cumulative Pb exposures 

6 may have a greater effect on cognitive function in adults. 

5.3.3 Behavioral Effects

5.3.3.1 Epidemiologic Studies of Behavioral Effects in Children

7 Noncognitive effects of Pb are more complex to study relative to IQ tests. There are 

8 fewer objective tests of behavior, and existing tests do not have as strong psychometric 

9 properties and are less rigorously standardized compared with IQ tests. In several studies, 

10 behaviors are assessed frequently using teacher and/or parent ratings and thus are subject 

11 to greater measurement error. However, domain-specific neuropsychological assessments 

12 are advantageous as they may provide greater insight into the underlying CNS damage 

13 that may be associated with exposures (e.g., structural, neural system, neurotransmitter)

14 (White et al., 2009). Several epidemiologic studies reviewed in the 2006 Pb AQCD 

15 reported associations between blood Pb levels and a wide range of behavioral effects, 

16 with the weight of evidence supporting associations with inattention and hyperactivity 

17 and smaller bodies of evidence indicating associations with misconduct and delinquent 

18 behaviors and withdrawn and depressive behaviors (Bellinger and Rappaport, 2002; 

19 Needleman et al., 2002; Dietrich et al., 2001; Burns et al., 1999; Wasserman et al., 1998; 

20 Needleman et al., 1996; Bellinger et al., 1994a). Coherence was provided by similar 

21 findings of inattention, impulsivity, and changes in social behavior in Pb-exposed 

22 animals (Section 5.3.3.2). Epidemiologic studies found that blood Pb levels were 

23 associated with decrements in cognitive function and behavioral problems within the 

24 same population of children, which demonstrates the strong relationship between the two 

25 neurodevelopmental domains. The strong relationship between cognitive function and 

26 behavior in children also is demonstrated by the fact that the schedule controlled behavior 

27 tests in animals measure both memory and inattention. Thus, behavioral problems 

28 associated with Pb exposure may contribute to problems with learning, which may 

29 progress to antisocial and delinquent behavior later in life.

30 Most previous epidemiologic studies found that blood or dentin Pb levels measured at an 

31 early age (e.g., 2-6 years of age) were associated with behavioral problems later in 

32 childhood and early adulthood (e.g., 7-22 years of age). Most studies examined 
February 2012 5-119 Draft – Do Not Cite or Quote

1 associations with blood Pb levels assessed at a single time point; however, even the 

2 prospective studies with serial measurements of blood Pb levels, found a range of 

3 behavioral problems to be associated with both prenatal and early childhood blood Pb 

4 levels (Dietrich et al., 2001) and lifetime average blood Pb levels (Burns et al., 1999).

5 Thus, evidence did not conclusively identify a single lifestage of Pb exposure that was 

6 associated with the greatest risk of behavioral problems and misconduct. Recent studies 

7 strengthened the evidence for associations of blood Pb levels with inattention and 

8 aggression (Figure 5-14 and Table 5-9). Based on a smaller number of studies, new 

9 evidence demonstrates associations of blood Pb levels with ADHD diagnosis and 

10 diagnostic indices as well as with conduct disorders (Table 5-10 and Table 5-11). As with 

11 cognitive function, the epidemiologic evidence for each category of outcomes was 

12 evaluated separately in order of increasing weight of evidence. Emphasis was placed on 

13 prospective studies with repeated assessments of blood Pb levels and behavior, studies 

14 assessing effects relevant to blood Pb levels in contemporary U.S. children (i.e., less than 

15 5 μg/dL), and studies of younger children whose blood Pb levels are less influenced by 

16 higher past Pb exposures.

17 Similar to cognitive function, associations between blood Pb levels and behavioral 

18 outcomes may be potentially confounded by factors such as parental SES, parental 

19 education, parental IQ, quality and stability of the caregiving environment, and 

20 nutritional status. Accordingly, in assessing whether blood Pb-behavior associations were 

21 independent of the effects of the other variables, greater weight was given to studies that 

22 accounted for potential confounding in the study design or in statistical analyses. 

Inattention and Hyperactivity in Children

23 Consistent with previous evidence, recent studies provided strong evidence that blood Pb 

24 level were associated with various endpoints related to inattention, hyperactivity, and 

25 impulsivity in children after adjusting for potential confounding by multiple SES-related 

26 variables and co-exposures (Figure 5-14 and Table 5-9). Evidence was equally consistent 

27 for inattention assessed using teacher and parent ratings and objective tests that measure 

28 sustained attention such as the continuous performance test (CPT). The associations 

29 observed with CPT, in particular, provide strong coherence with findings in animals for 

30 Pb-induced impairments in homologous tests of response inhibition in Schedule 

31 Controlled Behavior Tests (Section 5.3.3.2). Both tests measure reactions to stop signals, 

32 i.e., premature responses, reaction time. Most of the recent evidence is derived studies of 

33 non-U.S. children. Whereas previous studies primarily examined associations with blood 

34 Pb levels measured earlier in childhood, recent studies indicated associations with 

35 concurrent blood Pb levels. Further, many earlier studies of inattention and impulsivity 

36 included children with higher blood Pb levels than those observed in contemporary 
February 2012 5-120 Draft – Do Not Cite or Quote

1 children. Recent studies provided new evidence of blood Pb-associated inattention and 

2 impulsivity in populations of children with mean blood Pb levels ranging from 2 to 

3 5 μg/dL (Cho et al., 2010; Plusquellec et al., 2010; Chiodo et al., 2007; Plusquellec et al., 

4 2007).

5 Previous results from prospective studies indicated that early childhood Pb exposures 

6 were associated with attention deficits in adolescence and young adulthood (Ris et al., 

7 2004; Bellinger et al., 1994a). Bellinger et al. (1994a) found that compared with young 

8 adults (ages 19-20 years) who had deciduous tooth Pb levels 2.9-5.9 ppm, adults with 

9 deciduous tooth Pb levels > 19.9 ppm committed more errors on the CPT and performed 

10 worse on Stroop and WCST, which measure the ability to shift focus and execute a 

11 different task. In the CLS cohort, Ris et al. (2004) found increases in attention as 

12 measured by CPT in association with prenatal maternal, 3-60 month average, and 

13 78 month blood Pb levels in adolescents ages 15-17 years, adjusting for maternal IQ, 

14 SES, HOME, and adolescent marijuana use. Prenatal blood Pb level was associated with 

15 slightly larger increases in attention, and larger associations were estimates for males 

16 (Figure 5-14 and Table 5-9).

17 Studies in children in Korea with relatively low blood Pb levels (means < 3 μg/dL) both 

18 demonstrated associations with measures of inattention (Cho et al., 2010; Min et al., 

19 2007). In a population of children ages 8-11 years, Cho et al. (2010) found relationships 

20 of concurrent blood Pb levels with ADHD symptoms (i.e., inattentiveness, hyperactivity, 

21 and total score) rated by teachers and parents, with the association with teacher ADHD 

22 rating attaining statistical significance. In addition to the low blood Pb levels in this study 

23 (mean: 1.9 μg/dL [range: 0.53-6.16]), a strength of this study was the comparison of 

24 effect estimates with and without adjustment for potential confounders such as age, sex, 

25 paternal education, maternal IQ, child IQ, city of residence, birth weight, and urinary 

26 cotinine. In multivariate models, effect estimates decreased by 2 to 14%; however, 

27 associations remained statistically significant. Mean ADHD ratings by teacher and 

28 parents were similar (both 9.1); however, parental ratings had greater variability (SD: 

29 11.5 for parents and 8.6 for teachers), which may have contributed to differences in 

30 association. Although higher blood Pb levels were associated with more errors 

31 (responding to a nontarget) on the CPT test, they were not consistently associated other 

32 indicators of inattention on the CPT or Stroop test (Figure 5-14 and Table 5-9). Further, 

33 effect estimates lost statistical significance when urinary cotinine was included in models.

34 Cho et al. (2010) did not examine potential confounding by HOME score but they did 

35 examine parental history of neuropsychiatric disease (e.g., ADHD, learning disability, 

36 depression, obsessive-compulsive disorder). Mean blood Pb levels were similar in 

37 children with and without parental history of neuropsychiatric disease (1.80 and 
February 2012 5-121 Draft – Do Not Cite or Quote

1 1.93 μg/dL, respectively, p = 0.32). Thus, the relationship between blood Pb level and 

2 ADHD-related symptoms in the child was not likely confounded by parental 

3 neuropsychiatric disease. Results from this study also indicated that the relationship 

4 between blood Pb level and ADHD-related symptoms was independent of the 

5 relationship between blood Pb level and IQ. Min et al. (2007) examined a population of 

6 children (ages 7-16 years) in Korea with a mean concurrent blood Pb level of 2.9 μg/dL 

7 and adjusted for maternal score on attention tests. Investigators found that a 1 μg/dL 

8 increase in concurrent blood Pb level was associated with a 16.8 ms increase (95% CI: -

9 1.08, 0.06) in reaction time, a test of attention. Although this study did not consider 

10 confounding by SES and caregiving environment, the findings was consistent with those 

11 from Cho et al. (2010).

12 Similar results were obtained in a study of children, ages 8-12 years, in Romania, except 

13 that concurrent blood Pb level was associated with similar magnitudes of increase in 

14 inattention, hyperactivity, and impulsivity as assessed by both parents and teachers

15 (Nicolescu et al., 2010). Results for parent and teacher ratings were strengthened by 

16 observations that blood Pb levels also were associated with increased false-alarm rates in 

17 responses to stop signals (Figure 5-14 and Table 5-9). Blood Pb levels in this study also 

18 were relatively low (median: 3.7 [95% CI: 1.7, 11.1]), and removing five children with 

19 blood Pb levels at or above 10 μg/dL had minimal impact on observed associations. 

20 Exposures to aluminum and mercury, other neurotoxic metals, also were examined, and 

21 blood Pb level was associated with the largest, statistically significant increases in 

22 inattention, impulsivity, and hyperactivity.

23 The aforementioned studies did not consider confounding by the caregiving environment, 

24 i.e., HOME score. Studies that did adjust for HOME score, also found associations of 

25 blood Pb level with inattention and hyperactivity (Chandramouli et al., 2009; Chiodo et 

26 al., 2007). In their longitudinal study of children in the U.K., Chandramouli et al. (2009)

27 found association of higher blood Pb level at age 30 months with hyperreactivity at ages 

28 7 and 8 years; however, this association was observed primarily in children with blood Pb 

29 levels greater than 10 μg/dL (Table 5-9) and the strongest for ratings given by teachers. 

30 Children with elevated blood Pb levels did not consistently have elevated odds of 

31 inattention as assessed using stop signal tasks. Similar to Cho et al. (2010), this 

32 association was not influenced by the inclusion of child IQ in the model. Therefore, these 

33 findings add support for increasing blood Pb levels having effects on behavior 

34 independent of effects on cognitive function. Chiodo et al. (2007; 2004) found that 

35 concurrent blood Pb level was associated with inattention in 7 year-old children in 

36 Detroit, MI, as assessed using teacher ratings and CPT (Figure 5-14 and Table 5-9). 

37 Potential confounders were selected based on the association of each with a particular 

38 endpoint, thus model covariates varied among endpoints. HOME score was associated 
February 2012 5-122 Draft – Do Not Cite or Quote

1 with inattention teacher ratings (Chiodo et al., 2007) but not with all CPT indices of 

2 inattention (2007; Chiodo et al., 2004). Blood Pb level was associated with inattention 

3 alone and adjusted for covariates. These findings indicated that HOME score did not 

4 account for all of the association between blood Pb level and behavioral outcomes. As 

5 was discussed in Section 5.3.2.1, this cohort comprised large proportions of children with 

6 prenatal exposure to alcohol and drugs. While this may limit the generalizability of 

7 findings, it is important to note that alcohol and drug use were not associated with every 

8 inattention index or in each study. Blood Pb level remained associated with inattention in 

9 a model that included alcohol and marijuana use along with other demographic and SES10 related variables.

11 A series of studies conducted in Inuit children living in northern Quebec, Canada 

12 reported associations between blood Pb levels and measures of inattention after extensive 

13 consideration of potential confounding variables (Plusquellec et al., 2010; Plusquellec et 

14 al., 2007; Fraser et al., 2006). Plusquellec et al. (2007) found higher cord blood Pb level 

15 to be associated with higher ratings by investigators for distractibility in completing 

16 tasks, frenetic movement, and duration of off task behaviors at age 11 months (Figure 

17 5-14 and Table 5-9). In the same cohort of children, concurrent blood Pb level but not 

18 cord blood Pb level was associated with impulsivity, irritability, and duration of off task 

19 behavior at ages 4-6 years (Plusquellec et al., 2010). Fraser et al. (2006) additionally 

20 indicated that at ages 4-6 years, the relationship between concurrent blood Pb level and 

21 motor function (i.e., transversal sway, reaction time) may be mediated by the effects of 

22 blood Pb level on an inattention/impulsivity index. These studies conducted in Inuit 

23 children evaluated confounding by examining associations of demographic, SES, and co24 exposures with outcomes independently and with blood Pb level in models. HOME score 

25 was not associated with inattention or impulsivity. These findings further indicate that 

26 confounding by HOME score cannot fully account for the collective body of evidence for 

27 associations between blood Pb level and measures of inattention and impulsivity.

28 Consistent with findings in a group of children in Korea (Cho et al., 2010) and children in 

29 the U.K. (Chandramouli et al., 2009), Nigg et al. (2008) found that associations of blood 

30 Pb level with a hyperactivity/impulsivity index in a group of U.S. children, ages 

31 8-17 years, were independent of associations with IQ. Whereas other studies assessed 

32 direct and indirect effects of blood Pb level on behavioral outcomes by including IQ in 

33 the model as a covariate, Nigg et al. (2008) specifically used regression-based path 

34 analysis, a more rigorous method to characterize the impact of one variable on the 

35 association of another in the model after controlling for other previous variables. After 

36 adjusting for sex and income, investigators found that concurrent blood Pb level had a 

37 direct association with hyperactivity/impulsivity that was not completely mediated by the 

38 blood Pb-IQ association. Instead, the association between blood Pb level and IQ was 
February 2012 5-123 Draft – Do Not Cite or Quote

1 found to be mediated via the association with hyperactivity/impulsivity. Based on 

2 associations between hyperactivity/impulsivity slower stop signal reaction time and 

3 response variability, the authors concluded that the association between blood Pb level 

4 and hyperactivity/impulsivity was mediated via poorer response on the stop task. It is 

5 important to note that other potential confounders including parental IQ and HOME score 

6 were not examined

7 Several other recent studies found associations between concurrent blood Pb levels and 

8 inattention; however, their results should be interpreted with caution due to their limited 

9 consideration of confounding by SES-related variables (Li et al., 2008d), their 

10 examination of children living near Pb sources who likely have limited applicability to 

11 contemporary children in the U.S. general population (Bao et al., 2009; Kordas et al., 

12 2007), or both (Liu et al., 2011a).
February 2012 5-124 Draft – Do Not Cite or Quote

Note: Test scores were standardized to their standard deviation to facilitate comparisons among tests with different scales. Studies 

generally are presented in order of increasing mean blood Pb level. Effect estimates are standardized to a 1 μg/dL increase in blood 

Pb level within the 10th-90th percentile interval. Black diamonds, orange triangles, and blue circles represent associations with 

concurrent, prenatal (maternal), and earlier childhood blood Pb levels, respectively.

Figure 5-14 Associations of blood Pb levels with behavioral indices of 

inattention, hyperactivity, and impulsivity.

Study

Nigg et al. (2008)

Cho et al. (2010)

Plusquellec et al. (2007)

Chiodo et al. (2007)

Chiodo et al. (2004)

Fraser et al. (2006)

Plusquellec et al. (2010)

Kordas et al. (2007)

Ris et al. (2004)

Mean (SD)

blood Pb (μg/dL)

1.04 (0.54)

1.9 (0.67)

4.64 (3.54)

5.0 (3.0)

5.4 (3.3)

5.3 (4.9)

5.4 (5.0)

5.4 (5.0)

11.5 (6.1)

NR

10th-90th

percentile

0.5-1.7

1.2-2.8

1.4-8.3

2.1-8.7

2.3-9.5

1.4-10.6

1.4-10.8

1.4-10.8

5.4-19.2

NR

Blood Pb timing

Concurrent

Concurrent

Prenatal

Concurrent

Concurrent

Concurrent

Concurrent

Concurrent

Prenatal (maternal)

3-60 mo. avg

78 mo.

Outcome

Hyperactivity/Impulsivity

ADHD rating scale, teacher

ADHD rating scale, parent

Comission Errors, CPT

Response Time, CPT

Word Reading Score

Distraction

Off task duration

Hyperactivity

Attention problems

Comission Errors, CPT

Omission Errors

Number of errors, CPT

Comission Errors, CPT

Reaction Time

Impulsivity/activity

Impulsivity 

Off task duration

Off-task passive behavior

Inattention

►

-0.5 -0.3 -0.1 0.1 0.3 0.5

Standardized Effect Estimate per 1 μg/dL increase in

blood Pb level within the 10th-90th percentile interval
February 2012 5-125 Draft – Do Not Cite or Quote

Table 5-9 Additional characteristics and quantitative results for studies 

presented in Figure 5-14

Study Population/Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)a

Nigg et al.

(2008)

150 children ages 8-17 

yr.

Year of birth and location 

NR

Concurrent

Mean (SD)

8-11 yr: 1.04 

(0.53)

12-17 yr: 1.03 

(0.54)

Regression-based path analysis 

adjusted for sex and income.

No parental IQ or education

Hyperactivity/impulsivity

using Teacher ADHD Rating 

Scale and Kiddie Schedule for 

Affective Disorders and 

Schizophrenia at ages 8-17 yr.

0.21 (0, 0.42)b

Cho et al.

(2010)

667 children ages 8-11 

yr in 2008

Five Korean cities

Concurrent

Mean (SD): 1.9 

(0.67)

10th-90th: 1.2-2.8

Log linear regression model 

adjusted for age, sex, parental 

education, maternal IQ, child IQ, 

city, birth weight, urinary cotinine

ADHD rating scale, teacher

ADHD rating scale, parent

Korean ADHD Rating Scale IV

commission errors, CPT

Response time, CPT

Word reading score, Stroop

assessed at ages 8-11 yr.

0.042 (0.017, 0.067)

0.010 (-0.013, 0.033)

0.03 (-0.01, 0.07)

-0.01 (-0.05, 0.03)

-0.02 (-0.06, 0.02)c

Plusquellec et 

al. (2007)

148-164 children ages 

11 mo born 1995-2002

Inuit communities

Quebec, Canada

Prenatal (cord)

Mean (SD): 4.6 

(3.5)

10th-90th: 1.4-8.3

Log linear regression model 

adjusted for delivery complication, 

home organization, low birth weight 

(distractibility) and sex, maternal 

anxiety, prenatal alcohol exposure 

(off task duration) 

Several more examined

Distraction

Off task duration

using Bayley Behavioral Rating 

Scale at age 11 mo

0.06 (0.021, 0.098)

0.03 (-0.01, 0.073)

Chiodo et al.

(2007)

506 African-American 

children in Detroit, MI 

area followed from birth 

(1989-1991) to age 7 yr.

Large proportions of 

children with prenatal 

exposure to cocaine or 

marijuana 

Concurrent

Mean (SD): 5.0 

(3.0)

10th-90th: 2.1-8.7

Linear regression model adjusted 

for child age, sex

Several more examined

Hyperactivity

Attention problems

using PROBS-14 and Conners' 

Teacher Rating Scale-39 at age 

7 yr

Comission Errors (%)

Omission Errors (%)

0.13 (0.03, 0.23)b

0.13 (0.03, 0.23)b

-0.08d

0.18 (0.07, 0.29)c

Chiodo et al.

(2004)

246 African-American 

children in Detroit, MI 

area

followed from birth (not 

reported) to age 7.5 yr.

Large proportions of 

children with prenatal 

exposure to cocaine or 

marijuana

Concurrent

Mean (SD): 5.4 

(3.3)

10th-90th: 2.3-9.5

Log linear regression adjusted for 

SES, maternal vocabulary score 

(Number of errors and errors of 

commission); Number of children > 

18 yr (Number of errors); Child age 

(errors of commission);

Education, sex, prenatal cocaine 

exposure (reaction time)

Several more examined

Number of errors

Comission Errors

Reaction time

Using CPT at age 7.5 yr

0.35 (0, 0.69)b,c

0.05 (p > 0.05)d

0.25 (p > 0.05)d

Fraser et al.

(2006)

90 children ages 5-6 yr 

born 1993-1996

Inuit communities

Quebec, Canada

Concurrent Mean 

(SD): 5.3 (4.9)

10th-90th: 

1.4-10.6

Linear regression model adjusted 

for binge drinking in pregnancy

Several more examined

Impulsivity/activity

using modified Infant Behavioral 

Rating Scale at ages 5-6 yr

0.25 (0.04, 0.46)

Plusquellec et 

al. (2010)

95-98 children ages 5-6 

yr born 1993-1996

Inuit communities

Quebec, Canada

Concurrent Mean 

(SD): 5.4 (5.0)

10th-90th: 

1.4-10.8

Log linear regression model 

adjusted for birth weight, sex, 

parity, caregiver education 

(impulsivity) and birth weight, SES, 

child blood hemoglobin (off task 

duration)

Several more examined

Impulsivity

Off task duration

using modified Infant Behavioral 

Rating Scale at ages 5-6 yr.

0.019 (0.001, 0.036)

0.02 (0, 0.039)
February 2012 5-126 Draft – Do Not Cite or Quote

Study Population/Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)a

Kordas et al.

(2007)

168 children ages 6-8 yr. 

born 1993-1995

Torreon, Mexico

Residence near metal 

foundry

Concurrent

Mean (SD): 11.5 

(6.1)

10th-90th: 

5.4-19.2

Linear regression model adjusted 

for age, sex, SES, home 

ownership, crowding in home, 

maternal education, family 

structure, forgetting homework

Off task passive behavior

using instrument developed by 

investigator at ages 6-8 yr

0.034 (0.005, 0.063)

Ris et al.

(2004)

195 children, ages 15-17 

years, born 1979-1985

NR Linear regression adjusted for sex, 

maternal IQ, HOME, SES, 

adolescent marijuana use

Several more examined

Inattention using CPT at ages 

15-17 yr

Prenatal (maternal)

3-60 mo avg

78 mo

0.16 (0.04, 0.27)

0.11 (0.04, 0.19)

0.12 (0.02, 0.22)

Studies not included in figure because of categorical analysis of Pb biomarker or outcome:

Chandramouli 

et al. (2009)

488 children ages 7-8 yr 

born 1991-1992

Avon, UK

Age 30 mo

Mean (SD): NR

Linear regression model adjusted 

for maternal education, home 

ownership, maternal smoking, 

HOME score, maternal SES, family 

adversity index, parenting attitudes 

at 6 mo.

Hyperactivity, teacher

using Strengths and Difficulties

Questionnaire at ages 7-8 yr

Selective inattention

using Test of Everyday Attention

for Children at ages 7-8 yr

OR: 0.84 (0.47, 1.52)e

blood Pb 2-5 vs. 0-2 μg/dL

1.25 (0.67, 2.33)e

blood Pb 5-10 vs. 0-2 μg/dL

2.82 (1.08, 7.35)e

blood Pb > 10 vs. 0-2 μg/dL

OR: 0.97 (0.62, 1.52)e

blood Pb 2-5 vs. 0-2 μg/dL

1.01 (0.64, 1.61)e

blood Pb 5-10 vs. 0-2 μg/dL

0.88 (0.42,1.85)e

blood Pb > 10 vs. 0-2 μg/dL

Nicolescu et 

al. (2010)

83 children ages 8-12 yr 

born 1995-1999

Bucharest and 

Pantelimon, Romania

Concurrent 

Median (IQR): 3.7 

(2.6)

10th-90th: 2.0-8.5

Log linear regression model 

adjusted for city, sex, age, 

computer experience, handedness, 

eye problems, number of siblings, 

parental education, prenatal 

smoking, parental 

psychological/psychiatric problem

Inattention Parent rating

Inattention Teacher rating

Premature Response

using German version of Test 

battery for attention 

performance at ages 8-12 yr.

1.3% (-4.0, 6.9)f

4.3% (-0.02, 10.4)

8.3% (-0.02, 19.0)

Bellinger et al.

(1994a)

79 young adults, ages 

19-20 yr, born 1970, 

Boston, MA area

Deciduous tooth 

(age 5-8 yr)

Q1: 2.9-5.9 ppm

Q2: 6.0-8.7 ppm

Q3: 8.8-19.8 ppm

Q4: 

19.9-51.8 ppm

Regression model adjusted for 

parental IQ, sex, SES, current drug 

use, current alcohol use, current 

illicit drug use, maternal education, 

maternal age, birth order

Several more examined

Correct Responses on CPT Q1: 98.0 (1.0)g

Q2: 97.6 (1.1)

Q3: 96.9 (1.1)

Q4: 94.6 (1.1)

a

Effect estimates are standardized to a 1 μg/dL increase in blood Pb level within the 10th to 90th percentile interval and standardized to the standard 

deviation of the test score to facilitate comparisons among tests that are scored on different scales. b

Standard error was estimated from the reported p-value.

c

Direction of effect estimate was changed to indicate an improvement in performance.

d

Sufficent data were not provided to calculate 95% CIs.

e

Odds in higher quantile of blood Pb level compared to that in lowest quantile of blood Pb level.

f

Results represent the change in false alarm rate.

g

Results represent the mean (SD) score in each quartile.
February 2012 5-127 Draft – Do Not Cite or Quote

Attention Deficit Hyperactivity Disorder in Children

1 Studies of ADHD were included in the 2006 Pb AQCD; however, previous studies did 

2 not conclusively demonstrate that blood Pb level was associated with ADHD itself. 

3 Importantly, previously studies examining ADHD were fewer in number and limited by 

4 smaller sample sizes (Gittleman and Eskenazi, 1983; David et al., 1972). Recent studies 

5 consistently reported associations of blood Pb levels with ADHD in children. Coherence 

6 and biological plausibility for these associations are provided by the consistent body of 

7 epidemiologic and toxicological evidence indicating Pb-associated increases in 

8 inattention, hyperactivity, and impulsivity. 

9 Several recent studies reported associations between concurrent blood Pb level and 

10 ADHD diagnosis or diagnostic indices in children between the ages of 8 and 17 years

11 (Table 5-10). While none of the studies examined the potential for confounding by 

12 HOME score, they did evaluate confounding by several other demographic and SES13 related variables, as well as parental history of psychopathology, including ADHD (Cho 

14 et al., 2010; Nicolescu et al., 2010). Recent studies also provided evidence of association 

15 between blood Pb level and ADHD in populations born in the mid- to late-1990s with 

16 relatively low concurrent blood Pb levels (means: 1.9 and 3.7 μg/dL) (Cho et al., 2010; 

17 Nicolescu et al., 2010). However, the temporal trends in Pb exposure in these non-U.S. 

18 populations may have differed from those in U.S. children of the same age.
February 2012 5-128 Draft – Do Not Cite or Quote

Table 5-10 Associations between blood Pb level and ADHD diagnosis or 

diagnostic indices in children

Study Population/Location

Blood Pb Levels 

(μg/dL)a Statistical Analysis Outcome

Effect Estimate

(95% CI)b

Nigg et al.

(2008)

150 children ages 

8-17 yr.

Year of birth and 

location NR

Concurrent

Mean (SD)

8-11 yr: 1.04 

(0.53)

12-17 yr: 1.03 

(0.54)

10th-90th: 0.5-1.7

Three-group analysis of covariance 

with sex and income.

No parental IQ or education

ADHD Dx

by two licensed clinicians

F(2,145) = 6.08, p = 0.04

Braun et al.

(2006)

4,704 children ages 

4-15 yr born 

1984-1998

U.S. NHANES 

1999-2002

Concurrent 

3rd quintile: 

1.1-1.3

Logistic regression model adjusted for 

postnatal ETS, prenatal ETS, age, sex, 

race, childcare attendance, health 

insurance coverage, ferritin levels

ADHD Dx or medication use 

at age 4-15 yr

1.4 (0.4, 3.4) blood Pb 

0.8-1.0 vs. <0.8 μg/dLc

2.1 (0.7, 6.8), blood Pb 

1.1-1.3 vs. <0.8 μg/dLc

2.7 (0.9, 8.4), blood Pb 

1.4-2.0 vs. <0.8 μg/dLc

4.1 (1.2, 14.0), blood Pb > 

2.0 vs. <0.8 μg/dLc

Froehlich et al.

(2007)

2,588 children, ages 

8-15 yr born 

1986-1996

U.S. NHANES 

2001-2004

Concurrent

2nd quartile: 

0.9-1.3

Logistic regression model adjusted for 

current household ETS exposure, sex, 

age, race/ethnicity, income, preschool 

attendance, maternal age, birth weight, 

and interaction terms for Pb and 

prenatal ETS interaction

ADHD Dx 8.1 (3.8, 18.7), blood Pb 

level > 2.0 μg/dL plus 

prenatal ETS exposure vs. 

blood Pb level <0.8 μg/dL 

and no prenatal ETS 

exposurec

Cho et al.

(2010)

667 children ages 

8-11 yr born 

1997-2000

Five Korean cities

Concurrent

Mean (SD): 1.9 

(0.67)

10th-90th: 1.2-2.8

Log linear regression model adjusted 

for age, sex, parental education, 

maternal IQ, child IQ, city, birth weight, 

urinary cotinine

Total ADHD rating, teacher

Total ADHD rating, parent

using Korean ADHD Rating 

Scale IV at ages 8-11 yr.

0.042 (0.017, 0.067)

0.010 (-0.013, 0.033)

Nicolescu et al.

(2010)

83 children ages 8-12 

yr born 1995-1999

Bucharest and 

Pantelimon, Romania

Concurrent 

Median (IQR): 3.7 

(2.6)

10th-90th: 1.8-7.1

Log linear regression model adjusted 

for city, sex, age, computer experience, 

handedness, eye problems, number of 

siblings, parental education, prenatal 

smoking, family psychopathology

ADHD score, parent

ADHD score, teacher

using German version of 

Conner’s scales at ages 8-12 

yr.

OR: 1.04 (1.00, 1.10)

OR: 1.06 (1.00, 1.12)

Roy et al.

(2009a)

756 children ages 3-7 

yr tested 2005-2006

Chennai, India

Concurrent Mean 

(SD): 11.4 (5.3)

10th-90th: 

5.8-18.3

Log linear regression model adjusted 

for age, sex, hemoglobin, average 

monthly income, parental education, 

number of other children, clustering in 

school and classroom

ADHD index z-score

using Conners’ ADHD/DSM- IV Scales at ages 3-7 yr.

0.002 (0, 0.033)

Chen et al.

(2007)

780 children in TLC 

trial followed ages 2-7 

yr

Baltimore, MD; 

Cincinnati, OH; 

Newark, NJ; 

Philadelphia, PA

Children underwent 

chelation therapy

Concurrent Mean 

(SD): 12.0 (5.2)

10th-90th: 

6.5-18.7

Regression-based path analysis 

adjusted for city, race, sex, language, 

parental education, parental 

employment, single parent, age at 

blood Pb measurement, caregiver IQ

ADHD index

Using Conners' Parent Rating 

Scale-Revised at age 7 yr.

0.54 (-1.22, 2.30) Direct

0.90 (0.35, 1.45) Indirect

a

Studies are presented in order of increasing quantile or population mean blood Pb level. b

Except where noted, effect estimates represent regression coefficients. All effect estimates are standardized to a 1 μg/dL increase in blood Pb level within 

the 10th-90th percentile interval. c

Odds ratio in higher quantile of blood Pb level, with children in the lowest quantile of blood Pb level serving as the reference group.

1 In addition to finding associations of concurrent blood Pb level with inattention and 

2 hyperactivity, Cho et al. (2010) found a statistically significant relationship with a total 

3 ADHD index as rated by teachers in population of children ages 8-11 years in Korea. In a 
February 2012 5-129 Draft – Do Not Cite or Quote

1 multivariate model that included age, sex, paternal education, maternal IQ, child IQ, city 

2 of residence, birth weight, and urinary cotinine, the effect estimate decreased by 12%; 

3 however, the association remained statistically significant (Table 5-10). Neuropsychiatric 

4 disease (e.g., ADHD, learning disability, depression, obsessive-compulsive disorder) in 

5 parents was not associated with child blood Pb level, and thus, likely did not confound 

6 the relationship between blood Pb level and ADHD index in the child. Results from this 

7 study also indicated that the relationship between blood Pb level and ADHD index was 

8 independent of the relationship between blood Pb level and IQ.

9 Similar to Cho et al. (2010), Nicolescu et al. (2010) found higher concurrent blood Pb 

10 levels to be associated with greater inattention, impulsivity, and total ADHD score among 

11 children in Romania, ages 8-12 years. As with individual symptoms, only concurrent 

12 blood Pb level, and not aluminum or mercury, was associated with a higher total ADHD 

13 rating (4% increase [95% CI: 0, 10] per 1 μg/dL increase in concurrent blood Pb level 

14 within the 10th-90th percentile interval [1.8-7.1 μg/dL]). The association did not change 

15 substantially in an analysis that excluded the 5 children with blood Pb levels at or above 

16 10 μg/dL. Adjustment for potential confounding by city, sex, age, computer experience, 

17 handedness, eye problems, number of siblings, parental education, prenatal smoking 

18 exposure, prenatal alcohol exposure, and parental history of psychological or psychiatric 

19 problems resulted in a decrease in statistical significance for blood Pb level associations, 

20 although investigators did not report whether the magnitude of association changed. It is 

21 important to note that model covariates were selected a priori but not all were associated 

22 with both blood Pb level and behavioral endpoint. Only sex, number of siblings, and 

23 maternal education were reported to be significantly associated with both blood Pb level 

24 and total ADHD rating. Parental history of psychological or psychiatric problems was 

25 significantly correlated with parental but not teacher rating of ADHD total score; 

26 however, ORs were fairly similar for ADHD score rated by teachers and parents. Despite 

27 the potential for over-adjustment in this study, higher blood Pb level was associated with 

28 a higher ADHD total score rating.

29 A recent analysis of data from NHANES 1999-2002 found a relationship between higher 

30 blood Pb level and greater odds of ADHD (parent-report of a diagnosis of ADHD or use 

31 of stimulant medication) among children ages 4-15 years (Braun et al., 2006). Strengths 

32 of this study included the large sample representative of U.S. children (n = 4704) and the 

33 low concurrent blood Pb levels at which associations were observed. These authors 

34 reported a monotonic increase in ORs from the lowest to highest quintile of blood Pb 

35 level, adjusting for age, race, prenatal smoking exposure, smoker in the home, 

36 preschool/child care attendance, health insurance coverage, and ferritin levels (Figure 

37 5-15). With children in the lowest quintile serving as the reference group (<0.8 μg/dL), 

38 children in the fifth quintile of concurrent blood Pb level (>2.0 μg/dL, maximum not 
February 2012 5-130 Draft – Do Not Cite or Quote

1 reported) had the highest prevalence of ADHD (OR: 4.1 [95% CI: 1.2, 14]). A similar 

2 OR was estimated when children with blood Pb levels > 5.0 μg/dL were excluded from 

3 the highest group. Children in the other three higher quintiles of blood Pb level (range: 

4 0.8-2.0 μg/dL) also had a higher prevalence of ADHD relative to the reference group 

5 (Table 5-10). 

Source: Braun et al. (2006)

Note: Adjusted for child age, gender, race/ethnicity, preschool attendance, serum ferritin, prenatal tobacco smoke exposure, smoker 

in the household, and insurance status (p for trend = 0.012). Data from ages 4-15 years from NHANES 1999-2002 by quintile of 

concurrent blood Pb level.

Figure 5-15 Adjusted odds ratios for Attention Deficit Hyperactivity Disorder 

(ADHD) among U.S. children.

6 In the NHANES 2001-2004 dataset restricted to children ages 8-15 years, Froehlich and 

7 colleagues (2009) found an interaction between prenatal tobacco smoke exposure 

8 (maternal report) and concurrent blood Pb levels. Investigators found ADHD to be 

9 independently associated with prenatal tobacco smoke exposure (OR: 2.4 [95% CI: 1.5, 

10 3.7]) and concurrent blood Pb levels (OR: 2.3 [95% CI: 1.5, 3.8]) in children with blood 

11 Pb levels >1.3 μg/dL compared with children with blood Pb levels < 0.8 μg/dL. These 

12 results were adjusted for current household smoke exposure, sex, age, race/ethnicity, 

13 income, preschool attendance, maternal age, and birth weight. As in the younger 

14 NHANES dataset, a similar OR was estimated when children with blood Pb levels > 

15 5.0 μg/dL were excluded from the highest tertile. The strongest association was observed 

16 in children with both high blood Pb level and prenatal tobacco smoke exposure. 

17 Compared to children in the lowest tertile of blood Pb levels with no exposure to prenatal 

18 tobacco smoke, children in the highest tertile of blood Pb level with exposure to prenatal 

19 tobacco smoke had the highest prevalence of ADHD (OR: 8.1 [95% CI: 3.5, 18.7]). 

20 Although ADHD was associated with low concurrent blood Pb levels (1.3-5 μg/dL), it is 
February 2012 5-131 Draft – Do Not Cite or Quote

1 important to acknowledge that the adolescents in both NHANES analyses were born in 

2 the late 1980s and may have had higher Pb exposures earlier in childhood that 

3 contributed to the observed associations.

4 Consistent with findings in the NHANES population, Nigg et al. (2008) found an 

5 association between concurrent blood Pb level and ADHD diagnosis among children 

6 8-17 years. Concurrent blood Pb levels in this population also were low, ranging between 

7 0.40 and 3.47 μg/dL. However, the examination of adolescents adds uncertainty 

8 regarding the relative contributions of higher past Pb exposures and current exposures to 

9 the observed associations. A key strength of this study over other studies was the 

10 verification of parental reports of ADHD by licensed clinicians using the same diagnostic 

11 criteria. Although the results were adjusted for household income, other potential 

12 confounders including parental IQ and HOME score were not examined.

13 Roy and colleagues (2009a) examined associations between concurrent blood Pb level 

14 and a range of behavioral problems in 756 children, ages 3-7 years, in Chennai, India. In 

15 this population, the mean blood Pb level was higher than that in most other studies (11.4 

16 [SD: 5.3] μg/dL); however, this study demonstrated associations of concurrent blood Pb 

17 level with inattention, hyperactivity, and a total ADHD index (assessed by teachers using 

18 Conners’ ADHD/Diagnostic and Statistical Manual for Mental Disorders) in younger 

19 children compared with other studies. In generalized estimating equations, higher blood 

20 Pb level was associated with a higher ADHD index score adjusting for age, sex, 

21 hemoglobin, average monthly income, parental education, number of other children, and 

22 clustering at school and classroom levels (Table 5-10). In analyses that did not adjust for 

23 potential confounding variables, mean ADHD scores were similar in the first three 

24 quartiles of blood Pb level and elevated in children with blood Pb levels > 18.71 μg/dL, 

25 suggesting that in this population, associations may have been driven by children with the 

26 highest blood Pb levels.

27 Whereas several studies indicated the direct effects of increasing blood Pb level on 

28 inattention and hyperactivity indices, Chen et al. (2007) found stronger indirect effects of 

29 blood Pb level on an ADHD index among children at age 5 years (Table 5-10). Important 

30 limitations of this study include the high blood Pb levels of children at ages 12 to 

31 33 months (20-44 μg/dL) that made them eligible for a randomized controlled trial of 

32 chelation. Because this study only enrolled children with Pb poisoning, it is difficult to 

33 extend findings to children with lower blood Pb levels. Blood Pb levels of study subjects 

34 remained high at age 5 years (mean: 12.0 μg/dL) and out the range of the current U.S. 

35 general population of children.

36 In recent commentaries to studies reporting associations between blood Pb level and 

37 ADHD in children, Brondum (2011, 2007) asserted the need for studies to consider 
February 2012 5-132 Draft – Do Not Cite or Quote

1 confounding by parental history of ADHD. Given the highly heritable nature of ADHD, 

2 parental history of ADHD is a strong risk factor for ADHD in children (Faraone and 

3 Doyle, 2001). Therefore, the risk associated with parental history of ADHD may mask 

4 the smaller magnitudes of risk associated with environmental exposures, including Pb. 

5 However, little information is available on an association between ADHD in the parent 

6 and blood Pb level in the child, thus, it is uncertain whether parental ADHD confounds 

7 and fully accounts for the associations observed between blood Pb level and ADHD in 

8 children. It should be noted that for parental ADHD to be a confounder, parental Pb 

9 levels would have to be highly associated with ADHD in the parent and with blood Pb 

10 level in the child. Among children in Korea, Cho et al. (2010) found that parental history 

11 of psychopathology (ADHD or several other conditions) was not associated with child 

12 blood Pb level. Among children in Romania, parental history of psychopathology was 

13 associated with a higher ADHD rating in the child as assessed by parents but not teachers

14 (Nicolescu et al., 2010). Higher blood Pb level was associated with higher ADHD ratings 

15 in children in models with blood Pb level alone and in models that adjusted for parental 

16 psychopathology plus other covariates. Further, associations of blood Pb level with 

17 ADHD rating assessed by parents and teachers were similar in magnitude (Table 5-10). 

18 While available data are limited, they do not provide strong evidence that parental 

19 psychopathology fully accounts for the associations observed between blood Pb level and 

20 ADHD in children.

Social Misconduct and Delinquent Behavior

21 The 2006 Pb AQCD described several studies in which higher levels of blood and bone 

22 Pb were associated with higher frequency or risk of misconduct in children or delinquent 

23 behavior in adolescents (U.S. EPA, 2006b). Previous studies primarily indicated 

24 associations with measures of cumulative Pb exposure, including early childhood average 

25 blood Pb level, lifetime average blood Pb level, or bone Pb level. Recent studies added to 

26 the collective body of evidence by demonstrating associations for concurrent blood Pb 

27 level and for a variety of conduct problems including aggression (Chiodo et al., 2007),

28 conduct disorder (Braun et al., 2008), oppositional defiant disorder (Nigg et al., 2008)

29 and more serious behaviors such as criminal arrests (Wright et al., 2008) (Table 5-11). 

30 Associations were found with early childhood and concurrent blood Pb levels and were 

31 demonstrated in populations with lower blood Pb levels (means: 1-8 μg/dL) than those 

32 examined in the 2006 Pb AQCD. Nonetheless, the weight of cumulative evidence 

33 supports associations in populations with mean blood Pb levels between 5 and 11 μg/dL. 

34 However, because some studies examined outcomes in adolescents and young adults born 

35 in the 1970s and 1908s, there is greater uncertainty regarding the level, timing, 

36 frequency, and duration of Pb exposure that contributed to the observed associations. 
February 2012 5-133 Draft – Do Not Cite or Quote

1 Several other studies examined children and adolescents born in the 1990s, who likely 

2 did not have as high historical Pb exposures earlier in childhood (Chandramouli et al., 

3 2009; Braun et al., 2008; Chiodo et al., 2007).

Table 5-11 Associations between blood Pb level and misconduct and delinquent 

behavior in children and young adultsa

Study Population/Location

Blood Pb 

Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)b

Blood Pb level as a continuous variable

Chiodo et al.

(2007)

506 African-American 

children in Detroit, MI 

area followed from birth 

(1989-1991) to age 7 yr.

Large proportions of 

children with prenatal 

exposure to cocaine or 

marijuana 

Concurrent

Mean (SD): 

5.0 (3.0)

10th-90th: 

2.1-8.7

Linear regression model adjusted for 

HOME, child sex, current marijuana 

use (both outcomes)

Caretaker education, alcohol use during pregnancy, (delinquent behavior

Age, caretaker psychopathology, 

maternal IQ (inappropriate behavior)

Several more examined

Delinquent behavior

Inappropriate behavior

using Achenbach Teacher 

Report Form at age 7 yr

0.09 (0, 0.18)c

0.09 (0, 0.18)c

Dietrich et al.

(2001)

195 children followed 

from birth (1979-1985) to 

age 15-17 yr

Cincinnati, OH

0-6 yr avg: NR Linear regression model adjusted for 

HOME score, parental IQ, current SES

Delinquent behavior

using the Self-Report of 

Delinquent Behavior at ages 

15-17 yr.

1.21 (1.08, 1.37)

Wright et al.

(2008)

250 adults followed from 

birth (1979-1985) to age 

19-24 yr

Cincinnati, OH 

Age 6 yr

Mean (SD): 

8.3 (4.8)

10th-90th: 

3.9-14.5

Negative binomial regression models 

adjusted for maternal IQ, sex, SES, 

maternal education

Several more examined

Criminal arrests

Violent arrests

assessed from county records 

at ages 19-24 yr

RRs: 1.05 (1.00, 1.09) 

prenatal

1.05 (1.01, 1.09) 6 yr

1.01 (0.97, 1.05) 0-6 yr avg

1.06 (0.97, 1.15) prenatal

1.08 (1.03, 1.14) 6 yr

1.05 (1.01, 1.10) 0-6 yr avg

Chen et al.

(2007)

780 children participating 

in TLC trial followed 

between ages 2-7 yr

Baltimore, MD; 

Cincinnati, OH; Newark, 

NJ; Philadelphia, PA

Children underwent 

chelation therapy

Concurrent 

Mean (SD): 

12.0 (5.2)

10th-90th: 

6.5-18.7

Regression-based path analysis 

adjusted for city, race, sex, language, 

parental education, parental 

employment, single parent, age at 

blood Pb measurement, caregiver IQ

Externalizing behavior, parent

Externalizing behavior, teacher

using Behavior Assessment 

System for Children at age 7 

yr.

1.024 (0.996, 1.053) Direct

1.008 (1.002, 1.014) Indirect

1.036 (1.003, 1.069) Direct

1.004 (0.998, 1.010) Indirect
February 2012 5-134 Draft – Do Not Cite or Quote

Study Population/Location

Blood Pb 

Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)b

Blood Pb level as a categorical variable

Braun et al.

(2008)

Children ages 8-15 yr

U.S. NHANES 

2001-2004

2nd quartile: 

0.8-1.0

Logistic regression with sample 

weights applied to produce national 

estimates, adjusted for oversampling 

of minorities and young children and 

adjusted for age, poverty income ratio, 

maternal age, sex, race, prenatal ETS, 

cotinine, blood Pb levels

Conduct disorder

Using Diagnostic Interview 

Schedule for ChildrenCaregiver Module at age 8-15 

yr

OR: 7.24 (1.06, 49.47), blood 

Pb level 0.8-1.0 μg/dL vs. 

blood Pb <0.8 μg/dLd

12.37 (2.37, 64.56), blood Pb 

level 1.1-1.4 μg/dL vs. blood 

Pb <0.8 μg/dLd

8.64 (1.87, 40.04), blood Pb 

level 1.5-10 μg/dL vs. blood 

Pb <0.8 μg/dLd

Chandramouli 

et al. (2009)

488 children ages 7-8 yr 

born 1991-1992

Avon, UK

Age 30 mo

Mean (SD): 

NR

Linear regression model adjusted for 

maternal education, home ownership, 

maternal smoking, HOME score, 

maternal SES, family adversity index, 

parenting attitudes at 6 mo.

Antisocial activities

using Antisocial Behavior 

Interview at ages 7-8 yr

OR: 0.93 (0.47, 1.83)d

blood Pb 2-5 vs. 0-2 μg/dL

1.44 (0.73, 2.84)d

blood Pb 5-10 vs. 0-2 μg/dL

2.90 (1.05, 8.03)d

blood Pb > 10 vs. 0-2 μg/dL

a

Results with continuous blood Pb are presented first followed by results with categorical blood Pb. Within each group, studies generally are presented in 

order of increasing mean blood Pb level.

b

Effect estimates are standardized to a 1 μg/dL increase in blood Pb level.

c

95% CI was estimated from a reported p-value of 0.05. d

Odds in higher quantile of blood Pb level compared to that in lowest quantile of blood Pb level.

1 The consistency of association of Pb biomarker levels with social misconduct and 

2 delinquent behavior was corroborated in a recent meta-analysis (Marcus et al., 2010) that 

3 included 19 studies (those reviewed in the 2006 Pb AQCD plus several recent studies) 

4 with a total of 8,561 children and adolescents (mean ages ranging from 3.5 years to 

5 18.4 years). Effect estimates were converted to Pearson correlation coefficients, and the 

6 combined effect estimate was r = 0.19 (95% CI: 0.14, 0.23). The key finding of this study 

7 was the robustness of associations to considerations of heterogeneity in study design, 

8 definition and assessment method of conduct problems, potential confounding variables 

9 examined, and blood Pb levels. The major source of heterogeneity in effect estimates was 

10 the biomarker of Pb examined. A larger magnitude of effect was estimated for hair Pb 

11 levels compared with bone or blood Pb levels; however, similar effect sizes were 

12 estimated for blood and bone Pb levels. Although the authors suggested that hair Pb may 

13 be a better indicator of cumulative Pb exposure compared to bone Pb levels, due to the 

14 high turnover of bone in throughout childhood and into adolescence, an empirical basis 

15 for interpreting hair Pb measurements in terms of body burden or exposure has not been 

16 firmly established (Section 4.3.4.2).

17 In the meta-analysis, effect sizes did not differ significantly between longitudinal and 

18 cross-sectional studies, among studies that examined different conduct problems 

19 (i.e., opposition defiance, delinquency, externalizing problems), or among studies that 

20 assessed conduct disorders using self-report, teachers report, or criminal records. 

21 Controlling for covariates such as SES, birth weight, parental IQ, and home environment 

22 did not attenuate the relationship between blood Pb level and conduct problems. In 
February 2012 5-135 Draft – Do Not Cite or Quote

1 addition to strengthening the evidence for the independent associations of Pb biomarker 

2 levels with conduct disorders, the results indicate that the lack of adjustment for any 

3 particular covariate, including HOME score, does not warrant limiting inferences from a 

4 particular study. 

5 Studies of children ages 7-8 years found associations between blood Pb level and 

6 misconduct adjusting for HOME score and several other potential confounding variables. 

7 They examined different lifestages of blood Pb level and did not clearly indicate 

8 associations between blood Pb level and misconduct with blood Pb levels less than 

9 5 μg/dL. In their longitudinal study of children in the U.K. who were born in the 1990s, 

10 Chandramouli et al. (2009) found associations of age 30 month blood Pb level not only 

11 with hyperreactivity but also with antisocial behavior at age 7-8 years. Results were 

12 adjusted for maternal education, home ownership, maternal smoking, HOME score, 

13 maternal SES, family adversity index, and parenting attitudes at 6 months. In analyses of 

14 blood Pb level as a categorical variable, greater antisocial activity was most clearly 

15 indicated in children with blood Pb levels greater than 10 μg/dL (Table 5-11). Chiodo et 

16 al. (2007) examined children, ages 7.5 years in Detroit, Michigan with a mean (SD) 

17 concurrent blood Pb level of 5.0 (3.0) μg/dL. Higher blood Pb was associated with higher 

18 teacher ratings of delinquent behaviors and inappropriate behaviors, adjusting for HOME 

19 score and other variables. The association with inappropriate behavior additionally 

20 adjusted for caregiver psychopathology. As with cognitive function and inattention 

21 outcomes, prenatal alcohol and drug exposure were not found to influence associations 

22 for blood Pb level. Blood Pb level was associated with behavioral problems in unadjusted 

23 and adjusted analyses, corroborating the independent associations for blood Pb level.

24 In the U.S. NHANES 2001-2004 dataset of children ages 8-15 years (born in the 1990s), 

25 Braun et al. (2008) analyzed blood Pb as a categorical variable and found higher 

26 prevalence of conduct disorder with concurrent blood Pb levels in the range of 0.8 to 1.0 

27 μg/dL, the lowest level among all studies examined. Compared with children with blood 

28 Pb levels less than 0.8 μg/dL, the OR (95% CI) in children with blood Pb levels 0.8-1.0 

29 μg/dL was 7.24 (1.06, 49.47). Odds ratios also were elevated in children with blood Pb 

30 levels higher than 1.0 μg/dL (Table 5-11). The wide 95% CIs likely were due to the small 

31 numbers of cases of conduct disorder, as assessed using DSM-IV criteria. For example, 

32 there were 22 cases of conduct disorder among children with blood Pb levels 0.8-1.0 

33 μg/dL. Poisson regression models showed that children with blood Pb levels 0.8-1.0 

34 μg/dL had 1.55 (95% CI: 1.09, 2.22) times as many conduct disorder symptoms as did 

35 children with blood Pb levels < 0.8 μg/dL. Investigators had data available on a limited 

36 set of potential confounders, and poverty income ratio was used to control for potential 

37 confounding by SES. 
February 2012 5-136 Draft – Do Not Cite or Quote

1 While other recent studies supported associations between biomarkers of Pb exposure 

2 and misconduct and delinquent behavior, they examined Pb levels in blood or tooth 

3 samples collected in the 1980s when Pb exposures were much higher (Fergusson et al., 

4 2008; Wright et al., 2008). Although this evidence may be less informative as to the 

5 relationship between current blood Pb levels and misconduct and delinquent behavior in 

6 adolescents and young adults, it is important to note the consistency of findings among 

7 populations with widely varying blood Pb levels and historical Pb exposures.

8 In the CLS cohort, prenatal and postnatal blood Pb levels previously were reported to be 

9 associated with self- and parent-reported delinquent and social acts at ages 16-17 years

10 (Dietrich et al., 2001). Wright et al. (2008) extended these findings to include 

11 associations of prenatal and postnatal blood Pb levels with criminal arrests (ascertained 

12 from county records) at ages 19-24 years. Mean blood Pb levels were 8.3 μg/dL (range 

13 1-26) for the prenatal period (maternal blood), 13.4 μg/dL (range 4-37) for the average 

14 between birth and age 6 years, and 8.3 μg/dL (range 2-33) at age 6 years. In models that 

15 adjusted for maternal IQ, sex, SES score, and maternal education, the relative risks (RRs) 

16 for total arrests per 1 μg/dL increment in blood Pb level were 1.07 (95% CI: 1.01, 1.13) 

17 for prenatal blood Pb level, 1.01 (95% CI: 0.97, 1.05) for average childhood blood Pb 

18 level, and 1.05 (95% CI: 1.01, 1.09) for blood Pb level at age 6 years. Blood Pb levels 

19 measured during these lifestages also were associated with increased risk of violent 

20 criminal arrests (Table 5-11). Although interactions terms for blood Pb by sex were not 

21 statistically significant, the attributable risk for males was considerably higher for males 

22 (0.85 arrests/year [95% CI: 0.48, 1.47]) than for females (0.18 [95% CI: 0.09, 0.33]). 

23 Results from the two CLS studies suggest that in addition to the prenatal blood Pb levels, 

24 early childhood blood Pb levels may also predict criminal behavior in adulthood. 

25 However, it is important to note that in these CLS studies, concurrent blood Pb levels 

26 were not analyzed. Therefore, these studies do not provide information on the potential 

27 effects of more recent Pb exposures or differences in association between earlier and 

28 more recent blood Pb levels.

29 A strength of Wright et al. (2008) was the detailed examination of potential confounding 

30 by a large number of variables. All of the examined covariates were weakly correlated 

31 with blood Pb levels (r = 0.24-0.35), thereby minimizing the potential for confounding. 

32 Nonetheless, variables such as maternal IQ, maternal education, and SES were included 

33 in the model because they were associated with arrests in the full multivariate model or 

34 changed the blood Pb level by more than 10%. HOME score was similar between 

35 subjects with and without criminal arrest records and did meet the criteria for inclusion in 

36 the final model.
February 2012 5-137 Draft – Do Not Cite or Quote

1 In the New Zealand cohort of children born in 1977, dentin Pb levels measured between 

2 ages 6 and 8 years were associated with self-reported and documented violent or property 

3 convictions at ages 14-21 years (Fergusson et al., 2008). Similar to those from the meta4 analysis, these findings pointed to an effect of cumulative childhood Pb exposure. 

5 Analyses were adjusted for SES, parental criminal record, parental education, and 

6 parental alcoholism, and although effects estimates decreased in adjusted models, 

7 associations remained statistically significant.

8 Several studies of misconduct and delinquent behavior aimed to characterize whether 

9 associations with biomarkers of Pb exposure were independent of effects on IQ and 

10 educational attainment. Most studies found that associations of Pb biomarkers with 

11 misconduct and delinquent behavior remained statistically significant in a model that 

12 additionally adjusted for child IQ or educational attainment, indicating that Pb exposure 

13 may have a direct effect on misconduct independent of its effect on IQ (Chandramouli et 

14 al., 2009; Fergusson et al., 2008; Burns et al., 1999). It is important to note that because a 

15 decrement in IQ may on the causal pathway to behavioral problems, including both IQ 

16 and behavioral problems in the same model may result in an underestimate of the effect 

17 on behavior. Chen et al. (2007) used path analysis to characterize the direct effects and 

18 indirect effects (mediated through child IQ) of blood Pb level on externalizing problems 

19 (i.e., outbursts of behavior); however, results were inconclusive. A direct effect was 

20 estimated for externalizing problems assessed by teachers and an indirect effect was 

21 estimated for problems assessed by parents. These findings may have limited 

22 applicability to the general population given that the children in the study population had 

23 been referred for chelation therapy at enrollment because of high blood levels, and it is 

24 uncertain whether the observed associations were due to the residual effect of high blood 

25 Pb levels (20-44 μg/dL) four years earlier.

5.3.3.2 Toxicological Studies of Behavior

Neurobehavioral Changes

26 The effects of Pb on neurobehavioral changes in animals are well characterized with 

27 various targeted sites including the prefrontal cerebral cortex, cerebellum, and 

28 hippocampus; affected functions include cognition, execution of motor skills, and 

29 memory/behavior. As discussed in earlier Pb AQCDs, young animals are especially 

30 susceptible to the effects of Pb due to the ongoing development of the nervous system 

31 with greater Pb absorption and retention. Pb exposure has been documented to induce 

32 neurobehavioral changes in exposed animals including effects on learning, social 

33 behavior, memory, attention, motor function, locomotor ability and vocalization. At the 
February 2012 5-138 Draft – Do Not Cite or Quote

1 cellular level, Pb impairs axon and dendritic development and contributes to 

2 neurochemical changes in proteins, membranes, redox/antioxidant balance, and 

3 neurotransmission through a multitude of mechanisms, many of which involve the 

4 capability of Pb to mimic calcium. Very early toxicological research on neurobehavioral 

5 endpoints failed to capture the disposition of Pb and its resulting body burden or blood Pb 

6 level and was thus difficult to use in risk assessment. 

7 The 1986 Pb AQCD reported findings of Pb-induced aberrant operant conditioning tasks 

8 in rodents and non-human primates (some with resulting blood Pb levels: 11 to 15 μg/dL) 

9 as well as hyperactive or inappropriate Pb-mediated responses (U.S. EPA, 1986b). It was 

10 indicated that these effects were possibly of hippocampal origin and showed a curvilinear 

11 response, decreasing at higher Pb doses possibly due to impairment of motor function

12 (Ma et al., 1999; Crofton et al., 1980). Pb exposure in laboratory animals resulted in 

13 distractibility, reduced to adapt to changes in behavior, impaired ability to inhibit 

14 inappropriate responses, and perseveration (U.S. EPA, 2006b). Pb has been shown to 

15 impair learning in Fixed Interval tasks (FI) as indicated by premature responses in the 

16 absence of a fixed schedule of reinforcement or reward (Section 5.3.2.2). These findings 

17 in animals are consistent with observations in children that blood Pb levels are associated 

18 with poorer impulse control as rated by parents or teachers or assessed using the 

19 continuous performance test (Figure 5-14 and Table 5-9). Interresponse rates and overall 

20 run rate are the two subcomponents of FI response rate. The 2006 Pb AQCD reported 

21 consistent findings for Pb exposure (resulting in blood Pb levels: 58 to 94 μg/dL) 

22 affecting FI response rates, by means of decreased interresponse times. Some studies 

23 indicated decreased interresponse times in animals with blood Pb levels of 11 μg/dL

24 (U.S. EPA, 2006b). Discrimination reversal has been shown to be especially sensitive to 

25 Pb exposure. Spatial and non-spatial discrimination reversal or reversal of a previously 

26 learned habit was significantly affected after developmental Pb exposure and was 

27 exacerbated with distracting stimuli. Repeat-acquisition testing revealed that these 

28 deficits were likely not due to sensory or motor impairment at this dose. Together, the 

29 data from the 2006 Pb AQCD showed that social behavior and learning in rodents and 

30 nonhuman primates is significantly affected by Pb exposure that results in blood Pb 

31 levels 15-40 μg/dL. 

32 In the new literature, gestationally and early postnatally (gestation to PND10, G+P) Pb33 exposed male mice (low and high dose Pb: 10 and 42 μg/dL blood Pb level at PND10, 

34 respectively) were significantly less active than were control mice, and low dose mice 

35 were significantly less active than were high dose mice, demonstrating a nonlinear 

36 concentration-response relationship (Leasure et al., 2008). A similar nonlinear 

37 concentration-response relationship was observed for changes in corticosterone in male 

38 mice exposed post-weaning to Pb (Virgolini et al., 2005). Activity level of G+P Pb-
February 2012 5-139 Draft – Do Not Cite or Quote

1 exposed female mice versus controls was unaffected (Leasure et al., 2008).

2 Amphetamine-induced motor activity was monitored in male and female G+P Pb3 exposed mice at 1 year of age. Amphetamine-induced activity of male low and high dose 

4 G+P Pb-exposed offspring was significantly elevated over that in controls; G+P Pb5 exposed females had no change in sensitivity to amphetamine-induced motor activity

6 (Leasure et al., 2008).

7 Herring gull chicks injected with a single i.p. bolus dose of Pb (100 mg/kg Pb-acetate, a 

8 dose created to be similar to that which wild herring gulls are exposed in the wild) on 

9 PND2 were found to have neurobehavioral deficits and learning deficits. Pb-exposed 

10 chicks displayed multiple deficits related to impaired survival skills including decreased 

11 time spent begging the parent for food, decreased accuracy at pecking for food in the 

12 mouth of the parent bird, decreased time spent in the shade (behavioral 

13 thermoregulation), decreased learning in food location, decreased recognition of familiar 

14 individuals (caretaker or sibling), and slower development of motor skills (treadmill test) 

15 versus control birds (Burger and Gochfeld, 2005). The impaired thermoregulation with 

16 Pb exposure agrees with earlier work in Pb-exposed rat pups that also showed impaired 

17 thermoregulatory behavior, i.e., impaired ultrasonic vocalization (Davis, 1982). These 

18 studies in herring gull chicks demonstrate that a single dose of Pb early in life can induce 

19 neurobehavioral deficits that affect survival skills. 

20 Rhesus monkeys exposed to Pb in daily milk formula from PND8 to 1 or 2 years of age 

21 (Pb-acetate/50% glucose in 4 cc of commercial milk formula producing blood Pb of 

22 35-40 μg/dL) were assessed for tactile defensiveness using the Sensory Processing Scale 

23 for Monkeys, an adaptation of laboratory observational measures of sensory processing 

24 for children (Baranek and Berkson, 1994). Tactile defensiveness in children is defined as 

25 “feelings of discomfort and a desire to escape the situation when certain types of tactile 

26 stimuli are experienced” (Ayres, 1964) and is associated with emotional dysregulation, 

27 inattention, and difficult social relations. Other reports have shown that tactile 

28 defensiveness in monkey offspring is affected by prenatal stress alone without Pb 

29 exposure (Schneider et al., 2008), a factor that has been shown to affect other Pb-related 

30 outcomes.

Attention

31 Epidemiologic studies consistently have reported associations between higher blood Pb 

32 level and deficits in attention (inattention, distractibility, impulsivity, or ADHD) in 

33 children (Section 5.3.3.1). Animal toxicological studies in the 2006 Pb AQCD detailed 

34 attention deficits in animals undergoing various tests including FR/waiting-for-reward 
February 2012 5-140 Draft – Do Not Cite or Quote

1 testing and tests that employed signal detection with distraction, a test recording 

2 omissions after exposure to an external distraction. 

3 With FR testing, rats are trained on a FR/waiting-for-reward behavioral baseline, learning 

4 to produce food delivery by pressing a lever 50 times. Rodents can earn additional food 

5 by withholding lever presses by waiting; free food is given at increasing time intervals 

6 after completion of the FR. Brockel and Cory-Slechta (1998) exposed male Long-Evans 

7 rats to 0, 50, or 150 ppm Pb-acetate in water from weaning, which produced respective 

8 blood Pb levels of <5, 11, and 29 μg/dL after 3 months of exposure. After 40 days of 

9 exposure, rats were trained on a FR schedule, and the higher dose animals had 

10 significantly more frequent response rates and resets of the waiting period than did the 

11 low dose group and controls. In the waiting behavior component, wait time was 

12 significantly lower in both treated groups compared to controls. The higher dose group 

13 animals also had an increased number of reinforcers and a higher response to 

14 reinforcement ratio than did low dose and controls. Mechanistic understanding of the 

15 aforementioned FR deficits was studied using the same FR schedule with similar 

16 postweaning dosing of 0, 50, and 150 ppm Pb that yielded respective blood Pb levels of 

17 <5, 10, and 26 μg/dL after 3 and 7 months of exposure. Administration of dopamine 

18 receptor antagonists attenuated Pb-induced effects on FR schedule testing, suggested a 

19 role for D2 receptors in Pb-induced behavioral impairments. 

20 Another study used a similar postweaning Pb exposure as that in earlier studies (Cory21 Slechta et al., 1998) to yield blood Pb levels of <5, 16, and 28 μg/dL and evaluate 

22 sustained attention using testing of signal detection during a distraction (Brockel and 

23 Cory-Slechta, 1999). Rats earned food rewards by discriminating correctly between a 

24 target and distracter light. A 13 second time-out was given for incorrect responses. Pb 

25 exposure produced no deficits in attention with this testing. Further work since the 2006 

26 Pb AQCD affirmed these findings. Testing for signal detection with distraction showed 

27 no effect after postnatal Pb exposure in female rats (Stangle et al., 2007). The two dose 

28 groups (20 ppm or 300 ppm Pb-acetate in drinking water with lactational, and drinking 

29 water exposure, PND1-PND30) yielded blood Pb levels of 13 μg/dL and 31 μg/dL

30 (Stangle et al., 2007). Another recent study reported that impaired auditory threshold task 

31 related behavioral testing was likely due to inattention in Pb-exposed animals. The 

32 inability of some of the monkeys to engage or focus on the task at hand yielded fewer 

33 available measurements in Pb-exposed animals versus control animals (Laughlin et al., 

34 2009). These rhesus monkeys were exposed to Pb-acetate gestationally (dam drinking 

35 water, 3 months prior to mating) through age 5.5 months (weaning) and had resulting 

36 bone Pb levels at 11 years of 7 and 13 μg/dL for prenatal and postnatal groups, 

37 respectively, and blood Pb levels during Pb exposure of 35 and 46 μg/dL, respectively. 

38 Animals were tested at age 13 years when blood Pb levels had returned to baseline levels. 
February 2012 5-141 Draft – Do Not Cite or Quote

1 Thus, the inattention literature demonstrates that impulsivity, waiting-for-reward 

2 behavior, and sustained attention may all be affected by Pb with the first two possibly 

3 affected to a greater extent than is sustained attention.

5.3.3.3 Epidemiologic Studies of Mood in Children

4 A majority of investigation of the effects of Pb on behavior in children has focused on 

5 externalizing behaviors such as inattention, hyperactivity, aggression, and delinquency. 

6 However, studies also have linked biomarkers of Pb exposure with internalizing 

7 behaviors related to emotional dysfunction, including withdrawn behavior, depression, 

8 fearfulness, and anxiety. These behaviors were assessed frequently in school-aged 

9 children using the Child Behavior Checklist, and as with inattention, hyperactivity, and 

10 misconduct, often were rated by parents and/or teachers.

11 Several earlier results were provided by the prospective cohort studies in Boston, 

12 Cincinnati, Port Pirie, Australia, and Yugoslavia. Collectively, these studies found higher 

13 ratings for internalizing behaviors in association with concurrent blood Pb level as well 

14 as biomarkers of cumulative Pb exposure such as multiyear average blood, tooth, and 

15 bone Pb levels (Burns et al., 1999; Wasserman et al., 1998; Bellinger et al., 1994b; 

16 Dietrich et al., 1987b). While several studies demonstrated associations with biomarkers 

17 of long-term, cumulative exposures, results from the Cincinnati cohort provided support 

18 for the effects of shorter-duration exposures. Among CLS infants, blood Pb level at age 

19 3 months and average blood Pb level at age 6 months was associated with less positive 

20 mood in white infants at age 6 months (Dietrich et al., 1987b). In the Boston cohort, 

21 subjects were examined at age 8 years and 19-20 years. Pb levels measured in deciduous 

22 teeth but not cord blood were associated with a higher rating of internalizing behaviors at 

23 age 8 years but not in adulthood, indicating the lack of persistence of effects of early 

24 exposure (Bellinger et al., 1994a; Bellinger et al., 1994b). Wasserman et al. (1998) also 

25 examined blood Pb levels measured at various lifestages in the Yugoslavia cohort and 

26 found that concurrent blood Pb was associated more strongly with anxious-depressed and 

27 withdrawn behaviors at age 3 years than were blood Pb levels measured prenatally or 

28 between ages 6 months and 2 years. Thus, while biomarkers of Pb exposure were 

29 consistently associated with poorer mood and emotional state in children, there was no 

30 clear indication of differences in association among biomarkers measured at various 

31 lifestages. Associations between Pb biomarkers with mood also were consistently 

32 reported in recent studies, with most studies examining concurrent blood Pb level (Liu et 

33 al., 2011a; Bao et al., 2009; Roy et al., 2009b; Chiodo et al., 2004). However, several had 

34 limited generalizability to the general population of U.S. children due to the examination 
February 2012 5-142 Draft – Do Not Cite or Quote

1 of children residing near Pb sources or high prevalence of prenatal alcohol or drug 

2 exposure (Liu et al., 2011a; Bao et al., 2009; Chiodo et al., 2004).

3 Associations of Pb biomarkers with mood and emotional state in children were observed 

4 after adjusting for a wide range of potential confounding variables including parental 

5 education and HOME score. Several studies that each adjusted for a different set of 

6 covariates found similar effect estimates in univariate and multivariate models (Burns et 

7 al., 1999; Wasserman et al., 1998; Bellinger et al., 1994b), supporting the robustness of 

8 associations with Pb biomarkers. Associations between blood Pb level and internalizing 

9 behaviors also were observed in studies that adjusted for maternal psychopathology

10 (Burns et al., 1999; Wasserman et al., 1998). Thus, the evidence indicates that 

11 associations observed between Pb biomarkers and internalizing behaviors in children are 

12 not driven by confounding by any particular measured variable. 

13 In the collective body of evidence, associations between blood Pb level and internalizing 

14 behaviors were observed in populations of school-aged children with mean blood Pb 

15 levels between 8 and 28 μg/dL. Mean blood Pb levels were not related to the age of the 

16 study population. In the limited investigation of populations with mean blood Pb levels of 

17 approximately 5 μg/dL, results were inconclusive. Chiodo et al. (2004) found an 

18 association with internalizing behaviors in children (age 7 years) in Detroit, in whom the 

19 prevalence of prenatal alcohol and drug use was high. Neither exposure was found to 

20 influence associations with blood Pb level. Another study that examined Inuit children 

21 (age 5 years) in Quebec, Canada, did not find an association between concurrent blood Pb 

22 level and internalizing behaviors (Plusquellec et al., 2010).

23 A common observation across studies was finding that Pb biomarkers were associated 

24 with multiple indices of neurodevelopmental function, i.e., FSIQ, executive function, 

25 externalizing, and internalizing behaviors, within the same population. Whereas some 

26 studies found stronger associations for externalizing behaviors than for internalizing 

27 behaviors (Plusquellec et al., 2010; Bellinger et al., 1994a; Sciarillo et al., 1992), most 

28 did not find a clear difference in the strength of association (Roy et al., 2009b; Chiodo et 

29 al., 2004; Wasserman et al., 1998; Bellinger et al., 1994b). In the Port Pirie population, 

30 Burns et al. (1999) found that lifetime average blood Pb levels were associated with 

31 externalizing behaviors more strongly in boys and with internalizing behaviors more 

32 strongly in girls, indicating potential sex-based differences. These findings demonstrating 

33 Pb effects on a wide spectrum of cognitive and behavioral indices are not surprising 

34 given that Pb exposure is linked to changes in the HPA axis and dopaminergic and 

35 GABAergic systems, which are involved in mediating cognitive function, behavior, and 

36 mood. Dietrich et al. (1987b) provided support for Pb exposure affecting a wide range of 

37 neurodevelopmental outcomes by characterizing the direct and indirect effects of Pb 
February 2012 5-143 Draft – Do Not Cite or Quote

1 using structural equations. The associations of prenatal Pb exposure (maternal and infant 

2 day 10 blood) with poorer mood in infants aged 6 months were found to be mediated 

3 through lower birth weight and/or shorter gestation. These results suggested that Pb may 

4 exert its effects by impairing nervous system development. It is well known that the fetal 

5 period is an active period for neuronal differentiation, dendritic branching, and 

6 synaptogenesis. Thus, it is not surprising that prenatal Pb exposure effects on nervous 

7 system development could result in a wide range of neurodevelopmental effects assessed 

8 later in childhood.

5.3.3.4 Epidemiologic Studies of Mood and Psychiatric Effects in 

Adults

9 Examination of the potential effects of Pb exposure on mood and psychiatric outcomes in 

10 adults has received far less attention than that in children and cognitive function in adults. 

11 Nonetheless, evaluation of mood states is an integral part of the neurocognitive test 

12 battery of the World Health Organization (WHO), and it has been suggested that indices 

13 of the Profile Of Mood States may be particularly sensitive to toxicant exposures

14 (Johnson et al., 1987). As with other nervous system endpoints in adults, several early 

15 studies of Pb-exposed workers (mean blood Pb levels ranging from 23.5 to 64.5 μg/dL) 

16 found higher prevalence of symptoms related to mood disorders and anxiety among Pb17 exposed workers than unexposed controls (mean blood Pb levels ranging from 

18 15-38 μg/dL) (Schwartz et al., 2005; Maizlish et al., 1995; Parkinson et al., 1986; Baker 

19 et al., 1985; Baker et al., 1984; Lilis et al., 1977).

20 While comprising a smaller body of evidence, studies of adults without occupational 

21 exposures demonstrated associations of blood and bone Pb level with mood. Analyses of 

22 men ages 48-70 years in the NAS indicated associations of both concurrent blood (mean: 

23 6.3 μg/dL [SD: 4.16]) and tibia (mean: 21.9 μg/g [SD: 13.5]) Pb levels with greater self24 reported symptoms of depression and anxiety (Rhodes et al., 2003). As bone Pb is a 

25 major contributor to blood Pb levels in adults without current occupational Pb exposure, 

26 associations with both biomarkers may indicate effects of cumulative Pb exposure. In a 

27 subsequent analysis of the same dataset, Rajan et al. (2007) found effect modification of 

28 the associations for patella and tibia Pb levels by ALAD genotype, although results were 

29 not in a consistent direction for a particular genotype. For a majority of the mood 

30 symptoms considered, tibia bone Pb levels were associated with larger ORs among men 

31 with the ALAD 1-1 genotype. In contrast, ORs for associations between patella Pb levels 

32 and several mood symptoms such as depression and positive symptom distress index 

33 were larger among ALAD 1-2/2-2 carriers. In the NAS, inconsistent effect modification 
February 2012 5-144 Draft – Do Not Cite or Quote

1 by ALAD genotype also was observed for associations between tibia Pb levels and 

2 cognitive performance (Rajan et al., 2008) (Section 5.3.2.5). 

3 A cross-sectional study of 1,987 adults age 20-39 years participating in NHANES 

4 1999-2004 was the largest study of mood and included both men and women (Bouchard 

5 et al., 2009). However, only concurrent blood Pb levels were available for analysis. 

6 Investigators assessed major depressive disorder (MDD), panic disorder and generalized 

7 anxiety disorder (GAD) using the WHO Composite International Diagnostic Interview, 

8 which follows criteria defined in DSM. Adults with concurrent blood Pb levels higher 

9 than 0.7 μg/dL had higher prevalence of all three symptoms. Adults in the highest 

10 quintile of blood Pb level (≥ 2.11 μg/dL) had the highest OR for MDD (OR: 2.32 [95% 

11 CI: 1.13, 4.75]) and panic disorder (OR: 4.94 [95% CI: 1.32, 18.48]) compared with 

12 adults with blood Pb levels < 0.7 μg/dL. For all endpoints, ORs were larger in analyses 

13 excluding current smokers. While this study demonstrated associations with relatively 

14 low concurrent blood Pb levels, it is important to note the uncertainty of the magnitude, 

15 timing, frequency, and duration of Pb exposure that contributed to the observed 

16 associations.

17 In analyses of cohorts in California and New England, Opler et al. (2008; 2004) reported 

18 associations between higher prenatal levels of plasma δ-ALA and subsequent diagnosis 

19 of schizophrenia spectrum disorder (ascertained using DSM-IV criteria) in adolescence 

20 and adulthood. In the absence of blood Pb levels, investigators measured δ-ALA levels in 

21 stored serum samples to serve as a surrogate for Pb exposure citing previous observations 

22 of a high correlation (0.90) between δ-ALA levels ≥ 9.05 ng/mL and blood Pb levels ≥ 

23 15 μg/dL. In the California cohort, a δ-ALA level ≥ 9.05 ng/mL was associated with 

24 schizophrenia spectrum disorder with an OR (95% CI) of 2.43 (0.99, 5.96), adjusting for 

25 maternal age at delivery. In analyses combining the California and New England cohorts, 

26 a δ-ALA level ≥ 9.05 ng/mL was associated with schizophrenia spectrum disorder with 

27 an OR (95% CI) of 1.92 (1.05, 3.52), adjusting for maternal age and education. A 

28 covariate-adjusted OR was not presented for the New England cohort alone, and it 

29 appeared that the association in the combined cohorts was driven by that observed in the 

30 California cohort. Studies in other populations with direct measurements of blood Pb 

31 levels are warranted to characterize the potential effects of Pb exposure on schizophrenia. 

32 These limited available data indicate associations with higher blood Pb levels (≥ 

33 15 μg/dL) in individuals born in the 1950s and 1960s who likely had higher early Pb 

34 exposures. Thus, these findings may have limited relevance to current lower levels of Pb 

35 exposure.
February 2012 5-145 Draft – Do Not Cite or Quote

5.3.3.5 Toxicological Studies of Mood, Emotional, and Psychotic 

Changes

1 As in epidemiologic studies, neurotoxicological studies often focus more on motor, 

2 sensory, behavioral or cognitive outcomes and less on psychological pathologies. As 

3 described in Section 5.3.3.3 above, epidemiologic studies indicating associations of Pb 

4 biomarkers with mood disorders in children and adults have grown in recent years. 

5 Likewise, studies increasingly have found that developmental Pb exposure (gestation and 

6 lactation) affects emotional state and mood disorder-like behavior in adult animal 

7 offspring. Offspring of Wistar rats treated with 10 mg of Pb-acetate daily by gavage 

8 during pregnancy (G) or pregnancy plus lactation (G+L) were tested in the open field test 

9 and the forced swimming test also known as the Porsolt test (de Souza Lisboa et al., 

10 2005). Blood Pb levels in the pups at PND70 were 5-7 μg/dL. The open field test 

11 monitors activity levels and movements of animals in three dimension and is a measure 

12 of hyperactivity, emotion (grooming or freezing), and exploratory behavior (rearing). In 

13 the forced swim test, animals are placed in a tank and monitored for helplessness, a 

14 response common in animal models of depression. (G+L) Pb-treated male rats had 

15 increased emotionality with the open field test as indicated by decreased rearing and 

16 increased immobility. (G+L) Pb-exposed female offspring had a significantly increased 

17 depressive phenotype in the forced swim test as indicated by increased immobility

18 (de Souza Lisboa et al., 2005). It is interesting to note that this is one of many Pb-induced 

19 changes that seem to be sex-specific with males showing increased emotionality and 

20 females showing elevated depressive-like symptoms (Figure 5-16). In another study, 

21 anxiety was assessed in offspring after G+L Pb exposure. Dams received Pb-enriched 

22 drinking water (2.84 mg/mL Pb-acetate trihydrate) and produced pups with blood Pb 

23 levels of 698 ng/g at PND25. Using the elevated plus maze test, Pb-exposed animals 

24 displayed no signs of anxiety (Molina et al., 2011). However, it is important to note the 

25 high concentrations of Pb exposure used in this study. Another study found that female 

26 pups exposed to Pb-acetate (0.2% to pups PND1-30) had increased anxiety at PND30 as 

27 measured by a decreased percentage of entries and time spent in the open arms of the 

28 plus-maze (Fox et al., 2010).

29 Depression may seem initially like an unexpected comorbidity for immune inflammatory 

30 dysfunction, but many forms of depression are linked with the same cytokine imbalances 

31 that occur with Pb-induced innate immune dysfunction (Maes; Pace and Miller, 2009).

32 Some researchers use sickness behavior and its associated malaise as a model for 

33 depression. Sickness behavior is characterized by overall malaise, decreased food intake, 

34 immobility and changes in core body temperature. Dyatlov and Lawrence (2002)

35 observed in mice that sickness behavior, which is due to an interaction of the immune

36 system and the CNS, was potentiated by Pb exposure (resulting in blood Pb level: 
February 2012 5-146 Draft – Do Not Cite or Quote

1 17 μg/dL) and was correlated with depletions in specific thymic T-cell populations 

2 (Figure 5-16). Pb exposure also potentiated the infection-dependent elevation in IL-1β, a 

3 cytokine that has been shown to inhibit hippocampal glutamate release in young but not 

4 aged animals. Sickness behavior was induced with Listeria monocytogenes infection; Pb 

5 exposure occurred from birth through lactation and was continued for a brief period after 

6 weaning until the experiment was terminated. 

Note: The blue headings provide four independent possible contributors to development of psychiatric disorders in Pb-exposed 

individuals.

Figure 5-16 Animal toxicology evidence of possible Pb-induced contributors 

to the development of mood and psychotic disorders.

7 Schizophrenia is associated with a shortened lifespan in humans as reflected by increased 

8 standardized mortality ratio (McGrath et al., 2008). An environmental origin of 

9 schizophrenia was proposed years ago (Tsuang, 2000), and prenatal Pb exposure, 

10 assessed using ALAD activity as a biomarker, was linked to schizophrenia in a group of 

11 adolescents and young adults in California (Opler et al., 2008; Opler et al., 2004).

12 Because of these observations, the animal toxicological field is beginning to explore the 

13 mechanisms that may contribute to schizophrenia development and has proposed two 

14 explanations. These are Pb-induced NMDA receptor (NMDAR) hypofunction and Pb15 induced decreases in hippocampal neurogenesis (Figure 5-16 to Figure 5-18). Pb may 

16 bind a divalent cation site in the NMDAR and allosterically inhibit glycine binding

17 (Hashemzadeh-Gargari and Guilarte, 1999); human studies of patients with schizophrenia 
February 2012 5-147 Draft – Do Not Cite or Quote

1 have shown aberrations at this site (Coyle and Tsai, 2004). These findings are consistent 

2 with the glutamatergic hypothesis of schizophrenia which purports that NMDAR 

3 noncompetitive antagonist use in patients with schizophrenia exacerbates their psychotic 

4 symptoms and that administration of antagonist to non-psychotic subjects can induce a 

5 schizophrenic phenotype. The second mechanistic hypothesis for Pb-associated 

6 schizophrenia induction is decreased hippocampal degenerate gyrus (DG) neurogenesis, 

7 which is seen in patients with schizophrenia (Kempermann et al., 2008; Reif et al., 2006),

8 in animal models of schizophrenia (Maeda et al., 2007) and in animal models with 

9 developmental Pb exposure (Verina et al., 2007; Jaako-Movits et al., 2005) (Figure 5-16

10 to Figure 5-18). Animal models of schizophrenia (i.e., phencyclidine administration) 

11 show decreased hippocampal DG neurogenesis that can be reversed by treatment with 

12 clozapine, which is often used to treat schizophrenia (Maeda et al., 2007). These DG 

13 pathways are also NMDAR-dependent. Studies cited in this section are further detailed in 

14 other sections of the ISA. Another study gives insight into pathways involved in Pb15 induced changes in neurogenesis in the developing brain. Exposure of zebrafish embryos 

16 to Pb (50–700 μM Pb-acetate in embryo medium from 0 to 6 days post hatch) caused 

17 significant apoptosis (increased TUNEL positive brain cells) and impaired neurogenesis 

18 as shown by decreased levels of gfap and huC in the brain; however two other genes 

19 involved in neurogenesis, crestin and neurogenin1, were unaffected in brains of Pb 

20 exposed embryos (Dou and Zhang, 2011). Thus, toxicological studies indicate that Pb 

21 exposure may result in mood disorders via behavioral, neurochemical, and ultrastructural 

22 changes.
February 2012 5-148 Draft – Do Not Cite or Quote

Figure 5-17 Schematic representation of the contribution of Pb exposure to 

the development of a phenotype consistent with schizophrenia.

Rodent 

Studies

Pb exposure and Schizophrenia

Associated in Human Cohorts

(Epidemiologic)

NMDA Receptor

Hypofunction

(Neurochemical)

Glutamatergichypothesis of 

schizophrenia

Human Populations or Cohorts

Decreased Hippocampal

Neurogenesis

(Structural)

Pharmacologicallyinduced Schizophrenia in 

Animal models

Pb exposure NMDAR 

Antagonism
February 2012 5-149 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Elsevier Science, Fox et al. (2010)

Note: (A) Control; (B) Pb exposed light micrograph pictures of Brd-U positive cells; (C) Counts of Brd-U positive cells (proliferating 

cells); and (D) Volume of dentate gyrus. *p <0.05 vs. control.

Figure 5-18 Neurogenesis (production of new cells) in the rat dentate gyrus 

after postnatal Pb exposure.

5.3.3.6 Integrated Summary of Behavior and Mood

1 Epidemiologic studies in children demonstrate associations of higher blood Pb levels 

2 with a range of behavioral problems, with the weight of evidence demonstrating 

3 associations with inattention and hyperactivity as rated by parents or teachers and as 

4 assessed using objective neuropsychological tests. Previous studies found associations 

5 with early childhood blood or tooth Pb level (i.e., ages 2-6 years), and recent studies 

6 expanded evidence to include associations with concurrent blood Pb level. Recent 

7 epidemiologic studies consistently found associations with inattention and hyperactivity 

8 in children ages 1 to 12 years with mean concurrent blood Pb levels of 2 to 5 μg/dL (Cho 

9 et al., 2010; Nicolescu et al., 2010; Plusquellec et al., 2010; Chiodo et al., 2007; 

10 Plusquellec et al., 2007), similar to those associated with cognitive function decrements. 

11 The epidemiologic findings are strengthened by observations in animals of Pb-induced 
February 2012 5-150 Draft – Do Not Cite or Quote

1 inattention and impulsivity in homologous tests of response inhibition (e.g., DSA, FI, 

2 DR). Tests in children such as CPT, Stroop, WCST share homology with tests in animals 

3 such as DSA, DR, and FR tasks in that they all test the ability to inhibit inappropriate 

4 responses, organize behavior in response to varying stimuli, and learn from the 

5 consequences of previous actions. Inattention and impulsivity in animals are most clearly 

6 indicated with gestational and early postnatal Pb exposures that result in blood Pb levels 

7 of 10 to 40 μg/dL. These findings in children and animals for Pb-associated dysfunction 

8 in response inhibition and response variability may be explained by observations that Pb 

9 affects dopaminergic neurons of the frontal striatum of the brain by altering dopamine 

10 release and receptor density . The circuitry in this regions is thought to mediate response 

11 inhibition. Whereas previous evidence was inconsistent, several recent epidemiologic 

12 studies indicate associations between higher concurrent blood Pb level and higher 

13 prevalence or incidence of ADHD diagnosis and its contributing diagnostic indices in 

14 children ages 8–17 years (Cho et al., 2010; Nicolescu et al., 2010; Roy et al., 2009a; Nigg 

15 et al., 2008; Braun et al., 2006). The biological plausibility for associations with ADHD 

16 is strongly supported by the large epidemiologic and toxicological evidence base 

17 demonstrating Pb-associated increases in inattention and impulsivity, both of which are 

18 primary symptoms of ADHD. A smaller but equally consistent body of evidence 

19 indicated associations of concurrent and early childhood blood Pb levels with social 

20 misconduct in children and delinquent behaviors in adolescents and young adults

21 (Chandramouli et al., 2009; Braun et al., 2008; Wright et al., 2008; Chiodo et al., 2007).

22 Associations of blood Pb levels with ADHD, misconduct, and delinquency were 

23 observed in populations of children with a wide range of blood Pb levels, 1 to 11 μg/dL, 

24 all similar in the strength of evidence.

25 While mood and emotional state have been examined less frequently compared with 

26 inattention and misconduct, several studies found associations of biomarkers of 

27 cumulative Pb exposure (i.e., tooth or childhood average blood Pb) and concurrent blood 

28 Pb levels with parental or teacher reports of withdrawn behavior or depression in children 

29 with mean blood Pb levels 8-28 μg/dL (Section 5.3.3.3). These findings in children are 

30 supported by a small body of toxicological studies in which prenatal plus lactational Pb 

31 exposure resulted in depression-like behavior in rodents.

32 Rather than examining externalizing behaviors and criminal behavior, a small body of 

33 studies of behavior in nonoccupationally-exposed adults examined and found 

34 associations of blood (Bouchard et al., 2009) and tibia (Rajan et al., 2008) Pb levels with 

35 depression and anxiety symptoms. All of these studies used single assessments of Pb 

36 biomarker levels and outcomes and analyzed associations in a cross-sectional manner. 

37 Thus, there is uncertainty regarding the critical level, timing, frequency, and duration of 

38 Pb exposure associated with mood and psychiatric symptoms. The associations with bone 
February 2012 5-151 Draft – Do Not Cite or Quote

1 Pb levels indicate an effect of cumulative Pb exposure. Concurrent blood Pb levels in 

2 adults reflect both cumulative and recent exposure, and it is uncertain what are the 

3 relative contributions of past versus recent Pb exposures to the observed associations. 

4 Associations of Pb biomarkers with withdrawn behavior and anxiety in children and 

5 adults may by explained by evidence for Pb-induced changes in the HPA axis, 

6 dopaminergic and GABAergic CNS processes. These processed have been shown to 

7 mediate anxiety and depression. While it may seem that Pb exposure affects different 

8 behaviors in children and adults, it is important to acknowledge differences in the 

9 evidence base. Studies of behavior in children and young adults have focused on 

10 inattention and misconduct; studies of older adults did not examine externalizing 

11 behaviors or misconduct. Differential effects in children and adults also may be expected 

12 given the predominance of different neurophysiological processes operating at different 

13 ages, for example, neurogenesis and brain development in children and 

14 neurodegeneration in adults.

5.3.4 Sensory Organ Function

5.3.4.1 Epidemiologic Studies of Sensory Organ Function in 

Children

15 Although not as widely examined as cognitive and behavioral outcomes, several studies 

16 found associations of higher blood Pb level with higher hearing thresholds and poorer 

17 auditory processing in children (U.S. EPA, 2006b). Such evidence is limited largely to 

18 studies described in the 2006 Pb AQCD. The prospective CLS with repeat measurements 

19 of blood Pb prenatally to age 5 years provided information on potentially important 

20 lifestages of exposure. In this cohort with higher blood Pb levels than those in 

21 contemporary U.S. children (lifetime average mean: 17.4 [SD: 8.8] μg/dL), poorer 

22 auditory processing was associated with higher prenatal (maternal), neonatal (10-day), 

23 early childhood, and lifetime average blood Pb levels, with the strongest associations 

24 observed for neonatal blood Pb level. A 1 μg/dL higher neonatal blood Pb level was 

25 associated with a 0.20-point (p ≤ 0.01) and 0.26-point (p≤ 0.10) lower score on the total 

26 and left ear Filtered Word test (indicative of incorrectly identified filtered or muffled 

27 words), after adjusting for hearing screen, social class, HOME score, birth weight, 

28 gestational age, obstetrical complications, alcohol consumption, and prenatal and 

29 postnatal blood Pb levels (Dietrich et al., 1992). Overall, the findings pointed to an effect 

30 of early Pb exposure during infancy.
February 2012 5-152 Draft – Do Not Cite or Quote

1 Large U.S. studies, including NHANES II (Schwartz and Otto, 1987) and the Hispanic 

2 Health and Nutrition Examination Survey (HHANES) also found associations between 

3 higher blood Pb level and lower auditory function; however, their cross-sectional design 

4 precluded comparisons among various lifestages of exposure (Schwartz and Otto, 1991).

5 In these studies, concurrent blood Pb level (median: 8 μg/dL) from 6 to 18 μg/dL was 

6 associated with a 2-db loss in hearing and an increase in the percentage (15%) of children 

7 with a substandard hearing threshold (2,000 Hz). Higher concurrent blood Pb level also 

8 was associated with higher hearing thresholds across several frequencies in a population 

9 of children in Poland with similar blood Pb levels (median: 7.2 μg/dL [range: 1.9 to 28])

10 (Osman et al., 1999). In the HHANES and Polish studies, associations persisted in 

11 analyses restricted to subjects with concurrent blood Pb levels below 10 μg/dL.

12 Mechanistic support for these observations in children was provided by Otto and 

13 colleagues (Otto and Fox, 1993; Otto et al., 1985b), who found associations of blood Pb 

14 level with lower brainstem auditory evoked potentials in children. Rothenberg et al.

15 (1994b) and Rothenberg et al. (2000) reported similar findings; however, the direction of 

16 association differed between prenatal (maternal) and postnatal (ages 1-4 years) blood Pb 

17 level. Postnatal blood Pb level was associated with lower interpeak intervals in auditory 

18 evoked potentials at age 5-7 years. Prenatal maternal blood Pb level showed a biphasic 

19 relationship, with a lower evoked potentials at blood Pb levels of 1-8 μg/dL and higher 

20 evoked potentials at blood Pb levels of 8-30 μg/dL. Recent studies also aimed to identify 

21 the locus in the auditory system where Pb may exert its effects on auditory function. 

22 Investigation was limited to a population of children with high blood Pb levels (means 33 

23 and 37 μg/dL) living in Pb glazing communities in Ecuador (Buchanan et al., 2011; 

24 Counter et al., 2011). In these studies, concurrent blood Pb level was not correlated with 

25 the acoustic stapedius reflex (Counter et al., 2011) or distortion product otoacoustic 

26 emissions (Buchanan et al., 2011), indicating lack of effect on the auditory brainstem or 

27 inner ear, respectively. Other loci were not examined. 

5.3.4.2 Epidemiologic Studies of Sensory Organ Function in 

Adults

28 Studies of auditory function reviewed in the 2006 Pb AQCD provided consistent 

29 evidence of association between blood Pb levels and changes in auditory evoked 

30 brainstem potentials in occupationally-exposed adults, but less consistent findings for 

31 hearing thresholds (U.S. EPA, 2006b). A few new studies of Pb workers found Pb32 associated increases in hearing thresholds. A recent study provided new evidence in 

33 nonoccupationally-exposed adults for associations of tibia Pb levels with hearing loss.
February 2012 5-153 Draft – Do Not Cite or Quote

1 Men in the NAS study were free of hearing loss at baseline and had hearing tested 

2 repeatedly (median 5 observations per subject) over a median of 23 years (Park et al., 

3 2010). Higher tibia Pb level, measured up to 20 years after initial hearing testing was 

4 associated with a faster rate of increase in hearing threshold for frequencies of 1, 2, and 8 

5 kHz and a pure tone average. Blood Pb was not examined in this study. In cross-sectional 

6 analyses adjusted for age, race, education, body mass index, pack-years of cigarettes, 

7 diabetes, hypertension and occupational noise (based on a job-exposure estimate from 

8 occupations), and presence of a noise notch (indicative of noise-induced hearing loss), 

9 higher patella bone Pb level (measured within 5 years of hearing test) was associated with 

10 a higher hearing thresholds for frequencies greater than 1 kHz. A 21 μg/g (IQR) increase 

11 in patella bone Pb level was associated with pure tone average hearing loss with an OR of 

12 1.48 (95% CI: 1.14, 1.91) in adjusted analyses. Similar, but slightly weaker associations 

13 were found for tibia bone Pb levels.

14 With new investigation, the weight of evidence indicates that blood Pb levels are 

15 associated increased hearing thresholds in adults with occupational Pb exposures. In a 

16 study of 183 Pb workers with relatively low blood Pb levels, 1 to 18 μg/dL, higher blood 

17 Pb level was correlated with higher hearing threshold at 4 kHz (Forst et al., 1997). Other 

18 studies examined workers with much higher blood Pb levels. In a study of 220 Pb battery 

19 workers with higher blood Pb levels (mean: 56.9 μg/dL [SD: 25.3]) (Wu et al., 2000),

20 although hearing impairment was associated with a measure of cumulative Pb exposure 

21 based on years of work and ambient Pb measurements, no association was found with 

22 blood Pb levels at the time of hearing testing in analyses adjusted for age, sex, and 

23 duration of employment. Another cross-sectional study examined 259 steel plant workers 

24 with no parental history of ear-related problems, no congenital abnormalities, no 

25 occupational organic solvent exposure, and hearing loss difference no more than 15 dB 

26 between both ears (Hwang et al., 2009). The participants had a mean (SD) concurrent 

27 blood Pb level of 54.3 (34.6) μg/dL. Average noise levels also were measured in work 

28 areas and dichotomized at 80dB. In analyses adjusted for age and work area noise 

29 (dichotomized at 80 dB), workers with blood Pb level ≥ 7 μg/dL had a statistically 

30 significant higher prevalence (range of ORs: 3.06 to 6.26) of hearing loss at frequencies 

31 of 3, 4, 6, and 8 kHz compared to workers with blood Pb levels ≤ 4 μg/dL.

32 A hospital-based case-control study recruited workers referred for hearing testing 

33 (average hearing thresholds above 25 dB) as cases and workers with normal hearing 

34 thresholds who were having occupational health examinations for other reasons as 

35 controls (Chuang et al., 2007). The 121 cases had a geometric mean blood Pb level of 

36 10.7 μg/dL, and the 173 controls had a geometric mean blood Pb level of 3.9 μg/dL based 

37 on measurements in samples collected at the time of the study. In models that adjusted for 

38 age, smoking, alcohol consumption, years of noise exposure, as well as Mn, As, and Se 
February 2012 5-154 Draft – Do Not Cite or Quote

1 levels in blood, higher blood Pb levels were associated with a statistically significant 

2 higher average hearing threshold (0.5-6 kHz). 

3 In summary, together, the findings from NAS (Park et al., 2010) and studies of Pb4 exposed workers indicate that biomarkers of Pb exposure are associated with lower 

5 auditory function in adults. Evidence for association with concurrent blood Pb levels is 

6 provided studies of adults with current occupational Pb exposures. Because only bone Pb 

7 levels measured after auditory testing were examined in the NAS study, further 

8 investigation is required to characterize the timing, level, frequency, and duration of Pb 

9 exposure contributing to lower auditory function in adults without occupational 

10 exposures.

5.3.4.3 Toxicological Studies of Sensory Organ Function

11 The 1986 and 2006 Pb AQCDs detailed the effects of Pb in animals on vision, including 

12 the retina and CNS visual processing areas, as well as the auditory system and described 

13 possible or known mechanisms of action where available. Pb exposure effects on sensory 

14 dysfunction and impaired sensory processing have been postulated to contribute to Pb15 associated effects on neurocognition and attention. In support of this hypothesis, Dietrich 

16 et al. (1992) found that higher prenatal (maternal) and postnatal blood Pb levels were 

17 associated with both lower performance on the screening test for auditory processing 

18 disorders and lower cognitive function in 5 year-old CLS children (Section 5.3.4.1). New 

19 research in this area expands upon the extant evidence by exploring sensory function in 

20 animals with lower Pb exposures and blood Pb levels.

Auditory Effects

21 The 2006 Pb AQCD discussed impaired auditory function in nonhuman primates exposed 

22 to Pb from gestation through age 8-9 years (resulting in blood Pb levels 33-56 μg/dL 

23 during Pb exposure period). Brainstem auditory evoked responses (BAER), which are 

24 used as a general test to assess neurological auditory function, revealed Pb-related effects 

25 that persisted even after Pb exposure had ceased and blood Pb levels had returned to 

26 baseline. In Pb-exposed animals (birth to age 13 years), half of the pure tone detection 

27 thresholds were outside of the control range at certain frequencies (Rice, 1997). In 

28 concordance with the data from developmentally Pb-exposed laboratory animals, 

29 elevated auditory thresholds, as measured with brainstem auditory evoked response 

30 (BAER), have also been associated with blood Pb levels in children (Rothenberg et al., 

31 2000; Rothenberg et al., 1994a). In addition to indicating hearing loss, BAER can 

32 indicate impaired synaptic maturation and incomplete neuron axon myelination leading to 
February 2012 5-155 Draft – Do Not Cite or Quote

1 impaired neuronal conduction (Schwartz and Otto, 1991; Gozdzik-Zolnierkiewicz and 

2 Moszyński, 1969. Thus, the findings from Rice (1997) indicated that early life Pb 

3 exposure impaired auditory function. The cochlear nerve in both developing and mature 

4 humans appeared to be especially sensitive to the Pb insult. At low to moderate Pb 

5 exposures, elevated thresholds and increased latencies were observed in brainstem 

6 auditory evoked potentials.

7 In a recent study, Laughlin et al. (2009) studied rhesus monkeys exposed to Pb-acetate

8 gestationally through age 5.5 months (dam drinking water, 3 months prior to mating until 

9 weaning, resulting in bone Pb levels at 11 years of 7 and 13 μg/dL for prenatal and 

10 postnatal groups, respectively and blood Pb levels during Pb exposure of 35 and 

11 46 μg/dL, respectively) and conducted auditory threshold testing and threshold task 

12 testing at 13 years of age after blood Pb levels had returned to baseline. At birth, animals 

13 were cross fostered, creating a control group, a prenatal Pb group, and a postnatal Pb 

14 group; however, Pb exposed animals were analyzed as a single group. Pb exposure 

15 induced small elevations in auditory thresholds in animals. Auditory threshold task16 related behavioral testing was also impaired in Pb-exposed animals. This study has 

17 multiple limitations which likely contributed to its not finding statistically significant 

18 aberrations. These limitation included limited power with the examination of 5 animals 

19 per group, the inability of some of the monkeys to engage or focus on the task at hand 

20 and thus had fewer available measurements, differences between the sexes in inattention, 

21 and mixing of the postnatal Pb and prenatal Pb animals into one group (Pb-exposed 

22 animals).

23 In summary, studies in nonhuman primates have shown lower BAER due to chronic Pb 

24 exposure (gestation through adulthood) that result in blood Pb levels in the range of 

25 33-56 μg/dL. In concordance with these findings, previous epidemiologic studies 

26 described lower BAER in children with higher prenatal maternal blood Pb levels and 

27 concurrent blood Pb levels. Collectively, these findings provide support for the 

28 associations observed between higher blood Pb levels and increased hearing thresholds 

29 (concurrent blood Pb) and lower auditory processing in children (neonatal) 

30 (Section 5.3.4.1). The animal evidence for long-term Pb exposure effects provide 

31 coherence with findings in adults with occupational Pb exposures for associations 

32 between blood Pb levels and increasing hearing thresholds and in nonoccupationally33 exposed adults, for associations with biomarkers of cumulative Pb exposure (i.e., tibia 

34 and patella Pb level) (Section 5.3.4.2). 
February 2012 5-156 Draft – Do Not Cite or Quote

Effects on Vision

1 In toxicological studies, Pb has been shown to affect multiple aspects of the visual system 

2 from the retina, to the sensory processing areas of the brain, to neurons involved in 

3 vision. The selective action of Pb on retinal rod cells and bipolar cells is well documented 

4 in earlier Pb AQCDs (Fox et al., 1997; Fox and Sillman, 1979) and research in this area 

5 continues to date. Pb exposure during perinatal development and adulthood has also been 

6 shown to affect the visual cortex (Costa and Fox, 1983) and subcortical neurons (Cline et 

7 al., 1996). Extensive work in nonhuman primates with various Pb exposure paradigms 

8 (development or lifetime Pb) showed sensory impairment, i.e., dysfunction of spatial and 

temporal visual function (Rice, 1998). Environmentally-relevant doses of Pb (10-

3 9 μM) 

10 administered to tadpoles inhibited the growth of developing neurons in the subcortical 

11 retinotectal pathway, the main efferent from the retina (Cline et al., 1996). Functional 

12 tests like electroretinograms (ERGs) show Pb-related aberrations in children, rodents and 

13 nonhuman primates. 

14 The animal toxicological data show that developmental windows and the dose of Pb 

15 contribute to the complex and variable effects of Pb with the retina. Table 5-12

16 summarizes Pb-related effects in retinal ERG studies. Female rats exposed postnatally to 

17 moderate and high levels of Pb (0.02 or 0.2% Pb-acetate exposure in dam drinking water 

18 from birth through weaning, resulting in weaning blood Pb levels of 19 and 59 μg/dL, 

19 respectively) had subnormal scotopic ERGs (decreased A- and B-wave amplitudes) with 

20 decreased sensitivity and temporal resolution when assessed at 90 days of age (Fox et al., 

21 1991) (Table 5-12). Similar results were obtained in multiple studies conducted in in vitro 

22 models (Otto and Fox, 1993; Fox and Farber, 1988; Fox and Chu, 1988). Monkeys 

23 exposed to moderately high levels of Pb continuously from the prenatal period to age 

24 7 years (350 or 600 ppm Pb-acetate, resulting in blood Pb levels of 40 and 50 μg/dL, 

25 respectively) had persistently increased maximal retinal ERG amplitude (B-wave only, 

26 supernormality) and increased mean ERG latency when assessed 2 years after 

27 termination of Pb exposure when blood Pb levels were <10 μg/dL (Lilienthal et al., 1988)

28 (Table 5-12). 
February 2012 5-157 Draft – Do Not Cite or Quote

Table 5-12 Summary of Pb-related retinal ERG studies

Study Species Sexa Pb Exposure Protocol/Dose

Maximal 

Blood Pb 

Level (μg/dL)

ERG 

Abnormality

Progenitor 

cell 

proliferation

Retinal Cellular 

Apoptosis

Retinal Dopamine 

Levels

Retinal 

Cell Layer 

Thickness

Fox et al.

(2008)

Long- Evans 

Rat

F Prenatal-PND10b

DW

Low 27 ppm

Moderate, 55 ppm

High, 109 ppm

12

24

46

Supernormal

Supernormal

Subnormal

Yes

Yes

No

Not affected

Not affected

Yes

Dose-dependent ↓

Dose-dependent ↓

Dose-dependent ↓

↑

↑

↓

Lilienthal et 

al., (1988)

Rhesus 

Monkey 

M 

& F

Pre- and Post-natal 

(lifetime), DW

350 ppm

600 ppm

~50

~115

Supernormal

Supernormal —

—

—

—

—

—

—

Fox et al.

(1997)

LongEvans 

Rat

F PND1-PND21

0.02% DW

0.2% DW

19

59

Subnormal

Subnormal — Yes

Yes

↓

↓

Rothenberg 

et al.

(2002a)

Human

children

M

& F

Prenatal 

1st trimester ≥ 10.5 Supernormal — — — —

Guguchkova 

et al. (1972)

Human M Occupational Subnormal — — — —

Otto and Fox

(1993)

Human M Occupational Subnormal — — — —

a F: Females; M: Males

b

PND: postnatal day

“—“ Denotes not measured.

1 A recent study exposed female Long-Evans rats to low (27 ppm), moderate (55 ppm), and 

2 high (109 ppm) levels of Pb-acetate in drinking water beginning 2 weeks before mating, 

3 throughout gestation, and until PND10 (G+P exposure) (Fox et al., 2008) (Table 5-12). 

4 Blood Pb levels in G+P Pb-exposed pups in the three groups were 10-12 μg/dL (lower), 

5 21-24 μg/dL (moderate), and 40-46 μg/dL (higher). This developmental window in the 

6 retina of the rat is equivalent to gestational human retinal development. Results of this 

7 rodent study demonstrated persistent supernormal scotopic rod photoreceptor-mediated 

8 ERGs (lower and moderate Pb exposure) similar to the associations observed between 

9 ERG and prenatal maternal blood Pb levels ≥ 10.5 μg/dL in male and female children

10 (Rothenberg et al., 2002a). Supernormal scotopic ERGs may be recorded without other 

11 overt opthalogical changes and are rarely seen in the clinical setting (Terziivanov et al., 

12 1983). Lower and moderate levels of G+P Pb exposure increased neurogenesis of rod 

13 photoreceptors and rod bipolar cells without affecting Müller glial cells and statistically 

14 significantly increased the number of rods in central and peripheral retina. Higher-level 

15 G+P Pb exposure (109 ppm, blood Pb level of 46 μg/dL) or moderate to higher level 

16 postnatal exposure (PND1-21, blood Pb levels 19 and 59 μg/dL) statistically significantly 

17 decreased the number of rods in central and peripheral retina, induced scotopic ERG 

18 subnormality in adult rats (Table 5-12), and statistically significantly decreased the retinal 
February 2012 5-158 Draft – Do Not Cite or Quote

1 Zn concentration. Pb exposure induced concentration-dependent decreases in adult rat 

2 retinal dopamine synthesis and utilization/release (Fox et al., 2008). Thus, the dose of Pb 

3 and the exposure window produce complex interactions in the retina that differentially 

4 affect retinal histology and functional tests, i.e., ERG as summarized in Table 5-12. 

Note: *p <0.05

Source: Fox et al. (2008)

Figure 5-19 Retinal a-wave and b-wave ERG amplitude in adult rodents after 

prenatal and early postnatal Pb exposure.

5 A recent study of adult zebrafish that were exposed as embryos (2 to 24 hours post6 fertilization) to water containing 0.03 μM PbCl2 showed fish with impaired response to 

7 visual stimulation (visual response to a rotating bar) under low light conditions. These 

8 zebrafish also failed to respond normally to mechanosensory stimulation (0.01 and 0.03 

9 μM PbCl2), showing a significantly impaired startle response. These data show that low10 level developmental Pb exposure causes sensorimotor deficits in fish (Rice et al., 2011).

11 Another study evaluating visual spatial acuity in rhesus monkeys (Laughlin et al., 2008)

12 exposed to Pb-acetate postnatally (PND8-26 weeks of age via commercial milk formula, 

13 achieving a target blood Pb of 35-40 μg/dL) found no effects of Pb exposure on spatial 

14 acuity as assessed with modified the Teller preferential looking paradigm.

15 Mechanistic understanding of the effect of Pb on the visual system includes its capability 

16 to displace divalent cations, act as an inhibitor of physiological enzymes, regulate cell 

17 proliferation and apoptosis, impair and perturb normal neuroanatomy formation, 

18 i.e., cytoarchitecture in the brain, and affect neurotransmitters. The effects of Pb on the 

19 retina have been shown to by mediated by its capability to act as a cGMP 

20 phosphodiesterase (PDE) inhibitor (Srivastava et al., 1995; Fox and Farber, 1988). The 

21 drug sildenafil citrate, another cGMP PDE inhibitor, can also cause visual problems 

22 including alterations in scotopic ERGs (Laties and Zrenner, 2002). With postnatal 

23 exposure of animals or in vitro Pb exposure of isolated rods, Pb has been shown to induce 

24 elevated cGMP which contributes to elevated rod calcium concentration (Fox and Katz, 

25 1992) and subsequently to apoptotic cell death in a concentration-dependent fashion. In 
February 2012 5-159 Draft – Do Not Cite or Quote

1 separate work in mice, low and moderate doses of Pb (27 or 55 ppm Pb-acetate in dam 

2 drinking water from gestation to PND10, resulting in blood Pb levels of 12 and 25 μg/dL, 

3 respectively), induced greater rod and rod bipolar cell neurogenesis (proliferation) and 

4 greater thickness and cell number of the outer and inner neuroblastic layers of the retina

5 (Giddabasappa et al., 2011; Fox et al., 2008). Rodents with moderate dose G+P Pb 

6 exposure (resulting in blood Pb level of 25 μg/dL) had 27-fold greater and prolonged 

7 retinal progenitor cell proliferation (Giddabasappa et al., 2011); at higher doses of Pb 

8 (109 ppm Pb-acetate, resulting in blood Pb levels of 56 μg/dL) there was no rod 

9 neurogenesis. Nitric oxide has been shown to regulate retinal progenitor cell proliferation 

10 in chick embryos (Magalhaes et al., 2006). Thus, these authors postulated that impaired 

11 NO production may contribute to aberrant retinal cell proliferation (Giddabasappa et al., 

12 2011). Pb exposure has been shown to impair NO synthase activity in other organs 

13 (Section 5.2.4.5). Pb has been shown to affect a plethora of neurotransmitters in the brain 

14 and it has recently been shown to affect neurotransmitters in the retina. In the 

15 aforementioned model of G+P rodent Pb exposure through PND10, Pb decreased 

16 dopamine (DA) synthesis and use in a concentration-dependent manner (Fox et al., 2008)

17 (Figure 5-20). These new data provide further insight into retinal changes by showing 

18 increased proliferation of Pb-exposed retinal progenitor cells without changes in 

19 apoptosis in G+P exposed rats (Fox et al., 2008).
February 2012 5-160 Draft – Do Not Cite or Quote

Note: *p <0.05

Source: Fox et al. (2008)

Figure 5-20 Retinal dopamine metabolism in adult control and gestationally 

lead-exposed (GLE) rats.

1 As discussed in the 2006 Pb AQCD, Pb-induced decreased Na/K+ ATPase activity have 

2 been reported in vitro and in vivo. Also, structural changes from chronic Pb exposure 

3 (birth to age 6) included cytoarchitectural changes in visual projection areas of the brain 

4 of monkeys; maximum blood Pb level in the low and high dose group reached 20 μg/dL 

5 and 220 μg/dL, respectively (Reuhl et al., 1989). Within the visual system, Pb has been 

6 shown to affect multiple pathways including signaling, enzyme inhibition, 

7 neurotransmitter levels, neuroanatomy formation, and cell proliferation and apoptosis.

8 In summary, developmental windows of exposure as well as exposure dose (Table 5-12) 

9 are important factors that contribute to Pb-induced changes in retinal function. The retina 

10 is a sensitive target for Pb-induced altered histology and retinal function. Prenatal, 

11 postnatal, lifetime or occupational exposures provide windows with different effects on 

12 the retina, effects that are modified by dose of Pb.
February 2012 5-161 Draft – Do Not Cite or Quote

5.3.5 Motor Function

1 Some studies in children have assessed fine motor function, i.e., response speed, 

2 dexterity, eye-hand coordination, as part of a battery of cognitive function and behavioral 

3 testing, and most have found associations with blood Pb level. Fewer studies have 

4 examined gross motor function, i.e., postural balance, action tremor, agility but also have 

5 found associations with blood Pb level.

6 Multiple studies were conducted in the CLS cohort at age 6 years and in adolescence at 

7 ages 12 and 15-17 years. Assessments in younger children indicated associations of 

8 concurrent and lifetime average blood Pb levels but not prenatal blood Pb levels with 

9 poorer visuomotor control and upper limb dexterity (Dietrich et al., 1993b) and poorer 

10 postural balance (Bhattacharya et al., 1995). Associations with both fine and gross motor 

11 function were adjusted for HOME score and race. Additional covariates included 

12 maternal IQ, SES, and sex for fine motor functions (Dietrich et al., 1993b) and height, 

13 BMI, birth weight, bilateral ear infection, and foot area for postural balance

14 (Bhattacharya et al., 1995). Blood Pb levels were associated with fine and gross motor 

15 function in unadjusted and adjusted analyses, indicating that bias from the measured 

16 confounders was not driving the observed associations. Subsequent analyses in the CLS 

17 demonstrated associations of earlier childhood average blood Pb levels (0-5 year average 

18 or 78 month average) with poorer fine (Ris et al., 2004) and gross motor function

19 (Bhattacharya et al., 2006) assessed in adolescence. While these findings suggest the 

20 persistence of Pb effects or early, long-term exposure effects, it is important to note that 

21 blood Pb levels measured later in childhood or concurrently with motor function were not 

22 examined.

23 Studies in other populations found associations of blood Pb level (primarily concurrent) 

24 with poorer fine motor function in children ranging in age from 3 to 16 years

25 (Palaniappan et al., 2011; Min et al., 2007; Wasserman et al., 2000). Among children in 

26 New England participating in NECAT (described in Section 5.3.2.1) Surkan et al. (2007)

27 found higher blood Pb levels to be associated with lower FSIQ but better fine motor 

28 function as indicated by faster finger tapping speed. Gross motor function also was 

29 assessed in other populations of children. In the Yugoslavian cohort, lifetime average 

30 blood Pb level was not associated with gross motor function (Wasserman et al., 2000).

31 Concurrent blood Pb level was associated with greater sway oscillation, alternating arm 

32 movements, and action tremor in a group of Inuit preschool children in Quebec, Canada

33 (Despres et al., 2005). Investigators considered potential confounding by several 

34 variables including HOME score, maternal education and nutrient levels. In animal 

35 studies, Pb exposure has shown mixed effects on endurance, balance and coordination as 

36 measured by rotarod performance. G+P Pb-exposed male mice had significantly shorter 
February 2012 5-162 Draft – Do Not Cite or Quote

1 mean latencies to fall from the rotarod compared with controls; females were unaffected. 

2 Further, low dose G+P Pb-exposed male mice had significantly poorer rotarod 

3 performance (i.e., fell off more quickly) than did high dose G+P male mice (resulting 

4 blood Pb levels at PND10: 10 μg/dL and 42 μg/dL, respectively), indicative of a 

5 nonlinear concentration-dependent relationship (Leasure et al., 2008). Other rotarod 

6 experiments at higher doses of Pb exposure and at various speeds of rotarod rotation 

7 yielded mixed results (Kishi et al., 1983; Grant et al., 1980; Overmann, 1977).

8 The collective body of evidence demonstrates that within the same population of 

9 children, blood Pb levels are associated with decrements in cognitive function, attention, 

10 and fine and gross motor function. Studies found associations with concurrent and 

11 childhood average blood Pb levels mostly in populations with mean levels ranging from 

12 11 to 28 μg/dL. Despres et al. (2005) found impaired gross motor function in a children 

13 with a mean blood Pb level of 5 μg/dL. Min et al. (2007) found impaired fine motor 

14 function in children with a mean concurrent blood Pb level of 2.9 μg/dL; however, the 

15 results were not adjusted by SES-related variables. motor skills are the result of the 

16 coordination of complex cognitive and physical processes in the cortex, cerebellum, 

17 vestibular systems, and visual system, and biological plausibility for the associations of 

18 blood Pb levels with fine motor skills observed in children is provided by observations 

19 that Pb exposure affects development and function of these systems.

5.3.6 Seizures in Animals

20 One neurological sign of high dose Pb exposure has been the development of epileptic 

21 form activity or seizures in animals (Krishnamoorthy et al., 1993). However, earlier 

22 studies in the animal toxicological literature exploring the effects of Pb on seizure 

23 activity and threshold showed mixed results. Pb-acetate (250, 500, or 1,000 ppm for 30 

24 days in drinking water to PND60 male Wistar rats, resulting in blood Pb levels of ~20, 

25 35, and 42 μg/dL, respectively) significantly decreased the elapsed time required to 

26 develop the first myoclonic jerk and tonic-clonic seizure (Arrieta et al., 2005); also, the 

27 dose of pentylenetetrazol (PTZ) required to induce seizures was significantly decreased

28 across all Pb dose groups. Some studies showed no effect of Pb on kindled animals

29 (Schwark et al., 1985; Alfano and Petit, 1981). Other studies showed differential 

30 susceptibility to convulsant-inducing agents in developmentally Pb-exposed rats (Chen 

31 and Chan, 2002). Sprague Dawley rats were exposed to Pb-acetate (0.2% w/v in drinking 

32 water from PND1-25, followed by 25 days with no Pb exposure). Seizures were induced 

33 at PND25 or PND50. At PND25, Pb exposure significantly decreased (PTZ)-, picrotoxin 

34 (PIC)-, and strychnine (STRY)-induced convulsion thresholds, but increased N-methyl35 D-aspartate (NMDA) and 4-aminopyridine (4-AP)-induced convulsion thresholds. At 
February 2012 5-163 Draft – Do Not Cite or Quote

1 PND50, the effects of PTZ, NMDA, and 4-AP remained similar to those at PND25, but 

2 PIC and STRY-dependent convulsion thresholds were reversed and significantly 

3 increased (Chen and Chan, 2002). Chen and Chan (2002) hypothesized that this 

4 differential effect may be due to selective effects on inhibitory and excitatory 

5 neurotransmission as an effect of age and blood Pb level.

6 Recent investigation expands on the work by Arrieta et al. (2005) by showing similar 

7 effects in another rodent species, BALB/c mice. Adult male BALB/c mice were exposed 

8 to Pb for 30 days via drinking water (resulting in blood Pb levels in control and 50, 100, 

9 200 and 400 ppm Pb groups of 0.02, 6, 11, 15 and 18 μg/dL, respectively) (Mesdaghinia 

10 et al., 2010). Exposure to 50 ppm Pb did not affect PTZ-induced seizure threshold, but all 

11 other doses significantly reduced the thresholds of face and forelimb clonus, myoclonic 

12 twitch, running and bouncing clonus, and tonic hindlimb extension. These studies show 

13 that Pb exposure may modulate seizure activity in animals. 

5.3.7 Neurodegenerative Diseases

5.3.7.1 Epidemiologic Studies of Neurodegenerative Diseases in 

Adults

14 The 2006 Pb AQCD described several studies examining associations of blood and bone 

15 Pb levels with neurodegenerative diseases such as Alzheimer’s disease and dementia. 

16 Among NAS men, higher bone Pb levels were associated with lower MMSE scores

17 (Weisskopf et al., 2004; Wright et al., 2003b), which indirectly pointed to a potential 

18 association with dementia, given that the MMSE is widely used as a screening tool for 

19 dementia. Overall, studies had sufficient limitations (e.g., indirect assessment of 

20 dementia, comparison of Alzheimer's Disease patients and healthy controls, lack of Pb 

21 biomarker data), and findings were inconclusive (U.S. EPA, 2006b). New studies on 

22 dementia are not available to assess further the associations with Pb biomarkers. 

23 Similarly, new studies examining Alzheimer’s disease are not available, and as in 2006, 

24 the evidence is still inconclusive regarding the association with Pb biomarkers. In 

25 contrast, there has been additional investigation of ALS, Parkinson’s disease (PD), and 

26 essential tremor, which is described below.

Amyotrophic Lateral Sclerosis

27 Most studies of the association between Pb and ALS have relied on indirect methods of 

28 assessing Pb exposure and overall, have produced inconsistent results. Case-control 
February 2012 5-164 Draft – Do Not Cite or Quote

1 studies that measured blood Pb levels produced contrasting results. A study of 16 ALS 

2 cases (mean blood Pb level: 12.7 μg/dL) and 39 controls (mean blood Pb level: 

3 10.8 μg/dL) found a small difference in the mean concurrent blood Pb level (Vinceti et 

4 al., 1997). Another case-control study that examined concurrent blood and bone Pb levels 

5 in a New England-area population found higher odds of ALS among subjects with 

6 concurrent blood Pb levels ≥ 3 μg/dL (e.g., OR: 14.3 [95% CI: 3.0, 69.3] for blood Pb 

7 levels 3-4 μg/dL) (Kamel et al., 2002). In analyses of tibia or patella Pb tertiles, subjects 

8 in the highest two tertiles (≥ 10 μg/g patella Pb and ≥ 8 μg/g tibia Pb) had higher ALS 

9 prevalence. For example, the ORs (95% CIs) for patella Pb levels 10-20 μg/g and tibia Pb 

10 levels 8-14 μg/g were 2.1 (0.6, 7.4) and 1.6 (0.5, 5.6), respectively. Also in this 

11 population, an estimate of cumulative Pb exposure based on occupational history was 

12 found to be associated with ALS (Kamel et al., 2002). The stronger findings for blood Pb 

13 level were surprising given that bone Pb level is a better biomarker of cumulative Pb 

14 exposure. One explanation for these findings is that the association could be the result of 

15 reverse causality since the half-life of blood Pb is only about 30 days, and blood was 

16 collected from people who already had ALS. If, for example, reduced physical activity 

17 among those with ALS led to more bone turnover, then more Pb would be released from 

18 bones into circulation leading to elevations in blood Pb levels among cases as a result of 

19 effects of the disease. 

20 Since the 2006 Pb AQCD, a few additional studies have been conducted with the same 

21 New England-area case-control study population. Kamel et al. (2005) reported that the 

22 association between blood Pb level and ALS was not modified by the ALAD genotype

23 (Kamel et al., 2005). Another report examined survival of ALS among 100 of the original 

24 110 ALS cases (Kamel et al., 2008). Higher tibia Pb levels were associated with longer 

25 survival time. Findings were similar for patella and blood Pb levels, although they were 

26 associated with smaller increases in survival time. These paradoxical findings raise the 

27 concern that in a case-control study of ALS, the association between bone Pb levels and 

28 ALS may be biased because the case group may comprise more individuals with longer 

29 survival time. Consequently, their bone Pb levels may be higher because they reflect a 

30 longer period of cumulative exposure. On the other hand, the decreased mobility due to 

31 the disease itself would tend to increase bone resorption and lower bone Pb levels over 

32 time. This process might mitigate this effect for bone Pb but would tend to increase blood 

33 Pb levels among cases. However, this was not observed in the one study that had bone 

34 and blood Pb biomarkers (Kamel et al., 2002). Because the strongest findings for survival 

35 were found for tibia Pb, it is unlikely that the findings were biased due to increased 

36 survival of cases. 

37 Another case-control study examined concurrent blood Pb levels and ALS among 184 

38 cases (33 were either progressive muscular atrophy or primary lateral sclerosis, mean 
February 2012 5-165 Draft – Do Not Cite or Quote

1 blood Pb level: 2.41 μg/dL) and 194 controls (mean blood Pb level: 1.76 μg/dL) (Fang et 

2 al., 2010). The cases were recruited from the National Registry of U.S. Veterans with 

3 ALS, and controls were recruited from among U.S. Veterans without ALS and frequency 

4 matched by age, gender, race, and past use of the Veterans Administration system for 

5 health care. A doubling of blood Pb levels was associated with ALS with an OR (95% 

6 CI) of 2.6 (1.9, 3.7). Associations did not differ substantially by indicators of bone 

7 turnover but were slightly higher among ALAD 1-1 carriers. The association with blood 

8 Pb level was similar in analyses that excluded the progressive muscular atrophy and 

9 primary lateral sclerosis cases. The similar results by degree of bone turnover suggest that 

10 reverse causation is not likely driving the association between blood Pb level and ALS. 

11 Whether other types of reverse causality are contributing, however, cannot be ruled out. 

12 This study did not have measures of bone Pb and therefore could not assess the 

13 association with biomarkers of cumulative Pb exposure.

14 In summary, studies have found associations of blood and bone Pb levels with ALS but in 

15 relatively few different cohorts. The case-control design of most studies and issues of 

16 reverse causality and bias due to survival time make it difficult to draw firm conclusions. 

17 Studies in additional cohorts using designs other than case-control comparisons are 

18 needed to address the limitations of the available studies and characterize better the 

19 potential effects of Pb on ALS.

Parkinson’s Disease

20 A few previous studies, some ecological (Rybicki et al., 1993; Aquilonius and Hartvig, 

21 1986) and some case-control relying on questionnaire data or occupational history

22 (Gulson et al., 1999; Gorell et al., 1997; Tanner et al., 1989) indicated associations 

23 between exposure to heavy metals, particularly Pb, and risk of PD. Available evidence 

24 was limited and far from conclusive. A recent large case-control study (330 cases, 308 

25 controls) recently reported on associations between biomarkers of Pb and PD in a 

26 population with virtually no occupational exposures to Pb (Weisskopf et al., 2010).

27 Subjects in the highest quartile of tibia Pb level (>16.0 μg/g) had higher odds of PD 

28 compared to those in the lowest quartile (≤ 5 μg/g) (OR: 1.91 [95% CI: 1.01, 3.60]). In 

29 this study, cases and controls were recruited from several different sources including 

30 movement disorder clinics and community-based cohorts, which could have introduced 

31 some biases. However, when analyses were restricted to cases recruited from movement 

32 disorder clinics and to their spouse, in-law, or friend as controls, the results were even 

33 stronger (OR: 3.21 [95% CI: 1.17, 8.83]). Although the use of spouse, in-law, and friend 

34 controls can introduce bias, this is expected to be toward the null as these groups are 

35 likely to share many exposures.
February 2012 5-166 Draft – Do Not Cite or Quote

1 Manganese exposure has been associated with Parkinsonian symptoms and could 

2 potentially confound associations between Pb and PD. Weisskopf et al. (2010) did not 

3 adjust for Mn exposure in analyses. However, unlike a setting of occupational exposure 

4 to Pb, general environment exposure to Pb is much less likely to be correlated with 

5 environmental Mn exposure. Thus, the likelihood of any associations with Pb being 

6 confounded by co-occurring Mn exposure is less likely. 

7 Coon et al. (2006) conducted a smaller case-control study of 121 PD patients and 414 

8 age-, sex-, and race-, frequency-matched controls all receiving health care services from 

9 the Henry Ford Health System. Subjects in the highest quartile of both tibia (OR: 1.62 

10 [95% CI: 0.83, 3.17] for levels ≥ 15 μg/g) and calcaneus (OR: 1.50 [95% CI: 0.75, 3.00] 

11 for levels ≥ 25.29 μg/g) bone Pb levels had higher odds of PD compared to those in the 

12 lowest quartiles (0-5.91 μg/g for tibia and 0-11.70 μg/g for calcaneus). The highest OR 

13 for PD was estimated for subjects in the highest quartile of whole-body lifetime exposure 

14 to Pb (≥ 80.81 μg/g) estimated using PBPK modeling), compared to the lowest quartile of 

15 exposure (0-40.04 μg/g) (OR: 2.27 [95% CI:1.13, 4.55] for levels ≥ 80.81 μg/g). These 

16 analyses did not adjust for Mn either; however, in this study it was not clear what the 

17 extent of occupational exposure to Pb was among the participants. Thus, it is uncertain 

18 whether the observed associations were specifically related to Pb exposure or could have 

19 resulted from co-occurring Mn exposure.

20 In summary, a small number of recent studies expand on previous evidence by finding 

21 associations of bone Pb levels, biomarkers of cumulative Pb exposure, with PD in adults. 

22 Nonetheless, additional investigation is warranted to establish the temporality between Pb 

23 exposure and development of PD and to assess potential confounding by Mn exposure.

Essential Tremor

24 In a relatively small body of literature, concurrent blood Pb levels have been consistently 

25 associated with essential tremor, although studies have had relatively small sample sizes 

26 and have produced imprecise effect estimates. The 2006 Pb AQCD described case27 control studies that found associations between concurrent blood Pb levels and essential 

28 tremor in New York City metropolitan area populations (Louis et al., 2005; Louis et al., 

29 2003). In Louis et al. (2005), the magnitude of association was larger among carriers of 

30 an ALAD2 allele than among adults with only ALAD1 alleles.

31 Since 2006, Dogu et al. (2007) reported on a case-control study of 105 essential tremor

32 cases from a movement disorder clinic in Turkey and 105 controls (69 spouses and 36 

33 other relatives living in the same district). After adjusting for age, sex, education, 

34 cigarette smoking, cigarette pack-years, and alcohol use, a 1 μg/dL higher blood Pb level 
February 2012 5-167 Draft – Do Not Cite or Quote

1 (measured at the time of study recruitment) was associated with essential tremor with an 

2 OR (95% CI) of 4.19 (2.59, 6.78). This OR was much larger than that obtained in the 

3 New York area study (OR: 1.19 [95% CI: 1.03, 1.37]) (Louis et al., 2003). The 

4 magnitude of association in Dogu et al. (2007) is even more striking because so many of 

5 the controls were spouses who are expected to share many environmental exposures as 

6 cases. Most of the essential tremor cases were retired at the time of the study, but past 

7 occupational history was not reported. Occupational Pb exposures were less likely in the 

8 New York area population.

9 In this small body of studies, associations were observed between blood Pb level and 

10 essential tremor in adults. However, due to the case-control design of studies, temporality 

11 between exposure and development of essential tremor cannot be established. Further, the 

12 level, timing, frequency, and duration of Pb exposure associated with PD is uncertain as 

13 all studies examined blood Pb level at the time of study recruitment. Occupational 

14 histories were not reported in these studies. Thus, it is not clear what past exposure to Mn 

15 may have contributed to the associations observed with blood Pb levels. 

5.3.7.2 Toxicological Studies of Neurodegenerative Disease

16 Although epidemiologic studies have provided weak evidence for associations of blood 

17 Pb level and ALS in adults, toxicological studies have found that Pb exposure induces 

18 neurophysiologic changes consistent with ALS. For example, chronic Pb exposure 

19 (Pb-acetate in drinking water at 200 ppm from weaning onward, resulting blood Pb level: 

20 27 μg/dL) reduced astrocyte reactivity and induced increased survival time in the 

21 superoxide dismutase transgenic (SOD1 Tg) mouse model of severe ALS (Barbeito et al., 

22 2010). In this model, Pb exposure did not significantly increase the onset of the ALS 

23 disease but increased survival time in SOD1 Tg mice (Barbeito et al., 2010). This finding 

24 does provide biological plausibility for the association observed between blood Pb level 

25 and longer survival time in patients diagnosed with ALS (Kamel et al., 2008). In mice, 

26 astrocyte of vascular endothelial growth factor (VEGF) also was examined to understand 

27 its possible contribution to Pb effects on increasing survival time in ALS models. 

28 Baseline levels VEGF were elevated in astrocytes from the ventral spinal cord of 

29 untreated SOD1 Tg mice versus untreated nontransgenic animals. VEGF was not induced 

30 in the astrocytes of Pb-treated nontransgenic mice. Pb-exposed SOD1 Tg mice had 

31 significant elevations of astrocyte VEGF versus vehicle-treated SOD1 Tg animals

32 (Barbeito et al., 2010). Nontransgenic animals exposed to Pb showed no elevation in 

33 VEGF expression above that in nontransgenic vehicle-treated animals (Barbeito et al., 

34 2010). Other research has suggested that ALS initiation is dependent on motor neuron 

35 function and ALS progression is dependent on astrocyte and microglia function
February 2012 5-168 Draft – Do Not Cite or Quote

1 (Yamanaka et al., 2008; Boillee et al., 2006). Thus, the aforementioned findings for Pb2 induced effects on astrocytes provide a mechanistic explanation for Pb effects on ALS 

3 progression in animals.

4 Others have reported that VEGF administration to the SOD1 Tg mice significantly 

5 reduced glial reactivity, a marker or neuroinflammation (Zheng et al., 2007). Using a 

6 cell-based co-culture system of neurons and astrocytes, Barbeito et al. (2010) found that 

7 an up-regulation of VEGF production by astrocytes in the Pb-exposed SOD1 Tg mice 

8 was protective against motor neuron death in the SOD1 Tg cells (Barbeito et al., 2010).

9 Chronic Pb exposure in a mouse model of ALS was associated with increased survival 

10 time and was correlated with higher spinal cord VEGF levels, making astrocytes less 

11 cytotoxic to surrounding motor neurons (Barbeito et al., 2010). Also, in another study the 

12 metal chelators DP-109 and DP-460 were neuroprotective in the ALS mouse model or 

13 Tg(SOD1-G93A) (Petri et al., 2007).

14 Improper activation of microglia and release of inflammatory cytokines and metabolites 

15 can contribute to neurodegeneration (Zhang et al., 2010b; Qian and Flood, 2008). These 

16 two cell types are known to accumulate or sequester Pb in the nervous system. 

17 Researchers have implicated dysfunctional astrocytes as playing an important role in the 

18 chain of misregulated inflammation leading to neurodegenerative conditions (Barbeito et 

19 al., 2010; De Keyser et al., 2008).

Cell Death Pathways

20 Earlier work has documented that Pb exposure can induce cell death or apoptosis in 

21 various models including rat brain (Tavakoli-Nezhad et al., 2001), retinal rod cells (He et 

22 al., 2003; He et al., 2000), cerebellar neurons (Oberto et al., 1996), and PC12 cells

23 (Sharifi and Mousavi, 2008). These observations indicate that Pb-induced cell apoptosis 

24 may mediate its neurodegenerative effects. A recent study reported that chronic (40 days) 

25 Pb exposure induced hippocampal apoptosis in young (exposure starting at 2-4 weeks of 

26 age) and adult (exposure starting at 12-14 weeks of age) male rats exposed to 500 ppm Pb 

27 by drinking water (resulting in blood Pb levels of 98 μg/dL); apoptosis was verified by 

28 light and electron microscopy, and increased pro-apoptotic Bax protein levels (Sharifi et 

29 al., 2010). Another study followed the developmental profile of changes in various 

30 apoptotic factors in specific brain regions of animals exposed to Pb-acetate (0.2% dam 

31 drinking water) during lactation. Male offspring blood Pb level at the end of lactation or 

32 PND20 was 80 μg/dL. The data showed that hippocampal mRNA for various apoptotic 

33 factors including caspase-3, Bcl-x and Brain-derived neurotrophic factor (BDNF) was 

34 significantly upregulated on PND12, PND15 and PND20. The cortex of these male pups 

35 also showed upregulation of Bcl-x and BDNF on PND 15 and PND20 (Chao et al., 
February 2012 5-169 Draft – Do Not Cite or Quote

1 2007). The cerebellum did not have elevated apoptotic mRNA levels in this model. This 

2 study showed temporal and regional changes in activation of death protein message levels 

3 in male offspring.

4 Pb exposure has also been shown to induce apoptosis during spinal cord development in 

5 chicks exposed to 150 or 450 μg in ovo Pb-acetate exposure at embryonic day 3 or 5 and 

6 visualized six days later; TUNEL positive cells were at significantly higher levels in Pb7 exposed animals and were visualized in all layers of the developing spinal cords (Müller 

8 et al., In Press). Also, levels of glial fibrillary acidic protein (GFAP), a factor important in 

9 neuronal migration and cellular differentiation during nervous system development, was 

10 significantly attenuated in spinal cords of Pb-exposed chicks. Another recent study 

11 examined Pb treatment animals (i.p. injections of 350 mg/kg, twice daily, blood Pb levels 

12 at PND14 of 0.15 μg/dL in controls and 8.10 μg/dL in Pb-treated animals) during 

13 PND7-14, a known period of sensitivity to NMDA antagonist-dependent neuronal 

14 apoptosis (Ikonomidou et al., 1999). Liu et al. (2010b) examined apoptotic effects in 30 

15 day-old male rats that were exposed to Pb-acetate 1x/d for 6 weeks via intragastric 

16 infusion. Four dose groups: control, low (2), medium (20), and high (200 mg/kg BW) had 

17 blood Pb levels of 1.0 to 7.5 μg/dL; 4.5 to 11 μg/dL; 9 to 42 μg/dL; and 48 to 73 μg/dL, 

18 respectively. Liu et al. (2010b) reported Pb-induced apoptosis in the brain with 

19 hippocampal XIAP (statistically significant at high dose only) and Smac (statistically 

20 nonsignificant trend) downregulation and associated histopathology showing 

21 hippocampal neuronal apoptosis (TUNEL positive staining, significant at all doses) at the 

22 termination of the 6 week treatment. In another study, Pb exposure (500 ppm Pb-acetate

23 in drinking water for 8 weeks) of adult male rats induced regional-specific changes in 

24 brain apoptotic proteins poly(ADP-ribose) polymerase, Bcl-2, caspase-3) with a greater

25 effect observed in the hippocampus and cerebellum and a lesser effect observed in the 

26 brainstem and the frontal cortex (Kiran Kumar et al., 2009)

27 Collectively, studies have shown that Pb exposure induces neuronal apoptosis in animals 

28 during various developmental windows: early postnatal and adulthood. The new data 

29 continue to show that Pb exposure induced apoptosis in brains of animals. 

Lead-Induced Neuronal Plaque Formation

30 Epidemiologic studies have not provided compelling evidence that Pb exposure is 

31 associated with Alzheimer's Disease in adults (Section 5.3.7.1); however, Pb exposure in 

32 early life has been shown to promote Alzheimer’s Disease-like pathologies in the brains 

33 of aged adult animals. Alzheimer’s disease is characterized by amyloid-beta peptide (Ab) 

34 accumulation, hyper-phosphorylation of the tau protein, neuronal death and synaptic loss. 

35 In the last decade, the developmental origins of adult health and disease (DoHAD) 
February 2012 5-170 Draft – Do Not Cite or Quote

1 paradigm and the similar Barker hypothesis have indicated that early life exposures can 

2 result in aberrant adult outcomes. Bolin et al. (2006) demonstrated the connection 

3 between developmental exposure to Pb in the rat with early life programming and the 

4 resulting inflammation-associated DNA damage with neurodegenerative loss in the adult 

5 brain. Wu and colleagues (2008a) had similar findings in a study using infantile exposure 

6 to Pb in monkeys. The investigations reinforce the need to directly examine the long-term 

7 effects of developmental exposure to toxicants rather than relying on adult exposure 

8 alone to predict probable health risks from prenatal, neonatal or juvenile exposure

9 (Dietert and Piepenbrink, 2006). Mechanistically, some of these pathologies have been 

10 associated with changes in the epigenome. 

11 The fetal basis of amyloidogenesis has been examined extensively by the Zawia 

12 laboratory in both rodents and nonhuman primates. Mechanistically, amyloid plaques 

13 originate from the cleavage of the amyloid precursor protein (APP) to Ab, which 

14 comprises the plaque. In rodents exposed to Pb as neonates or as adults, neonatal Pb 

15 exposure induced amyloidogenesis in the aged animal brains; adult exposure to Pb did 

16 not contribute to plaque formation. Basha et al. (2005) exposed male rodents neonatally 

17 via lactation to Pb (PND1-PND20 exposure, dam drinking water Pb-acetate 200 ppm, 

18 resulting in pup PND20 blood Pb level of 46 μg/dL and cortex 0.41 μg/g wet weight of 

19 tissue) and examined cortical APP gene expression over the lifetime. A bimodal response 

20 was observed, with a significant increase in APP expression above that in control animals 

21 first manifesting neonatally and second manifesting in old age (82 weeks of age) (Basha 

22 et al., 2005). A concomitant bimodal response was observed in specificity protein 1 

23 (Sp1), a transcription factor known to be related to APP expression. Ab, the amyloid 

24 plaque constituent, was also significantly elevated in these aged animals developmentally 

25 exposed to Pb. A subset of rodents exposed to Pb only as aged adults (18-20 weeks of 

26 age) was unresponsive in APP or Sp1 expression or Ab production after Pb exposure, 

27 indicating the developmental lifestage and not adult lifestage as the susceptible period for 

28 Pb-induced amyloidogenesis. The Zawia lab (Wu et al., 2008a) produced similar findings 

29 for amyloid plaques in the brains of monkeys that were exposed to Pb as infants 

30 (PND1-PND400), i.e., significantly higher gene expression of APP, and Sp1 and 

31 significantly higher protein expression of APP and Ab in aged female monkey cortex 

32 tissue (23 year-old Macaca fascicularis) from a cohort of animals established in the 

33 1980s by Rice (1992, 1990). After weaning but with continued Pb exposure, the monkeys 

34 had blood Pb levels of 19-26 μg/dL. However, in old age when amyloid plaques had 

35 manifested, blood Pb levels and brain cortex Pb levels had returned to control or baseline 

36 levels. Together, the rodent and nonhuman primate toxicological studies concur and show 

37 that developmental Pb exposure induced elevations in neuronal plaque proteins in aged 

38 animals. 
February 2012 5-171 Draft – Do Not Cite or Quote

1 Mechanistic understanding of Ab production and elimination after Pb exposure was 

2 explored in human SH-SY5Y neuroblastoma cells exposed to Pb concentrations of 0, 5, 

3 10, 20, and 50 μM for 48 hours. These studies showed that Pb affected two separate 

4 pathways to contribute to elevated Ab. Pb exposure induced both the overexpression of 

5 APP and repression of neprilysin, a rate-limiting enzyme involved in Ab metabolism or 

6 removal (Huang et al., 2011a). Further mechanistic understanding of how Ab peptide 

7 formation is affected by Pb exposure was examined by Behl et al. (2009). The choroid 

8 plexus is capable of removing beta-amyloid peptides from the brain extracellular matrix; 

9 however, Pb was shown to impair this function, possibly via the metalloendopeptidase, 

10 insulin-degrading enzyme (IDE), which metabolizes Ab (Behl et al., 2009). In another 

11 study, the effect of Pb on transcription factors essential in the regulation of the 

12 developing brain was explored. Pb exposure has been shown to perturb DNA binding of 

13 transcription factors including SP1 at essential sites like zinc finger proteins. In Long14 Evans hooded rat pups exposed to Pb during lactation, these Pb-induced developmental 

15 perturbations of SP1 DNA binding were found to be ameliorated by exogenous zinc 

16 supplementation (Basha et al., 2003).

17 An additional study with developmental Pb exposure (gestational plus lactational, dam 

18 drinking water solutions of 0.1%, 0.5% or 1%, blood Pb level 40, 80 and 1,00 μg/dL) 

19 showed that the hippocampus contained neurofibrillary changes as early as PND21. 

20 These changes manifested with Tau hyper-phosphorylation, and increased tau and beta 

21 amyloid hippocampal protein levels in Pb-exposed offspring (Li et al., 2010b).

22 In summary, the multiple recent studies showed that developmental Pb exposure induced 

23 significant increases in neuronal plaque associated proteins, which is the pathology found 

24 in humans with Alzheimer’s disease. Adult exposure to Pb did not generate this 

25 neurofibrillary pathology, further demonstrating that early life Pb is a sensitive window 

26 for Pb-induced pathology including Ab-peptide accumulation, activation of Ab27 supporting transcription factors, as well as tau hyperphosphorylation. Epidemiologic 

28 evidence for Pb-associated Alzhemier's disease is weak, and although several study 

29 design limitations were noted (Section 5.3.7.1), the animal evidence indicates that 

30 epidemiologic studies assessing concurrent bone or brain Pb levels or occupational Pb 

31 exposure may not have examined the etiologically relevant exposure period. Notably, 

32 animals were not behaviorally assessed for dementia.
February 2012 5-172 Draft – Do Not Cite or Quote

5.3.8 Modes of Action for Lead Nervous System Effects

5.3.8.1 Effects on Brain Physiology and Activity

1 A small body of available epidemiologic evidence with small sample sizes demonstrated 

2 associations of Pb biomarkers with electrophysiologic changes in the brains of young 

3 adults as assessed by magnetic resonance imaging (MRI) or spectroscopy (MRS) (Yuan 

4 et al., 2006; Cecil et al., 2005; Meng et al., 2005; Trope et al., 2001). By characterizing 

5 underlying mechanisms by which Pb exposure may disrupt brain function, these studies 

6 have provided biologically plausible evidence for the effects of Pb exposure on cognitive 

7 and psychological and behavioral function observed in adults. 

8 Findings such as lower levels of N-acetylaspartate (NAA), creatine (Cr), and choline are 

9 linked to decreased neuronal density and alteration in myelin. Notably, Trope et al.

10 (2001) and Meng et al. (2005) reported that all subjects had normal MRIs with no 

11 evidence of structural abnormalities. Thus, the clinical significance of the observed 

12 physiological changes was unclear. Additionally, these studies compared subjects with 

13 relatively high childhood blood levels (23-65 μg/dL) to those with childhood blood Pb 

14 levels <10 μg/dL. Therefore, it is unclear whether physiological changes would be 

15 observed in association with lower blood Pb levels. Cecil et al. (2005) and Yuan et al.

16 (2006) conducted functional MRI in 42 adult (ages 20-23 years) participants from the 

17 CLS cohort during a verb generation language task and found that childhood average 

18 blood Pb level was associated with decreased activation in the left frontal gyrus and left 

19 middle temporal gyrus, regions traditionally associated with semantic language function. 

20 Although these findings were in adults, they were consistent with findings in the same 

21 cohort of subjects that indicated associations of blood Pb level with other indices of 

22 language skills in childhood.

23 Since the 2006 Pb AQCD, studies examining MRI data were largely limited to CLS 

24 cohort participants as adults (ages 19-24), and recent results continue to support 

25 associations of childhood blood Pb levels with physiological changes in the brain of 

26 adults. These recent studies expanded on previous studies by including larger sample 

27 sizes, aiming to characterize important lifestages of Pb exposures, and evaluating 

28 potential links between changes in brain activity and functional neurodevelopmental 

29 deficits. Whereas previous CLS analyses focused on activity in specific regions of the 

30 brain, Cecil et al. (2011) examined brain metabolites in 159 subjects and found that 

31 childhood average blood Pb levels were associated with lower levels of NAA and Cr in 

32 the basal ganglia and lower levels of choline in white matter. These results were adjusted 

33 for age and FSIQ. A recent analysis of 31 men in the NAS cohort similarly reported an 
February 2012 5-173 Draft – Do Not Cite or Quote

1 association between biomarkers of cumulative, long-term Pb exposure and changes in 

2 brain metabolites in older adults. Weisskopf et al. (2007a) found higher tibia and patella 

3 Pb levels to be associated with a higher myoinositol/Cr ratio, which may be indicative of 

4 glial activation and is a signal reportedly found in the early stages of HIV-related 

5 dementia and Alzheimer’s disease.

6 Other studies in CLS young adults found that childhood average blood Pb levels were 

7 associated with altered brain architecture. Brubaker et al. (2009) reported associations of 

8 childhood average blood Pb levels with diffusion parameters that were indicative of less 

9 organization of fibers throughout white matter. Results were adjusted for potential 

10 confounding by variables such as maternal IQ, prenatal alcohol and tobacco exposure, 

11 and adult marijuana use. In regions of the corona radiata, higher blood Pb levels were 

12 associated with less myelination axonal integrity. In regions of the corpus callosum, 

13 higher blood Pb levels were associated with greater myelination and axonal integrity. The 

14 differential impact on different neural elements may be related to the stage of myelination 

15 development at various periods of exposure.

16 Another study of 157 CLS participants provided evidence of region-specific reductions in 

17 adult gray matter volume in association with childhood blood Pb levels (Cecil et al., 

18 2008). The most affected regions included frontal gray matter, specifically the anterior 

19 cingulate cortex and the ventrolateral prefrontal cortex (i.e., areas traditionally related to 

20 executive functions, mood regulation, and decision-making). Further, investigators found 

21 that fine motor factor scores positively correlated with gray matter volume in the 

22 cerebellar hemispheres; adding blood Pb level as a variable to the model attenuated this 

23 correlation. These findings suggested that MRI changes associated with blood Pb levels 

24 may be indicative of decrements in cognitive function. The functional relevance of these 

25 structural changes in the brain also is supported by observations from other studies that 

26 link changes in brain architecture and activity with changes in cognitive function 

27 (e.g., visuoconstruction, visual memory, eye-hand coordination) (Schwartz et al., 2007)

28 and behavior (impulsivity, aggression, violence) (Yang et al., 2005; Raine et al., 2000).

29 In a subsequent comparison of blood Pb levels measured at various lifestages, Brubaker 

30 et al. (2010) found that blood Pb levels at older ages were associated with greater losses 

31 in gray matter volume than were childhood average or maximum blood Pb levels. Both 

32 Cecil et al. (2008) and Brubaker et al. (2010) found that Pb-associated reductions in gray 

33 matter were more pronounced in CLS males than females.

34 Studies of Pb-workers also found associations of blood and bone Pb levels with changes 

35 in brain structure and physiology, adding support for the effects of chronic Pb exposure. 

36 Pb-associated changes included white matter lesions, smaller brain volumes, less total 

37 gray matter, and lower levels of brain metabolites such as NAA and Cr (Hsieh et al., 
February 2012 5-174 Draft – Do Not Cite or Quote

1 2009b; Jiang et al., 2008; Bleecker et al., 2007a; Stewart et al., 2006). In a few of these 

2 occupational groups, Pb-associated brain changes were linked to poorer performance in 

3 cognitive function tests (Caffo et al., 2008; Bleecker et al., 2007a).

4 In summary, results in a few different populations indicated associations of blood or bone 

5 Pb levels with changes in brain structure and physiology as assessed by MRI or MRS. A 

6 majority of evidence was derived from the CLS study. Several studies linked these 

7 changes to functional changes in cognitive performance or motor skills. While the 

8 collective findings pointed to the effects of long-term or past Pb exposures, it is important 

9 to recognize that other lifestages of exposure, including recent adult, were not widely 

10 examined.

5.3.8.2 Oxidative Stress

11 Pb has been shown to induce oxidative stress in multiple animal models, and this 

12 oxidative stress can contribute to DNA damage, which can be measured with the adduct 

13 8-hydroxy-2’-deoxyguanosine (8-oxo-dG). The contribution of reactive oxygen or 

14 nitrogen species to these Pb induced changes was assayed by examining the ratio of 

15 8-oxo-dG to 2-deoxyguanosine (2-dG). 2-dG is product of oxidative cleavage that can 

16 generate 8-oxo-dG from a parent compound forming a DNA adduct during conditions of 

17 nitrosative or oxidative stress. The 8-oxo-dG to 2-dG ratio data from rodent male 

18 offspring were similar to the amyloid data with significant biphasic elevations in 

19 developmentally Pb-exposed animals (0.2% Pb-acetate in dam drinking water from 

20 PND1-20) versus control, nonPb-exposed animals at early (PND5) and late life time 

21 points (80 weeks of age) (Bolin et al., 2006). Activity of the base-excision DNA repair 

22 enzyme oxoguanine glycosylase or Ogg1 was unaffected by Pb exposure (Bolin et al., 

23 2006). Interestingly, similar findings were reported in a monkey study. The ratio of 

24 8-oxo-dG to 2-dG in the brains of aged monkeys (23 years) after being exposed to Pb as 

25 infants, was significantly elevated above that in controls (Wu et al., 2008a). Several lines 

26 of evidence indicate that oxidative stress is involved in neurodegenerative pathologies 

27 including Alzheimer's disease; hydrogen peroxide-induced oxidative stress has been

28 shown to induce intracellular accumulation of amyloid beta-protein (Abeta) in human 

29 neuroblastoma cells (Misonou et al., 2000). Similar to the amyloid findings in Pb30 exposed animals the oxidative stress markers showed no significant changes above 

31 baseline when animals were exposed to Pb only as aged adults (Wu et al., 2008a; Bolin et 

32 al., 2006). Thus, the data for biomarkers of oxidative stress concur with the 

33 amyloidogenesis data with both demonstrating kinetically similar biphasic significant 

34 elevations in markers of oxidative stress and amyloidogenesis with early life Pb exposure 

35 and an absence of effect with adult only exposure.
February 2012 5-175 Draft – Do Not Cite or Quote

1 Because the brain has the highest energy demand and metabolism of any organ, energy 

2 homeostasis is of utmost importance. Pb has been shown to inhibit various enzymes 

3 involved in energy production or glucose metabolism including glyceraldehydes-3 

4 phosphate dehydrogenase, hexokinase, pyruvate kinase, and succinate dehydrogenase

5 (Verma et al., 2005; Yun and Hoyer, 2000; Regunathan and Sundaresan, 1984; Sterling et 

6 al., 1982). Mitochondria produce ATP or energy through oxidative phosphorylation. 

7 Aberrant mitochondrial function can decrease the energy pool and contribute to ROS 

8 formation via electron transport chain disruption. ATP depletion can also affect synaptic 

9 and extracellular neurotransmission. The mitochondrial Na/K ATPase is important in 

10 maintaining the inner mitochondrial membrane potential (delta omega sub m) and the 

11 functioning of the mitochondria.

12 To characterize the effect of Pb exposure on these mitochondrial parameters, brains from 

13 the offspring of Pb-exposed mice were collected at PND8 (Baranowska-Bosiacka et al., 

14 2011b). Cerebellar granular cells were harvested from PND 8 control and Pb-exposed 

15 animals (0.1% Pb-acetate in dam drinking water, resulting in blood Pb levels of 4 μg/dL 

16 and cerebella Pb levels of 7.2 μg/g dry weight). These neuronal cells were cultured for 5 

17 days in vitro, at which point various mitochondrial parameters were measured. With Pb 

18 exposure, reactive oxygen species were significantly increased in both the cortical 

19 granule cells and in the mitochondria. Intracellular ATP concentration and adenylate 

20 energy charge values were significantly decreased in cells of Pb-exposed mice versus 

21 controls. Neuronal Na/K ATPase activity was significantly lower in cortical granule cells 

22 from Pb-exposed mice versus cells from controls. Mitochondrial mass was unaffected 

23 with Pb treatment, but mitochondrial membrane potential was significantly decreased 

24 with Pb exposure. Pb-exposed crayfish that were placed under hypoxic conditions 

25 adapted to the situation by decreasing their metabolism (Morris et al., 2005) and showed 

26 mitochondrial changes consistent with those observed in granule cells (Baranowska27 Bosiacka et al., 2011b). These studies showed impaired mitochondrial function and 

28 energy production in neuronal cells from mice with gestational and lactational Pb 

29 exposure with concomitant increases in mitochondrial and cellular ROS production. 

30 The effects of co-administration of flaxseed oil (FSO) with Pb-acetate on oxidative stress 

31 and neurotoxicity were examined in adult male Wistar albino rats (Abdel Moneim et al.).

32 Animals were administered Pb-acetate i.p. for 5 days (20mg/kg, resulting in blood Pb 

33 level ~31 μg/dL the day after the last Pb injection) or FSO (oral gavage 1000mg/kg body 

34 weight for 5 days, 1 hour prior to Pb dosing, resulting in blood Pb level ~12 μg/dL). 

35 Administration of the polyunsaturated fatty acid FSO significantly attenuated the blood 

36 Pb level of Pb-exposed animals and control animals, indicating that FSO may alter Pb 

37 toxicokinetics in animals. As would be expected with a lower blood Pb level, FSO+Pb 

38 exposed animals had significant attenuations in Pb-induced histological neuronal damage 
February 2012 5-176 Draft – Do Not Cite or Quote

1 and brain tissue DNA fragmentation. Pb exposure effects on indicators of oxidative stress 

2 included decreased antioxidant GSH levels, elevated lipid peroxidation, and decreased 

3 antioxidant enzyme activity in brain tissue. These Pb-induced effects were attenuated in 

4 FSO+Pb exposed animals. These results indicated that FSO likely attenuates Pb-related 

5 neurotoxicological damage and oxidative stress via its action on altering Pb 

6 toxicokinetics. 

7 Green tea extract (GTE) was shown to be protective against Pb-induced oxidative stress 

8 in the brains of adult male rats (1.5% GTE +/- 0.4% Pb-acetate for 6 weeks) (Hamed et 

9 al., 2010). GTE+Pb-exposed animals had significantly less brain Pb than did Pb-exposed 

10 animals (1.2 versus 1.9 ppm). Levels of pro-oxidants or antioxidants including lipid 

11 peroxides (LPO), nitric oxides (NO), total antioxidant capacity (TAC), glutathione 

12 (GSH), glutathione-S-transferase (GST), and superoxide dismutase (SOD) were 

13 measured in brain tissue homogenate 24 hours after the termination of Pb exposure. 

14 Comparing the Pb-exposed animals to controls, brain LPO was significantly elevated and 

15 brain GSH, GST and NO were significantly decreased. GTE and Pb co-exposure 

16 significantly attenuated the Pb-related changes. This study reported a positive correlation 

17 between whole blood Pb levels and brain tissue LPO levels and a negative correlation 

18 between whole blood Pb and NO levels. The antioxidant quercetin (Que) rescued chronic 

19 Pb exposure-related (0.2% Pb-acetate in drinking water from birth to PND67, 30 mg/kg 

20 BW Que for one week from PND60-67) impaired synaptic plasticity in adult male and 

21 female Wistar rat dentate gyrus (DG) (Hu et al., 2008a). Pb-related impaired long-term 

22 potentiation, paired-pulse reactions, and input/output functions were significantly 

23 attenuated with Que treatment at PND67. Que treated animals had significantly less 

24 hippocampal Pb than did the Pb-exposed animals and this decreased Pb brain burden 

25 likely contributed to the attenuated response observed with Que exposure.

26 The vulnerability of the highly energetic brain tissue to stressors and cell death can be 

27 exacerbated with an energy imbalance. Baranowska-Bosiacka et al. (2011b) showed that 

28 Pb exposure induced energy imbalances in immature rat brains exposed to Pb. Wistar rat 

29 pups (PND15) of both sexes were injected daily for 2 weeks with Pb-acetate (15mg/kg 

30 BW, i.p., resulting in blood Pb levels of 3 and 30 μg/dL, control and Pb-exposed, 

31 respectively) and thereafter sacrificed for analysis of regional brain purines and 

32 purinergic receptors. ATP and ADP were significantly decreased in various brain regions 

33 with Pb exposure, with the cerebellum and hippocampus more strongly affected than the 

34 forebrain cortex. Also, enhanced expression of the proinflammatory P2XR receptor was 

35 observed in the glial fraction, indicating the astrocyte pool may be involved in the 

36 pathological changes found in Pb-exposed immature rat brains. 
February 2012 5-177 Draft – Do Not Cite or Quote

5.3.8.3 Nitrosative Signaling and Nitrosative Stress

1 The nitric oxide system is increasingly being recognized as a signaling system in addition 

2 to its more classical role as a marker of cellular stress. In studies of learning and memory 

3 using the Morris water maze, hippocampal changes in NO were noted in Pb-exposed 

4 animals after completion of the test. Pb exposure has been repeatedly shown to increase 

5 the escape latency in Pb-exposed animals (Section 5.3.2.2). Chetty (2001) initially 

6 reported decreased hippocampal neuronal nitric oxide synthase (NOS) with perinatal Pb 

7 exposure. Namely, with repeated swim tests, control animals more quickly found a 

8 submerged platform (i.e., escaped), than did Pb-exposed animals. After either 4 or 

9 8 weeks of Pb exposure to weanling male rats (resulting in blood Pb level of 6.2 μg/dL), 

10 hippocampal NOS and NO were significantly decreased. Dietary supplementation with 

11 taurine or glycine concomitant with 8 weeks of Pb exposure induced significant increases 

12 in hippocampal NOS, whereas Pb plus dietary supplementation with vitamin C, 

13 methionine, tyrosine, or vitamin B1 decreased hippocampal NOS. Supplementation also 

14 changed Pb-related effects on hippocampal NO. Pb-induced NO increased with taurine 

15 and decreased with vitamin C, tyrosine or glycine. Dietary supplementation with tyrosine,

16 methionine, or ascorbic acid after 4 weeks of Pb exposure in weanling males (4-week 

17 blood Pb level of 47.6 μg/dL and 8-week blood Pb level of 8.1 μg/dL), induced 

18 significant increases in NO. Zinc supplementation in this model had no effect on the NO 

19 system. The investigators concluded various combinations of nutrients significantly 

20 attenuate Pb-related decreases in NO/NOS. Specifically, nutrients prevented (8 weeks Pb 

21 plus concomitant exposure to methionine, zinc, ascorbic acid, and glycine) or restored 

22 (4 weeks Pb exposure followed by 4 weeks nutrient exposure, taurine and thiamine) Pb23 related decrements in NO/NOS concentrations (Fan et al., 2009a).

5.3.8.4 Synaptic Changes

24 Previous toxicological studies point to an effect of developmental Pb exposure on 

25 synapse development, which mechanistically may contribute to multiple Pb-related 

26 aberrant effects, including changes in long-term potentiation (LTP) and facilitation. 

27 Earlier work has shown that developmental Pb exposure results in altered density of 

28 dendritic hippocampal spines (Király and Jones, 1982; Petit and LeBoutillier, 1979),

29 aberrant synapse elimination (Lohmann and Bonhoeffer, 2008), and abnormal long-term 

30 and short-term plasticity (MacDonald et al., 2006). Newer research using the Drosophila

31 larval neuromuscular junction model has shown that compared with unexposed controls, 

32 Pb-exposed larvae had significant increases in intracellular calcium and significant delays 

33 in calcium decays back to baseline levels at the pre-synaptic neuronal bouton (as 

34 stimulated with multiple action potentials, also called AP trains). Pb-exposed larvae had 
February 2012 5-178 Draft – Do Not Cite or Quote

1 reduced activity of the plasma membrane calcium ATPase, which is responsible for 

2 extravasations of calcium from the synaptic terminal (He et al., 2009). Intracellular 

3 calcium in Pb exposed-larvae was no different from that in controls under resting 

4 conditions or in neurons with stimulation by a single action potential. Pb media 

5 concentrations in these experiments were 100 or 250 μM with the low-dose body burden 

6 (100 μM) of Pb calculated to be 13-48 μM per larvae. Facilitation of a neuronal terminal 

7 is defined as the increased capability to transmit an impulse down a nerve due to prior 

8 excitation of the nerve. After stimulation of the axon, facilitation of the excitatory post9 synaptic potential, which is dependent on residual terminal calcium, was significantly 

10 elevated in Pb-exposed larvae versus control (He et al., 2009). The data from this synapse 

11 study demonstrate that developmental Pb exposure affected the plasma membrane 

12 calcium ATPase, induced changes in the intracellular calcium levels during impulse 

13 activation, and produced changes in facilitation of the neuronal networks of Drosophila. 

14 Thus, the neuromuscular junction is a potential site of Pb interaction. 

15 A study by Li et al. (2009c) focused on inflammatory endpoints and synaptic changes 

16 after gestational plus lactational dam drinking water Pb exposure (solutions of 0.1%, 

17 0.5% or 1%, resulting in offspring blood Pb levels of 40, 80 and 100 μg/dL, respectively 

18 at PND 21). Hippocampal TNF-α was significantly elevated with Pb exposure and 

19 proteins that comprise the SNARE complex were all changed with Pb exposure. The 

20 SNARE complex of synaptic proteins includes SNAP-25, VAMP-2 and Syntaxin 1a and 

21 is essential in exocytotic neurotransmitter release at the synapse (Li et al., 2009c). Thus, 

22 Li et al. (2009c) found significant differences in hippocampal synaptic protein 

23 composition and increased pro-inflammatory cytokine levels in the brains of Pb-exposed 

24 offspring.

25 Neurotransmission is an energy-dependent process as indicated by the presence of 

26 calcium-dependent ATP releases at the synaptic cleft. At the synapse, ATP is 

27 metabolized by ectonucleotidases. In heme synthesis, Pb is known to substitute for the 

28 cation zinc in another nucleotidase, pyrimidine 5′-nucleotidase, and thus, the nucleotidase 

29 is used as a biomarker of Pb exposure. Acute exposure (96 hours) of male and female 

30 zebrafish to Pb-acetate (2 μg/dL) in their water induced significant decreases in ATP 

31 hydrolysis in brain tissue. This dose is deemed to be environmentally relevant. With 

32 chronic exposure (30 days), Pb-acetate promoted the inhibition of ATP, ADP and AMP 

33 hydrolysis; these findings were consistent with findings in rodents (Baranowska34 Bosiacka et al., 2011b). The authors hypothesized that at 30 days, this Pb-induced change 

35 in nucleotide hydrolysis was likely due to post-translational modification because 

36 expression of enzymes responsible for the hydrolysis, NTPDase1 and 5′-nucleotidase, 

37 were unchanged (Senger et al., 2006). Thus, Pb has been shown to affect nucleotidase 
February 2012 5-179 Draft – Do Not Cite or Quote

1 activity in the central nervous system of zebrafish, possibly contributing to aberrant 

2 neurotransmission.

3 Another enzyme important in synaptic transmission at cholinergic junctions in the CNS 

4 and at neuromuscular junctions peripherally is acetylcholinesterase (AChE). After 24 

5 hours of exposure to Pb-acetate (2 μg/dL water), AChE activity was significantly 

6 inhibited in zebrafish brain tissue. In Pb-exposed fish, AChE activity returned to baseline 

7 by 96 hours and maintained baseline activity after chronic exposure of 30 days. Thus, Pb 

8 was shown also to affect synaptic homeostasis of AChE in the brains of zebrafish

9 (Richetti et al., 2010).

10 Pb has been shown to act as an antagonist of the NMDA receptor (NMDAR). The 

11 NMDAR is essential for proper presynaptic neuronal activity and function. Primary 

12 cultures of mouse hippocampal cells were exposed to Pb (10 or 100 μM solutions in 

13 media) during the period of synaptogenesis (Neal et al., 2010a). This exposure induced 

14 the loss of two proteins necessary for presynaptic vesicular release, synaptophysin (Syn) 

15 and synaptobrevin (Syb), without affecting a similar protein synaptotagmin (Syt). This 

16 deficit was found in both GABAergic and glutamatergic neurons. Pb also induced an 

17 increase in number of presynaptic contact sites. But, these sites may be nonfunctional as 

18 they lack the protein receptor complexes necessary for proper vesicular exocytosis. 

19 Another factor involved in growth and signaling of presynaptic neurons is BDNF, which 

20 is synthesized and released by postsynaptic neurons. BDNF is regulated by the NMDAR 

21 and acts in a retrograde fashion, participating in presynaptic maturation. In hippocampal 

22 cells, both pro-BDNF and BDNF release were significantly attenuated with Pb exposure

23 (Neal et al., 2010a). Further, exogenous BDNF administration rescued the 

24 aforementioned Pb-related presynaptic effects. Thus, this cell culture model showed that 

25 Pb-related presynaptic aberrations are controlled by NMDAR-dependent BDNF effects 

26 on synaptic transmission.

27 Animals exposed to Pb postnatally (Wistar Albino rats, drinking water 300 mg/L 

28 Pb-acetate, resulting in blood Pb levels of 17 μg/dL at 6 weeks of age), from birth 

29 through lactation and through 12 weeks of age showed decreased learning ability and 

30 decreased hippocampal glutamate at 6, 8, 10 and 12 weeks of age (Niu et al., 2009) as 

31 well as significant decrements in the hippocampal glutamate synthesis-related enzymes 

32 aspartate aminotransferase and alanine aminotransferase.

5.3.8.5 Blood Brain Barrier

33 Two barrier systems exist in the body to separate the brain or the central nervous system 

34 from the blood. These two barriers are the blood brain barrier (BBB) and the blood 
February 2012 5-180 Draft – Do Not Cite or Quote

1 cerebrospinal fluid barrier (BCB). The BBB, formed by tight junctions at endothelial 

2 capillaries forming the zonulae occludens (occludins, claudins, and cytoplasmic proteins), 

3 separates the brain from the blood and its oncotic and osmotic forces, allowing for 

4 selective transport of materials across this barrier.

5 Pb exposure during various developmental windows has been shown to increase the 

6 permeability of the BBB (Dyatlov et al., 1998; Struzynska et al., 1997b; Moorhouse et 

7 al., 1988; Sundstrom et al., 1985). Possibly due the underdevelopment of the BBB early 

8 in life, prenatal and perinatal Pb exposure has been found to result in higher brain Pb 

9 accumulation than have similar exposures later in life (Moorhouse et al., 1988). Studies 

10 reviewed in earlier Pb AQCDs have shown that the chemical form of Pb and its 

11 capability to interact with proteins and other blood components affects its capability to 

12 penetrate the BBB (U.S. EPA, 2006b). Pb also has been shown to compromise the 

13 function of the BCB and decrease the CSF level of transthyretin, a thyroid binding 

14 protein made in the choroid plexus. The choroid plexus and cerebral endothelial cells that 

15 form the BBB and BCS tight junctions have been shown to accumulate Pb more than 

16 other cell types and regions of the CNS do. Hypothyroid status can contribute to impaired 

17 learning and IQ deficits (Lazarus, 2005).

18 Recent research with weanling rats exposed to Pb-acetate via drinking water showed 

19 leaky cerebral vasculature, an indication of a compromised BBB, as detected 

20 histologically with lanthanum nitrate staining of the brain parenchyma. Cerebral 

21 vasculature leakiness was ameliorated or resembled controls after iron supplementation. 

22 These weanlings also had significant Pb-induced decreases in the BBB tight junction 

23 protein occludin in the hippocampus, brain cortex, and cerebellum that were rescued to 

24 control levels with iron supplementation (Wang et al., 2007b). These data demonstrate 

25 that Pb induced a leaky BBB in weanling rats with associated decreases in the junctional 

26 protein occludin; dietary supplementation with iron ameliorated these Pb-induced 

27 impairments of the BBB in male rats. This loss of integrity at the junctional protein level 

28 was affirmed with additional experiments using the rat brain vascular endothelial cell line 

29 RBE4, in which 10 μM Pb-acetate exposure for 2, 4, 8, 16 and 24 hours resulted in 

30 decreases in junctional proteins occludin and claudin 5 as well as scaffold proteins ZO1 

31 and ZO2 (Balbuena et al., 2011). Because expression of these junctional and scaffold 

32 proteins did not show decrements, it was determined that these protein decrements were 

33 due to post-translational modifications.

34 A study examined the effects of Pb on transendothelial electrical resistance (TEER), a 

35 marker of BBBB integrity, in an in vitro co-culture system employing endothelial cells 

36 (RBE4 or bovine brain microvascular endothelial cells) and astrocytes (primary Sprague37 Dawley neonatal pup astrocytes, in utero day 21) as the barrier between Pb containing 
February 2012 5-181 Draft – Do Not Cite or Quote

1 exposure media and neurons. After 14 hours of exposure to Pb, TEER was significantly 

2 impaired in a concentration-dependent manner, with the lowest significant effects found 

3 with 1 μM Pb-acetate exposure. Thus, Pb exposure was found to contribute to leakiness 

4 of the BBB by decreasing the resistance across the junction. 

5 Adult male rats exposed to Pb-acetate in drinking water for 4 or 12 weeks (50 or 

6 500 ppm, resulting in blood Pb levels of 12 and 55 μg/dL, respectively) were assessed by 

7 diffusion weighted imaging for changes in apparent diffusion coefficent (ADC), a 

8 measure of tissue water diffusivity that changes under pathological conditions like 

9 cerebral edema. After 4 weeks of exposure to 500 ppm Pb, the water ADC was 

10 significantly increased in the hippocampus, mesencephalic reticular formation, and 

11 cerebellum but unaffected in other brain areas. After 12 weeks of 500 ppm Pb exposure, 

12 ADC was significantly increased in the corpus callosum and caudate putamen. Exposure 

13 to 50 ppm Pb for 12 weeks increased the ADC values in the cerebellum and 

14 mesencephalic reticular formation. The brain areas with elevated ADC also showed 

15 increased BBB permeability as measured with evans blue albumin complex. These data 

16 show that adult animals with chronic Pb exposure are susceptible to regional edema and 

17 regional increased permeability of the BBB, even with low blood Pb levels (12 μg/dL)

18 (López-Larrubia and Cauli, 2011).

5.3.8.6 Cell Adhesion Molecules

19 Classic cell adhesion molecules including NCAM and the cadherins are junctional or cell 

20 surface proteins that are critical for cell recognition and adhesion. Cell adhesion 

21 molecules, particularly the cadherins, are calcium-dependent and thus interaction from 

22 competing cations like Pb can potentially contribute to nervous system barrier function 

23 disruption, tissue development dysregulation, immune dysfunction, and affect learning 

24 and memory (Prozialeck et al., 2002).

5.3.8.7 Glial Effects

25 Astroglia and oligodendroglia are supporting cells in the nervous system that maintain the 

26 extracellular space in the brain and provide support and nutrition to neurons via nutrient 

27 transport, structural support to neurons, and myelination. Glial cells provide immune 

28 surveillance in the brain or contribute to inflammation-mediated pathologies. In the 

29 central nervous system, Pb treatment of Wistar rats 15 mg/kg of Pb-acetate, i.p.) during 

30 early postnatal maturation was observed to produce chronic glial activation with 

31 coexisting features of inflammation and neurodegeneration (Struzynska et al., 2007).
February 2012 5-182 Draft – Do Not Cite or Quote

1 Among the cytokines detected in the brains of these Pb-treated rats were IL-1β, TNF-α 

2 and IL-6. Glial cells have been shown to serve as Pb sinks in the developing and mature 

3 brain (Tiffany-Castiglioni et al., 1989) by sequestering Pb. This glial sequestration of Pb 

4 has been shown to decrease brain glutamine concentrations at doses of 0.25 ± 1.0 μM 

5 Pb-acetate via Pb-related reduction in glutamine synthetase activity in the astroglia; 

6 astroglia take up glutamate after its release and convert it to glutamine. Pb has been 

7 shown to induce hypomyelination and demyelination (Coria et al., 1984) mediated 

8 through the oligodendrocytes with younger animals being more susceptible to the effects 

9 of Pb (Tiffany-Castiglioni et al., 1989). Pb accumulation in young glial cells may 

10 contribute to a lifelong exposure of neurons to Pb as Pb is released from the sink over 

11 time. Thus, Pb accumulation in glial cells can contribute to continual damage of 

12 surrounding neurons (Holtzman et al., 1987).

Glial transmitters

13 To determine the contribution of the gliotransmitter serine to Pb-mediated changes in 

14 long-term potentiation (LTP), Sun et al. (2007) exposed pups to Pb-acetate in utero, 

15 lactationally, through PND28 via drinking water and collected hippocampal sections. 

16 CA1 section LTPs were examined using in vitro patch clamp monitoring. Chronic Pb 

17 exposure impaired the magnitude of hippocampal LTPs, but the magnitude of LTDs was 

18 restored with supplementation with D-serine (Sun et al., 2007), which is known to be 

19 regulated by the NMDAR (Bear and Malenka, 1994). The use of 7-chlorokynurenic acid, 

20 an antagonist of the glycine binding site of the NMDAR, which also is the binding site of 

21 D-serine, effectively abolished the rescue to LTP by D-serine. NMDAR-independent 

22 LTP hippocampal neurotransmission, which was examined in slices of Pb-exposed 

23 mossy-CA3 synapses, was not rescued by exogenous D-serine supplementation. These 

24 data indicate that glial transmission is affected with Pb exposure and that the NMDAR 

25 may also be involved in this aberrant glial transmission. 

5.3.8.8 Neurotransmitters

26 Pb has been shown to compete with calcium for common binding sites and second 

27 messenger activation. When Pb activates a calcium-dependent system in the nervous 

28 system, it can contribute to aberrant neurotransmitter regulation and release because this 

29 system intimately relies on calcium signaling for its homeostasis. Pb also has been shown 

30 to interfere with other physiological divalent cations. Pb-related alterations in 

31 neurotransmission are discussed in further detail below.
February 2012 5-183 Draft – Do Not Cite or Quote

Monoamine Neurotransmitters and Stress

1 Combined exposures of maternal stress and Pb exposure can synergistically enhance 

2 behavioral and neurotoxic outcomes in offspring of exposed animals and can sometimes 

3 potentiate an effect that would otherwise be sub-threshold. Virgolini et al. (2008a) found 

4 that CNS effects of developmental Pb exposure (50 or 150 ppm via drinking water, 

5 2 months prior to mating through lactation, resulting in blood Pb levels of 11 μg/dL and 

6 35 μg/dL, respectively) were enhanced with combined maternal and offspring stress. 

7 Offspring neurotransmitter concentrations were significantly affected with Pb exposure, 

8 but the most interesting findings were those of potentiated effects, i.e., effects that were 

9 not observed with Pb exposure alone or stress alone. These potentiated effects were only 

10 observed when Pb was combined with stress (maternal [MS] and/or offspring stress 

11 [OS]). Potentiation of serotonin (5HT) levels in females was significant in the frontal 

12 cortex in females and in the nucleus accumbens (NAC) in the male offspring (50 and 

13 150 ppm Pb drinking water exposure) (Cory-Slechta et al., 2009). Regional 5HT levels 

14 were unaffected in offspring with Pb exposure alone. The concentration 

15 of 5-Hydroxyindoleacetic acid (5HIAA), the main metabolite of 5HT, was significantly 

16 increased with Pb exposure alone in the striatum of male offspring with 150 ppb Pb 

17 exposure alone; with the remaining Pb-stress exposure combinations, Pb plus stress 

18 potentiated striatal and frontal cortex 5HIAA in males. Potentiated 5HIAA levels in 

19 females were significant in the NAC at both Pb doses; stress alone also significantly 

20 increased 5HIAA levels in females with no Pb exposure. Pb-induced changes in brain 

21 neurochemistry with or without concomitant stress exposure are complex with 

22 differences varying by brain region, neurotransmitter type and sex of the animal. 

Monoamine Neurotransmitters and Auditory Function

23 The monoamine neurotransmitters include DA, 5HT, and norepinephrine (NE). Earlier 

24 work has shown that perinatal Pb exposure of rats induced increased tyrosine 

25 hydroxylase, increased DA and increased cerebral cortex catecholamine 

26 neurotransmission (Devi et al., 2005; Leret et al., 2002; Bielarczyk et al., 1996). Earlier 

27 publications detailing important time windows of exposure, duration of exposure, and 

28 dose of Pb indicated varying effects on monoamine transmitters. In more recent work, 

29 these neurotransmitters, among others, have been implicated in Pb effects on auditory 

30 function in the brainstem in various integration centers there including the lateral superior 

31 olive (LSO), and the superior olivary complex (SOC). Among various functions, the SOC 

32 is vital for sound detection in noisy settings. Low-level Pb exposure has been associated 

33 with altered processing of auditory temporal signals in animal studies (Lurie et al., 2006; 

34 Finkelstein et al., 1998). Blood Pb levels for control, very low Pb (VLPb) and low Pb 

35 (LPb) exposure groups were 1.4, 8.0, and 42.2 μg/dL, respectively. Developmental Pb 
February 2012 5-184 Draft – Do Not Cite or Quote

1 exposure from the formation of breeding pairs to PND21, which is at the end of auditory 

2 development in the mouse, led to significant decreases in immunostaining of LSO and 

3 SOC brainstem sections for monoamine vesicular transporter VMAT2, and for 5HT and 

4 dopamine beta-hydroxylase (DbH), a marker for NE. Statistically significant alterations 

5 in VMAT2 and DbH were found with both VLPb and LPb exposure; however, 

6 decrements in 5HT were statistically significant only with VLPb. Immunostaining for TH 

7 and transporters including VGLUT1, VGAT, VAChAT indicated that they were 

8 unaffected by developmental Pb exposure. These data provide evidence that specific 

9 regions of the brainstem involved in auditory integration are affected by developmental 

10 Pb exposure via effects on the monoamine neurotransmitter system (Fortune and Lurie, 

11 2009). The Pb-induced effects on the auditory portion of the brainstem at the level of the 

12 monoamine system provide possible mechanistic understanding of the animal data 

13 showing Pb-induced impaired auditory processing.

Dopamine

14 The 2006 Pb AQCD detailed evidence for low-dose Pb-related decreased dopaminergic 

15 cell activity in the substantia nigra and ventral segmental areas. Earlier studies with 

16 moderate- to high-dose postnatal or adult Pb exposure have reported changes in DA 

17 metabolism, as indicated by changes in DA and DOPAC, a DA metabolite. Given these 

18 findings, a recent study measured DA and DOPAC in various brain regions of year-old 

19 male rodents to examine if GLE affected DA metabolism. Low- and high-dose GLE in 

20 male rodents induced significant elevations in DOPAC concentration and the DOPAC to 

21 DA ratio in the forebrain. In the forebrain, DA was significantly decreased in low-dose 

22 GLE males and significantly elevated in high-dose GLE males compared to controls. In 

23 the striatum, DOPAC was significantly elevated with both low- and high-dose GLE 

24 males, but DA concentration was only significantly elevated in high-dose GLE males. 

25 The striatum ratio of DOPAC to DA was not significantly different from that in controls. 

26 These new data expand upon the monoamine literature base in which indicates low 

27 concentration perinatal Pb exposure of rats was found to induce increased sensitivity of 

28 the dopamine receptors (D2 and D3) (Gedeon et al., 2001; Cory-Slechta et al., 1992),

29 produce higher DA levels (Devi et al., 2005; Leret et al., 2002), and enhance 

30 catecholamine neurotransmission in the cerebral cortex, cerebellum, and hippocampus

31 (Devi et al., 2005).

32 The interaction of DA and the nitric oxide system in the striatum was studied after 

33 prenatal Pb exposure (Nowak et al., 2008). Blood Pb levels were not reported in this 

34 study, but similarly treated Wistar rat pups had blood Pb levels at parturition in range of 

35 50-100 μg/dL (Grant et al., 1980). 7-nitroinidazole (7-NI), a selective inhibitor of nNOS, 

36 enhanced amphetamine-evoked DA release in the rat striatum (Nowak et al., 2008).
February 2012 5-185 Draft – Do Not Cite or Quote

1 Prenatal Pb exposure attenuated the facilitatory effect of 7-NI on DA release in the 

2 striatum. This interaction is ROS-independent; using spin trap measurements, there were 

3 no significant concentration changes in hydroxyl radical with Pb exposure (Nowak et al., 

4 2008). Thus, the neuronal NO system appears to be involved in specific aspects of Pb5 related dopaminergic changes. 

Dopamine and Vision

6 In various experimental animal models, the loss of retinal DA or zinc is associated with 

7 abnormal rod-mediated scotopic ERGs. These effects may explain observations of Pb8 associated retinal effects (Rothenberg et al., 2002a; Lilienthal et al., 1994; Lilienthal et 

9 al., 1988; Alexander and Fishman, 1984). In the human and animal toxicological 

10 literature, prenatal maternal blood Pb level (humans), gestational + early postnatal Pb 

11 exposure (rodents, low and moderate dose Pb only), or gestational continuing to lifetime 

12 (non-human primate) Pb exposure has been associated with increased amplitude 

13 (supernormality) of ERGs (Rothenberg et al., 2002a; Lilienthal et al., 1994; Lilienthal et 

14 al., 1988). In the animal toxicological literature, subnormality of the ERGs has been 

15 observed with postnatal Pb exposure (Fox et al., 2008; Otto and Fox, 1993; Fox et al., 

16 1991; Fox and Farber, 1988) and high-dose developmental (prenatal + early postnatal, 

17 rodent) Pb exposure (Fox et al., 2008). Producing results consistent with observations in 

18 humans (Rothenberg et al., 2002a), Fox et al. (2008) showed that low- (LPb) and 

19 moderate-level (MPb) gestational+early postnatal Pb exposure in the rat, a period 

20 equivalent to the human gestational retinal development period, produced supernormal 

21 retinal ERGs. In children, supernormal ERGs were associated with prenatal Pb maternal 

22 blood Pb levels >10.5 μg/dL (Rothenberg et al., 2002a). The animal data provide 

23 mechanistic information that may begin to explain these supernormal ERGs that are seen 

24 in children and rodents, i.e., significant increases in retinal neurogenesis and significant 

25 decreases in retinal DA use and dopamine turnover (DOPAC:DA ratio). High-dose 

26 developmental (rodent, gestational+early postnatal to PND10) Pb exposure (HPb) 

27 produced significant subnormal retinal ERGs. Subnormal ERGs also were found in 

28 occupationally-exposed adults (Otto and Fox, 1993; Guguchkova, 1972). Female rats 

29 were exposed to Pb-acetate in drinking water from 2 weeks prior to mating throughout 

30 gestation and lactation until PND 10, a period of developmental exposure that is 

31 equivalent to gestational exposure in humans. Peak blood Pb levels in the offspring at 

32 PND1-10 were 12, 24, and 46 μg/dL in the LPb, MPb, and HPb groups, respectively. LPb 

33 and MPb gestational exposure induced increased cellularity or retinal thickness in the 

34 outer nuclear layer, inner nuclear layer and total retina (Leasure et al., 2008). In 

35 summary, the retina is affected by low-dose Pb exposure and gestational Pb exposure, as 

36 indicated by concentration-dependent decreases in DA use and turnover. Inverted U-
February 2012 5-186 Draft – Do Not Cite or Quote

1 shaped Pb concentration-response relationship were reported for retinal endpoints 

2 including ERG and retinal thickness. 

NMDA

3 NMDA receptors (NMDAR) have been shown to contribute to synaptic plasticity, and Pb 

4 exposure at different developmental stages has been shown to contribute to aberrations in 

5 LTP or long term depression (LTD) in the hippocampus via reduced NMDA current, 

6 among other mechanisms (Liu et al., 2004). The 2006 Pb AQCD indicated that Pb 

7 induced decreases in stimulated glutamate release that affected LTP. Further, the 

8 relationships between Pb exposure and decreased magnitude and increased threshold of 

9 the LTP in the hippocampus were found to be biphasic or nonlinear. NMDAR subtypes 

10 have been shown to be significantly decreased with developmental Pb exposure (Guilarte 

11 and McGlothan, 1998). Recent work examining dietary supplement use, found that Pb12 related decreases in the gene expression and protein level of NMDAR subunit NR1 in 

13 weanling male rats were rescued with methioninecholine co-exposure (Fan et al., 2010).

14 Fan et al. (2010) found that Pb-related suppression of the NMDAR subunits NR2A and 

15 NR2B was not rescued with methioninecholine treatment. Other recent mechanistic 

16 studies found that pretreatment of primary fetal brain neuronal rat cultures with glutamic 

17 acid, a NMDAR agonist, reversed Pb-induced reductions in NMDAR subunits (Xu and 

18 Rajanna, 2006) whereas pretreatment with the NMDA antagonist MK-801 exacerbated 

19 Pb-induced NMDAR deficits (Xu and Rajanna, 2006).

20 Studies continue to show that Pb exposure affects neurogenesis or proliferation of new 

21 cells in the hippocampus. Earlier work by Schneider et al. (2005) showed that postnatal 

22 Pb exposure (30-35 days starting at PND25, 1,500 ppm Pb-acetate in chow, resulting in 

23 blood Pb level of 20 μg/dL) of male Lewis rats induced significant decrements in BrdU 

24 incorporation (proliferation) at PND50-55. Recent publications affirm this original 

25 finding with different sex of animals, dosing and exposure time windows. Postnatal Pb 

26 exposure to Wistar rat pups (0.2% Pb-acetate from PND1-30, resulting in blood Pb levels 

27 of 34 and 6.5 μg/dL at PND21 and PND80, respectively) induced a statistically 

28 significant decrement in the number of new cells (BrdU positive cells) in the dentate 

29 gyrus at PND80 (Fox et al., 2010) (Figure 5-18). In another study, developmental/lifetime 

30 Pb exposure (1,500 ppm Pb-acetate in chow from 10 days before mating to termination of 

31 experiment at PND50, resulting in blood Pb levels of 0.8 μg/dL in controls and 26 μg/dL 

32 in Pb-exposed animals) to female Long-Evans rats induced significant decrements in 

33 hippocampal granule cell neurogenesis or proliferation of new cells in adult rats (Verina 

34 et al., 2007), outcomes that affect LTP, spatial learning, neuronal outgrowth, and possibly 

35 mood disorders such as schizophrenia. NMDAR mediates the integration of new neurons 

36 into existing neuronal pathways in the adult hippocampal DG, which is important to 
February 2012 5-187 Draft – Do Not Cite or Quote

1 learning and memory. Lifetime Pb exposure (dam Pb-acetate chow exposure 10 days 

2 prior to mating through pregnancy to PND50 or PND78) induced significant decrements 

3 in hippocampal granule cell neurogenesis or proliferation of new cells in adult rats. Also, 

4 Pb-exposed animals had significant decreases in brain volume in the stratum oriens (SO) 

5 region of the hippocampus, specifically significant decreases in the mossy fiber terminals 

6 of the SO. Pb-exposed animals also showed a significant decrease in the length-density of 

7 immature or newly-formed neuron in the outer portion of the DG. These findings show 

8 that exposure to environmentally-relevant doses of Pb induced significant aberrations in 

9 adult hippocampus granule cell neurogenesis and morphology, providing mechanistic 

10 explanations for Pb-induced neuronal aberrations. Guilarte et al. (2003) demonstrated that 

11 Pb exposure of rats from an enriched environment was associated with reduced learning 

12 impairment, increased expression of hippocampal NMDA receptor subunit 1, and 

13 increased induction of brain derived neurotrophic factor mRNA (Guilarte et al., 2003).

Glutamate Receptor

14 Glutamate receptors including the ionotropic NMDAR and the metabotropic glutamate 

15 receptors (mGluR) are known targets of Pb toxicity with recent findings showing a role 

16 for mGluR5 in learning and memory. In vitro (GD18 fetal rat cultures, 100 μM, 1 μM, 

17 0.01 μM PbCl2 in culture media) and in vivo studies (gestational and lactational 

18 Pb-acetate exposure; control, 0.05, 0.2, or 0.5% in dam drinking water, with respective 

19 weanling blood Pb levels of 3, 18, 57, 186 μg/dL) showed that Pb exposure induced 

20 mGluR5 mRNA and protein decrements in a concentration-dependent manner (Xu et al., 

21 2009c).. The Pb-related attenuation of mGlu5 expression may contribute to the effect of 

22 Pb on LTP and LTD. 

5.3.8.9 Neurite Outgrowth

23 The 2006 Pb AQCD reported that Pb decreased neurite outgrowth at 20μg/dL and noted 

24 that Pb interfered with neurite outgrowth via protein kinase mediated pathways 

25 (MAPK/ERK); earlier work had documented decreased primary DA neuron outgrowth 

26 with 0.001 μM Pb exposure (Lidsky and Schneider, 2004). Recent studies have shown 

27 that exposure of dams to low-dose Pb (resulting in blood Pb level of 4 μg/dL) 

28 significantly decreased pup hippocampal neurite outgrowth (pup blood Pb level: 

29 12 μg/dL) and reduced the expression of hippocampal polysialylated neural cell adhesion 

30 molecule (PSA-NCAM), NCAM, and sialytransferase (Hu et al., 2008b). PSA-NCAM is 

31 transiently expressed in newly formed neurons during the period of neurite outgrowth 

32 from embryrogenesis until the early postnatal period and is down-regulated in the adults 
February 2012 5-188 Draft – Do Not Cite or Quote

1 except in areas known to exhibit synaptic plasticity (Seki and Arai, 1993). NCAM is

2 important for memory formation, plasticity and synapse formation, and early-life Pb 

3 exposure was found to affects its expression in laboratory rats. 

5.3.8.10 Epigenetics

4 Many investigators are beginning to show that environmental chemical exposures are 

5 associated with epigenetic changes. Air pollution exposure is being linked increasingly 

6 with epigenetic changes (Baccarelli and Bollati, 2009; Pavanello et al., 2009; Tarantini et 

7 al., 2009; Bollati et al., 2007). Epigenetic changes involve changes in DNA expression 

8 without changes in the DNA sequence, and these changes may be heritable. Epigenetic 

9 changes are mediated by histone modification, DNA methylation, miRNA changes, or 

10 pathways that affect these processes. Differential epigenetic modification has the 

11 potential to contribute to disease by silencing or activating genes in an aberrant manner. 

12 Monozygotic twins are often used to study epigenetic changes, and a recent study 

13 identified differential methylation of a specific locus in twins discordant for 

14 schizophrenia (Dempster et al., 2011); Pb was not examined in this study. 

15 DNA methyltransferases catalyze the transfer of a methyl group to DNA and are 

16 important in epigenetics (i.e., silencing of genes like tumor suppressors) and imprinting. 

17 DNA methyltransferase activity was significantly decreased in cortical neurons from Pb18 exposed monkeys (aged animals, blood Pb) and mouse brains (fetal cells exposed to Pb in 

19 culture, 0.1 μM Pb) (Wu et al., 2008b). Changes in DNA methyltransferases (Dnmt1, 

20 Dnmt3a) were noted in control primate brains as they aged and these changes were 

21 further exacerbated by Pb exposure (Bihaqi et al., 2011). Another enzyme involved in 

22 DNA methylation, methyl CpG binding protein 2 MECP2, showed a similar trend as the 

23 Dnmts. Profiles of the histone modifying gene H34mc2 increases with age in control 

24 animals. This age-related increase is significantly attenuated in Pb-exposed animals. The 

25 cerebral cortex tissue used in this experiment was obtained from female primates who 

26 had received 1.5 mg/kg • day Pb-acetate via diet from birth until 400 days of age 

27 (resulting in blood Pb levels 19-26 μg/dL at age 400 days) (Rice, 1990).

28 Methyltransferases catalyze biological methylation reactions and are dependent on the 

29 cofactor S-adenosyl methionine (SAM) for this transfer to acceptor molecules. SAM 

30 exposure after gestational and lactational Pb exposure (dam Pb-acetate exposure to 

31 1,500 ppm Pb-acetate followed by 20-22 days of daily 20 mg/kg BW SAM exposure) 

32 improved hippocampal LTP and Morris water maze performance at PND 44-54. Thus, 

33 the impaired cognition and synaptic plasticity induced by developmental Pb exposure 

34 were attenuated with SAM treatment (Cao et al., 2008).
February 2012 5-189 Draft – Do Not Cite or Quote

5.3.8.11 Cholesterol and Lipid Homeostasis

1 Various pathological conditions are associated with elevated plasma free fatty acids or 

2 elevated cholesterol. Adult male rats exposed to Pb-acetate (200, 300, or 400 ppm) in 

3 their drinking water for 12 weeks had increased cholesterogenesis and phospholipidosis 

4 in brain tissue (Ademuyiwa et al., 2009). Pb-induced changes in brain cholesterol showed 

5 an inverse U concentration-response relationship, with the largest increase in brain 

6 cholesterol observed with 200 ppm Pb followed by 300 ppm Pb. Animals exposed to 

7 400 ppm Pb did not have significant changes in brain cholesterol. Mechanistically, Pb 

8 exposure has been shown to depress the activity of cholesterol-7-a-hydroxylase, an 

9 enzyme involved in bile acid biosynthesis (Kojima et al., 2005); bile acids are the route 

10 by which cholesterol is eliminated from the body. Pb exposure produced significant 

11 increases in brain triglycerides with an 83% increase at 300 ppm and a 108% increase at 

12 400 ppm. At 200 ppm, Pb exposure induced a statistically nonsignificant decrease in 

13 brain triglycerides. Pb exposure across all three dose groups induced significantly 

14 increased brain phospholipids. Interestingly, plasma free fatty acids were significantly 

15 elevated in a concentration-dependent manner; plasma triglycerides and cholesterol were 

16 unaffected by Pb exposure. The molar ratio of brain cholesterol to phospholipids, an 

17 indicator of membrane fluidity (Abe et al., 2007), was significantly increased at 200 and 

18 300 ppm Pb exposure indicating increased membrane fluidity. Brain Pb in all dose groups 

19 was below the limit of detection (0.1 ppm). Blood Pb levels at 0, 200, 300, and 400 ppm

20 were 7, 41, 61, and 39 μg/dL, respectively. In summary, based on limited examination, 

21 Pb exposure significantly increased brain cholesterol, triglycerides, and phospholipids as 

22 well as significantly increased plasma free fatty acids. These effects were sometimes 

23 more prominent at lower doses of Pb. Future characterization of molecular and cellular 

24 pathways affected by Pb exposure may bring insight to these Pb-related changes in 

25 phospholipidosis and cholesterogenesis. 

5.3.9 Lifestage of Lead Exposure and Neurodevelopmental Deficits

26 Environmental exposures during critical lifestages can affect key physiological systems 

27 that orchestrate plasticity (Feinberg, 2007). Exposure to environmental toxins during 

28 prenatal and/or early postnatal development may alter the normal course of 

29 morphogenesis and maturation that occurs in utero and early in life, resulting in changes 

30 that affect structure or function of the central nervous system via altered neuronal growth 

31 and/or synaptogenesis/pruning structure (Rice and Barone, 2000; Landrigan et al., 1999).

32 Synaptic pruning, which is active throughout early childhood (ages 1-4 years), may 

33 underlie the elevated risk of young children to environmental exposures. MRI studies 

34 have provided understanding of brain development in normally developing children and 
February 2012 5-190 Draft – Do Not Cite or Quote

1 adults, ages 3-30 years (Giedd et al., 2009; Lenroot and Giedd, 2006). Total cerebral 

2 volume was found to peak at age 10.5 and 14.5 years in females and males, respectively. 

3 The volume of the cerebellum was found to peak 2 years after the cerebral volume peaks. 

4 Lateral ventricular volume showed the most inter-individual variation and was found to 

5 increase throughout childhood and adolescence. White matter volume generally increased 

6 throughout childhood and adolescence. Gray matter volume and associated structures 

7 show inverted U-shaped developmental trajectories with peak volumes found in late 

8 childhood or early adolescence. Females generally had gray matter volume peaks 1 to 

9 3 years earlier than did males. Thus, the anatomical development of the brain during 

10 childhood and adolescence is found to be a dynamic process with variation by age, brain 

11 region, and sex. Observations that brain development is active throughout childhood and 

12 in adolescence indicates that Pb exposure may affect neurodevelopment throughout this 

13 period.

14 The elevated risk of Pb-associated neurodevelopmental deficits in children is well 

15 supported by findings in animals that prenatal and postweaning Pb exposure alters brain 

16 development via changes in synaptic architecture (Section 5.3.8.4) and neuronal 

17 outgrowth (Section 5.3.8.9) and leads to impairments in memory and learning 

18 (Section 5.3.2.2) and emotional and depressive changes postnatally (Section 5.3.3.5). 

19 Unlike other organ systems, the unidirectional nature of CNS development limits the 

20 capability of the developing brain to compensate for cell loss, and environmentally21 induced cell death can result in a permanent reduction in cell numbers (Bayer, 1989).

22 Hence, when normal development is altered, the early effects may persist into adult life 

23 even in the absence of current exposure, magnifying the public health impact. Supporting 

24 evidence is provided by a few available new toxicological studies that find that Pb 

25 exposure during neonatal development but not in adulthood leads to neurodegenerative 

26 amyloid plaque formation in the brains of aged rodents and monkeys (Section 5.3.7.2).

27 With repeated assessments of children prenatally to later childhood and early adulthood, 

28 the prospective cohort studies have aimed to distinguish among neurodevelopmental 

29 effects associated with blood Pb levels measured at different periods of development. In 

30 the collective body of evidence, cognitive function decrements in children have been 

31 associated with prenatal, early childhood, childhood average, and concurrent blood Pb 

32 levels, without clear indication that the risk of neurodevelopmental decrements is greatest 

33 for blood Pb levels measured at a particular lifestage. In these studies, the identification 

34 of developmental periods when children are at increased risk of Pb-associated 

35 neurodevelopmental decrements has been complicated by the high degree of correlation 

36 in the blood Pb levels of children over time and the confounding of age and peak blood 

37 Pb levels (Lanphear et al., 2005; Dietrich et al., 1993b; Needleman et al., 1990).
February 2012 5-191 Draft – Do Not Cite or Quote

1 As described in detail in the 2006 Pb AQCD, several studies with varying lengths of 

2 follow-up demonstrated associations of prenatal blood Pb levels (maternal and umbilical 

3 cord) with neurodevelopmental deficits throughout childhood and into early adulthood

4 (U.S. EPA, 2006b). Pb exposure during the prenatal lifestage may be associated with

5 increased risk of neurodevelopmental effects not only because of the nervous system 

6 developmental processes that are active as described above but also because of factors 

7 that result in elevated Pb exposures. Substantial fetal Pb exposure may occur from 

8 mobilization of maternal skeletal Pb stores that may be related to past Pb exposures

9 (Gulson et al., 2003; Hu and Hernandez-Avila, 2002). Pb can cross the placenta to affect 

10 the developing fetal nervous system (Rabinowitz, 1988). Maternal and cord blood Pb 

11 levels generally have been shown to be highly correlated, indicating that blood Pb levels 

12 in a newborn infant reflects that of the mother (Schell et al., 2003).

13 Prenatal blood Pb levels (maternal and cord) were consistently associated with cognitive 

14 function decrements and behavioral problems assessed between infancy and age 3 years 

15 (Table 5-4 and Table 5-13). Among studies that had blood Pb measurements at both 

16 lifestages, some found stronger associations for prenatal blood Pb levels (Hu et al., 2006; 

17 Bellinger et al., 1984), and other found stronger associations for concurrent blood Pb 

18 levels (Wasserman et al., 1998; Wasserman et al., 1992). Studies that found associations 

19 with concurrent blood Pb levels also tended to find associations with prenatal cord or 

20 maternal blood Pb levels. Thus, both postnatal child and maternal Pb exposures may 

21 contribute to lower cognitive function in young children. Several studies found that 

22 prenatal or neonatal blood Pb levels were associated with neurodevelopmental 

23 decrements assessed neonatally (within 30 days) or early in infancy (within 3 months), 

24 which indicated that relatively short durations of Pb exposure were associated with 

25 neurodevelopmental decrements (Shen et al., 1998; Rothenberg et al., 1989; Dietrich et 

26 al., 1987a; Ernhart et al., 1986). In the studies of neurodevelopmental effects in infancy, 

27 prenatal blood Pb level may be serving as a surrogate of postnatal blood Pb levels as both 

28 are expected to be highly correlated.
February 2012 5-192 Draft – Do Not Cite or Quote

Table 5-13 Associations of cognitive function and behavioral outcomes with 

blood Pb levels measured at different lifestagesa

Study Population/Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)b

Assessments in children up to age 3 years

Rothenberg et al.

(1989)

42 children followed 

prenatally to child age 30 

days

Mexico City, Mexico

Maternal week 36 

gestation mean (SD): 

15.0 (6.4)

Maternal at birth mean 

(SD): 15.5 (5.7)

Regression model adjusted 

for smoking, single mother, 

problems in pregnancy, 

alcohol use in previous 

month, use of spinal block, 

gravidity, income

Self-quieting ability 

(regulation of state) 

at age 30 days

Assessed using 

Newborn Brazelton 

Assessment System 

Prenatal: -0.091 (-0.18, 0)

Dietrich et al. (1986) 305 children followed 

prenatally to age 6 mo. 

in Cincinnati, OH

Prenatal (maternal) 

mean (SD): 8.0 (3.8)

Concurrent mean (SD): 

5.9 (3.4)

Log linear regression model 

adjusted for birth weight, 

gestation, sex

Bayley MDI assessed 

at age 6 mo

Prenatal: -0.6 (-1.1, -0.09)

Concurrent: -0.23 (-0.58, 0.12)

Bellinger et al.

(1987)

249 children followed from 

birth (1979-1981) to age 

36 mo

Boston area, MA

Prenatal (cord blood) 

mean (SD): 6.6 (3.2)

Regression and longitudinal 

analyses adjusted for the 

mother’s age, race, IQ, 

education, number of years 

of cigarette smoking, number 

of alcoholic drinks per week 

in the third trimester, mean 

family social class over the 

period of the study, quality of 

the care-giving environment, 

infant’s sex, birth weight, 

gestational age, birth order

Bayley MDI assessed 

at age 6, 12, 18, 24 

mo

Prenatal: -4.8 (-7.3, -2.3), 

blood Pb levels ≥ 15 μg/dL vs. 

blood Pb levels <3c

Hu et al. (2006) 146 children born 

1997-1999 followed 

prenatally to age 24 mo

Mexico City, Mexico

Prenatal (maternal 1st 

trimester) mean (range):

7.1 (1.5-43.6)

Early childhood (12 mo) 

mean (SD): 5.2 (3.4)

Concurrent mean (SD): 

4.8 (3.7)

Log linear regression model 

adjusted for concurrent blood 

Pb, sex, maternal age, 

current weight, height-for-age 

Z score, maternal IQ

Bayley MDI assessed 

at age 24 mo

Prenatal 1st trimester: -4.1 (-

8.1, -0.17)

Prenatal (avg): -3.5 (-7.7, 

0.63)

12 month: -2.4 (-6.2, 1.49)

Concurrent: -1.0 (-3.9, 1.9)

Gomaa et al. (2002) 197 children followed 

prenatally to age 24 mo

Mexico City, Mexico

Prenatal (cord blood) 

mean (SD): 6.7 (3.4)

Log linear regression model 

adjusted for maternal IQ, 

maternal age, sex, parental 

education, marital status, 

breastfeeding duration, child 

hospitalization status

Bayley MDI assessed 

at age 24 mo

Prenatal: -2.1 (-3.9, -0.39)

Wasserman et al.

(1992)

392 children followed 

prenatally to age 24 mo

Kosovo, Yugoslavia (K. 

Mitrovica, Pristina)

Prenatal (cord blood) 

mean (SD): 14.4 (10.4)

Concurrent means:

K. Mitrovica: 35.4, 

Pristina: 8.5

Log linear regression model 

adjusted for sex, birth order, 

birth weight, ethnic group, 

HOME score, years of 

maternal education, maternal 

age, maternal intelligence

Bayley MDI assessed 

at age 24 mo

Concurrent: -4.1 (-6.2, -2.0)

Prenatal: -3.2 (-7.2, 0.86)

Jedrychowski et al.

(2009b)

444 children born 

2001-2004 followed 

prenatally to age 36 mo

Krakow, Poland

Prenatal (cord blood) 

geometric mean (range): 

1.29 (0.44-5)

Linear regression model 

adjusted for maternal 

education, birth order, 

prenatal ETS, sex

Bayley MDI assessed 

at age 36 mo

Prenatal: -2.9 (-5.0, -0.75)

Cognitive function assessments at school age 

Wasserman et al.

(1994)

332 children followed 

prenatally to age 3-4 yr

Kosovo, Yugoslavia (K. 

Mitrovica, Pristina)

Prenatal (cord blood) 

mean (SD): 14.4 (10.4)

Concurrent means: 

K. Mitrovica: 39.9

Pristina: 9.6

Log linear regression model 

adjusted for HOME score, 

maternal age, maternal 

intelligence, maternal 

education, language, birth 

weight, sex

McCarthy GCI 

assessed at age 

3-4 yr

Concurrent: -4.1 (-6.2, -2.0)

Prenatal: -3.2 (-5.1, -1.2)
February 2012 5-193 Draft – Do Not Cite or Quote

Study Population/Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)b

Dietrich et al. (1992) 259 followed from birth 

(1979-1984) to age 5 yr

Cincinnati, OH

Prenatal (cord blood) 

mean (SD): 8.2 (3.8)

Neonatal (10 days) 

mean (SD): 4.8 (3.3)

Concurrent mean (SD): 

11.9 (6.4)

Linear regression model 

adjusted for fetal distress and 

growth, perinatal 

complications, postnatal 

indices of health and 

nutritional status, 

sociodemographic 

characteristics, HOME score

Total FWS 

assessed using 

KABC at age 5 yr

Neonatal: -0.38, p ≤ 0.01d

Prenatal: -0.25, p ≤ 0.01d

Concurrent: -0.19, p ≤ 0.01d

Lifetime avg: -0.16, p ≤ 0.01d

Bellinger et al.

(1991)

170 children followed from 

birth (1979-1981) to age 

57 mo

Boston area, MA

Early childhood (24 mo) 

mean (SD): 6.8 (6.3)

Early childhood tooth 

mean (SD): 2.8 (1.7) 

μg/g

Concurrent mean (SD): 

6.4 (4.1)

Log linear regression model 

adjusted for family social 

class, maternal IQ, marital 

status, preschool attendance, 

HOME score, out of home 

care, number of residence 

changes, recent medication 

use, number of adults in 

household, sex, race, birth 

weight, birth order

McCarthy GCI 

assessed at age 57 

mo

Early childhood blood: 

-3.0 (-5.7, -0.2)

Early childhood tooth: -2.5 

(-10.2, 5.2)

Concurrent blood: -2.3 (-6.0, 

1.4)

Dietrich et al.

(1993b)

245 children followed from 

birth (1979-1984) to age 6 

yr

Cincinnati, OH

Prenatal (cord blood) 

mean (SD): 8.4 (3.8)

Neonatal (10 days) 

mean (SD): 4.8 (3.1)

Concurrent mean (SD): 

10.1 (5.6)

Linear regression model 

adjusted for obstetric 

complications, perinatal 

status, sex, social class, 

maternal intelligence, quality 

of rearing environment, 

earlier measures of 

neurobehavioral status

Bruininks-Oseretsky 

Test of Motor 

Proficiency 

assessed at age 6 

yr

Concurrent: 

-0.18 (-0.26, -0.10)

Neonatal: -0.15 (-0.33, 0.03)

Lifetime avg: -0.11 

(-0.19, -0.03)

Prenatal: -0.04 (-0.20, 0.12)

Dietrich et al.

(1993b)

253 children followed from 

birth (1979-1985) to age 

6.5 yr

Cincinnati, OH

Prenatal (cord blood) 

mean (SD): 8.3 (3.7)

Neonatal (10 days) 

mean (SD): 5.0 (3.4)

Concurrent mean (SD): 

11.8 (6.3)

Linear regression model 

adjusted for fetal distress and 

growth, perinatal 

complications, prenatal 

maternal substance abuse, 

postnatal indices of health 

and nutritional status, 

sociodemographic 

characteristics, maternal IQ, 

HOME score

FSIQ assessed 

using WISC-R at 

age 6.5 yr 

Concurrent: 

-0.33 (-0.60, -0.06)

Lifetime avg: -0.13 (-0.35, 0.09)

Neonatal: -0.03 (-0.42, 0.36)

Prenatal: 0.15 (-0.26, 0.56)

Baghurst et al.

(1992)

494 children followed from 

birth (1979-1982) to age 

11-13 yr

Port Pirie, Australia

Prenatal mean of 

second quartile: 7.4

Early childhood (2 yr) 

mean of second quartile: 

16.6

Lifetime avg mean of 

second quartile: 15.7

Log linear regression model 

adjusted for sex, birth weight, 

birth order, feeding method, 

breastfeeding duration, 

parental education, maternal 

age, parental smoking, SES, 

quality of home environment, 

maternal IQ, parents living 

together

FSIQ assessed 

using WISC-R at 

age 7-8 yr 

Early childhood: 

-2.0 (-3.8, -0.21)

Lifetime avg: -1.6 (-3.7, 0.52)

Prenatal: 0.26 (-0.67, 1.5)

Lanphear et al.

(2005)

1,333 children pooled from 

Boston, Cincinnati, 

Cleveland, Mexico City, 

Port Pirie, Rochester, and 

Yugoslavia cohorts

Median (5th-95th)

Early childhood: 12.7 

(4.0-34.5)

Peak: 18.0 (6.2-47.0)

Lifetime avg: 12.4 

(4.1-34.8)

Concurrent: 9.7 

(3.5-33.2)

Log linear regression model 

adjusted for HOME score, 

birth weight, maternal IQ, 

maternal education

FSIQ measured at 

ages 4.8-10 yr

Concurrent: -

0.23 (-0.32, -0.14)

Peak: -0.20 (-0.29, -0.11)

Lifetime avg: 

-0.15 (-0.22, -0.09)

Early childhood: 

-0.14 (-0.23, -0.06)

Pocock et al. (1994) Meta-analysis of 5 

prospective (over 1,100 

children and 14 crosssectional studies (3,499 

children)

Early childhood (2 yr) 

range in means: 

6.8-21.2

Meta-analysis of combining 

effect estimates from 

individual studies

FSIQ assessed 

using various tests 

at ages 5-10 yr

Early childhood:

-2.7 (-4.1, -1.2)

Postnatal mean: -1.3 (2.9, 0.37)

Around birth: 0.26 (-1.5, 2.0)

Schnaas et al.

(2006)

150 children followed from 

prenatally (1987-1992) to 

age 6-10 yr

Mexico City, Mexico

Prenatal (maternal 28-36 

wk gestation): 7.8 (2.5, 

24.6)

Early childhood avg (1-5 

yr) mean (range): 9.8 

(2.8-36.4)

Later childhood avg 

(6-10 yr): 6.2 (2.2-18.6) 

Log linear mixed effects 

model adjusted for blood Pb 

levels at other lifestages, sex, 

birth weight, SES, maternal 

IQ, First FSIQ measurement

FSIQ assessed 

using WISC-R at 

ages 6-10 yr

Prenatal: -3.9 (-6.5, -1.4)

Early childhood avg: 

0.10 (-3.9, 4.1)

Later childhood avg: 0.17 (-1.4, 

1.8)
February 2012 5-194 Draft – Do Not Cite or Quote

Study Population/Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)b

Bellinger et al.

(1992)

148 children followed from 

birth (1979-1981) to age 

15-17 yr

Boston area, MA

Prenatal: NR

Early childhood (2 yr) 

mean (SD): 6.5 (4.9)

Concurrent mean (SD): 

2.9 (2.4)

Linear regression model 

adjusted for HOME score 

(age 10 and 5), child stress, 

race, maternal IQ, SES, sex, 

birth order, marital status

FSIQ assessed 

using WISC-R at 

age 10 yr 

Early childhood: 

-0.58 (-0.99, -0.17)

Prenatal: -0.48 (-5.7, 4.7), blood 

Pb >10 μg/dL vs. <3 μg/dLc

Concurrent: -0.46 (-1.5, 0.56)

Ris et al. (2004) 195 children in followed 

from birth (1979-1985) to 

age 

15-17 yr

Cincinnati, OH

NR Linear regression model 

adjusted for maternal IQ, sex, 

and average total HOME 

score

Learning/IQ 

composite 

assessed using 

WISC-III indices at 

age 15-17 yr

Prenatal: -0.08 (-0.18, 0.03)

Early childhood, 6.5 yr: 

-0.08 (-0.17, 0.003)

Early childhood avg: 

-0.03 (-0.18, 0.03)

Behavioral assessments 

Wasserman et al.

(1998)

379 children followed 

prenatally to age 3 yr

Kosovo, Yugoslavia 

(K. Mitrovica, Pristina)

Prenatal mean (SD): 

16.1 (2.6)

Concurrent mean (SD): 

25.8 (19.1)

Hierarchical log linear 

regression analyses adjusted 

for town, sex, ethnicity, 

maternal education, HOME score

Anxiety/depression 

assessed using 

Child Behavior 

Checklist at 3 yr

Concurrent: 1.45 (0.04, 2.86)

Prenatal: 1.16 (0.02, 2.3)

Leviton et al. (1993) 1,923 children followed 

from birth (1979-1980) to 

age 8 yr

Boston area, MA

Prenatal blood 2nd 

quartile: 4.8-6.3

Early childhood (tooth) 

second quartile: 2.0-2.9 

μg/g

Log linear regression model 

adjusted for single-parent 

family, gestational age <37 

wk, mother not a college 

graduate, self-identification 

as black, only child, daycare 

during first 3 yr

Hyperactivity 

assessed using 

Boston Teacher 

Questionnaire at 

age 8 yr

Prenatal, girls: 0.26 (-0.69, 1.13

Early childhood, girls: 

0.10 (-0.92, 1.1)

Bellinger et al.

(1994a)

1,782 children followed 

from birth (1979-1980) to 

age 8 yr

Boston area, MA

Prenatal (cord blood) 

mean (SD): 6.8 (3.1)

Early childhood (tooth) 

mean (SD): 3.4 (2.4) 

ppm

Log linear regression 

analyses adjusted for 

prepregnant weight, race, 

delivery by cesarean section, 

marital status, paternal and 

maternal education, sex, birth 

weight, maternal smoking, 

prenatal care beginning after 

the first trimester, recipient of 

public assistance, number of 

children in family, child 

currently on medication

Problem behaviors 

(t-scores) assessed 

using Teacher 

Report Form of the 

Child Behavior 

Profile at age 8 yr

Early childhood: 

1.8 (0.49, 3.1)

Prenatal: -0.31 (-1.7, 1.07)

Ris et al. (2004) 195 children in followed 

from birth (1979-1985) to 

age 15-17 yr

Cincinnati, OH

NR Linear regression model 

adjusted for maternal IQ, sex, 

and average total HOME 

score

Inattention 

composite 

assessed using 

Continuous 

Performance Test

at 15-17 yr

Prenatal: 0.16 (0.04, 0.27)

Early childhood, 6.5 yr: 

0.12 (0.02, 0.22)

Early childhood avg: 

0.11 (0.03, 0.19)

Dietrich et al. (2001) 195 children followed from 

birth (born1979-1985) to 

age 

15-17 yr

Cincinnati, OH

NR Linear regression model 

adjusted for birth weight, 

HOME score, SES, parental 

IQ

Parental report of 

delinquent behavior

at 15-17 yr 

Prenatal: 0.19 (0.02, 0.37)

Early childhood, 6.5 yr: 

0.13 (-0.01, 0.27)

Early childhood avg: 

0.09 (-0.02, 0.20)

MDI = Mental Developmental Index, ETS = Environmental tobacco smoke, HOME = Home Observation for Measurement of the Environment, GCI = 

General Cognitive Index, FWS = Filtered Word Test, KABC = Kaufman Assessment Battery of Children, FSIQ = Full-scale IQ, WISC = Wechsler Intelligence 

Scale for Children, NR = Not reported

a

Studies are organized by age of neurodevelopmental assessment. Effect estimates are presented in order of increasing magnitude, with statistically 

significant results in bold.

b

Effect estimates are standardized to a 1 μg/dL increase in blood Pb level in analyses of blood Pb as a continuous variable.

c

Effect estimate represent comparisons between children in different categories of blood Pb level, with children in the lower blood Pb category serving as 

the reference group.

d

Sufficient data were not provided in order to calculate 95% CI.

1 Prenatal and neonatal (10 days after birth) blood Pb levels also were associated with 

2 cognitive function and behavioral outcomes in children examined at school-age (ages 

3 4-17 years) (Table 5-13). Concurrent blood Pb levels generally were estimated to have 

4 similar or larger magnitudes of effect (Wasserman et al., 1998; Wasserman et al., 1994; 
February 2012 5-195 Draft – Do Not Cite or Quote

1 Dietrich et al., 1993a; Dietrich et al., 1993b; Bellinger et al., 1992; Dietrich et al., 1992).

2 Studies conducted in the Cincinnati cohort examined diverse neurodevelopmental effects 

3 and found that prenatal and neonatal blood Pb levels were associated with impairments in 

4 behavior and auditory processing (Ris et al., 2004; Dietrich et al., 2001; Dietrich et al., 

5 1992) but not cognitive function or motor function in children at age 6 years or in 

6 adolescence (Bhattacharya et al., 2006; Bhattacharya et al., 1995; Dietrich et al., 1993a; 

7 Dietrich et al., 1993b). These findings suggest that the effects of prenatal Pb exposure 

8 may vary among different populations and specific endpoints.

9 Early childhood blood Pb levels were associated with diverse neurodevelopmental effects 

10 assessed later in childhood and into early adulthood in both recent (Chandramouli et al., 

11 2009; Min et al., 2009; Miranda et al., 2009) and previous studies that did not compare

12 various lifestages of Pb exposure (Yuan et al., 2006; Cecil et al., 2005; Tong et al., 2000).

13 In a meta-analysis of results from five cohort studies (Pocock et al., 1994), a larger 

14 decrease in FSIQ was estimated for an increase in peak (around age 2 years) blood Pb 

15 level than for blood Pb level measured around birth or after age 2 years (Table 5-13). 

16 This lag effect may be the result of a toxicological process in which some period of time 

17 is required for past Pb exposure to affect CNS function. Alternatively, associations with 

18 early childhood or peak blood Pb levels may reflect the greater reliability of 

19 neurodevelopmental assessments at later ages when the processes modalities of children 

20 are more highly differentiated. Early testing may lead to false negative results and fail to 

21 identify a child who is at risk for later neurodevelopmental dysfunction. Further, due to 

22 the correlation of blood Pb levels over time, it is difficult to assess whether early 

23 childhood or peak blood Pb levels were serving as surrogates of concurrent or cumulative 

24 blood Pb levels.

25 As presented in Table 5-13, several studies of school-aged children estimated larger 

26 blood Pb-associated decreases in neurodevelopmental function for concurrent or lifetime 

27 average (range: 5 to 13 years) blood Pb levels than for blood Pb levels at other lifestages. 

28 These findings were substantiated in the analysis pooling data from seven prospective 

29 studies, in which concurrent, peak, average lifetime (5- to 10-year average), and age 

30 2 year blood Pb levels all were negatively associated with IQ measured between ages 5 

31 and 10 years, with the larger magnitude of decrease associated with increases in 

32 concurrent and peak blood Pb levels (Lanphear et al., 2005). Childhood average blood Pb 

33 levels (Lanphear et al., 2005; Dietrich et al., 1993a; Dietrich et al., 1993b) and deciduous 

34 tooth Pb levels (Bellinger et al., 1994a) have been associated with neurodevelopmental 

35 effects. While the ages of children varied among these studies as did the years over which 

36 blood Pb levels were averaged, the results nonetheless indicate cumulative Pb exposure 

37 over multiple years may contribute to neurodevelopmental effects in children. 

38 Associations with concurrent blood Pb level also were demonstrated consistently in 
February 2012 5-196 Draft – Do Not Cite or Quote

1 studies without comparisons to blood Pb levels during other lifestages (Figures and 

2 Tables in Sections 5.3.2.1 and 5.3.3.1).

3 Some studies have aimed to improve the characterization of important lifestages of Pb 

4 exposure by examining children in whom blood Pb levels are weakly correlated over time 

5 (i.e., children whose blood Pb level ranking changed over time) (Hornung et al., 2009; 

6 Schnaas et al., 2006; Chen et al., 2005; Tong et al., 1998; Bellinger et al., 1990).

7 Collectively, the results did not conclusively demonstrate stronger findings for concurrent 

8 blood Pb level. However, with the exception of Schnaas et al. (2006), the results did not 

9 preclude an association with concurrent blood Pb level. Schnaas et al. (2006) followed 

10 children in Mexico City prenatally through age 10 years and found prenatal blood Pb 

11 levels (maternal blood at 28-36 weeks of pregnancy) to be weakly correlated with 

12 repeated measures of blood Pb between ages 1 and 10 years (Pearson r ≤ 0.25). In a 

13 mixed effects model that included prenatal and multiple postnatal blood Pb measures, 

14 only prenatal blood Pb level was associated with a decrement in FSIQ (Table 5-13). 

15 Analysis of variance inflation factors indicated a lack of collinearity among the serial 

16 blood Pb measures.

17 Tong et al. (1998) found that higher early-life blood Pb level was associated with a larger 

18 deficit in IQ (Figure 5-21 and Table 5-14). As part of the Port Pirie, Australia cohort 

19 study, investigators separately examined intellectual attainment in groups of children 

20 with different degrees of decline in blood Pb levels between ages 2 and 11-13 years. 

21 Although the mean blood Pb level in the study population declined overall from 

22 21.2 μg/dL at age 2 years to 7.9 μg/dL at age 11-13 years, the magnitude of decline 

23 varied among children. In comparisons of tertiles of change in blood Pb level between 

24 age 2 and 11-13 years, investigators found that FSIQ at ages 2, 4, 7, and 11-13 years did 

25 not significantly differ between children with the largest declines (>16 μg/dL) in blood 

26 Pb level and children with a smaller decline (<10 μg/dL). These findings indicated an 

27 influence of higher blood Pb levels early in life despite declines in blood Pb with age and 

28 a persistence of Pb effects. The results do not preclude an independent association with 

29 concurrent blood Pb level.

30 In several different cohorts of U.S. children, larger decrements in cognitive function were 

31 estimated for concurrent blood Pb levels with consideration of blood Pb levels measured 

32 earlier in childhood (Hornung et al., 2009; Chen et al., 2005; Bellinger et al., 1990)

33 (Figure 5-21 and Table 5-14). In the Boston cohort, Bellinger et al. (1990) found that at 

34 age 57 months, FSIQ, as assessed by McCarthy GCI, was similar between children with 

35 higher (≥ 10 μg/dL) and lower (<3 μg/dL) prenatal cord blood Pb levels. Additionally, 

36 higher concurrent blood Pb level (age 57 months) was associated with the largest decline 

37 in GCI score over time (score at age 57 months-score at age 24 months) among children 
February 2012 5-197 Draft – Do Not Cite or Quote

1 with high prenatal blood Pb levels (≥ 10 μg/dL), which pointed to a larger decrease in 

2 FSIQ in children with both high early and concurrent blood Pb levels (Figure 5-21 and 

3 Table 5-14). The findings indicated that by age 5 years, children with higher prenatal 

4 blood Pb levels appeared to recover the Pb-associated decrements in cognitive function 

5 unless concurrent blood Pb levels remained high. The investigators also demonstrated 

6 that positive home and caregiving environment (e.g., HOME score >52, higher SES, 

7 higher maternal IQ) may also protect against decrements in cognitive function associated 

8 with higher postnatal Pb exposures. Collectively, these results suggest that cognitive 

9 development is not fixed early in childhood and can be affected negatively or positively 

10 by postnatal influences.

a

Values represent the ratio of blood Pb level at age 6 years to that at age 2 years.

b

Results represent the decrease in FSIQ per 1 μg/dL increase in concurrent blood Pb level in children in different groups of prenatal

cord blood Pb levels. FSIQ scores were standardized to their standard deviation. Effect estimates in blue represent associations for 

higher prenatal or early childhood blood Pb levels relative to concurrent blood Pb levels.

Note: Effect estimates represent associations between concurrent blood Pb level and cognitive function (standardized to standard 

deviation) in children. Studies are presented in order of increasing prenatal level. 

Figure 5-21 Associations of cognitive function in children with different 

degrees of changes in blood Pb levels over time.

Study

Tong et al. (1998)

Bellinger et al. (1990)

Chen et al. (2005)

Hornung et al. (2009)

Age of FSIQ 

Assessment

11-13 yr

24 and 57 mo

7 yr.

6 yr.

age 2 to 11-13

age 2 to 11-13

age 2 to 11-13

prenatal < 3b

prenatal 3-10b

prenatal ≥ 10b

Low 7 yr (< 7.2)

High 7 yr (> 7.2)

Low 7 yr (< 7.2)

High 7 yr (> 7.2)

Blood Pb (μg/dL) 

variable examined

< 10.2 decline

10.2-16.2 decline

> 16.2 decline

Concurrent, 

Concurrent, 

Concurrent, 

Low 2 yr (< 24.9), 

Low 2 yr (< 24.9), 

High 2 yr. (> 24.9), 

High 2 yr. (> 24.9), 

0.5 ratio age 6 to 2 yra

2.0 ratio age 6 to 2 yra

-1 -0.5 0

Change in FSIQ (95% CI)
February 2012 5-198 Draft – Do Not Cite or Quote

Table 5-14 Additional characteristics and quantitative results for studies 

presented in Figure 5-21

Study

Population/

Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Outcome

Effect Estimate

(95% CI)a

Hornung et 

al. (2009)

462 children followed 

from birth 

(1979-1984) to age 6 

yr

Rochester, NY and 

Cincinnati, OH

Geometric mean 

(5th-95th):

Peak: 13.6 

(4.6-34.4)

Early childhood: 

8.9 (3.0-23.8)

Lifetime mean: 8.5 

(3.0-22.1)

Concurrent: 6.0 

(1.9-17.9)

Linear regression model adjusted for 

city, HOME score, birth weight, 

maternal IQ, maternal education

FSIQ assessed using 

WISC-R at age 6 yr

0.5 ratio of blood Pb level at age 

6 to age 2: 0 (reference)

2.0 ratio of blood Pb level at age 

6 to age 2 yr: -0.70 (-1.0, -0.40)

Bellinger et 

al. (1990)

170 children followed 

prenatally to age 57 

mo

Boston area, MA

NR Log linear regression adjusted for 

HOME score, social class, maternal 

IQ, maternal age, sex, ethnicity

Change in McCarthy GCI 

score (z-score) between 

age 57 and 24 mo

For concurrent blood Pb levelb

Prenatal <3 μg/dL: -0.16 

(-0.43, 0.11)

Prenatal 3-10 μg/dL: -0.14 

(-0.57, 0.29)

Prenatal ≥ 10 μg/dL: -0.46 

(-0.81, -0.11)

Tong et al.

(1998)

375 children followed 

from birth 

(1979-1982) to age 

11-13 yr

Port Pirie, Australia

Means: 21.2 (age 

2 yr), 7.9 (age 

11-13 yr)

Log linear regression model adjusted 

for sex, birth weight, birth rank, feeding 

style, breastfeeding duration, maternal 

IQ, maternal age, SES, HOME score, 

parental smoking, parents living 

together.

ANOVA to assess association of 

change in IQ with change in blood Pb 

across time intervals

Change in cognitive 

function (z-scores) using 

Bayley MDI at age 2 yr, 

McCarthy GCI at age 4 yr, 

WISC-R at ages 7, and 

11-13 yr

<10.2 μg/dL decline: 0.03 

(-0.15, 0.21)c

10.2-16.2 μg/dL decline: 0.04 

(-0.15, 0.23)c

>16.2 μg/dL decline: -0.01 

(-0.20, 0.18)c

Chen et al.

(2005)

780 children 

participating in the 

TLC trial from age 

12-33 mo to age 7 yr

Baltimore, MD; 

Cincinnati, OH; 

Newark, NJ; 

Philadelphia, PA

Children underwent 

chelation therapy

Mean (SD):

Age 2 yr: 26.2 

(5.1)

Age 5 yr: 12.0 

(5.2)

Age 7 yr: 8.0 (4.0)

Low age 2 yr: 

<24.9

Low age 7 yr: < 

6.2

Linear regression model adjusted for 

city, race, sex, language, parental 

education, parental employment, single 

parent, age at blood Pb measurement, 

caregiver IQ

WISC-III at age 7 yr Low age 2, Low age 7: 0d

Low age 2, High age 7: -0.27 

(-0.48, -0.05)

High age 2, Low age 7: 0 

(-0.21, 0.20)

High age 2, High age 7: -0.28 

(-0.47, -0.10)

a

Effect estimates represent the cognitive function score or change in score over time standardized to its standard deviation.

b

Effects are estimated for concurrent blood Pb level (continuous variable) in children in different categories of prenatal blood Pb level: <3 μg/dL, 3-10 μg/dL, 

and ≥ 10 μg/dL.

c

Investigators estimated changes in IQ in groups of children with different degrees of decline in blood Pb levels over the study period: children with 

<10.2 μg/dL decline, children with a 10.2-16.2 μg/dL decline, and children with >16.2 μg/dL. d

Investigators compared IQs among children with different categories of blood Pb level early and later in childhood: low levels at age 2 (<11.4 μg/dL) and 

age 7 (<7.2 μg/dL), low levels at age 2 (<11.4 μg/dL) and high levels at age 7 (>7.2 μg/dL), high levels at age 2 (>11.4 μg/dL) and low levels at age 7 

(<7.2 μg/dL), and high levels at age 2 (>11.4 μg/dL) and age 7 (>7.2 μg/dL). Cutoffs were based on the median blood Pb levels.

1 Pooling the Cincinnati and Rochester cohorts (n = 397), Hornung et al. (2009) created a 

2 new indicator of Pb exposure: the ratio of blood Pb level at 6 years of age to that at 

3 2 years of age. The greatest decrease in cognitive and behavioral development was 

4 observed for children with blood Pb ratios greater than 1 (indicating an increase in blood 

5 Pb level from 2 to 6 years of age) (Figure 5-21, Figure 5-22, and Table 5-14). Presumably 

6 areas under the curve would be similar among children with blood Pb level ratios of 1, 
February 2012 5-199 Draft – Do Not Cite or Quote

1 greater than 1, and less than 1, indicating that cumulative blood Pb levels would not be 

2 predictive. 

3 As part of the multicenter TLC, Chen et al. (2005) also found higher concurrent blood Pb 

4 level (≥ median 7.2 μg/dL) to be associated with lower IQ at age 7 years, regardless of 

5 whether blood Pb level at age 2 years was low or high (less than or greater than the 

6 median of 24.9 μg/dL, respectively). Blood Pb levels at ages 2 and 7 years were weakly 

7 correlated (r = 0.27). It is important to note that children participating in TLC had 

8 undergone chelation therapy due to high blood Pb levels (20-44 μg/dL) at age 12 to 

9 33 months, and the findings may have limited application to the general population of 

10 children currently living in the U.S. In fact, in all of the studies that examined weakly 

11 correlated serial blood Pb measurements, blood Pb levels were higher than those 

12 currently measured in U.S. children. Additionally, in several study populations, children 

13 experienced large decreases in blood Pb levels over time. It is unclear whether these 

14 findings would apply to children in the U.S. who currently are within the same age range 

15 and who would be expected to have smaller decreases in blood Pb levels over time.

Source: Hornung et al. (2009)

Note: All three patterns have an identical mean blood Pb level of 10 μg/dL.

Figure 5-22 Estimated IQ in combined Cincinnati and Rochester cohorts, for 

three patterns of blood Pb level levels from 1 through 6 years of 

age: peak at 2 years (blue diamonds), peak at 5 years (black 

triangles), and constant blood Pb level (white squares).
February 2012 5-200 Draft – Do Not Cite or Quote

1 To conclude, in the collective body of epidemiologic evidence in children, it is difficult to 

2 ascertain which lifestage of Pb exposure is associated with the greatest risk of Pb3 associated neurodevelopmental effects. Associations have been observed with prenatal, 

4 early-childhood, lifetime average, and concurrent blood Pb levels as well as with 

5 childhood tooth Pb levels. Comparisons among different lifestages of exposure is 

6 complicated further by the fact that blood Pb levels in children, although highly affected 

7 by recent exposure, are also influenced by Pb stored in bone due to rapid growth-related 

8 bone turnover in children relative to adults. Thus, concurrent blood Pb level in children 

9 also may reflect body burden (Section 4.3.4.6). Nonetheless, while the evidence indicates 

10 that prenatal and early-childhood blood Pb levels are associated with neurodevelopmental 

11 deficits, subsequent exposures that are reflected in concurrent, cumulative blood Pb 

12 levels or tooth Pb levels also are demonstrated to contribute to neurodevelopmental 

13 deficits throughout school-age and into adolescence. With additional results from studies 

14 described in Sections 5.3.2.1 and 5.3.3.1, the weight of epidemiologic evidence supports 

15 associations of concurrent blood Pb level with neurodevelopmental effects in children. 

16 These findings are consistent with the understanding that the nervous system continues to 

17 develop throughout childhood.

5.3.10 Examination of the Lead Concentration-Response Relationship

18 With each successive Pb AQCD and supplement, epidemiologic and toxicological studies 

19 find that progressively lower blood Pb levels are associated with cognitive deficits and 

20 behavioral impairments. For example, among children, such effects were observed in 

21 association with blood Pb levels in the range of 10-15 μg/dL in the 1986 Addendum and 

22 1990 Supplement and 10 μg/dL and lower in the 2006 Pb AQCD (U.S. EPA, 2006b).

23 Furthermore, in the 2006 Pb AQCD, several individual studies, pooled analyses, and 

24 meta-analyses estimated a supralinear blood Pb concentration-response relationship in 

25 children, i.e., greater decrements in cognitive function per incremental increase in blood 

26 Pb level among children in lower strata of blood Pb levels compared with children in 

27 higher strata of blood Pb levels (Figure 5-23 and Table 5-15). As lower concentrations of 

28 Pb exposure are being used experimentally, the toxicological literature also has reported 

29 nonlinear concentration-response relationships for various endpoints, including those 

30 coherent with cognitive function decrements observed in children. Also consistent with 

31 the epidemiologic literature, some toxicological studies have shown larger magnitudes of 

32 effect (absolute effects) in lower Pb exposure groups (relative to control groups) than in 

33 the higher exposure groups.

34 In epidemiologic studies, a supralinear concentration-response relationship was observed 

35 for concurrent, early childhood, and lifetime average blood Pb levels. Most 
February 2012 5-201 Draft – Do Not Cite or Quote

1 epidemiologic studies used a blood Pb level of 10 μg/dL to define lower and higher blood 

2 Pb levels. Several of these observations were made in groups of children aged 2 to 

3 10 years, and the mean blood Pb levels in the lower strata of blood Pb levels were in the 

4 range of 3-5 μg/dL (Bellinger, 2008; Canfield, 2008; Hornung, 2008; Rojo-Tellez, 2008).

5 Except for the pooled analysis, lower strata of blood Pb levels comprised at least 55% of 

6 the study population and consequently the concentration-response relationships 

7 calculated for these lower strata likely are not outliers comprised a minimum 55% of the 

8 study population and consequently the concentration-response relationships calculated for 

9 these lower strata are not outliers or unrepresentative of the overall study population. 

10 Using data pooled from seven prospective studies, Lanphear et al. (2005) fit various types 

11 of models to the data and observed that a cubic spline, log-linear model, and piece-wise 

12 linear model all supported a more negative concentration-response relationship at 

13 concurrent blood Pb levels <10 μg/dL. A linear model was found to be inadequate as the 

14 polynomial terms for concurrent blood Pb were statistically significant. These findings 

15 were corroborated in a separate analysis by Rothenberg and Rothenberg (2005) who 

16 found that the log-linear model fit the relationship between blood Pb level and IQ better 

17 than a linear model did.

18 A few studies demonstrated larger Pb-associated decreases in cognitive function with 

19 blood Pb levels < 5 μg/dL. Tellez-Rojo et al. (2006) estimated a larger decrement in IQ 

20 per unit increase in blood Pb level for children (age 2 years) with concurrent blood Pb 

21 levels <5 μg/dL compared with children with levels 5-10 μg/dL, and > 10 μg/dL (Figure 

22 5-23 and Table 5-15). In an NHANES analysis 1989-1994, Lanphear et al. (2000) found 

23 larger decrements in reading and math skills and memory per unit increase in blood Pb

24 level in children with concurrent blood Pb levels <2.5 μg/dL compared with children 

25 with levels <5 μg/dL, <7,5 μg/dL, <10 μg/dL, and all subjects. However, as the study 

26 population included adolescents, the oldest of whom were 16 years of age and born 

27 1972-1978, higher Pb exposures earlier in childhood may have contributed to 

28 associations.
February 2012 5-202 Draft – Do Not Cite or Quote

Note: Studies are presented in order of increasing mean blood Pb level. Strata refer to peak blood Pb level measured in child at any 

point during follow up. FSIQ = full-scale IQ, MDI = mental development index. Effect estimates are standardized to a 1 μg/dL 

increase in blood Pb level. Black symbols represent effect estimates among all subjects or in the highest blood Pb stratum. Blue 

symbols represent effect estimates in lower blood Pb strata

Figure 5-23 Comparison of associations between blood Pb level and cognitive 

function among various blood Pb strata.

Study

Lanphear et al. (2000)

Bellinger and Needleman

(2003)

Canfield et al. (2003)

Tellez-Rojo et al. (2006)

Jusko (2011)

Kordas et al. (2006)

Lanphear et al. (2005)

Schwartz (1994)

Blood Pb Timing

Concurrent

Peak

Lifetime avg

Concurrent

Peak

Concurrent

Concurrent

Early childhood

Outcome

Reading score

FSIQ

FSIQ

Bayley MDI

FSIQ

Math score

FSIQ

FSIQ

Blood Pb

stratum (μg/dL)

All

<10

<7.5

<5

<2.5

>10

<10

All

<10a

≥10

<10

5-10

<5

All

< 10

all

<10

≥10a

<10a

≥7.5a

<7.5a

>15

≤15

◄

◄

◄

►

Sample size

4853

4681

4526

4043

2467

NR

NR

172

101

90

294

101

193

174

96

532

293

1089

244

1230

103

NR

NR

-4 -3 -2 -1 0 1

Change in Cognitive Function Score per 1 μg/dL

increase in blood Pb level (95% CI)
February 2012 5-203 Draft – Do Not Cite or Quote

Table 5-15 Additional characteristics and quantitative results for studies 

presented in Figure 5-23

Study Population/Location

Blood Pb Levels 

(μg/dL) Statistical Analysis Outcome

Blood Pb 

stratum

(μg/dL)

Effect Estimate

(95% CI)a

Lanphear et al.

(2000)

4,853 children ages 6-16 

yr

NHANES 1988-1994

Concurrent mean 

(SE): 1.9 (0.1)

Linear regression model 

adjusted for sex, race/ethnicity, 

poverty index ratio, reference 

adult education level, serum 

ferritin level, serum cotinine 

level

WRAT reading 

subtest at ages 6-16 

yr

All subjects

<10

<7.5

<5

<2.5

-0.70 (-1.03, -0.37)

-0.89 (-1.52, -0.26)

-1.06 (-1.82, -0.30)

-1.06 (-2.00, -0.12)

-1.28 (-3.20, -0.64)

Tellez-Rojo et 

al. (2006)

294 children followed 

from birth (1994-1995, 

1997-1999) to age 2 yr

Mexico City, Mexico

Concurrent (age 2 

yr) mean (SD): 4.28 

(2.25)

Linear regression model 

adjusted for sex, birth weight, 

maternal IQ

Bayley MDI at age 2 

yr

≥ 10

<10

5-10

<5

0.07 (-10, 9.2) b

-1.04 (-1.8, -0.30) b

-0.94 (-2.1, 0.2) b

-1.71 (-3.0, -0.42) b

Bellinger et al.

(1992)

Bellinger and 

Needleman

(2003)

148 children followed 

from birth (1979-1981) to 

age 10 yr

Boston area, MA

Early childhood 

(age 2 yr) mean 

(SD): 6.5 (4.9)

Linear regression model 

adjusted for HOME score (age 

10 and 5), child stress, race, 

maternal IQ, SES, sex, birth 

order, marital status

WISC-R at age 10 yr >10

<10

-0.58 (-1.0, -0.2)b

-1.56 (-2.9, -0.2)b

Canfield et al.

(2003a)

172 children born 

1994-1995 followed from 

infancy to age 3-5 yr

Rochester, NY

Lifetime avg (3 or 5 

yr) mean (SD): 7.4 

(4.3)

Mixed effects models adjusted 

for sex, maternal race, parental 

smoking, child iron status, 

maternal income, maternal IQ, 

HOME score

Stanford-Binet at age 

3 or 5 yr

All

<10

-0.46 (-0.76, -0.15)

-1.37 (-2.56, -0.17)

Lanphear et al.

(2005)

1,333 children pooled 

from Boston, Cincinnati, 

Cleveland, Mexico City, 

Port Pirie, Rochester, 

and Yugoslavia cohorts

Concurrent

Median (5th-95th)

9.7 (2.5-33.2)

Linear regression model 

adjusted for HOME score, birth 

weight, maternal IQ, maternal 

education

FSIQ measured at 

ages 4.8-10 yr

≥ 10

<10

≥ 7.5

<7.5

-0.13 (-2.3, -0.03)

-0.80 (-1.74, -0.14)

-0.16 (-2.4, -0.08)

-2.94 (-5.16, -0.71)

Jusko et al.

(2011)

194 children in 

Rochester, NY followed 

from age 6 mo 

(1994-1995) to age 6 yr.

Peak

Mean (SD): 11.4 

(7.3)

Linear regression model 

adjusted for sex, birth weight, 

transferrin saturation, maternal 

race, maternal IQ, maternal 

education, HOME score, family 

income, and maternal prenatal 

smoking

WPPSI-R at age 6 yr All

< 10

-0.19 (-0.46, 0.07)

-1.66 (-3.1, -0.23)

Kordas et al.

(2006)

602 children in 1st grade

Torreon, Mexico

Concurrent mean 

(SD): 11.4 (6.1)

Linear regression model 

adjusted for sex, age, 

hemoglobin, family 

possessions, forgetting 

homework, house ownership, 

crowding, maternal education, 

birth order, family structure, 

arsenic exposure, tester, 

school

Math achievement 

test in 1st grade

all

<10

-0.17 (-0.28, -0.06)

-0.42 (-0.92, 0.08)

Schwartz

(1994)

Meta-analysis of 7 

studies with sample sizes 

75-579 children

Early childhood (2-3 

yr) range in study 

means: 6.5-23

Meta-analysis of combining 

effect estimates from individual 

studies

FSIQ measured at 

school-age

Studies with 

mean >15

Studies with 

mean ≤ 15

-2.32 (-3.10, -1.54)

-3.23 (-5.70, -0.76)

a

Effect estimates are standardized to a 1 μg/dL increase in blood Pb level. b

95% CIs calculated from reported p-value.

1 Several studies also found a nonlinear blood Pb-cognitive function relationship in 

2 nonparametric regression analyses using lowess with smoothing parameters or using 

3 splines (Min et al., 2009; Jusko et al., 2008; Schnaas et al., 2006). Min et al. (2009)

4 performed a formal test of nonlinearity using nonparametric regression by testing the 

5 statistical significance of a restricted cubic spline term for blood Pb level. Although the 
February 2012 5-204 Draft – Do Not Cite or Quote

1 term was not statistically significant, the concurrent blood Pb level-FSIQ curve appeared 

2 to be more negative at blood Pb levels < 7 μg/dL. In a similar analysis in a Mexico City 

3 cohort, Schnaas et al. (2006) found a more negative blood Pb-FSIQ slope at concurrent 

4 blood Pb levels < 6 μg/dL and found a test of nonlinearity to be statistically significant. 

5 Among children in India ages 3-7 years, Palaniappan et al. (2011), most associations of 

6 concurrent blood Pb level with various indices of executive function in nonparametric 

7 regression analyses were found to be linear.

8 Studies of adults have not assessed widely whether the relationship between blood or 

9 bone Pb level and cognitive performance is described better with a linear or nonlinear 

10 function. In the various NHANES analyses, only log-linear models were used to fit the 

11 data (Krieg et al., 2010; Krieg and Butler, 2009; Krieg et al., 2009). Nonlinearity in the 

12 BMS and NAS cohorts was examined with the use of quadratic terms, penalized splines, 

13 or visual inspection of bivariate plots (Bandeen-Roche et al., 2009; Weisskopf et al., 

14 2007a; Shih et al., 2006). There was some evidence for nonlinearity for some (Figure 

15 5-12 and Figure 5-13) but not all cognitive tests or all subjects in the NAS cohort. For 

16 example, Wang et al. (2007a) found that among NAS men with an HFE variant, there 

17 was a steeper Pb-associated decline in MMSE score at higher tibia Pb levels (20-25 μg/g, 

18 Figure 5-13). In the BMS cohort, observations of a statistically nonsignificant quadratic 

19 term (Shih et al., 2006) or spline (Bandeen-Roche et al., 2009) for tibia Pb indicated that 

20 a linear model adequately fit the relationship between tibia Pb level and various tests of 

21 cognitive performance.

22 Attenuation of the concentration-response relationships at higher exposure or dose levels 

23 has been reported in the occupational literature, and explanations have included greater 

24 exposure measurement error, competing risks, and saturation of biological mechanisms at 

25 higher levels; larger proportions of at-risk populations at lower exposure levels; and 

26 variations in other risk factors among exposure levels (Stayner et al., 2003). Other 

27 explanations for nonlinearity include differential activity of mechanisms at different 

28 exposure levels, confounding by omitted or misspecified variables, and the lower 

29 incremental effect of Pb due to covarying risk factors such as low SES, poor caregiving 

30 environment, and higher exposure to other environmental factors.

31 The contribution of these factors to the supralinear relationship between blood Pb levels 

32 and cognitive function in children has not been examined widely in epidemiologic studies 

33 to date. However, in several different populations, higher blood Pb levels have been 

34 measured in potentially at-risk groups such as those with higher poverty, greater exposure 

35 to tobacco smoke, lower parental education, and lower birth weight, which argues against 

36 a larger proportion of at-risk populations at lower blood Pb levels (Lanphear et al., 2005; 

37 Lanphear et al., 2000). It has been suggested that in populations of low SES, poorer 
February 2012 5-205 Draft – Do Not Cite or Quote

1 caregiving environment, and greater social stress, the incremental effect of Pb exposure 

2 may be attenuated due to the overwhelmingly larger effects of these other risk factors

3 (Schwartz, 1994). Several studies found statistically significant associations of these 

4 sociodemographic risk factors with neurocognitive deficits, and Miranda et al. (2009)

5 found that indicators of SES (i.e., parental education and enrollment in a free/reduced fee 

6 lunch program) accounted for larger decrements in EOG scores than did blood Pb level 

7 (Figure 5-7). Few studies have compared blood Pb level effect estimates among groups in 

8 different sociodemographic strata, and the limited data are mixed. Greater Pb-associated 

9 neurocognitive deficits were reported in low-SES groups by Bellinger et al. (1990). In a 

10 meta-analysis of eight studies, Schwartz (1994) found a smaller decrement in IQ per 

11 1 μg/dL increase in blood Pb level for studies in disadvantaged populations (-2.7 points 

12 [95% CI: -5.3, -0.07]) than for studies in advantaged populations (-4.5 points [95% CI: 

13 -5.6, -2.8]). It is important to note that blood Pb level is associated with deficits in 

14 cognitive function in both higher and lower SES groups; however, it is unclear what 

15 differences there are between groups in the decrement per unit increase in blood Pb and 

16 whether these differences can explain the observed nonlinear concentration-response 

17 relationship.

18 Rothenberg and Rothenberg (2005) formally assessed the influence of residual 

19 confounding on the nonlinear blood Pb-FSIQ concentration-response relationship by 

20 comparing model fit between linear and spline transformations (df = 2) of covariates such 

21 as maternal IQ, HOME score, and maternal education. Inclusion of covariates as spline 

22 functions did not significantly improve model fit either with a linear blood Pb term or log 

23 blood Pb term, which indicated that their inclusion as linear functions was adequate. 

24 These findings demonstrate that the improved model fit with log-specification of blood 

25 Pb level was not due to residual confounding by covariates.

26 Bowers and Beck (2006) postulated that a supralinear slope necessarily will be found 

27 when modeling a relationship between a log-normally distributed variable and a normally 

28 distributed variable. However, as discussed in the 2006 Pb AQCD, this modeling strategy 

29 was not employed in the epidemiologic analyses showing a supralinear concentration30 response function. IQ scores generally were not forced into a normal distribution. Four of 

31 the seven studies included in the pooled analysis by Lanphear et al. (2005) did not use 

32 normalized IQ scores, and scores were not normalized in the pooled analysis (Hornung et 

33 al., 2006). Further, a log-linear model (a linear relationship between IQ and the log of 

34 blood Pb) provided the best fit of the pooled data.

35 In support of the nonlinear associations between blood Pb levels and cognitive function 

36 observed in children, toxicological studies provided some evidence of nonlinear 

37 relationships between Pb exposure and effects related to impaired learning and memory 
February 2012 5-206 Draft – Do Not Cite or Quote

1 in animals. Multiple studies showed U- or inverse U-shaped curves with lower exposures 

2 of Pb having different or often the opposite effect from higher exposures. Coherent with 

3 epidemiologic findings, results summarized across multiple studies in multiple species 

4 demonstrated that lower Pb exposures increased FI response rates relative to controls and 

5 higher Pb exposures decreased FI response rates (Cory-Slechta, 1994). Consistent with 

6 these findings, Rossi-George et al. (2011) found that 50 ppm gestational plus lactational 

7 Pb exposure when combined with stress increased FI responses of 2-month old rats 

8 whereas 150 ppm Pb exposure with stress did not affect FI responses. It is important to 

9 note that the larger effects observed with lower Pb exposures were less consistently 

10 observed with longer duration exposures (e.g., 8-11 months) (Rossi-George et al., 2011; 

11 Cory-Slechta, 1990). As discussed in Section 5.3.2.2, FI responses measure learning and 

12 memory by reflecting the ability of animals to learn to respond to one schedule of 

13 reinforcement and change responses according to a change in reinforcement.

14 Toxicological studies provided additional support for nonlinear relationships between Pb 

15 exposure and neurodevelopmental effects by finding that lower and higher Pb exposures 

16 differentially activate mechanisms underlying such effects. Gilbert et al. (1999) found 

17 reduced LTP with Pb exposures ranging between 0.1 and 0.5% but not 1.0%. LTP is one 

18 indication of synaptic plasticity (Section 5.3.8.4) and is considered to contribute to 

19 learning and memory. Likewise, glutamate release in the hippocampus was reduced in 

20 animals with blood Pb levels 27-40 μg/dL but not with blood Pb levels of 62-117 μg/dL

21 (Lasley and Gilbert, 2002). Glutaminergic neurotransmission via its NMDA receptor has 

22 been implicated in learning and memory (Section 5.3.8.8).

23 Dopaminergic neurotransmission is involved in many CNS processes including 

24 cognition, behavior, and motor function. The shape of the Pb-DA concentration-response 

25 relationship varied among toxicological studies. Some studies found that lower Pb 

26 exposures (~ 50 ppm) did not affect or increased DA activity relative to controls and 

27 higher Pb exposure (109-250 ppm) (Leasure et al., 2008; Virgolini et al., 2005; Lewis and 

28 Pitts, 2004). However, compared with control and lower Pb exposures, higher Pb 

29 exposures (109 or 150 ppm) were found both to increase and impair DA activity (Leasure 

30 et al., 2008; Virgolini et al., 2005). These differential responses of DA may be related to 

31 the diverse actions of DA in different regions of the brain and on a range of CNS effects. 

32 For example, the increased dopamine turnover with 50 ppm Pb exposure may explain the 

33 greater spontaneous and amphetamine-induced motor activity in males induced by 

34 50 ppm GLE (Leasure et al., 2008).

35 Lower and higher Pb exposures also were found to differentially affect calcineurin 

36 enzyme activity; activity was inhibited by higher Pb exposure and stimulated by lower Pb 

37 exposure (Kern and Audesirk, 2000). While calcineurin activity has been found to 
February 2012 5-207 Draft – Do Not Cite or Quote

1 modulate learning, LTP, and behavior in animals, studies have found lower calcineurin 

2 activity to be associated with both impaired and improved effects related to learning

3 (Zeng et al., 2001). Thus, it is uncertain whether altered calcineurin activity contributes to 

4 the nonlinear relationships observed between Pb exposure and learning. Nonetheless, 

5 results indicated that lower and higher Pb exposures may have different modes of action. 

6 At lower concentrations, Pb may displace calcium at its binding sites on calmodulin and 

7 by acting as a calmodulin agonist at the catalytic A subunit of calcineurin, stimulate 

8 calcineurin activity. At higher Pb exposure, Pb may bind directly to a separate calcium9 binding B subunit, override the calmodulin-dependent effect and turn off the activity of 

10 calcineurin. Similarly, Lasley and Gilbert (2002) also indicated that at higher 

11 concentrations (i.e., 1%), Pb stimulated gluatamate release by acting as a calcium 

12 mimetic.

13 While epidemiologic studies have not examined widely the shape of the concentration14 response relationship for other nervous system effects, toxicological studies have found 

15 nonlinear relationships for diverse outcomes. U-shaped Pb concentration-response 

16 relationships were found for rotarod performance, amphetamine-induced motor activity, 

17 and latency to fall from rotarod (Leasure et al., 2008). Inverted U-shaped Pb 

18 concentration-response relationships were found for histological parameters such as the 

19 numbers of rod photoreceptors and bipolar cells, activity level, and adult body weight

20 (Leasure et al., 2008) as well as ERG wave amplitudes (Fox et al., 2008) and 

21 hippocampal neurogenesis (Fox et al., 2008; Gilbert et al., 2005). Additional evidence 

22 points to differences in hormonal homeostasis by Pb exposure level. In male mice with 

23 chronic Pb exposure (PND21-9 months of age), basal corticosterone levels were 

24 significantly lower in the 50 ppm exposure group than in the control or 150 ppm Pb 

25 exposure group.

26 Sensory organ functions in animals also have shown to be differentially affected by lower 

27 versus higher Pb exposure (developmental Pb exposurefrom gestation to PND10, pup 

28 blood Pb levels 12, 24, and 46 μg/dL. Inverted U-shaped dose-response curves have been 

29 observed for rod photoreceptor numbers or neurogenesis (Giddabasappa et al., 2011) and 

30 retinal thickness (Fox et al., 2010). Thus, these dichotomous histological findings may 

31 give insight to the complex sensory organ findings that vary by exposure window and 

32 exposure dose (Section 5.3.4.3).

33 The supralinear concentration-response relationship widely documented for Pb is 

34 consistent with the lack of a threshold for Pb-associated neurodevelopmental effects as a 

35 smaller effect estimate would be expected at lower blood Pb levels if a threshold existed. 

36 Schwartz (1994) explicitly assessed evidence for a threshold in data from the Boston 

37 prospective cohort by regressing FSIQ and blood Pb level on potential confounders 
February 2012 5-208 Draft – Do Not Cite or Quote

1 including age, race, maternal IQ, SES, and HOME score and fitting a nonparametric 

2 smoothed curve to the residuals of both regression models (variation in FSIQ or blood Pb 

3 level not explained by covariates). A 7-point decrease in IQ was observed over the range 

4 of blood Pb residuals below 0, which corresponds to the mean blood Pb level in the study 

5 (6.5 μg/dL). Thus, in the Boston study, the association between blood Pb level and FSIQ 

6 was clearly demonstrated at blood Pb levels below 5 μg/dL.

7 An important limitation of previous studies in terms of characterizing the concentration8 response relationship, in particular, identifying whether a threshold exists, was the limited 

9 examination of associations in populations or subgroups with blood Pb levels more 

10 comparable to the current U.S. population mean. While Schwartz (1994) did not find 

11 evidence for a threshold in the Boston study data, the mean blood Pb in that population 

12 was 6.5 μg/dL, and 56% of subjects had a blood Pb level >5 μg/dL. Recent studies 

13 indicate a downward shift in the distribution of blood Pb levels. In the various NHANES 

14 analyses of children, a large proportion of children had blood Pb levels <1 μg/dL, for 

15 example, 50% of subjects in the 2001-2004 NHANES population (Braun et al., 2008).

16 More sensitive quantification methods have improved the detection limits, for example, 

17 from 0.6 μg/dL in 1999-2002 NHANES to 0.025 μg/dL in 2003-2004 NHANES. In 

18 analyses of children in multiple blood Pb quantiles below 1 μg/dL, Braun et al. (2008)

19 found higher odds ratios for conduct disorder and ADHD among children with blood Pb 

20 levels 0.8-1.0 μg/dL (2nd quartile) compared with children with blood Pb levels 0.2-

21 0.7 μg/dL (1st quartile). Despite the availability of large proportions of subjects at blood 

22 Pb levels below 1 μg/dL, the ability to discern a threshold for Pb-associated 

23 neurodevelopmental effects is limited due to the large proportions of adolescents in 

24 NHANES analyses who were born the 1970s and whose higher past exposures may have 

25 contributed to associations observed with concurrent blood Pb levels. Nonetheless, 

26 several recent studies reported associations between blood Pb levels and deficits in 

27 cognitive and behavioral endpoints in children ages 8-11 years with mean or quantile 

28 blood Pb levels <2 μg/dL (Cho et al., 2010; Kim et al., 2009b; Miranda et al., 2009). In 

29 comparisons of various quantiles of blood Pb, Miranda et al. (2009) reported lower EOG 

30 scores in children in North Carolina with blood Pb levels of 2 μg/dL compared with 

31 children with blood Pb levels of 1 μg/dL. Collectively, these new findings in children, do 

32 not provide evidence for a threshold for the neurodevelopmental effects of Pb in the 

33 range of blood Pb levels examined to date.

34 It is important to note, however, that the lack of a reference population with blood Pb 

35 levels reflecting pre-industrial Pb exposures limits the ability to identify a threshold. 

36 Estimates of “background” blood Pb levels have been garnered from the analysis of 

37 ancient bones in pre-industrialized societies. These studies suggest that the level of Pb in 
February 2012 5-209 Draft – Do Not Cite or Quote

1 blood in preindustrial humans was approximately 0.016 μg/dL (Flegal and Smith, 1992),

2 approximately 65-fold lower than that currently measured in U.S. populations and lower 

3 than the levels at which neurodevelopmental effects have been observed (1 μg/dL). Thus, 

4 the current evidence does not preclude the possibility of a threshold for 

5 neurodevelopmental effects in children existing in the large range of blood levels 

6 between 1 μg/dL and preindustrial “background” levels.

7 To conclude, several studies found a supralinear blood Pb-cognitive function 

8 concentration-response relationship in children based on comparisons of effect estimates 

9 in lower and higher strata of blood Pb level and nonparametric regression. Supporting 

10 evidence was provided by some toxicological studies that showed that lower Pb 

11 exposures induced learning and memory impairments in animals compared to control 

12 exposures or higher Pb exposures. While explanations for this supralinear relationship 

13 have not been well characterized by epidemiologic studies, recent toxicological studies 

14 that examined lower concentrations of experimental Pb exposure suggest that 

15 mechanisms may be differentially activated at lower and higher Pb exposures. 

16 Nonlinear concentration-response relationships were found for effects such as LTP, 

17 hippocampal glutamate release, and calcineurin expression. Observations of associations 

18 between blood Pb levels and deficits in cognitive and behavioral endpoints in children 

19 (ages 8-11 years) with mean or range of blood Pb levels ≤ 2 μg/dL (Cho et al., 2010; Kim 

20 et al., 2009b; Miranda et al., 2009) do not provide evidence for a threshold for 

21 neurodevelopmental effects of Pb in the range of blood Pb levels examined to date.

5.3.11 Confounding in Epidemiologic Studies of Nervous System Effects

22 In addition to Pb exposure, many factors influence cognitive function and behavior in 

23 children, including parental IQ and education, SES of the family, quality of the 

24 caregiving environment, and other environmental exposures (Wasserman and Factor25 Litvak, 2001). These other risk factors often are correlated with blood, tooth, and bone Pb 

26 levels, thus, a major challenge to observational studies examining associations of Pb 

27 biomarker levels with cognitive and behavioral function in children has been the 

28 assessment and control for potential confounding factors. By definition, a confounder is 

29 associated with both the exposure and the outcome and consequently has the potential to 

30 bias the association between the exposure of interest and the outcome. Epidemiologic 

31 studies of Pb biomarkers in children have most commonly examined potential 

32 confounding by quality of the caregiving environment (i.e., HOME score), parental IQ, 

33 and SES-related variables such as parental education, household income, and the 

34 Hollingshead Four-Factor Index of Social Position, which incorporates education and 
February 2012 5-210 Draft – Do Not Cite or Quote

1 income of both parents. A relatively smaller number of studies have considered 

2 confounding by other environmental exposures, measures of parental substance abuse, 

3 and psychopathology. Studies have varied with respect to the number of potential 

4 confounders examined, with some studies considering multiple SES-related variables and 

5 other studies focusing on a smaller set. It is important to note that the extent of 

6 confounding by a particular variable likely is study-specific, i.e., dependent on the 

7 population examined. Thus, the impact of adjustment for specific covariates on the Pb 

8 effect estimate also is likely to be study-specific.

9 Studies have used various methods to control for confounding, including examining a 

10 homogeneous population with respect to SES, examining populations in which factors are 

11 not correlated, conducting multivariate regression, characterizing the magnitude of 

12 change in the blood Pb level effect estimate with adjustment for a covariate, and 

13 examining associations in different strata of a covariate. While no single method is 

14 without limitation, the consistency of findings among different methodologies 

15 substantiates independent associations of blood Pb level with cognitive function 

16 decrements and behavioral problems in children. The evidence derived from each of 

17 these control strategies is discussed below.

18 In the Boston Prospective Study, potential confounding by SES was largely controlled for 

19 by study design. The study subjects were generally middle- to upper-middle-class 

20 children with married, college-educated parents. Hence, the potential for confounding by 

21 SES in this study was considerably less compared to other studies examining the same 

22 associations. Yet, in this cohort, blood Pb levels measured prenatally, at age 2 years, and 

23 integrated over ages 2 and 5 years were associated decrements in full-scale IQ and 

24 various measures of executive function (Stiles and Bellinger, 1993); (Bellinger et al., 

25 1992; Bellinger et al., 1990). In some analyses of this cohort, larger effects were 

26 estimated as compared with other studies.

27 Studies also have demonstrated associations between blood Pb levels and cognitive 

28 function in populations in which blood Pb levels were not associated with SES-related 

29 variables (Factor-Litvak et al., 1999; Bellinger et al., 1987). In the Boston cohort, 

30 parental education, social class, and HOME score were similar among low (<3 μg/dL), 

31 medium (6-7 μg/dL), and high (≥ 10 μg/dL) cord blood Pb level groups. Nonetheless, 

32 adjusting for these and other demographic variables, Bellinger et al. (1987) found that 

33 children in the high cord blood Pb group had a 4.8-point lower Bayley MDI score at age 

34 24 months than did children in the low cord blood Pb group. In the Yugoslavia cohort, 

35 blood Pb levels at age 4 years were higher in groups with higher maternal education, 

36 maternal IQ, and HOME score in one city and were lower in another city. Among all 

37 children, higher blood Pb level was associated with lower FSIQ and specific indices of 
February 2012 5-211 Draft – Do Not Cite or Quote

1 learning and memory and with greater depression and withdrawn behavior (Factor-Litvak 

2 et al., 1999).

3 The primary method used by epidemiologic studies to control for potential confounding 

4 has been multivariate regression. This was the main method employed in recent studies 

5 that examined children with blood Pb levels in the range of current U.S. levels. Some 

6 studies selected a set of covariates based on a priori knowledge, whereas others selected 

7 particular covariates based on their association with the outcome in a model with all 

8 potential covariates and/or a greater than 10% change in the blood Pb level effect 

9 estimate. Studies also varied in the number of variables included in models as potential 

10 confounders. Some included multiple SES-related variables, whereas others analyzed one 

11 or two variables. Regardless of the method used to select model covariates or the number 

12 of covariates included, studies consistently demonstrated associations of higher blood Pb 

13 level with lower cognitive function and greater behavioral problems. This consistency 

14 suggests that confounding by particular variables may be specific to the population 

15 examined and that no single measured variable or set of variables fully accounts for the 

16 associations observed with blood Pb levels and neurodevelopmental effects in children.

17 The consistency of associations across populations with different SES and across studies 

18 examining different covariates was reinforced in pooled and meta-analyses (Marcus et 

19 al., 2010; Lanphear et al., 2005; Schwartz, 1994). Pooling data from seven international 

20 longitudinal cohorts, Lanphear et al. (2005) found similar blood Pb-FSIQ effect estimates 

21 (-2.6 to 8.6% difference) among models, each with one study omitted. These results 

22 indicated that the pooled estimate is relatively stable despite between-study differences in 

23 population characteristics, including SES. In a meta-analysis, Schwartz (1994) found that 

24 individual study blood Pb-IQ effect estimates fell within a relatively narrow range despite 

25 large differences among studies in the correlation between blood Pb level and SES. A 

26 wider range of effect estimates would be expected if omitted SES factors confounded the 

27 blood Pb level association. A recent meta-analysis examined the association between 

28 blood Pb level and conduct problems in older and recent studies of children (Marcus et 

29 al., 2010). Among the studies included, adjustment for variables such as SES and home 

30 environment did little to attenuate the association between Pb and conduct problems. 

31 Among studies that provided both unadjusted and adjusted effect estimates, most 

32 indicated statistically significant associations of blood Pb level with neurodevelopmental 

33 outcomes in children before and after adjusting for potential confounders. Although 

34 examples exist where blood Pb level was estimated to have a weaker, statistically 

35 nonsignificant effect after adjustment for potential confounders (Tong and Lu, 2001; 

36 Ernhart et al., 1989), most notably in multiple analyses of the Cleveland cohort, studies 

37 also reported stronger blood Pb level associations in covariate-adjusted models (Canfield 
February 2012 5-212 Draft – Do Not Cite or Quote

1 et al., 2003b; Factor-Litvak et al., 1999). In recent studies with population mean blood Pb 

2 levels between 5 and 10 μg/dL, associations between blood Pb levels and cognitive 

3 function (e.g., FSIQ, executive function, learning, and memory) remained statistically 

4 significant after adjusting for covariates (Min et al., 2009; Chiodo et al., 2007; Froehlich 

5 et al., 2007; Kordas et al., 2006; Schnaas et al., 2006). Although most effect estimates 

6 changed by 20-50% in multivariate models, they remained within the 95% CI of the 

7 unadjusted estimate. In particular, Schnaas et al. (2006) and Froehlich et al. (2007) found 

8 some adjusted effect estimates to be larger in magnitude compared with unadjusted 

9 estimates. Although information was not provided on the magnitude of change in effect 

10 estimates after adjustment for confounding, Chiodo et al. (2007) found statistically 

11 significant associations between blood Pb level and behavioral problems 

12 (e.g., inattention, hyperactivity, social problems, impulsivity) in models with blood Pb 

13 level alone and with covariates. These findings demonstrate that SES-related and 

14 demographic factors may partially account for but do not fully explain the associations 

15 observed between higher blood Pb level and neurodevelopmental impairments. 

16 A challenge to separating the effects of Pb exposure from those related to SES and 

17 quality of the caregiving environment is the high correlation typically observed among 

18 these measures. In such cases, it is difficult to know how much variation in the outcome 

19 to attribute to each of the various risk factors (Needleman and Bellinger, 2001). For 

20 example, the high correlation between blood Pb level and SES may lead to an 

21 underestimation of the Pb effect when SES is added to the model. A reduction in the 

22 magnitude and statistical significance of the Pb effect estimate following adjustment for 

23 some measure of SES may be the result of the misattribution of the variance in outcome 

24 due to Pb to the variance due to SES. SES may be a proxy for Pb exposure rather than a 

25 confounder of the association of interest. This misattribution may be exacerbated when 

26 several correlated variables are included in the same model (i.e., overcontrol).

27 Instead of being a confounder, Pb may be on the causal pathway of the association 

28 between social class and IQ. Lower social class in urban children is closely linked to 

29 residence in older housing in poor condition that, in turn, increases exposure of children 

30 to environmental Pb and increases their risk of cognitive deficits (Clark et al., 1985). In 

31 such cases, statistical adjustment for SES will lead to the underestimation of the Pb effect

32 (Bellinger, 2004a). One extreme example of overcontrol of this nature can be found in the 

33 New Zealand studies where investigators regularly adjusted for residence in older 

34 wooden housing, which is associated with higher exposure to Pb paint and accumulated 

35 dust and soil (Fergusson et al., 1988a, b). However, it is worth noting that, even in the 

36 models including this variable, Pb remained a statistically significant predictor of 

37 intellectual and academic under-attainment in the Christchurch Health Study. Variables 

38 related to SES have been shown to be effect modifiers in studies of Pb and child 
February 2012 5-213 Draft – Do Not Cite or Quote

1 development (Tong et al., 2000; Bellinger et al., 1990). That is, increases in blood Pb 

2 level have been associated with larger decreases in cognitive function among children 

3 from less advantaged (e.g., lower SES, lower HOME score, lower maternal IQ) 

4 household than among those from more advantaged households. Similarly, a larger 

5 decrease in cognitive function in association with higher blood Pb level was found among 

6 children with mothers with lower self-esteem, an indicator of stress and less connection 

7 with the child (Surkan et al., 2008).

8 In the collective body of epidemiologic evidence, higher blood Pb levels are consistently 

9 associated with cognitive function decrements and behavioral problems in children, with 

10 the weight of evidence indicating decrements in FSIQ and diverse measures of executive 

11 function and inattention and hyperactivity, respectively. These associations have been 

12 observed in diverse populations in the U.S., Mexico, Europe, Asia, and Australia. 

13 Associations have been observed across studies that use different methods of control for 

14 confounding and adjust for different potential confounders. While no single method is 

15 without limitation, the consistency of findings among different methodologies and sets of 

16 covariates substantiates independent associations of blood Pb level with cognitive 

17 function decrements and behavioral problems in children. In addition, Pb exposure has 

18 been extensively studied in animals that produce blood Pb levels in the range of those 

19 examined in children. Experimental animal studies are not vulnerable to confounding by 

20 such factors as social class and correlated environmental factors. Adding further support 

21 for the independent associations of blood Pb levels with neurodevelopmental outcomes in 

22 children is the coherence of findings in animal studies for Pb-induced impairments in 

23 tests of learning, inattention, and impulsivity, especially tests that are directly 

24 homologous to those in children, i.e., spatial memory, rule learning and reversal, and 

25 response inhibition. Further, an extensive body of toxicological evidence for Pb-induced 

26 changes in brain physiological processes that mediate cognition and behavior, including 

27 changes in neurogenesis, synaptic pruning, and neurotransmitter function in the frontal 

28 lobe and striatum of the brain provides biological plausibility for observations in 

29 children.

5.3.12 Public Health Significance of Associations between Lead Biomarkers 

and Neurodevelopmental Effects

30 As described in Section 5.3.2.1, most studies found that a 1 μg/dL increase in blood Pb 

31 level was associated with fractional decrements in FSIQ in school-aged children (Figure 

32 5-2 and Table 5-3) and Bayley MDI in infants (Table 5-4). Similarly, a 1 μg/dL increase 

33 in blood Pb level typically was associated with decreases in specific cognitive abilities 

34 (Figure 5-5 and Table 5-5) and increases in inattention and hyperactivity (Section 5.3.3.1, 
February 2012 5-214 Draft – Do Not Cite or Quote

1 Figure 5-14 and Table 5-9) on the order of less than 1 standard deviation. These findings 

2 have generated discussion of the public significance of blood Pb level-associated changes 

3 in cognitive function and behavior in children, specifically, whether these fractional 

4 changes have consequences on the health and life-success of individuals. The World 

5 Health Organization definition of health is the following: “Health is a state of complete 

6 physical, mental and social well-being and not merely the absence of disease or 

7 infirmity” (WHO, 1948). By this definition, even decrements in health status that are not 

8 severe enough to meet diagnostic criteria might be undesirable if they reflect a decrement 

9 in the well-being of an individual. Deficits in health indices or life-success may not be 

10 observable except in aggregate, at the population level. The American Thoracic Society 

11 discussed the need to consider the prevalence of exposures in the population and 

12 exposure to other risk factors in evaluating whether shifts in the population-level risk are 

13 adverse (ATS, 2000). It should also be noted that these deficits when measured in 

14 children may set affected children on trajectories more prone toward lower educational 

15 attainment and financial well-being. Thus, early deficits in children may have lifetime 

16 consequences.

17 It has been argued that fractional decrements in IQ points are meaningless given that 

18 these Pb-associated changes are within the 3- to 4-point standard error on a single test 

19 (i.e., the statistic that defines the range within which the true value of an individual is 

20 likely to lie) (Kaufman, 2001). However, this argument incorrectly assumes that 

21 conclusions drawn from individual-level data apply to populations. It is important to note 

22 that evidence does not indicate that the standard error is nonrandom, i.e., biased in one 

23 direction. In particular, evidence has not indicated that children with higher blood Pb 

24 levels systematically test lower than their true IQ value and that children with lower 

25 blood Pb levels test higher than their true IQ value. Thus, in a population of children, on a 

26 given assessment, some children will test lower than their true value and others will test 

27 higher than their true value. In such cases, between-group differences will be 

28 measureable on a population basis. Error in the measurement of IQ in an individual will 

29 contribute nondifferential error on a population-level and bias the association to the null. 

30 The issue of individual-level versus population-level risk also pertains to the relevance of 

31 the magnitude of decrease in cognitive function or increase in behavioral problems per 

32 incremental increase in blood Pb level. Although fractional changes in IQ, memory, or 

33 inattention may not be consequential for an individual, they may be consequential on a 

34 population level, especially in the two tails of the distribution (Bellinger, 2007, 2004b).

35 Weiss (1990) predicted, on purely statistical grounds, that a downward shift of five points 

36 in mean IQ, if the amount of dispersion in the distribution remained the same, should be 

37 accompanied by a doubling of the numbers of individuals with scores two or more 

38 standard deviations below the mean and a reduction by half of the number of individuals 
February 2012 5-215 Draft – Do Not Cite or Quote

1 with scores two or more standard deviations above the mean. Thus, for an individual 

2 functioning in the low range of the IQ distribution, a Pb-associated decline of several 

3 points might be sufficient to drop that individual into the range associated with increase 

4 risk of educational, vocational, and social failure. 

5 This hypothesis is supported by several observations. In a recent study of fourth graders 

6 across the entire state of North Carolina, Miranda et al. (2009) found that higher blood Pb 

7 level measured once in each child between age 9 months and 3 years was associated with 

8 larger decreases in EOG scores in lower distribution of EOG scores (Figure 5-7). 

9 Needleman et al. (1982) found that a downward shift in the mean IQ value was associated 

10 not only with a substantial increase in the percentage of individuals achieving very low 

11 scores, but also with substantial decreases in percentages achieving very high scores. 

12 Based on the study by Bellinger et al. (1987) examining associations between blood Pb 

13 level and IQ scores in children, Weiss (1988) discussed the shift in the population 

14 distribution of IQ from a mean of 100 and a standard deviation of 15 to a mean of 95, a 

15 5% reduction. When the mean IQ level is 100, 2.3% of the individuals in a given 

16 population would score above 130. However, with the population distribution shift and 

17 the resulting mean decline in IQ, only 0.99% of the individuals would score above 130. 

18 Weiss (1988) stated that the implication of such a loss transcends the current 

19 circumscribed definitions of risk. In a similar analysis presented in the 2006 Pb AQCD, 

20 using a blood Pb-IQ effect estimate of −0.9 points/μg/dL (based on the median of effect 

21 estimates for blood Pb levels <10 μg/dL), the fraction of the population with an IQ level 

22 less than 80 more than doubles from 9% with a blood Pb level of 0 μg/dL to 23% with a 

23 blood Pb level of 10 μg/dL (Figure 5-24). The proportion with an IQ level below 70, a 

24 level often requiring community support to live (WHO, 1992) increases from a little over 

25 2% with a blood Pb level of 0 μg/dL to about 8% with a blood Pb level of 10 μg/dL 

26 (Figure 5-24) (U.S. EPA, 2006b).
February 2012 5-216 Draft – Do Not Cite or Quote

Source: U.S. EPA (2006) Pb AQCD

Figure 5-24 Effect of blood Pb level on the proportion of the population with 

IQ levels <70 and <80 points.

1 Evidence also has demonstrated larger changes in neurodevelopmental outcomes in 

2 children with additional risk factors such as lower SES (Tong et al., 2000; Bellinger et 

3 al., 1990), mothers with low self-esteem (Surkan et al., 2008), or co-exposures to 

4 manganese (Kim et al., 2009a). Moreover, interventions that shift the population mean, in 

5 a beneficial direction, by an amount that is without clinical consequence for an individual 

6 have been shown to produce substantial decreases in the percentage of individuals with 

7 values that are clinically significant (Bellinger, 2007, 2004b).

8 Also supporting the public health significance of blood Pb level-associated changes in 

9 cognitive function and behavior are observations within the same cohorts that higher 

10 blood Pb level are associated with decrements in IQ and measures of executive function 

11 earlier in age and with lower academic performance, antisocial behavior, or delinquent 

12 behavior assessed later in adolescence or in early adulthood (Chandramouli et al., 2009; 

13 Wright et al., 2008; Kordas et al., 2006; Canfield et al., 2004; Kordas et al., 2004; 

14 Canfield et al., 2003b; Dietrich et al., 2001; Dietrich et al., 1993a; Bellinger et al., 1992).
February 2012 5-217 Draft – Do Not Cite or Quote

1 Thus, the blood Pb-associated deficits measured in children may set affected children on

2 trajectories more prone toward lower educational attainment and life success. Studies also 

3 found that higher blood Pb level is associated with measures of inattention and 

4 hyperactivity as rated by teachers or parents and by objective tests and with ADHD 

5 diagnosis or diagnostic indices (Nicolescu et al., 2010; Roy et al., 2009a; Nigg et al., 

6 2008). These findings demonstrate that associations of blood Pb level with small changes 

7 in a health index can be markers or indicators of other changes that are likely to have 

8 occurred whose significance is more certain.

5.3.13 Summary and Causal Determination

9 Recent epidemiologic and toxicological studies substantiated the strong body of evidence 

10 presented in the 2006 Pb AQCD for the role of Pb in nervous system effects in two 

11 domains: cognitive function and behavior in children. Specifically, new studies of 

12 cognitive function focused on indices of learning, memory, and measures of executive 

13 function in children consistently showed Pb-related impairments. Coherence for these 

14 findings in children was provided by extensive toxicological evidence for Pb-induced 

15 impairments in homologous tests of learning and memory in juvenile rats and monkeys. 

16 New epidemiologic studies of behavior focused on and found evidence for Pb-related 

17 effects on inattention and impulsivity in children. These findings were supported by a 

18 large historical evidence base in juvenile animals. Consistent with these observations, 

19 new evidence demonstrated associations with ADHD in children. Recent studies of adults 

20 without current occupational Pb exposures continued to find associations between bone 

21 Pb levels, a biomarker of cumulative Pb exposure, and poorer cognitive function. 

22 Additional toxicological evidence for Pb-induced inhibition of neurotransmitter release, 

23 decline in synaptic plasticity, and decreases in the magnitude of LTP strengthened the 

24 biological plausibility for Pb exposure effects on decrements in both cognitive function 

25 and behavior in children. This section presents a summary of the collective body of 

26 evidence and identifies the new insights provided by recent studies.

5.3.13.1 Cognitive Function in Children

27 Epidemiologic studies provide robust evidence for higher blood Pb levels being 

28 associated with lower FSIQ in children ages 3-11 years (Figure 5-2 and Table 5-3), with 

29 the strongest evidence demonstrated by the consistency of association in prospective 

30 studies in diverse populations (e.g., varying distributions of blood Pb levels, SES, 

31 parental intelligence, and quality of caregiving) and the persistence of associations after 

32 adjustment for potential confounding by SES, parental intelligence, caregiving 
February 2012 5-218 Draft – Do Not Cite or Quote

1 environment, and other environmental exposures. Recent epidemiologic studies in 

2 children focused on and found associations of concurrent blood Pb levels with specific 

3 indices of cognitive function such as reading and verbal skills, memory, learning, and 

4 visuospatial processing. Several new studies shifted the weight of evidence for 

5 associations with cognitive performance to lower blood Pb levels (primarily concurrent), 

6 with populations means in the range of 2-7 μg/dL (2011; Kim et al., 2009b; Min et al., 

7 2009; Miranda et al., 2009; Zailina et al., 2008; Chiodo et al., 2007). Evidence clearly 

8 indicates that prenatal cord and concurrent blood Pb levels are associated with lower 

9 cognitive function in younger children ages 6 months to 3 years (Table 5-4). Coherence 

10 for findings in children is derived from extensive evidence in animals that gestational and 

11 early postnatal Pb exposures resulted in impaired learning and memory, in particular, in 

12 homologous tests of spatial memory and rule learning and reversal. Reflecting the 

13 tendency to examine higher Pb exposures, toxicological studies most clearly 

14 demonstrated neurodevelopmental effects with blood Pb levels of 20-40 μg/dL in 

15 animals. However, several new studies added to the evidence for impaired learning and 

16 memory in animals with lower blood Pb levels, 8-17 μg/dL (Cory-Slechta et al., 2010; Li 

17 et al., 2009c; Niu et al., 2009; Virgolini et al., 2008a; Stangle et al., 2007).

18 The large body of evidence indicating associations between blood Pb levels and 

19 decrements in the diverse set of indices related to learning, memory, and other executive 

20 functions provides coherence with findings for FSIQ, a global measure of cognitive 

21 function that reflects the integration of these individual domains. Further, evidence for 

22 effects on various diverse measures of cognitive function provides biological plausibility 

23 for associations observed between blood Pb levels and factors that may be indicators of 

24 life success, including the level of educational attainment and academic performance.

5.3.13.2 Behavior in Children

25 Epidemiologic studies in children demonstrate associations of higher blood Pb levels 

26 with a range of behavioral problems, with the weight of evidence demonstrating 

27 associations with inattention and hyperactivity as rated by parents or teachers and as 

28 assessed using objective neuropsychological tests. Previous studies found associations 

29 with early childhood blood or tooth Pb level (i.e., ages 2-6 years), and recent studies 

30 expanded evidence to include associations with concurrent blood Pb level. Recent 

31 epidemiologic studies consistently found associations with inattention and hyperactivity 

32 in children ages 1 to 12 years with mean concurrent blood Pb levels of 2 to 5 μg/dL (Cho 

33 et al., 2010; Nicolescu et al., 2010; Plusquellec et al., 2010; Chiodo et al., 2007; 

34 Plusquellec et al., 2007), similar to those associated with cognitive function decrements. 

35 The epidemiologic findings are strengthened by observations in animals of Pb-induced 
February 2012 5-219 Draft – Do Not Cite or Quote

1 inattention and impulsivity in homologous tests of response inhibition. Such effects in 

2 animals are most clearly indicated with gestational and early postnatal Pb exposures that 

3 result in blood Pb levels of 10 to 40 μg/dL. Whereas previous evidence was inconsistent, 

4 several recent epidemiologic studies indicate associations between higher concurrent 

5 blood Pb level and higher prevalence or incidence of ADHD diagnosis and its 

6 contributing diagnostic indices in children ages 8–17 years (Cho et al., 2010; Nicolescu et 

7 al., 2010; Roy et al., 2009a; Nigg et al., 2008; Braun et al., 2006). The biological 

8 plausibility for associations with ADHD is strongly supported by the large epidemiologic 

9 and toxicological evidence base demonstrating Pb-associated increases in inattention and 

10 impulsivity, both of which are primary symptoms of ADHD. A smaller but equally 

11 consistent body of evidence indicated associations of concurrent and early childhood 

12 blood Pb levels with social misconduct in children and delinquent behaviors in 

13 adolescents and young adults (Chandramouli et al., 2009; Braun et al., 2008; Wright et 

14 al., 2008; Chiodo et al., 2007). Associations of blood Pb levels with ADHD, misconduct, 

15 and delinquency were observed in populations of children with a wide range of blood Pb 

16 levels, 1 to 11 μg/dL, all similar in the strength of evidence.

17 While the different behavioral indices are examined, Pb exposure also was found to affect 

18 behavior (decreased ability to escape predators or capture prey) in aquatic and terrestrial 

19 species (Sections 7.2 and 7.3).

5.3.13.3 Other Nervous System Effects in Children

20 A few new toxicological studies augmented the evidence for Pb-related effects to the 

21 visual system by demonstrating retinal changes in male rodent offspring in association 

22 with lower blood Pb levels (<15 μg/dL) than previously examined (20 to > 100 μg/dL) 

23 (Section 5.3.4.3). A small body of epidemiologic evidence together with a large historical 

24 base of toxicological evidence indicated associations of Pb biomarkers or exposure with 

25 impaired auditory function. Associations were found in children ranging from 4 to 

26 19 years in age and with mean concurrent blood Pb levels of 7-12 μg/dL Section 5.3.4.1). 

27 While mood and emotional state have been examined less frequently compared with 

28 inattention and misconduct, several studies found associations of biomarkers of 

29 cumulative Pb exposure (i.e., tooth or childhood average blood Pb) and concurrent blood 

30 Pb levels with parental or teacher reports of withdrawn behavior or depression in children 

31 with mean blood Pb levels 8-28 μg/dL (Section 5.3.3.3). These findings in children are 

32 supported by a small body of toxicological studies in which prenatal plus lactational Pb 

33 exposure resulted in depression-like behavior in rodent models. Studies also reported 

34 associations of early childhood average and concurrent blood Pb levels with lower fine 

35 and gross motor function in children ages 3 to 17 years (Section 5.3.5). A common 
February 2012 5-220 Draft – Do Not Cite or Quote

1 observation across studies was finding that biomarkers of Pb exposure were associated 

2 with decrements in multiple neurodevelopmental outcomes, including cognitive function, 

3 externalizing behaviors, internalizing behaviors, and motor function, within the same 

4 population. These findings indicate that Pb exposure affects a broad spectrum of 

5 neurodevelopmental effects in children.

5.3.13.4 Factors that Modify Risk in Children

6 Several host and environmental factors were examined for their modification of 

7 associations between blood Pb levels and nervous system effects in children. Although 

8 each particular factor was examined in only one to two epidemiologic studies, it is 

9 important to note that several findings are supported by a larger base of toxicological 

10 evidence. Interactions of blood Pb levels with race/ethnicity and SES are not well 

11 characterized. Most investigation focused on sex-based differences. Cumulative 

12 epidemiologic evidence does not conclusively demonstrate increased risk of males or 

13 females for Pb-associated cognitive function decrements. Animal studies continue to 

14 demonstrate differential effects in males and females that vary depending on the 

15 endpoint. 

16 The weight of evidence from animal studies continues to support interactions between 

17 developmental Pb exposure and stress. New animal studies find a potentiating effect of 

18 stress, whereby lower concentration Pb exposures impact behavior and memory with co19 exposures to stress than with Pb exposures alone. In comparison, epidemiologic evidence 

20 for such interactions is sparse; however, a recent study indicated that among children 

21 with a positive social environment, as characterized by maternal self-esteem, blood Pb 

22 level is not associated with a decrease in cognitive function (Surkan et al., 2008). This 

23 finding was consistent with a previous study in rats, in which Pb-exposed animals reared 

24 in cages with enriched environments (i.e., toys) performed better in a test of spatial 

25 learning and memory than did their Pb-exposed littermates reared in traditional caging, 

26 Studies have examined modification of Pb-associated cognitive function by genes that 

27 affect Pb toxicokinetics and/or function in neurophysiological and neurochemical 

28 processes that mediate cognition. Most investigation has focused on ALAD variants but 

29 has not consistently found to be in a consistent direction. Inconsistencies also have been 

30 observed for VDR and dopamine receptor variants. In addition to host factors, recent 

31 studies suggested that associations between blood Pb levels and cognitive function in 

32 children are greater with co-exposures to environmental tobacco smoke (Froehlich et al., 

33 2007) and manganese (Claus Henn et al.; Kim et al., 2009b)..
February 2012 5-221 Draft – Do Not Cite or Quote

5.3.13.5 Nervous System Effects in Adults

1 Among adults, associations of blood Pb level with the spectrum of nervous system effects 

2 (e.g., impairments in memory, attention, mood, balance, motor function) were most 

3 consistently observed in occupationally-exposed adults with blood Pb levels ≥ 14 μg/dL. 

4 Evidence in adults without occupational Pb exposures is derived primarily from cross5 sectional analyses in a few different cohorts, thus the important magnitude, timing, 

6 frequency, and duration of Pb exposures contributing to the observed associations are 

7 uncertain. The weight of evidence demonstrated associations of bone Pb levels and not 

8 blood Pb levels with poorer cognitive performance, including declines in function over 

9 time (Bandeen-Roche et al., 2009; Weisskopf et al., 2007a). These findings point to a 

10 stronger effect of cumulative Pb exposure, including likely higher past exposures. One 

11 explanation for the overall weaker body of evidence in adults may be that cognitive 

12 reserve may compensate for the effects of Pb exposure on learning new information. 

13 Compensatory mechanisms may be overwhelmed with age, which may provide an 

14 explanation for more consistent associations observed for higher tibia Pb levels, 

15 representing higher long-term or cumulative Pb exposure.

16 Based on a smaller body of epidemiologic studies, blood and bone Pb levels were 

17 associated with essential tremor and PD, respectively, in adults (Section 5.3.7.1). 

18 However, in these case-control studies, it is difficult to establish temporality between Pb 

19 exposure and disease. Support for epidemiologic findings for PD is provided by 

20 toxicological evidence for Pb-induced decreased dopaminergic cell activity in the 

21 substantia nigra, which contributes to the primary symptoms of Parkinson’s disease. 

22 Whereas evidence for association with Alzheimer's Disease in adults is weak, 

23 developmentalbut not adult-only Pb exposures of monkeys (early postnatal, PND 1-400) 

24 and rats (lactational) has been shown to induce formation of amyloid plaques, pathology 

25 that underlies Alzheimer's Disease (Section 5.3.7.2). Thus, epidemiologic studies that 

26 examined concurrent bone or brain Pb levels or occupational Pb exposure may not have 

27 examined the etiologically relevant exposure period.

28 Rather than examining externalizing behaviors and criminal behavior, a small body of 

29 studies of behavior in adults examined and found associations of blood (Bouchard et al., 

30 2009) and tibia (Rajan et al., 2008) Pb levels with depression and anxiety symptoms. It is 

31 not surprising that Pb exposure may increase the risk of different nervous system 

32 endpoints in children and adults given the predominance of different neurophysiological 

33 processes operating at different ages, for example, neurogenesis and brain development 

34 in children and neurodegeneration in adults.
February 2012 5-222 Draft – Do Not Cite or Quote

5.3.13.6 Neurophysiological and Neurochemical Changes

1 Extensive evidence from toxicological studies clearly provides the coherence and 

2 biological plausibility for effects observed in epidemiologic and toxicological studies on 

3 cognitive function, behavior, mood, and neurodegenerative conditions by characterizing 

4 underlying mechanisms (Section 5.3.8). Dopamine plays a key role in cognitive functions 

5 mediated by the prefrontal cortex and also motor functions mediated by the substantia 

6 nigra. Thus, extensive evidence for Pb-induced dopaminergic changes in animals 

7 provides mechanistic support for associations in humans between blood Pb levels and 

8 cognitive deficits and in adults for associations with Parkinson’s Disease. Current 

9 toxicological research has been expanded to document that early-life Pb exposure can 

10 contribute to neurodegeneration and neurofibrillary tangle formation in the aged brains of 

11 animals. Pb has been shown to induce complex neurochemical changes in the brain that 

12 differ by region of the brain, neurotransmitter type, age and sex of the organism. These 

13 changes remain aberrant over time and are dynamic in nature. Pb exposure affects 

14 NMDA receptors, which may explain findings in animals and humans for Pb-associated 

15 symptoms of depression and withdrawn behavior. Recent toxicological studies continue 

16 to document Pb exposure effects on synapse formation, adhesion molecules, and 

17 nitrosative stress. A new study of epigenetics details that Pb exposure affects methylation 

18 patterns in rodent brains, which may provide a mechanism by which prenatal Pb exposure 

19 leads to impaired neurodevelopmental function later in life. Biological plausibility also is 

20 provided by a small body of evidence in young adults in which childhood blood Pb levels 

21 are associated with altered structure and activity in regions of the brain (assessed by MRI 

22 or MRS) that mediate cognitive processes and behavior.

5.3.13.7 Lifestages and Duration of Lead Exposure

23 Toxicological studies clearly demonstrate that in utero and early postnatal exposure to Pb 

24 results in impaired learning, memory, and behavior. This evidence is well supported by 

25 knowledge that processes such as neurogenesis, synaptogenesis, and synaptic pruning are 

26 most active during this developmental period. In epidemiologic studies reviewed in the 

27 2006 Pb AQCD, biomarkers of prenatal, early life, concurrent, and cumulative Pb 

28 exposures were associated with decrements in neurodevelopmental function in children, 

29 with no clear indication that blood Pb levels measured at a particular lifestage was more 

30 strongly associated with neurodevelopmental effects (Section 5.3.9). Distinguishing 

31 among the effects of Pb exposures at different lifestages is difficult in epidemiologic 

32 studies due to the high correlations among blood Pb levels within children over time. 

33 Recent studies in children primarily examined concurrent blood Pb levels but also found 

34 association with prenatal cord (Pilsner et al., 2010; Jedrychowski et al., 2009b; 
February 2012 5-223 Draft – Do Not Cite or Quote

1 Plusquellec et al., 2007) and early childhood blood Pb levels (Chandramouli et al., 2009; 

2 Min et al., 2009; Miranda et al., 2009). Thus, while biomarkers of Pb exposure across 

3 lifestages were associated with neurodevelopmental decrements in children, the weight of 

4 cumulative evidence demonstrated associations with concurrent blood Pb levels. In 

5 adults, the weight of evidence supported associations of cumulative Pb exposure 

6 (i.e., tibia Pb) with poorer cognitive performance.

7 With regards to duration of exposure, several epidemiologic studies indicated that Pb 

8 exposures of less than 1 year, represented by blood Pb levels prenatally (maternal or 

9 cord) and at age 6, 12, or 24 months, were associated with lower cognitive function in 

10 children ages 6 months to 3 years (Table 5-4). Toxicological studies supported these 

11 observations with effects observed with gestational and early postnatal Pb exposures.

5.3.13.8 The Lead Concentration-Response Relationship

12 In the 2006 Pb AQCD, several individual epidemiologic studies, pooled analyses, and 

13 meta-analyses estimated a supralinear blood Pb concentration-response relationship in 

14 children, i.e., greater decrements in cognitive function per incremental increase in blood 

15 Pb level or more negative slope among children in lower strata of blood Pb levels 

16 compared with children in higher strata of blood Pb levels (Figure 5-23 and Table 5-15).

17 Although a majority of epidemiologic evidence used a blood Pb level of 10 μg/dL to 

18 define lower and higher blood Pb levels, some found that among children less than age 

19 10 years with mean concurrent blood Pb levels of 3 μg/dL, the blood Pb-cognitive 

20 function slope was more negative with blood Pb levels <7.5 (Lanphear et al., 2005) or 

21 <5 μg/dL (Tellez-Rojo et al., 2006).

22 While explanations for this supralinear relationship have not been well characterized by 

23 epidemiologic studies, recent toxicological studies that examined lower concentrations of 

24 experimental Pb exposure suggest that mechanisms may be differentially activated at 

25 lower and higher Pb exposures. Multiple studies showed U- or inverse U-shaped curves 

26 with lower Pb exposures having different levels or often the opposite effect from higher 

27 exposures. Nonlinear concentration-response relationships were found for effects such as 

28 learning (Section 5.3.2.2), adult forebrain dopamine levels (Leasure et al., 2008), and 

29 neurogenesis (Fox et al., 2008). Sensory organ parameters in animals, namely, the 

30 numbers of rod photoreceptors and bipolar cells and ERG wave amplitudes also show 

31 vastly different changes with low versus higher Pb exposure (Table 5-12).

32 The examination of populations of children with large proportions of subjects at very low 

33 blood Pb levels has improved the ability to discern a threshold for Pb-associated effects 

34 on cognitive function and behavior. Several recent epidemiologic studies reported 
February 2012 5-224 Draft – Do Not Cite or Quote

1 associations between blood Pb levels and deficits in cognitive and behavioral endpoints 

2 in children (ages 8-11 years) with mean or range of blood Pb levels ≤ 2 μg/dL (Cho et al., 

3 2010; Kim et al., 2009b; Miranda et al., 2009). Collectively, these new findings in 

4 children do not provide evidence for a threshold for neurodevelopmental effects of Pb in 

5 the ranges of blood Pb levels examined to date.

5.3.13.9 Evidence that Forms the Basis of the Causal 

Determination

6 In summary, recent findings strengthen epidemiologic and toxicological evidence 

7 indicating that Pb exposure is associated with nervous system effects. The weight of 

8 epidemiologic and toxicological evidence clearly supports associations of higher blood 

9 Pb levels with lower cognitive function in children, i.e., full-scale IQ and various 

10 measures of learning and memory. In epidemiologic studies, these associations were 

11 substantiated in children ages 1 to 11 years and in populations with mean blood Pb levels 

12 between 2 and 7 μg/dL. Observation of a supralinear concentration-response relationship 

13 and associations with mean (or quantile) blood Pb levels < 2 μg/dL do not provide 

14 evidence for a threshold for the neurodevelopmental effects of Pb exposure. 

15 Epidemiologic and toxicological evidence clearly demonstrates Pb-associated increases 

16 in behavioral problems, in particular, inattention and impulsivity. Associations are 

17 substantiated in children ages 1 to 12 years with mean concurrent blood Pb levels of 2 to 

18 5 μg/dL. In animals, the weight of evidence demonstrates effects on cognition and 

19 behavior with prenatal and early postnatal Pb exposures that resulted in blood Pb levels 

20 of 10 to 40 μg/dL. In epidemiologic studies, associations with cognitive function and 

21 behavior were observed after adjustment for a range of potential confounding variables, 

22 but most commonly, parental IQ, parental education, and other SES-related variables. In 

23 children, the weight of evidence supports cognitive function decrements and behavioral 

24 problems in association with concurrent blood Pb levels. Associations also are observed 

25 with prenatal, early childhood, and childhood average blood Pb levels, thus uncertainty 

26 remains regarding the lifestage of exposure within childhood that is associated with the 

27 greatest risk. The weight of toxicological evidence demonstrates neurodevelopmental 

28 effects with prenatal and early postnatal Pb exposures that can have effects persisting to 

29 adulthood. The biological plausibility for epidemiologic and toxicological findings for 

30 effects on cognitive function and behavior is provided by evidence characterizing 

31 underlying mechanisms, including Pb-induced changes in neurogenesis, synaptogenesis 

32 and synaptic pruning, long term potentiation, and neurotransmitter function. In adults, the 

33 timing, level, frequency, and duration of Pb exposure implicated in nervous system 

34 effects remain uncertain. Among occupationally-exposed adults, a spectrum of nervous 

35 system effects is associated with concurrent blood Pb level (≥ 14 μg/dL), which reflects 
February 2012 5-225 Draft – Do Not Cite or Quote

1 both current and cumulative exposure. However, in adults without occupational exposure, 

2 cognitive performance is more strongly associated with tibia Pb levels than blood Pb 

3 levels, which indicates an effect of long-term, cumulative Pb exposures. Based most 

4 heavily on cognitive function decrements and inattention in children, the collective body 

5 of evidence integrated across epidemiologic and toxicological studies is sufficient to 

6 conclude that there is a causal relationship between Pb exposures and nervous system 

7 effects.

5.4 Cardiovascular Effects

5.4.1 Introduction

8 The 2006 Pb AQCD (U.S. EPA, 2006b) concluded that both epidemiologic and animal 

9 toxicological studies support the relationship between increased Pb exposure and 

10 increased cardiovascular effects, in particular, increased BP and increased incidence of 

11 arterial hypertension. Although fewer in number, epidemiologic studies demonstrated 

12 associations of blood and bone Pb levels with other cardiovascular diseases (CVDs) in 

13 adults, such as ischemic heart disease, cerebrovascular disease, peripheral vascular 

14 disease, and CVD-related mortality. As the cardiovascular and renal systems are 

15 intimately linked, cardiovascular effects can arise secondarily to Pb-induced renal injury 

16 (Section 5.5). Toxicological studies also provided compelling evidence supporting the 

17 biological plausibility for Pb-associated cardiovascular effects by characterizing a 

18 number of the underlying mechanisms by which Pb exposure can lead to human 

19 cardiovascular health effects. Such studies demonstrated that the Pb content in heart 

20 tissue of animals reflects the increases in blood Pb levels (Lal et al., 1991), indicating that 

21 the cardiovascular morbidity associated with blood Pb levels may represent the effects of 

22 the bioavailable Pb in the target tissue. The strongest evidence supported the role of 

23 oxidative stress in the pathogenesis of Pb-induced hypertension. Additionally, several 

24 toxicological studies characterized other pathways or cellular, molecular, and tissue 

25 events promoting the Pb-induced increase in BP. These mechanisms included 

26 inflammation, adrenergic and sympathetic activation, renin-angiotensin-aldosterone 

27 system (RAAS) activation, vasomodulator imbalance, and vascular cell dysfunction. 

28 With regard to the concentration-response relationship, meta-analysis of human studies 

29 found that each doubling of blood Pb level (between 1 and >40 μg/dL measured 

30 concurrently in most studies) was associated with a 1 mmHg increase in systolic BP and 

31 a 0.6 mmHg increase in diastolic BP (Nawrot et al., 2002). On a population-wide basis, 

32 the estimated effect size could translate into a clinically significant increase in the 
February 2012 5-226 Draft – Do Not Cite or Quote

1 segment of the population with the highest BP. In a moderately-sized population, a 

2 relatively small effect size thus has important health consequences for the risk of 

3 sequelae of increased BP, such as stroke, myocardial infarction, and sudden death. It was 

4 also noted that most of the reviewed studies examining bone Pb levels, biomarkers of 

5 cumulative Pb exposure, also showed increased BP (Cheng et al., 2001; Hu et al., 1996b)

6 or increased hypertension with increasing bone Pb level (Lee et al., 2001a). Across 

7 studies, over a range of bone Pb concentrations (<1.0 to 96 μg/g), every 10 μg/g increase 

8 in bone Pb was associated with increased odds ratios of hypertension between 1.28 and 

9 1.86. Studies observed an average increase in systolic BP of ~0.75 mmHg for every 10 

10 μg/g increase in bone Pb concentration over a range of <1 to 52 μg/g.

11 With regard to etiologically-relevant timing of Pb exposure, the weight of toxicological 

12 evidence demonstrated increases in BP after long-term (> 4 weeks) Pb exposure. In 

13 epidemiologic studies, as cardiovascular outcomes were most often examined in cross14 sectional studies with one or a limited number of Pb biomarker measurements, 

15 uncertainty exists as to the specific Pb exposure level, timing, frequency, and duration 

16 that contributed to the observed associations. While associations of adult bone Pb 

17 (particularly tibia Pb) with health outcomes in adults are indicative of effects related to 

18 past or cumulative exposures, interpretation of similar associations involving adult blood 

19 Pb levels, especially those measured concurrently with outcomes, are complicated by the 

20 generally higher past exposures common in this population. Detailed interpretation of Pb 

21 in blood and bone are provided in Sections 4.3 and 4.7.3. Briefly, higher past Pb 

22 exposures in adults increased their bone Pb stores which contribute to current blood Pb 

23 levels through the normal process of bone remodeling, as well as during periods of 

24 increased bone remodeling and loss (e.g., osteoporosis and pregnancy). Due to the long 

25 latency period for the development of increased BP and CVD, associations of 

26 cardiovascular effects with low concurrent blood Pb levels (e.g., population means 1.6-4 

27 μg/dL) in adults may be influenced by higher past Pb exposures (Section 4.4.1).

28 This section reviews the published studies pertaining to the cardiovascular effects of Pb 

29 exposure in humans, experimental animals, isolated vascular tissues, and cultured 

30 vascular cells. With the large and strong existing body of evidence serving as the 

31 foundation, emphasis was placed on studies published since the 2006 Pb AQCD (U.S. 

32 EPA, 2006b). Epidemiologic and toxicological studies continued to augment the evidence 

33 for increases in BP and hypertension development associated with long-term Pb exposure 

34 and expanded the evidence for the biological pathways of these effects. Epidemiologic 

35 studies strengthened the evidence for associations between Pb biomarkers and 

36 cardiovascular effects after adjusting for potential confounding factors such as age, diet, 

37 alcohol use, BMI, comorbidities, and smoking. The epidemiologic evidence was 
February 2012 5-227 Draft – Do Not Cite or Quote

1 substantiated with results from several available prospective studies indicating 

2 associations between Pb biomarkers and the incidence of cardiovascular health effects. 

5.4.2 Blood Pressure and Hypertension

5.4.2.1 Epidemiology

3 The most commonly used indicator of cardiovascular morbidity was increased BP and its 

4 derived index, hypertension. Hypertension in these studies was defined as diastolic and/or 

5 systolic BP above certain cut-points or use of anti-hypertensive medicines. The BP cut6 points employed have been historically established by reference to informed medical 

7 opinion, and as medical knowledge has improved BP cut-points defining hypertension 

8 have been lowered over time. Consequently, different studies using “hypertension” as a 

9 cardiovascular outcome may have assigned different cut-points, depending on the year 

10 and location of the study and the individual investigator. All of the new studies in the 

11 current review used the same criteria for hypertension (e.g., systolic BP at or above 140, 

12 diastolic BP at or above 90, or use of anti-hypertensive medications). Studies in the 

13 medical literature show that elevated BP is associated with increased risk of CVD 

14 including coronary disease, stroke, peripheral artery disease, and cardiac failure. 

15 Coronary disease (i.e., myocardial infarction, angina pectoris, sudden death) is the most 

16 lethal sequela of hypertension (Ingelsson et al., 2008; Chobanian et al., 2003; Pastor17 Barriuso et al., 2003; Prospective Studies Collaboration, 2002; Kannel, 2000a, b; Neaton 

18 et al., 1995). Several recent general population and occupational cohort and cross19 sectional studies strengthened the evidence that blood and bone Pb level were associated 

20 consistently with measures of BP (Figure 5-25 and Table 5-16) as well as with the 

21 prevalence and incidence of hypertension (Figure 5-26 and Table 5-17). Further, recent 

22 studies expanded evidence by finding differences in association among racial/ethnic 

23 groups, perceived stress, diet, and genetic variants and thus, identified populations 

24 potentially at increased risk of Pb-associated cardiovascular effects.

25 In a cross-sectional analysis, Martin et al. (2006) examined the associations of concurrent 

26 blood and tibia Pb levels with BP and hypertension in a community-based study of older 

27 adults (n = 964, age ranging from 50 to 70 years) in Baltimore, MD. A key strength of 

28 this study was the extensive consideration of potential confounding variables. Four 

29 models evaluated associations for BP and hypertension. The base model included age, 

30 sex, BMI, sodium intake, potassium intake, total cholesterol, time of day, testing 

31 technician, and hypertensive medication use. Other models added SES, race/ethnicity, or 

32 both as covariates. Blood Pb but not tibia Pb level was a strong predictor of BP in all 
February 2012 5-228 Draft – Do Not Cite or Quote

1 models; a 1 μg/dL increase in concurrent blood Pb level was associated with an 

2 approximately 1 mmHg increase in systolic BP and an approximately 0.5 mmHg increase 

3 in diastolic BP. Tibia Pb but not blood Pb was associated with hypertension in logistic 

4 regression models. The authors applied propensity analysis to their models to better 

5 account for the effect of other risk factors for hypertension such as race/ethnicity, age, 

6 and SES that were strongly associated with tibia Pb level. The propensity score analysis 

7 and model adjustment did not substantially change the numerical findings and 

8 conclusions (e.g., tibia Pb and hypertension were positively associated independently of 

9 race/ethnicity and SES), indicating that neither SES nor race/ethnicity confounded the 

10 association between tibia Pb level and hypertension. No evidence for effect modification 

11 by race/ethnicity was found either. Martin et al. (2006) concluded that Pb in blood has an 

12 acute effect on BP and that Pb contributes to hypertension risk as a function of 

13 cumulative, chronic exposure (as represented as bone Pb in this population). While 

14 different aspects of Pb exposure may contribute differentially to increases in BP and 

15 hypertension, it is important to note that concurrent blood Pb levels in adults also reflect 

16 cumulative Pb exposure. Thus, its association with BP may not reflect an acute effect but 

17 may also reflect an effect of cumulative Pb exposure. 
February 2012 5-229 Draft – Do Not Cite or Quote

a

Pb distribution presents the median (IQR) that were estimated from the mean and SD assuming a normal distribution. 

b

Effect estimates were standardized to 1 μg/dL blood Pb or 10 μg/g bone Pb.

Note: In general, results are categorized by specific BP parameter, then by Pb biomarker. For categories with multiple studies, the 

order of the studies follows the order of discussion in the text. For associations of a 1 μg/dL increase in blood Pb level (closed 

circles) or 10 μg/g increase in bone Pb (open circles) with systolic BP (SBP; blue), diastolic BP (DBP; red), and pulse pressure (PP; 

purple) in adults.

Figure 5-25 Concentration-response relationships (95% CI).
February 2012 5-230 Draft – Do Not Cite or Quote

Table 5-16 Additional characteristics and quantitative data for associations of 

blood and bone Pb with BP measures for studies presented in 

Figure 5-25

Study

Population

/Location Parameter Pb Data Statistical Analysis

Effect Estimate

β (95% CI)

Martin et al.

(2006)

964 men and women, 50-

70 yr, 40% African 

American, 55% White, 5% 

other, in Baltimore, MD

BP Concurrent Mean

Blood Pb:

Mean (SD): 3.5 (2.3) μg/dL

African American: 3.4 (2.3)

White: 3.5 (2.4)

Tibia Pb:

Mean (SD): 18.8 (12.4) μg/g

African American: 21.5 (12.6)

White: 16.7 (11.9) 

Multiple linear regression base model 

adjusted for age, sex, BMI, 

antihypertensive medication use, 

dietary sodium intake, dietary 

potassium intake, time of day, testing 

technician, serum total cholesterol. 

SES, race/ ethnicity also included in 

select models that are presented in 

Figure 5-25 and tabulated here.)

Blood Pb 

SBP: 1.05 (0.53,1.58) 

DBP; 0.53 (0.25, 0.81)

Tibia Pb:

SBP: 0.07 (-0.05, 0.14)

DBP: 0.05 (-0.02, 0.08)

mmHg per μg/dL blood Pb

mmHg per μg/g bone Pb

Glenn et al.

(2006)

575 Pb exposed workers, 

age 18-65 yr, in South 

Korea (10/1997-6/2001)

BP Blood Pb mean (SD):

Visit 1: 20.3 (9.6), Women

Visit 2: 20.8 (10.8), Women

Visit 3: 19.8 (10.7), Women

Visit 1: 35.0 (13.5), Men

Visit 2: 36.5 (14.2), Men

Visit 3: 35.4 (15.9), Men

Tibia Pb, mean (SD):

Visit 1: 28.2 (19.7), Women

Visit 2: 22.8 (20.9), Women

Visit 1: 41.7 (47.6), Men

Visit 2: 37.1 (48.1), Men

Patella Pb, mean (SD):

Visit 3 49.5 (38.5) Women

Visit 3 87.7 (117.0)

Multivariable models using GEE were 

used in longitudinal analyses. Models 

were adjusted for visit number, 

baseline age, baseline age squared, 

baseline lifetime alcohol consumption, 

baseline body mass index, sex, 

baseline BP lowering medication use, 

alcohol consumption, body mass 

index, sex, BP lowering medication use.

Model 1 (short-term)

Blood Pb concurrent

0.08 (-0.01, 0.16)

Blood Pb (longitudinal)

0.09 (0.01, 0.16)

Model 4: short and longerterm)

Blood Pb concurrent

0.10 (0.01, 0.19)

Blood Pb longitudinal:

0.09 (0.01, 0.16)

mmHg per 10 μg/dL blood 

Pb

Weaver et al.

(2008)

652 current and former 

Pb workers in South 

Korea (12/1999-6/2001)

BP Concurrent Blood Pb: 

Mean (SD): 30.9 (16.7) 

μg/dL

Concurrent Patella Pb:

Mean (SD): 75.1 (101.1) 

μg/g

Linear regression model adjusted for 

age, gender, BMI, diabetes, 

antihypertensive and analgesic 

medication use, Pb job duration, work 

status, tobacco and alcohol use

SBP

Patella Pb

0.0059 (-0.008, 0.02)a

Blood Pb

0.1007 (0.02, 0.18)a

mmHg per 1 μg/dL blood Pb 

or 1 μg/g patella Pb

Interaction between blood 

Pb/patella Pb with ALAD and 

vitamin D receptor 

polymorphisms not 

significant.

Peters et al.

(2007)

513 elderly men (mean 

67 y) from NAS in Greater 

Boston, MA area

BP Tibia Pb: 

mean (SD): 21.5 (13.4) μg/g

Patella Pb:

Mean (SD): 31.5 (19.3) μg/g

Logistic and linear regression models 

adjusted for age, age squared, 

sodium, potassium, and calcium 

intake, family history of hypertension, 

BMI, educational level, pack-years of 

smoking, alcohol consumption, and 

physical activity

SBP

Tibia Pb/ High Stress:

3.57 (0.39, 6.75)

Low Stress:

0.21 (-1.70, 1.29)

per SD increase in tibia Pb

Patella Pb/ High Stress:

2.98 (-0.12, 6.08)

per SD increase in tibia Pb

Patella Pb/ Low Stress: NR
February 2012 5-231 Draft – Do Not Cite or Quote

Study

Population

/Location Parameter Pb Data Statistical Analysis

Effect Estimate

β (95% CI)

Scinicariello et 

al. (2010)

6,016 NHANES III (1988-

1994) participants ≥ 17 yr

BP Concurrent Blood Pb:

Overall Mean (SE): 2.99 

(0.09) μg/dL

Non-Hispanic Whites: 2.87 

(0.09)

Non-Hispanic Blacks 3.59 

(0.20)

Mexican American 3.33 

(0.11)

Multivariable linear regression of logtransformed blood Pb level adjusted 

for age, sex, education, smoking 

status, alcohol intake, BMI, serum 

creatinine levels, serum calcium, 

glycosylated hemoglobin, and 

hematocrit

SBP

Non-Hispanic whites: 

1.05 (0.32, 1.78)

Non-Hispanic blacks: 

2.55 (1.59, 3.51)

Mexican Americans: 

0.84 (-0.06, 1.74)

DBP

Non-Hispanic whites: 

-0.14 (-1.1, 0.82)

Non-Hispanic blacks: 

1.99 (1.13, 2.85)

Mexican Americans:

0.74 (-0.005, 1.48)

mmHg per unit increase in ln 

Blood Pb

Significant interactions with 

blood Pb and ALAD 

observed in relation to SBP 

for non-Hispanic whites and 

non-Hispanic blacks

Zhang et al.

(2010a)

619 older adult males 

(mean 67 yr) enrolled in 

the NAS in Greater 

Boston, MA area

PP Wild type HFE

Tibia Pb: Median (IQR):8(12-

27) μg/g

Patella Pb: Median 

(IQR):26(17-37) μg/g

C282Y HFE

Tibia Pb: Median (IQR):20 

(14-27) μg/g

Patella Pb: Median 

(IQR):25(17-37) μg/g

H63D HFE

Tibia Pb: Median 

(IQR):19(14-26) μg/g

Patella Pb: Median 

(IQR):27(19-37)μg/g

Linear mixed effects regression 

models with repeated measurements 

adjusted for age; education; alcohol 

intake; smoking; daily intakes of 

calcium, sodium, and potassium; total 

calories; family history of 

hypertension; diabetes; height; heart 

rate; high-density lipoprotein (HDL); 

total cholesterol:HDL ratio; and waist 

circumference

PP

mmHg per 13 μg/g Tibia Pb:

Wild Type HFE: 0.38 (0,1.96)

H63D HFE: 3.30 (0.16, 6.46)

C282Y HFE: 0.89 (0, 5.24)

Any HFE variant: 2.90 (0.31, 

5.51)

mmHg per 19 μg/g Patella 

Pb:

Wild Type HFE: 0.26 (0, 

1.78)

H63D HFE: 2.95 (0, 5.92)

C282Y HFE: 0.55 (0, 1.66)

Any HFE variant: 2.83 

(0.32,5.37)

Perlstein et al.

(2007)

593 predominantly white 

men from NAS in Greater 

Boston, MA area (1991-

1997)

PP Blood Pb: 

Overall mean (SD): 6.12 

(4.03) μg/dL

Mean (SD) quintiles: 

Q1: 2.3 (0.8) μg/dL

Q2: 3.9 (0.3) μg/dL

Q3: 5.4 (0.5) μg/dL

Q4: 7.4 (0.6) μg/dL

Q5: 12.4 (4.4) μg/dL

Tibia Pb: 

Median: 19 μg/g

Mean (SD) quintiles: 

Q1: 7.4 (3.2) μg/g 

Q2: 14.1 (1.4) μg/g 

Q3: 18.9 (1.4) μg/g 

Q4: 24.9 (2.2) μg/g 

Q5: 40.9 (14) μg/g 

BP association assessed using 

spearman correlation coefficients.

PP association(adjusted mean 

difference) assessed using multiple 

linear regression model adjusted for 

age, height, race, heart rate, waist 

circumference, diabetes, family history 

of hypertension, education level 

achieved, smoking, alcohol intake, 

fasting plasma glucose, and ratio of 

total cholesterol to HDL cholesterol

PP

4.2 (1.9, 6.5) mmHg higher 

in men with tibia Pb > 19 

μg/g (median) compared with 

men with tibia Pb < μg/g

Blood Pb (mean difference): 

Q5: -1.49 (–4.93, 1.94) 

Q4: -1.39 (–4.94, 2.15), 

Q3: -2.56 (–5.78, 0.67) 

Q2: -4.37 (–7.88,–0.86) 

Q1: Referent group

Tibia Pb (mean difference): 

Q5: 2.58 (–1.15, 6.33)

Q4: 2.64 (–0.93, 6.21) 

Q3: -0.73 (–4.27, 2.82) 

Q2: -3.02 (–6.48, 0.44) 

Q1: Referent group
February 2012 5-232 Draft – Do Not Cite or Quote

Study

Population

/Location Parameter Pb Data Statistical Analysis

Effect Estimate

β (95% CI)

Navas-Acien et 

al. (2008)

Meta-analysis of studies 

using bone Pb as an 

exposure metric and BP 

as the outcome (8 

studies)

BP Inverse variance weighted randomeffects meta-analysesBP Pooled Estimates mmHg 

per 10 μg/g Tibia Pb

Prospective/SBP

0.33 (-0.44, 1.11)

X-sectional SBP

0.26 (0.02, 0.50)

X-sectional DBP

0.02 (-0.15, 0.19)

Hypertension per 10 μg/g 

patella Pb

x-Sectional hypertension 

OR: 1.04 (1.01, 1.07)

Pooled Estimate 

hypertension

OR: 1.04 (0.96, 1.12)

Yazbeck et al.

(2009) b 971 pregnant women, age 

18-45 yr, in France

BP Midpregnancy Blood Pb:

PIH group mean (SD): 2.2 

(1.4)

No PIH group mean (SD): 

1.9 (1.2)

Multivariable logistic regression 

models adjusted for maternal age; 

cadmium, manganese, and selenium 

blood levels; hematocrit; parity; BMI; 

pregnancy weight gain; gestational 

diabetes; educational level; SES; 

geographic residence; and smoking 

status and alcohol consumption before 

and during pregnancy

Log-transformed blood Pb at 

mid-pregnancy 

SBP: r = 0.08; p = 0.03

DBP: r = 0.07; p = 0.03 

Significant correlations also 

observed after 24 weeks of 

gestation and after 36 weeks 

of gestation.

Elmarsafawy 

et al. (2006) b 471 elderly men (mean 67 yr) from NAS in 

Greater Boston, MA area

BP Blood Pb:

Mean (SD): 6.6 (4.3) μg/dL

Tibia Pb:

Mean (SD): 21.6 (12.0) μg/g

Patella Pb:

Mean (SD): 31.7 (18.3) μg/g

Linear regression models adjusted for 

age, BMI, family history of 

hypertension, history of smoking, 

dietary sodium intake, and cumulative 

alcohol ingestion

Tibia Pb 

High calcium group (>800 

mg/d):

SBP: 0.40.(0.11, 0.70) 

Low calcium group (<800 

mg/d):

SBP: 0.19 (0.01, 0.37)

mmHg per μg/g tibia Pb

a

95% CIs estimated from given p-value.

b

References not included in Figure 5-25.
February 2012 5-233 Draft – Do Not Cite or Quote

Note: Studies are categorized by Pb biomarker. Within each category, studies generally are presented in order of discussion in the 

text. (a) The outcomes plotted are hypertension prevalence with the exception of Yazbeck et al. (2009) which measured pregnancy 

induced hypertension and Peters et al. (2007) which measured hypertension incidence. (b) ALAD2 vs. 1 indicates comparison 

between ALAD 2 carriers (e.g., ALAD1-2 and ALAD2-2) and ALAD 1 homozygotes (e.g., ALAD1-1). (c) Effect estimates were 

standardized to a 1 μg/dL increase in blood Pb. (d) Effect estimates were standardized to a 10 μg/g increase in bone Pb.

Figure 5-26 Odds ratios (95% CI) for associations of blood and bone Pb with 

hypertension prevalence and incidence.

►

Reference

Martin et al. (2006)

Peters et al. (2007)

Elmarsafawy et al. (2006)

Yazbeck et al. (2009)

Muntner et al. (2005)

Scinicariello et al. (2010)

age ≥ 17 y

mn BLL= 2.99 μg/dL

Park et al. (2009)

mn BLL= 3.52 μg/dL

Population

High Stress

High Stress

Low Calcium

High Calcium

Pregnant Women

Non-Hispanic Whites

Non-Hispanic Blacks

Mexican Americans

Non-Hispanic Whites

Non-Hispanic Blacks

Mexican Americans

Overall

White Men

Black Men

White Women

Black Women

Men <50

Men >50

Women <50

Women >50

Study Location

Baltimore, MD

Boston, MA

NAS

France

NHANES III

NHANES III

1998-1994

NHANES III

Blood Pb (μg/dL)

3.5 (2.3)

Tibia Pb (μg/g)

18.8 (12.4)

21.5 (13.4)

Patella Pb (μg/g)

31.5 (19.3)

6.6 (4.3)

21.6 (12.0)

31.7 (18.3)

6.6 (4.3)

21.6 (12.0)

31.7 (18.3)

1.2-1.7

1.71-2.30

>2.30

<1.06

1.06-1.63

1.63-2.47

>2.47

<1.06

1.06-1.63

1.63-2.47

>2.47

<1.06

1.06-1.63

1.63-2.47

>2.47

0.7-1.4

1.5-2.3

2.4-3.7

3.8-52.9

2.4-3.7

0.7-1.4

1.5-2.3

2.4-3.7

3.8-52.9

2.4-3.7

0.7-1.4

1.5-2.3

2.4-3.7

3.8-52.9

2.4-3.7

3.52 (0.10)

Comparison

per 2.5 μg/dL 

per 12.9 μg/g

per 11.6 μg/g

per 17.1 μg/g

per 1 μg/g

per 1 μg/g

per 1 μg/g

per 1 μg/g

per 1 μg/g

per 1 μg/g

Reference

Q2 v Q1

Q3 v Q1

Q4 v Q1

Reference

Q2 v Q1

Q3 v Q1

Q4 v Q1

Reference

Q2 v Q1

Q3 v Q1

Q4 v Q1

Reference

Q2 v Q1

Q3 v Q1

Q4 v Q1

Reference

Q2 v Q1

Q3 v Q1

Q4 v Q1

ALAD2 v 1 b

Reference

Q2 v Q1

Q3 v Q1

Q4 v Q1

ALAD2 v 1 b

Reference

Q2 v Q1

Q3 v Q1

Q4 v Q1

ALAD2 v 1 b

Continuous

per 0.75 μg/dL 

increase

0 1 2 3 4 5 6

Odds Ratio (95% CI)
February 2012 5-234 Draft – Do Not Cite or Quote

Table 5-17 Additional characteristics and quantitative data for associations of 

blood and bone Pb with hypertension measures for results 

presented in Figure 5-26

Study

Population/

Location Parameter Pb Data Statistical Analysis

Effect Estimate

(95% CI)

Martin et al.

(2006)

964 men and women, 

50-70 y, 40% African 

American, 55% White, 

5% other, in Baltimore, 

MD

Hypertension 

(current use of 

antihypertensive 

medication, mean 

SBP ≥ 140 mmHg 

or DBP ≥ 90 

mmHg)

Blood Pb:

Mean (SD): 3.5 (2.3) μg/dL

Tibia Pb:

Mean (SD): 18.8 (12.4) μg/g

Logistic regression models 

adjusted for age, sex, BMI, 

antihypertensive medication use, 

dietary sodium intake, dietary 

potassium intake, time of day, 

testing technician, and serum 

homocysteine 

Blood Pb level

OR=1.02 (0.87, 1.19)

Tibia Pb

OR=1.24 (1.05, 1.47)

mmHg per μg/dL blood Pb

mmHg per μg/g bone Pb

Weaver et al.

(2008) a 652 current and former 

Pb workers in South 

Korea (12/1999- 6/2001)

Hypertension

(mean SBP ≥ 140 

mmHg, DBP ≥ 90 

mmHg; and/or use 

of antihypertensive 

medications; or 

physician 

diagnosis) 

Blood Pb: 

Mean (SD): 31.9 (14.8) μg/dL

Patella Pb:

Mean (SD): 37.5 (41.8) μg/g

Logistic regression models 

adjusted for age, gender, BMI, 

diabetes, antihypertensive and 

analgesic medication use, Pb job 

duration, work status, tobacco 

and alcohol use

None of the Pb exposure 

metrics examined were 

(blood, patella, and ln 

patella) were significantly 

associated with 

hypertension (results not 

reported) 

Peters et al.

(2007)

513 elderly men 

(mean 67 y) from NAS 

in Greater Boston, MA 

area

Hypertension

(mean SBP >140 

mmHg, DBP >90 

mmHg; or physician 

diagnosis)

Tibia Pb: 

mean (SD): 21.5 (13.4) μg/g

Patella Pb:

Mean (SD): 31.5 (19.3) μg/g

Cox proportional hazards models 

adjusted for age, age squared, 

sodium, potassium, and calcium 

intake, family history of 

hypertension, BMI, educational 

level, smoking, alcohol 

consumption, baseline SBP and 

DBP, and physical activity

Hypertension Incidence

High Stress

RR=2.66 (1.43, 4.95) per 

SD increase in tibia Pb 

RR=2.64 (1.42, 4.92) per 

SD increase in patella Pb

Elmarsafawy et 

al. (2006)

471 elderly men 

(mean 67 y) from NAS 

in Greater Boston, MA 

area

Hypertension

(mean SBP ≥ 160 

mmHg, DBP ≥ 95 

mmHg; and/or 

physician diagnosis 

with current use of 

antihypertensive 

medications)

Blood Pb:

Mean (SD): 6.6 (4.3) μg/dL

Tibia Pb:

Mean (SD): 21.6 (12.0) μg/g

Patella Pb:

Mean (SD): 31.7 (18.3) μg/g

Logistic regression models 

adjusted for age, BMI, family 

history of hypertension, history of 

smoking, dietary sodium intake, 

and cumulative alcohol ingestion

Low calcium group (<800 

mg/d):

Blood Pb: 1.07 (1.00, 1.15)

Tibia Pb: 1.02 (1.00, 1.04)

Patella Pb: 1.01 (1.00, 

1.03)

High calcium group (>800 

mg/d):

Blood Pb: 1.03 (0.97, 1.11)

Tibia Pb: 1.01 (0.97, 1.04)

Patella Pb: 1.01 (0.99, 

1.03)

Per μg/dL blood Pb or μg/g 

tibia or patella Pb

Yazbeck et al.

(2009)

971 pregnant women, 

age 18-45 y, in France

PIH

(SBP ≥ 140 mmHg 

or DBP ≥ 90 mmHg 

after the 22nd wk of 

gestation)

Blood Pb:

PIH group mean (SD): 2.2 (1.4) 

μg/dL

No PIH group mean (SD): 1.9 

(1.2) μg/dL

Q1: <1.20 μg/dL

Q2: 1.20-1.70 μg/dL

Q3: 1.71-2.30 μg/dL

Q4: >2.30 μg/dL

Multivariable logistic regression 

models adjusted for maternal 

age, Cd, Mn, and Se blood 

levels, parity, hematocrit, BMI, 

gestational diabetes, educational 

levels, SES, geographic 

residence, and smoking status 

during pregnancy

PIH

Blood Pb 

OR=3.29 (1.11, 9.74) per 1 

unit increase in log 

maternal blood Pb level

Q1: Reference group

Q2: OR 1.84 (0.77, 4.41)

Q3: OR=2.07 (0.83, 5.13)

Q4: OR=2.56 (1.05, 6.22)
February 2012 5-235 Draft – Do Not Cite or Quote

Study

Population/

Location Parameter Pb Data Statistical Analysis

Effect Estimate

(95% CI)

Muntner et al.

(2005)

9,961 NHANES (1999-

2002) participants

Hypertension

(current use of 

antihypertensive 

medication, SBP ≥ 

140 mmHg, or DBP 

≥ 90 mmHg)

Concurrent Blood Pb:

Overall Mean (CI): 1.64 (1.59-

1.68) μg/dL

quartile 1: <1.06 μg/dL, quartile 

2: 1.06-1.63 μg/dL, quartile 3: 

1.63-2.47 μg/dL, and quartile 4: 

≥ 2.47 μg/dL

Multivariable logistic regression 

models adjusted for age, sex, 

diabetes mellitus, BMI, cigarette 

smoking, alcohol consumption, 

high school education, and 

health insurance status

Adjusted OR

Non-Hispanic white:

Q1: Reference group

Q2 OR=1.12 (0.83, 1.50) 

Q3 OR=1.03 (0.78, 1.37) 

Q4 OR=1.10 (0.87, 1.41)

Non-Hispanic black

Q1: Reference group

Q2 OR=1.03 (0.63, 1.67) 

Q3 OR=1.12 (0.77, 1.64) 

Q4 OR=1.44 (0.89, 2.32)

Mexican American

Q1: Reference group

Q2 OR=1.42 (0.75, 2.71) 

Q2 OR=1.48 (0.89, 2.48) 

Q3 OR=1.54 (0.99, 2.39)

Significant trend (p=0.04)

Scinicariello et 

al. (2010)

6,016 NHANES III 

(1988-1994) 

participants ≥ 17 y

Hypertension 

(current use of 

antihypertensive 

medication, SBP ≥ 

140 mmHg, or DBP 

≥ 90 mmHg)

Concurrent Blood Pb:

Mean (SE): 2.99 (0.09) μg/dL

Q1 0.7-1.4 μg/dL, 

Q2 1.5-2.3 μg/dL, 

Q3 2.4-3.7 μg/dL, 

Q4 3.8-52.9 μg/dL

Non-Hispanic Whites: 2.87 

(0.09)

Non-Hispanic Blacks 3.59 (0.20)

Mexican American 3.33 (0.11)

Multivariable logistic regression 

model adjusted for race/ethnicity, 

age, sex, education, smoking 

status, alcohol intake, BMI, 

serum creatinine levels, serum 

calcium, glycosylated

hemoglobin, and hematocrit

Non-Hispanic whites:

Q1: Reference group

Q2 POR=1.21 (0.66, 2.24)

Q3 POR=1.57 (0.88, 2.80)

Q4 POR=1.52 (0.80, 2.88)

ALAD1-2/2-2: POR= 0.76 

(0.17, 3.50)

ALAD-1: Reference group

Non-Hispanic blacks:

Q1 Reference

Q2 POR=1.83 (1.08, 3.09)

Q3 POR=2.38 (1.40, 4.06)

Q4 POR=2.92 (1.58, 5.41)

ALAD1-2/2-2: POR= 3.40 

(0.05, 219.03)

ALAD-1: Reference group

Mexican Americans:

Q1 Reference

Q2 POR=0.74 (0.24, 2.23)

Q3 POR=1.43 (0.61, 3.38)

Q4 POR=1.27 (0.59, 2.75)

ALAD1-2/2-2: POR= 0.49 

(0.08, 3.20)

ALAD-1: Reference group

POR for hypertension with 

ALAD2 carriers across 

quartiles of blood Pb level 

also reported. ALAD2

carriers associated with 

hypertension in nonHispanic whites.
February 2012 5-236 Draft – Do Not Cite or Quote

Study

Population/

Location Parameter Pb Data Statistical Analysis

Effect Estimate

(95% CI)

Park et al.

(2009a)

12,500 NHANES III 

(1988-1994) 

participants

Hypertension NHANES III Concurrent Blood 

Pb 

3.52 (0.10)

White men

<50 yr 4.02 (0.16)

≥ 50 yr 4.92 (0.18)

Black men

<50 yr 4.55 (0.15)

≥ 50 yr 7.57 (0.22)

White women

<50 yr 2.09 (0.07)

≥ 50 yr 3.53 (0.12)

Black women

<50 yr 2.52 (0.09)

≥ 50 yr 4.49 (0.16)

Logistic regression models 

adjusted for age, education, 

smoking status, cigarette 

smoking, BMI, hematocrit, 

alcohol consumption, physical 

activity, antihypertensive 

medication use, and diagnosis of

type-2 diabetes

OR per SD (0.75 μg/dL) in 

log blood Pb:

Overall: 1.12 (1.03, 1.23). 

White men: 1.06 (0.92, 

1.22) 

Black men: 1.17 (0.98, 

1.38) 

White women:1.16 (1.04, 

1.29) 

Black women: 1.19 (1.04, 

1.38) 

Men <50 yr: 0.98 (0.80, 

1.22)

Men >50 yr: 1.20 (1.02, 

1.41), 

Women <50 yr: 1.23 (1.04, 

1.46), 

Women >50 yr:1.09 (0.94, 

1.26).

a

Not included in Figure 5-26 because OR data were not reported.

1 In an occupational cohort in South Korea, Glenn et al. (2006) simultaneously modeled 

2 multiple Pb dose measures of individuals collected repeatedly over four years of follow 

3 up. Thus, through the assessment of cross-sectional and longitudinal relationships with 

4 BP, this study provided key insight on potentially important time periods of Pb exposure. 

5 The initial blood Pb level was used as a baseline covariate and the difference in blood Pb 

6 level between visits was computed for each subsequent visit. The bone Pb measures were 

7 used to indicate historical exposure and cumulative dose. Four models were specified: 

8 Model 1 was conceptualized to reflect short-term changes in BP associated with recent 

9 dose; Model 2 to reflect longer-term changes associated with cumulative dose controlling 

10 for association of baseline BP with recent dose; Model 3 to reflect longer-term changes 

11 associated with cumulative dose controlling for cross-sectional influence of cumulative 

12 dose on baseline BP; and Model 4 to reflect both short-term change with recent dose and 

13 longer-term change with cumulative dose. Concurrent blood Pb and increases in blood Pb 

14 between visits were associated with increases in systolic BP in Model 1 (short-term dose) 

15 and Model 4 (short- and longer-term dose). No association was observed between BP and 

16 tibia Pb at baseline while higher tibia Pb was associated with a decrease in systolic BP in 

17 each of the models.

18 These results indicate that circulating Pb (e.g., blood Pb) may act continuously on 

19 systolic BP and reduction in blood Pb may contribute to reductions in BP, while 

20 cumulative Pb exposure (represented by bone Pb in this study) may contribute to 

21 hypertension incidence by different mechanisms over longer time periods and in older 

22 subjects. This analysis in relatively young subjects with a low prevalence of hypertension 

23 suggests that at least one of the biological pathways that influences how systolic BP 
February 2012 5-237 Draft – Do Not Cite or Quote

1 responds to Pb operates over a relatively rapid timeframe and may reflect an immediate 

2 response to Pb at a biochemical site of action as a consequence of the biologically

3 available Pb circulating in blood. A persistent effect of cumulative doses over a lifetime 

4 may occur via other mechanisms. This study was strengthened by the analysis of 

5 associations between changes in blood Pb and changes in BP over time within individual 

6 subjects. Bone Pb level may exert influence on blood Pb levels and consequently on BP 

7 in an aging population with prolonged Pb exposure. Thus, there is strength in the 

8 conclusions drawn from this study that inform various short and long-term exposure 

9 relationships with increases in BP and hypertension. It is important to acknowledge the 

10 uncertainty regarding the applicability of these findings regarding short-term and long11 term effects in Pb workers with relatively high current Pb exposures contributing to blood 

12 Pb levels (mean blood Pb levels over time: 20-37 μg/dL) to adults in the U.S. general 

13 population whose concurrent blood Pb levels are influenced more by Pb mobilized from 

14 bone stores.

15 In a separate analysis of the same occupationally exposed group in year three of follow16 up, Weaver et al. (2008) examined cross-sectionally associations of concurrent patella Pb 

17 and blood Pb level with systolic BP, diastolic BP, and hypertension and effect 

18 modification by ALAD and vitamin D receptor (VDR) polymorphisms. None of the Pb 

19 biomarkers were associated with diastolic BP. Patella Pb alone was not significantly 

20 associated with systolic BP, while blood Pb, either alone or with patella Pb was 

21 significantly associated with higher systolic BP. The patella Pb-age and blood Pb-age 

22 interactions were not statistically significant. There were no significant associations of 

23 blood Pb or patella Pb with hypertension status or effect modification by age or sex. 

24 Further, interactions between polymorphisms of the VDR and of ALAD with blood Pb 

25 and patella Pb on systolic BP were not statistically significant. Mean blood Pb level was 

26 high (30.9 μg/dL) compared to non-occupational groups. 

27 Weaver et al. (2010) provided the results of further analysis of this Korean worker cohort, 

28 with a focus on determining the functional form of the concentration-response 

29 relationships. In a log linear model, the coefficient indicated that every doubling of blood 

30 Pb level was associated with a systolic BP increase of 1.76 mmHg. The J test, a statistical 

31 test for determining which, if either, of two functional forms of the same variable 

32 provides a superior fit to data in non-nested models (Davidson and MacKinnon, 1981),

33 returned a p-value of 0.013 in favor of the natural log blood Pb level over the linear blood 

34 Pb level specification. This analysis indicates that the systolic BP increase in this cohort 

35 is better described as a logarithmic function of blood Pb level within the blood Pb level 

36 range of the study than by a linear function. 
February 2012 5-238 Draft – Do Not Cite or Quote

1 Several analyses in the NAS cohort of predominantly white older men in the greater 

2 Boston area found associations of blood and bone Pb level with BP and hypertension and 

3 indicated effect modification by calcium intake, perceived stress, and HFE gene variants. 

4 In a cross-sectional analysis, Perlstein et al. (2007) found a statistically significant 

5 association between blood Pb and diastolic BP in adjusted models. The subjects in this 

6 study had at least one bone Pb measurement during the years 1991-1997 and were not on 

7 antihypertensive medication at the time of the measurement. While tibia Pb was not 

8 significantly associated with BP, it was associated with pulse pressure (PP). Men with 

9 tibia Pb above the median (19 μg/g) had a higher mean PP (4.2 mmHg [95%CI: 1.9, 6.5]) 

10 compared to men with tibia Pb below the median. The trend toward increasing PP with 

11 increasing quintile of tibia Pb was statistically significant although none of the 

12 confidence intervals for PP referenced to the lowest quintile of tibia Pb (< 7.4 μg/g) 

13 excluded the null value. 

14 Peters et al. (2007) examined cross-sectionally the modification of the associations of 

15 tibia and patella Pb with BP and hypertension by self-reported stress (assessed by 

16 questionnaire) in NAS men. High stress also has been linked with higher BP, potentially 

17 via activation of sympathetic pathways, ROS, and the HPA axis. Among all subjects, 

18 higher bone Pb level was associated (statistically nonsignificant) with greater odds of 

19 hypertension status and higher systolic BP. As indicated in Figure 5-27, the association 

20 between systolic BP and tibia Pb differed between those with high and low self-reported 

21 stress (β for tibia Pb x stress interaction = 3.77 [CI: 0.46, 7.09]) per SD increase in tibia 

22 Pb. Stress also was found to modify the patella Pb-BP association (β for patella Pb x 

23 stress interaction = 2.60 [CI: -0.95, 6.15] per SD increase in patella Pb). Neither bone, 

24 self-reported stress, nor their interaction was associated significantly with diastolic BP. 

25 Peters et al. (2007) also used Cox proportional hazards models to assess the interaction of 

26 stress and bone Pb level in the development of hypertension among those free of 

27 hypertension at baseline. The results of this analysis showed that increasing tibia and 

28 patella Pb were associated with greater risk of developing hypertension among those with 

29 high stress compared with those with lower perceived stress (RR of developing 

30 hypertension among those with high stress: 2.66 [CI: 1.43, 4.95] per SD increase in tibia 

31 Pb and 2.64 [CI: 1.42, 4.92] per SD increase in patella Pb). These results provide 

32 evidence supporting adults with higher stress as a population at increased risk of Pb33 associated cardiovascular effects. 
February 2012 5-239 Draft – Do Not Cite or Quote

Source: Peters et al. (2007)

Figure 5-27 The relationship between tibia Pb and estimated systolic BP 

(SBP) for those with high self-reported stress versus those with 

low self-reported stress.

1 Elmarsafawy et al. (2006) examined the modification of the relationship between Pb and 

2 hypertension risk by dietary calcium, with 467 subjects from the NAS. Responses on a 

3 semi-quantitative dietary frequency questionnaire with one-year recall were used to 

4 estimate calcium intake. Effect modification by calcium intake (dichotomized at 

5 800 mg/day) was examined using interaction terms in logistic regression models and by 

6 conducting analyses stratified on the calcium variable. Increasing bone and blood Pb 

7 increased the risk of hypertension, particularly among subjects with low dietary calcium. 

8 Zhang et al. (2010a) examined the effect of polymorphisms of the hemochromatosis gene 

9 (HFE) on the relationship of bone Pb with PP in NAS men. Subjects had up to three PP 

10 measurements during the 10 year study period. The overall results demonstrated a strong 

11 relationship between bone Pb and PP in this study, similar to an earlier cross-sectional PP 

12 study of many of the same subjects (Perlstein et al., 2007). Zhang et al. (2010a) extended 

13 these findings by demonstrating larger increases in PP per unit increase in tibia and 

14 patella Pb level among those with the H63D variant compared to those with the wild-type 

15 or the C282Y variant.

16 A small number of cross-sectional studies examined and found that blood Pb level was 

17 associated with hypertension in pregnancy. Yazbeck et al. (2009) examined a 

18 community-based group of pregnant women in France and unlike most other studies, 

19 adjusted for potential confounding by blood concentrations of cadmium, manganese, and 

20 selenium. Pregnancy induced hypertension (PIH) was defined as systolic BP >140 mmHg 

21 and/or diastolic BP >90 mmHg during at least two clinic visits after week 22 of gestation. 

22 Patients with pre-existing chronic hypertension were excluded. The mean (SD) blood Pb 
February 2012 5-240 Draft – Do Not Cite or Quote

1 levels measured during pregnancy were 2.2 (1.4 μg/dL) in PIH cases and 1.9 (1.2) μg/dL 

2 in normotensive women. An association between blood Pb and PIH was observed (OR 

3 3.29 [95% CI: 1.11, 9.74] per unit increase in log-transformed blood Pb level). Cadmium 

4 and selenium concentrations were comparable between PIH and no PIH groups. 

5 Adjustment for the metals slightly attenuated but did not eliminate the association 

6 between blood Pb levels and the risk of PIH. Investigators observed no significant 

7 interactions among blood Pb level, any of the other elements, and maternal characteristics 

8 in predicting the risk of PIH. Interaction between blood selenium and Pb concentrations 

9 was not significant, and the putative protection effects of selenium through antioxidative 

10 properties were not found in this study. 

11 Wells et al. (2011b) measured the relationship of cord blood Pb with BP in 285 women at 

12 admission to the Johns Hopkins Hospital in Baltimore, MD, during labor and delivery. 

13 Women with cord blood Pb levels in the highest quartile for the study group 

14 (>0.96 μg/dL) had significantly higher systolic and diastolic BP (upon admission and for 

15 maximum BP) compared to women in the first quartile (<0.46 μg/dL). The authors used 

16 Benchmark Dose Software V2.1, developed by the EPA, to estimate the blood Pb level 

17 (benchmark dose or BMD) and the associated lower confidence limit (BMDL) that was 

18 associated with one standard deviation (SD) increase in BP. In this study group, one SD 

19 is approximately equivalent to a 10% increase above the mean for the first quartile blood 

20 Pb reference group. The BMD approach was used only as a means of quantifying the 

21 relationship of blood Pb with BP in this population. This analysis indicated that the 95% 

22 lower bound confidence limit on the maternal blood Pb level (estimated from cord blood 

23 Pb levels) that was associated with a 1 SD increase in all blood pressure outcomes was 

24 about 1.4 μg/dL. These reported results are similar to those reported in the 2006 Pb 

25 AQCD as well as those found 25 years ago but with blood Pb levels in the more recent 

26 study an order of magnitude lower.

27 New analyses using NHANES data continued to indicate associations of Pb biomarkers 

28 with BP and hypertension. Muntner et al. (2005) previously used the NHANES 1999-

29 2002 data to indicate that concurrent blood Pb levels were associated with hypertension, 

30 peripheral artery disease (PAD), and chronic kidney disease. The PAD results are 

31 discussed later in Section 5.4.3.4, and chronic kidney disease results are discussed in 

32 Section 5.5.2.2. Blood Pb increased regularly with age (geometric means [95% CIs]: 

33 1.28 μg/dL [1.23, 1.33] in the 18-39 age group to 2.32 μg/dL [2.20, 2.44] in the 75 and 

34 older age group). Associations were observed between concurrent blood Pb level and 

35 hypertension across race/ethnicity groups with significant trends observed for 

36 non-Hispanic blacks and Mexican Americans.
February 2012 5-241 Draft – Do Not Cite or Quote

1 In the NHANES III 1988-1994 population, Scinicariello et al. (2010) found a gene2 environment interaction between blood Pb level and ALAD genotype in relation to SBP 

3 and DBP in a cross-sectional analysis. These interactions varied across race/ethnicity 

4 strata. The strongest associations were observed among non-Hispanic blacks (Figure 

5 5-25, Table 5-16). A statistically significant interaction was observed between concurrent 

6 blood Pb level and ALAD1-2/2-2b among non-Hispanic whites and non-Hispanic blacks. 

7 Scinicariello et al. (2010) also found an interaction between ALAD genotype and blood 

8 Pb level in the association with hypertension. Statistically significant associations 

9 between concurrent blood Pb level and hypertension were observed among non-Hispanic 

10 blacks and nonsignificant increases were observed among non-Hispanic whites and 

11 Mexican Americans (with the exception of Q2 association for Mexican Americans) 

12 (Figure 5-26, Table 5-17). In addition, non-Hispanic white ALAD2 carriers in the highest 

13 blood Pb level quartile 3.8-52.9 μg/dL) had a significantly higher association with 

14 hypertension compared with ALAD1 homozygous individuals in the highest quartile of 

15 blood Pb. In the same NHANES population, Park et al. (2009a) predicted bone Pb levels 

16 using a model developed with NAS data. Concurrent blood Pb was associated with 

17 hypertension overall in the NHANES population, with larger associations observed 

18 among black men and women as well as older adults Figure 5-26, Table 5-17). 

19 Associations also were observed with estimated bone Pb.

5.4.2.2 Toxicology

20 Studies on the effect of Pb (as blood Pb level) on systolic BP in unanesthetized adult rats 

21 consistently reported an increase in BP with increasing blood Pb level as shown in Figure 

22 5-28 (results summarized in Table 5-18). An array of studies have provided evidence that 

23 long-term Pb exposure (> 4 weeks), resulting in blood Pb levels below 10 μg/dL can 

24 result in the onset of hypertension (after a latency period) in experimental animals that 

25 persists long after the cessation of Pb exposure (U.S. EPA, 2006b). Tsao et al. (2000)

26 presented evidence for increased systolic and diastolic BP in rats with blood Pb levels 

27 relevant to those in humans (mean [SD]: 2.15 [0.92] μg/dL blood Pb; 140 [7] mmHg 

28 systolic BP, 98 [7] mmHg diastolic BP) compared to untreated controls (mean [SD]: 0.05 

29 [0.05] μg/dL blood Pb; 127 [7] mmHg systolic BP, 88 [7] mmHg diastolic BP). As this 

30 was the lowest Pb level tested, no evidence of a threshold was evident. Further, a test for 

31 linear trend revealed a statistically significant, positive trend for increasing BP with 

32 increasing blood Pb levels up to 56 μg/dL (e.g., mean [SD]: 5.47 [2.1] μg/dL blood Pb; 

33 143 [6] mmHg systolic BP, 97 [8] mmHg diastolic BP), with the effect leveling off at 

34 higher blood Pb levels. 
February 2012 5-242 Draft – Do Not Cite or Quote

Note: Crosses represent standard error for blood Pb and BP measurements. If no crossbar is present, error results were not 

reported. Arrows represent higher doses tested.

Figure 5-28 Changes in BP after Pb exposure (represented as blood Pb level) 

in unanesthetized adult rats across studies.
February 2012 5-243 Draft – Do Not Cite or Quote

Table 5-18 Characteristics of studies of blood Pb with BP measures in animals 

presented in Figure 5-28

Referencea Lifestage; Sex

Exposure 

Duration

Exposure Level; 

Route

Mean [SEM]b

Blood Pb 

Level (μg/dL) n

ΔSBP

(mmHg; lowest 

blood Pb level 

compared with 

control)c Comments

Fiorim et al. (2011) Adult; M 7 days 4 μg/100 g followed by 

0.05 μg/100 g daily; 

intramuscular

9.98 [1.7] 12 16

Nakhoul et al.

(1992)

Adult; M 8 weeks 100 ppm; drinking water 5.3 [3] 7 28 Spontaneously 

hypertensive rat 

model

Chang et al. (2005) Adult; M 8 weeks 2% (20,000 ppm) then 

removal and measurements 

1-7 mo after; drinking water 

Range: 4.5 – 83 5 13.8

Tsao et al. (2000) Adult 8 weeks 0.01 - 2% (100 - 20,000 

ppm); drinking water

Range of means: 

2.15 [0.29] –

85.76 [1.29]a

10 13

Rizzi et al. (2009) Adult; M 8 weeks 30 - 90 ppm; drinking water 7.6 [1.3], 19.3 

[3.4]

11 13.3

Chang et al. (1997) Adult; M 8 weeks 0.5% (500 ppm); drinking 

water

29.1 [0.6]a 10 58

Heydari et al. (2006) Adult; M 12 weeks 100 ppm; drinking water 26.8 [2.2] 6 25.8

Bravo et al. (2007) Adult; M 14 weeks 100 ppm; drinking water 23.7 [1.9]a 12 30

Zhang et al. (2009a) Adult; M 40 weeks 100 ppm; drinking water 28.4 [1.1]a 8-10 15.3

a

Studies are presented in order of increasing duration of exposure.

b

Standard deviation converted to SEM.

c

Difference in systolic BP (SBP) between group means not within one exposure group.

1 Experimental animal studies continued to provide evidence that long-term Pb exposure 

2 results in sustained arterial hypertension after a latency period. Systolic BP increased in 

3 rats after exposure to 90-10,000 ppm Pb (as Pb-acetate in drinking water) for various 

4 time periods that resulted in blood Pb levels between 19.3-240 μg/dL (Mohammad et al., 

5 2010; Zhang et al., 2009a; Badavi et al., 2008; Grizzo and Cordellini, 2008; Reza et al., 

6 2008; Bravo et al., 2007; Vargas-Robles et al., 2007; Heydari et al., 2006; Bagchi and 

7 Preuss, 2005). Past studies have shown statistically significant elevations in BP in rats 

8 with lower blood Pb levels. For example, long-term Pb exposure to spontaneously 

9 hypertensive rats (resulting in mean [SEM] blood Pb level: 5.3 [3] μg/dL) led to 

10 increased BP (Nakhoul et al., 1992). Consistent with measurements of systolic BP by tail11 cuff plethysmography, Pb exposure (100 ppm for 14 weeks; mean blood Pb level: 

12 24 μg/dL) also caused an increase in intra-aortic mean arterial pressure (Bravo et al., 

13 2007). In a study that tested low levels of Pb exposure (30 ppm; mean blood Pb level: 

14 7.6 μg/dL), a statistically significant increase in systolic BP was not observed despite 

15 elevated blood Pb level after 8 weeks of treatment. Nonetheless, there was a trend of 

16 higher BP with higher blood Pb levels (Rizzi et al., 2009).
February 2012 5-244 Draft – Do Not Cite or Quote

1 Studies found that Pb-induced increased BP persisted long after cessation of Pb exposure. 

2 Bagchi and Preuss (2005) found that elevated systolic BP was maintained for 210 days 

3 after Pb exposure cessation. However, chelation therapy using Na2CaEDTA returned 

4 systolic BP to levels comparable to those in rats not treated with Pb (Bagchi and Preuss, 

5 2005). Chang et al. (2005) reported a partial reversibility of effect after cessation of Pb 

6 exposure, where Pb-induced elevated BP decreased but did not return to control levels 7 

7 months post Pb exposure. After Pb exposure was removed, blood, heart, aorta, and 

8 kidney Pb levels decreased quickly within the first three months (Chang et al., 2005). Pb9 induced elevated systolic BP persisted for one month following Pb exposure cessation, 

10 followed by obvious decreases in BP until 4 months after Pb exposure cessation. Between 

11 4 and 7 months after Pb exposure cessation, the still-elevated BP did not decrease further, 

12 thus never returning to control BP levels. Decreases in BP were strongly correlated with 

13 decreases in blood Pb level after exposure cessation. 

14 The aforementioned studies all assessed the relationship between long-term exposure (> 4 

15 weeks) of rats to Pb and measures of BP. However, recent research also investigated BP 

16 elevation occurring after short-term treatment with Pb (< 4 weeks). Studies found 

17 increased systolic BP after 7 days of Pb treatment (daily injections resulting in mean 

18 [SEM] blood Pb levels of 9.98 [1.7] μg/dL) (Fiorim et al., 2011) and after 2 weeks of Pb 

19 exposure (100 ppm via drinking water) (Sharifi et al., 2004). A study utilizing intra20 arterial pressure measurements found that a single high-dose Pb injection (resulting in 

21 mean [SEM] blood Pb levels of 37 [1.7] μg/dL) increased systolic arterial pressure after 

22 only 60 minutes (Simões et al., 2011). These studies suggest that there is the potential for 

23 increase in BP following short-term Pb treatment. It is possible that the increases in BP 

24 following short- and long-term Pb exposures are occurring through separate mechanisms; 

25 however, studies using both short- and longer-term Pb exposure have correlated increased 

26 BP with an activation of the renin-angiotensin system (i.e., increase in angiotensin 

27 converting enzyme (ACE) activity) (Section 5.4.2.3). Several of these aforementioned 

28 studies used the injection route of Pb administration, and the relevance of these bolus 

29 doses over short periods of time to human routes of short-term exposure is uncertain. 

30 However, it is important to acknowledge that the results were similar to those from the 

31 study that examined short-term exposure to Pb via drinking water, 

5.4.2.3 Hypertension Modes of Action

32 The 2006 Pb AQCD examined a number of mechanisms leading to Pb-induced 

33 hypertension, including oxidative stress, hormonal and blood pressure regulatory system 

34 dysfunction, vasomodulation, and cellular alterations. As described below, recent studies 
February 2012 5-245 Draft – Do Not Cite or Quote

1 in experimental animals and cells further supported roles for these potential mechanisms 

2 in mediating hypertension from Pb exposure.

Oxidative Stress Response - Reactive Oxygen Species and Nitric Oxide

3 Several studies discussed in the 2006 Pb AQCD demonstrated a role for oxidative stress 

4 in the pathogenesis of Pb-induced hypertension, mediated by the inactivation of nitric 

oxide (

• 5 NO) and downregulation of soluble guanylate cyclase (sGC) (Dursun et al., 2005; 

6 Attri et al., 2003; Gonick et al., 1997; Vaziri et al., 1997; Khalil-Manesh et al., 1994; 

Khalil-Manesh et al., 1993b). Pb-induced reduction of biologically active • 7 NO was found 

not to be due to a reduction in • 8 NO-production capacity (Vaziri and Ding, 2001; Vaziri et 

al., 1999b); instead it was found to result from inactivation and sequestration of • 9 NO by 

10 ROS (Malvezzi et al., 2001; Vaziri et al., 1999a). Oxidative stress from Pb exposure in 

11 animals may be due to upregulation of NAD(P)H oxidase (Ni et al., 2004; Vaziri et al., 

12 2003), induction of Fenton and Haber-Weiss reactions (Ding et al., 2001; Ding et al., 

13 2000), and failure of the antioxidant enzymes, CAT and GPx, to compensate for the 

14 increased ROS (Farmand et al., 2005; Vaziri et al., 2003). Many biological actions of 

• 15 NO, such as vasorelaxation, are mediated by cGMP, which is produced by sGC from the 

16 substrate GTP. Oxidative stress also has been found to play a role in Pb-induced 

17 downregulation of sGC (Farmand et al., 2005; Courtois et al., 2003; Marques et al., 

2001). The reduction of the vasodilator • 18 NO leads to increased vasoconstriction and BP.

19 Pb-induced oxidative stress also has been found to induce renal tubulointerstitial 

20 inflammation which plays a crucial role in models of hypertension (Rodriguez-Iturbe et 

21 al., 2005; Rodriguez-Iturbe et al., 2004). Tubulointerstitial inflammation from treatment 

22 with Pb has been coupled with activation of the redox sensitive NF-κB (Ramesh et al., 

23 2001). Pb-induced hypertension, inflammation, and NF-κB activation can be ameliorated 

24 by antioxidant therapy (Rodriguez-Iturbe et al., 2004). There is mixed evidence to 

25 suggest that Pb-induced hypertension may also be promoted by activation of PKC leading 

26 to enhanced vascular contractility (Valencia et al., 2001; Watts et al., 1995).

Recent studies continued to provide evidence for the role of ROS and • 27 NO metabolism in 

28 Pb-induced hypertension and vascular disease. Increased systolic BP after Pb exposure 

was accompanied by increased superoxide (O2

-

) and O2

- 29 positive cells (Bravo et al., 2007; 

30 Vargas-Robles et al., 2007), elevated urinary malondialdehyde (MDA, a measure of lipid 

31 peroxidation) (Bravo et al., 2007), and increased 3-nitrotyrosine (Vargas-Robles et al., 

2007). Inhibition of NAD(P)H oxidase, an enzyme that generates O2

- 32 and hydrogen 

33 peroxide, was able to block Pb-induced (1 ppm) aortic contraction 

34 to 5-hydroxytryptamine (5-HT) (Zhang et al., 2005). Increases in systolic BP, intra-aortic 

35 mean arterial pressure, and MDA after Pb exposure (100 ppm; mean blood Pb level: 
February 2012 5-246 Draft – Do Not Cite or Quote

1 23.7 μg/dL) were also prevented by treatment with the immunosuppressant, 

2 mycophenolate mofetil (MMF) (mean blood Pb level in MMF-treated animals: 27 μg/dL)

3 (Bravo et al., 2007). MMF has been shown to inhibit endothelial NAD(P)H oxidase, 

4 which could explain how it decreases Pb-induced increases in oxidative stress and BP. 

5 MMF was not found to alter blood Pb levels of animals. Red grape seed extract and 

6 ascorbic acid supplementation were also able to protect rats from Pb-induced (100 ppm) 

7 increased BP and heart rate, perhaps through the antioxidant properties of the extract

8 (Badavi et al., 2008) and vitamin (Mohammad et al., 2010). Red grape seed extract did 

9 not alter the accumulation of Pb in blood, indicating that its protective effect was not 

10 mediated through altered Pb toxicokinetics; however, internal doses of Pb were not 

11 measured in the vitamin C study to clarify the mechanism of action of vitamin C. Another 

12 study found that the antioxidant, anti-inflammatory chemical, curcumin, as well as 

13 physical exercise training reversed Pb-induced increases in serum creatinine kinase-MB 

14 (CK-MB), low density lipoprotein (LDL), heart high-sensitivity C-reactive protein 

15 (hs-CRP), and MDA, and Pb-induced decreases in serum total antioxidant capacity, high 

16 density lipoprotein (HDL), and heart glutathione peroxidase (GPx); however, internal 

17 doses of Pb were not measured to clarify the mechanism of action in this study (Roshan 

18 et al., 2011).

19 Exposure to Pb can also affect the activity and levels of antioxidant enzymes. Male (♂) 

20 and female (♀) rats exposed to Pb for 18 weeks (100-1,000 ppm) had altered responses in 

21 antioxidant enzymes in heart tissue (Sobekova et al., 2009; Alghazal et al., 2008a). Pb 

22 exposure in female rats increased the activity of cardiac SOD, GST, GR, and GPx (> 

23 100 ppm) and increased cardiac thiobarbituric acid reactive substances (TBARS, measure 

24 of lipid peroxidation) (1,000 ppm). Pb exposure in male rats did not affect the activity of 

25 SOD or production of TBARS, however decreased the activity of GST and GR 

26 (>100 ppm). Male and female rats also accumulated different amounts of Pb in the 

27 cardiac tissue after similar exposure (♂ 100 ppm: 205% of control, 1,000 ppm: 379%; ♀ 

28 100 ppm: 246%, 1,000 ppm: 775%), which could explain the sex differences observed in 

29 antioxidant enzyme responses. 

30 Oxidative stress can trigger a cascade of events that promote cellular stress, renal 

31 inflammation, and hypertension. As was shown previously (Rodriguez-Iturbe et al., 

32 2005), Pb exposure can increase renal NF-κB, which was associated with 

33 tubulointerstitial damage and infiltration of lymphocytes and macrophages (Bravo et al., 

34 2007). These events could also be ablated by MMF treatment, likely due to its anti35 inflammatory and antioxidant properties. Pb also was found to induce inflammation in 

36 human endothelial cells as a model for vessel intima hyperplasia (Zeller et al., 2010). The 

37 pro-inflammatory cytokine, interleukin (IL)-8 protein and mRNA were increased, 

38 concentration- and time-dependently, after in vitro Pb exposure (5-50 μM). Enhanced IL-
February 2012 5-247 Draft – Do Not Cite or Quote

1 8 production was mediated through activation of the transcription factor Nrf2 (but not 

2 NF-κB, hypoxia inducible factor-1, or aryl hydrocarbon receptor), as shown through 

3 increased nuclear translocation and Nrf2 cellular knockdown experiments. Additionally, 

4 measures of endothelial stress, NQO1 and HO-1 protein, were induced by Pb exposure

5 (Zeller et al., 2010). Pb treatment (20 ppm, i.p., 3 days/week, 8 weeks) increased the 

6 inflammatory markers hs-CRP and CK-MB in rat hearts (Roshan et al., 2011).

7 Oxidative stress affects vascular reactivity and tone through inactivation and 

sequestration of •NO, causing a reduction in biologically active • 8 NO. Recent studies 

affirmed past conclusions on the interplay of ROS and • 9 NO metabolism in the 

10 cardiovascular effects of Pb. Elevated systolic BP and altered vasorelaxation after Pb 

11 exposure was accompanied by a decrease in total nitrates and nitrites (NOX) (Mohammad 

12 et al., 2010; Zhang et al., 2007a; Heydari et al., 2006). Serum NOX levels in Pb-treated 

13 rats remained depressed for 8 weeks and then reversed after 12 weeks, despite continued 

14 elevation in systolic BP (Heydari et al., 2006). This return of serum NOX levels to levels 

15 similar in controls could be a result of compensatory increases in endothelial NOS 

(eNOS) attempting to replenish an over-sequestered • 16 NO supply. With this in mind, 

17 studies showed increased eNOS protein expression after long-term Pb exposure in kidney

18 (Zhang et al., 2007a) and isolated cultured aorta (Vargas-Robles et al., 2007). No change 

19 in inducible NOS was observed in isolated cultured aorta after 1 ppm Pb exposure (Zhang 

20 et al., 2007a). In contrast to long-term exposure, Pb treatment over a short time period 

21 (daily injections resulting in mean [SEM] blood Pb levels of 9.98 [1.7] μg/dL) was found 

22 to increase iNOS and phosphorylated eNOS protein (Fiorim et al., 2011) which may 

cause an increase in •NO production and a short-term increase in • 23 NO bioavailability. 

This increase in • 24 NO bioavailability early after Pb exposure could be the immediate 

25 compensatory mechanism against the elevation in BP. 

• 26 NO, also known as endothelium-derived relaxing factor, is a potent endogenous 

27 vasodilator. Toxicological studies continued to investigate the effects of Pb on 

•

NO-dependent vascular reactivity by using • 28 NO stimulating vasodilators, such as 

acetylcholine (ACh) and sodium nitroprusside (SNP), and • 29 NO inhibiting 

30 vasoconstrictors, such as L-NAME. Studies provided mixed evidence; however, results 

suggested that Pb disrupts the vasorelaxant response to • 31 NO in the aorta due to damage to 

32 the endothelium. Pb exposure (1 ppm and 100 μM, 1 hour) decreased ACh-induced 

vasorelaxation, which triggers the release of • 33 NO from the endothelial cell, in isolated rat 

34 tail artery, suggesting damage to the endothelium (Silveira et al., 2010; Zhang et al., 

35 2007a). In aortic rings of perinatally exposed rats (1,000 ppm through pregnancy and 

36 lactation, mean blood Pb level: 58.7 μg/dL), blocking NOS with L-NAME abolished the 

37 relaxant response evoked by ACh (Grizzo and Cordellini, 2008). However, there was no 

38 change observed in the relaxation response to ACh by Pb alone (Fiorim et al., 2011; Rizzi 
February 2012 5-248 Draft – Do Not Cite or Quote

1 et al., 2009; Grizzo and Cordellini, 2008). Conversely, Skoczynska and Stojek (2005)

found that Pb exposure (50 ppm; blood Pb level 11.2 μg/dL) enhanced • 2 NO-mediated 

3 vasodilation by ACh in rat mesenteric arteries, and NOS inhibition enhanced the ACh 

4 relaxant response. A number of studies found that Pb exposure did not affect smooth 

5 muscle integrity since SNP-induced vasorelaxation, which is endothelium independent, 

6 was unchanged (Fiorim et al., 2011; Silveira et al., 2010; Rizzi et al., 2009; Grizzo and 

7 Cordellini, 2008).

• 8 NO also was found to play a role in the interaction between Pb and the vasoconstrictor 

9 response. Blocking NOS with L-NAME or inhibiting iNOS specifically, which decreases 

• 10 NO production, increased the contraction of aortic rings to the vasoconstrictor 

11 phenylephrine (PHE) and Pb exposure potentiated this response (Fiorim et al., 2011).

12 Also, L-NAME increased the Pb pressor response to PHE after perinatal Pb exposure 

13 (1,000 ppm through pregnancy and lactation, blood Pb level 58.7 μg/dL) (Grizzo and 

14 Cordellini, 2008). Conversely, in rat renal interlobar arteries, Pb exposure blunted the 

15 increase in renal angiotensin II (AngII)-mediated contraction from NOS inhibition by L16 NAME (Vargas-Robles et al., 2007). Treatment with the SOD mimetic tempol, which 

would increase • 17 NO bioavailability, decreased, but did not eliminate, the Pb pressor 

18 response (Silveira et al., 2010).

19 In summary, recent studies continued to provide evidence for the role of ROS in Pb20 induced hypertension and vascular disease by indicating Pb-induced increases in ROS 

21 and modulation of cardiovascular responses by antioxidant substances. Additionally, 

22 recent studies continued to show that Pb-induced hypertension and vascular responses are 

mediated primarily via inactivation of •NO not via inhibition of • 23 NO production.

Vascular Reactivity 

24 Alteration of the adrenergic system from Pb exposure, which can increase peripheral 

25 vascular resistance, and thereby arterial pressure, may be one cause of Pb-induced 

26 hypertension. Pb exposure in animals can increase stimulation of the sympathetic nervous 

27 system (SNS), as shown by increased plasma levels of norepinephrine (NE) and other 

28 catecholamines (Carmignani et al., 2000; Chang et al., 1997), and decreased β adrenergic 

29 receptor density and β agonist-stimulated cAMP production in the aorta and heart (Tsao 

30 et al., 2000; Chang et al., 1997). These stimulatory effects on the SNS paralleled the 

31 effects of Pb on BP, cardiac contractility, and carotid blood flow. Pb-induced elevations 

32 in arterial pressure and heart rate were abrogated by ganglionic blockade (Simões et al., 

33 2011; Lai et al., 2002). Arterial pressure and heart rate gradually decreased 7 months 

34 after Pb exposure cessation as did the Pb-induced SNS alterations (Chang et al., 2005).
February 2012 5-249 Draft – Do Not Cite or Quote

1 Increases in BP can be caused by activation of the SNS, which can lead to vascular 

2 narrowing, in turn, resulting in increased total peripheral resistance. In this neural 

3 mechanism, activation of the SNS leads to vasoconstriction, whereas inhibition leads to 

4 vasodilation. It has been suggested that Pb leads to increased vascular reactivity to 

5 catecholamines (i.e., epinephrine, NE, and dopamine), hormones of the SNS. Indeed, the 

6 isolated mesenteric vessel bed from Pb-treated rats (50 ppm with blood Pb level: 

7 11.2 μg/dL, but not 100 ppm with blood Pb level: 17.3 μg/dL) exhibited increased 

8 reactivity to NE (Skoczynska and Stojek, 2005). However, in another study, 100 ppm Pb 

9 did not affect the NE-induced contractile response after 10 months of exposure (Zhang et 

10 al., 2009a), suggesting a small range of Pb doses affects pressor response to NE. 

11 Catecholamines act primarily through the adrenergic and dopaminergic receptors. 

12 Antagonists of α1-adrenergic, α2-adrenergic, β-adrenergic, and dopamine D1 receptors 

13 were found to abolish Pb-induced aortic contraction (Fazli-Tabaei et al., 2006; Heydari et 

14 al., 2006). However, the α1-adrenergic receptor agonist PHE induced aortic contractions 

15 and these were enhanced by treatment with Pb (100 ppm; blood Pb level: 26.8 μg/dL), 

16 indicating a specific role for the α1-adrenergic receptor (Silveira et al., 2010; Grizzo and 

17 Cordellini, 2008; Heydari et al., 2006). Removal of the endothelium blunted the PHE18 induced contraction. Conversely, short-term Pb exposure (7 days) decreased the 

19 contractile response induced by PHE in rat aortas resulting in a decreased vascular 

20 reactivity (Fiorim et al., 2011). This decrease may be playing a compensatory role in 

21 attempting to correct the Pb-induced BP elevation. Additionally, Pb blunted the 

22 isoproterenol-induced relaxation, supporting a role for the β-adrenoceptors (Vassallo et 

23 al., 2008; Heydari et al., 2006).

24 Recently, there was mixed evidence for Pb disrupting vascular reactivity to other pressor 

25 agents. Pb (1 ppm) treatment of isolated rat thoracic aorta increased 5-HT induced 

26 contraction, which was endothelium dependent, but not due to 5-HT2B receptor 

27 expression (Zhang et al., 2005). Follow-up of this study in whole animals found, on the 

28 contrary, that Pb (100 ppm; blood Pb level: 28.4 μg/dL) decreased the maximum 

29 contractile response to 5-HT, but did not affect 5-HT plasma levels or 5-HT2B receptor 

30 expression (Zhang et al., 2009a). In addition, Pb exposure (100 ppm, 12 weeks) increased 

31 the renal vascular response to AngII in isolated perfused kidneys from Pb-exposed rats

32 (Vargas-Robles et al., 2007).

Studies continued to investigate the effects of Pb on • 33 NO-dependent vascular reactivity by 

using •NO stimulating vasodilators, such as ACh and SNP, and • 34 NO inhibiting 

35 vasoconstrictors, such as L-NAME. These studies were discussed in the preceding 

36 section (Oxidative Stress Response).
February 2012 5-250 Draft – Do Not Cite or Quote

Renin-Angiotensin-Aldosterone and Kininergic Systems

1 The adrenergic system also affects the renin-angiotensin-aldosterone system (RAAS), 

2 which is responsible for fluid homeostasis and BP regulation, and has been shown to be 

3 affected by Pb exposure. A meta-analysis found that Pb exposure (resulting in blood Pb 

4 levels: 30-40 μg/dL) increased plasma renin activity and renal tissue renin in young but 

5 not old rats (Vander, 1988). Exposure of experimental animals to Pb also induced 

6 increases in plasma, aorta, heart, and kidney angiotensin converting enzyme (ACE) 

7 activity; plasma kininase II, kininase I, and kallkrein activities; and renal AngII positive 

8 cells (Rodriguez-Iturbe et al., 2005; Sharifi et al., 2004; Carmignani et al., 1999). ACE 

9 activity declined over time while arterial pressure stayed elevated, suggesting that the 

10 RAAS may be involved in the induction, but not the maintenance of Pb-induced 

11 hypertension in rats.

12 Recent studies continued to implicate the RAAS in the development of Pb-induced 

13 hypertension, especially during early exposure in young animals. AngII, a main player in 

14 the RAAS, induces arteriolar vasoconstriction leading to increased BP. Pb exposure 

15 increased the vascular reactivity to AngII (Vargas-Robles et al., 2007). Acute 

16 (60 minutes) or short-term (7 days) exposure of Pb to rats increased the plasma ACE 

17 activity (Fiorim et al., 2011; Simões et al., 2011), and Fiorim et al. (2011) additionally 

18 found this increase to be correlated with the Pb-induced increase in systolic BP. 

19 However, at these short time points there were no changes in the AngII receptors 1 or 2 

20 protein levels or expression. Treatment with the AngII receptor (AT1R) blocker, losartan, 

21 or the ACE inhibitor, enalapril, blocked the Pb-induced systolic BP increase (Simões et 

22 al., 2011) and decreased the PHE-induced vasoconstrictor response in Pb-treated aortas

23 (Fiorim et al., 2011). Similarly, treatment with Losartan resulted in a greater decrease in 

24 systolic BP in highly Pb exposed rats (1% Pb, 40 days; blood Pb level >240 μg/dL after 

25 exposure, 12-13 μg/dL after chelation after 1 year) compared to control rats that 

26 continued into later periods of follow-up (day 283) (Bagchi and Preuss, 2005). Increased 

27 systolic BP after early exposure to Pb corresponded with increased water intake, urine 

28 output, potassium excretion, and decreased urinary sodium and urine osmolality. These 

29 functional changes in renal behavior are consistent with the actions of a stimulated 

30 RAAS. Lower level Pb (100 ppm, 14 weeks; range of blood Pb levels: 23.7-27 μg/dL) 

31 exposure increased renal cortical AngII content and the number of tubulointerstitial 

32 AngII-positive cells (Bravo et al., 2007). This heightened intrarenal angiotensin 

33 corresponded with sodium retention and increased systolic BP and was ablated by the 

34 anti-inflammatory antioxidant, MMF. Sodium reabsorption is important for the 

maintenance of BP, and Na+ 35 transporters play a key role in this process. In other studies, 

Pb exposure increased activity and levels of the α-1 subunit protein of Na+/K+- 36 ATPase, 

which plays a major role in Na+ 37 reabsorption and is regulated by the RAAS (Fiorim et al., 
February 2012 5-251 Draft – Do Not Cite or Quote

1 2011; Simões et al., 2011). These studies point to the activation of the RAAS in the 

2 course of Pb-induced hypertension, particularly in the early stages of elevated BP.

Vasomodulators 

3 The balance between production of vasodilators and vasoconstrictors is important in the 

4 regulation of BP and cardiovascular function. The 2006 Pb AQCD reported that Pb did 

5 not affect all vasomodulators in the same way. Urinary excretion of the vasoconstrictor, 

6 thromboxane (TXB2), and the vasodilatory prostaglandin, 6-keto-PGF1α, were 

7 unchanged in rats with Pb-induced hypertension (Gonick et al., 1998). However, in vitro 

8 Pb exposure promoted the release of the prostaglandin substrate, arachidonic acid, in 

9 vascular smooth muscle cells (VSMCs) via activation of phospholipase A2 (Dorman and

10 Freeman, 2002). Plasma concentration and urinary excretion of the vasoconstrictive 

11 peptide, endothelin (ET) 3 was increased after low (100 ppm), but not high-level 

12 (5,000 ppm) Pb exposure in rats (Gonick et al., 1997; Khalil-Manesh et al., 1994; Khalil13 Manesh et al., 1993b). Antagonism of the ET receptor A blunted the downregulation of 

14 sGC and cGMP production by Pb in isolated rat artery segments, suggesting that some of 

15 the hypertensive effects of Pb exposure may be mediated through ET (Courtois et al., 

16 2003). Additionally, Pb-exposed animals exhibited fluid retention and a 

17 concentration-dependent decline in the vasodilator, atrial natriuretic factor (ANF)

18 (Giridhar and Isom, 1990). Results from these studies suggest that Pb may interfere with 

19 the balance between vasodilators and vasoconstrictors that contribute to the complex 

20 hormonal regulation of vascular contraction and BP.

21 The imbalance in vasomodulators is one explanation for the concentration-dependent 

22 vasoconstriction observed in animals after Pb exposure (Valencia et al., 2001; Watts et 

23 al., 1995; Piccinini et al., 1977). Vasoconstriction after Pb exposure was not reported in 

24 all studies (Shelkovnikov and Gonick, 2001) and is likely varied depending on the type of 

25 vessel used, the Pb concentration employed, and the animal species being studied. 

Studies have reported Pb-induced attenuation of ACh- and • 26 NO-mediated vasodilation

27 (Marques et al., 2001; Oishi et al., 1996) in some, but not all vascular tissues and in 

28 some, but not all studies (Purdy et al., 1997). These effects have been variably attributed 

to Pb-mediated activation of PKC and direct action on the VSMCs through the Ca2+ 29

30 mimetic properties of Pb among other possibilities (Valencia et al., 2001; Watts et al., 

31 1995; Piccinini et al., 1977).

32 A recent study investigated the role of the endothelial-derived vasoconstrictor, ET-1, in 

33 Pb-induced hypertension. ET-1 from the endothelium acts on the ETA-type receptors 

34 located on the vascular smooth muscle layer and may be involved in vascular reactivity 

by • 35 NO and COX derivatives. Pb exposure (1 ppm, 24 hours) to rat aortic segments 
February 2012 5-252 Draft – Do Not Cite or Quote

decreased expression of sGC-β1 subunit, an enzyme involved in • 1 NO-induced 

2 vasodilation, and increased expression of COX-2 in an endothelium-dependent manner

3 (Molero et al., 2006). Even though Pb treatment did not alter ET-1 or ETA-type receptor 

4 protein expression in this system, blocking the ETA-type receptors partially reversed Pb5 induced changes in sGC and COX-2 in vascular tissue. This study suggests that the 

6 endothelium and ET-1 may contribute to Pb-induced hypertension through activation of 

7 ETA-type receptors that alter expression of COX-2 and sGC-β1 subunit, which affects 

• 8 NO signaling.

9 COX-2 blockade has been shown to prevent Pb-induced downregulation of sGC 

10 expression (Courtois et al., 2003). Inhibition of COX-2 also decreased the Pb-induced 

11 pressor response to ACh (Grizzo and Cordellini, 2008) and PHE (Silveira et al., 2010) in 

12 experimental animals. These studies suggest that Pb-induced vascular reactivity may 

13 depend on the participation of a COX-derived vasoconstrictor, such as prostaglandins, 

14 prostacyclins, or thromboxanes.

15 In summary, a small number of available recent studies continued to show that Pb 

16 exposure affect vasomodulatory pathways that are important for the maintenance of 

17 vascular tone; however, results indicated that not all vascular cell types are similarly 

18 affected by Pb exposure. Further, effects appeared to vary according to the concentration 

of Pb exposure. Pb exposure has been shown to interrupt baseline or endogenous • 19 NO20 mediated vasodilation of vessels via alterations in PKC, sGC, VSMC, endothelial cells, 

NADPH oxidase, and Ca+2 21 levels. Recent studies indicated that Pb exposure may affect 

22 vascular reactivity by increasing COX-2 and COX-2-dependent vasoconstrictors. Also, 

23 the vasoconstrictor endothelin may contribute to Pb-induced vasomodulation via similar 

pathways as • 24 NO including sGC and COX-2.

5.4.2.4 Summary of Blood Pressure and Hypertension

25 The 2006 Pb AQCD reported a clear association between higher blood Pb levels and 

26 higher BP. The effect was modest, but robust, as determined by a meta-analysis (Nawrot 

27 et al., 2002) of over 30 studies comprising over 40,000 adults (Figure 5-29). In the meta28 analysis, each doubling of concurrent blood Pb was associated with a 1 mmHg increase 

29 in systolic BP and a 0.6 mmHg increase in diastolic BP. Recent epidemiologic studies 

30 supported this conclusion at lower concurrent blood Pb levels (in populations with mean 

31 blood Pb levels < 2 μg/dL) and added to the evidence base regarding populations 

32 potentially at increased risk (i.e., high stress, genetic variants) and regarding associations 

33 of bone Pb levels with BP and hypertension in populations with mean bone Pb levels less 

34 than 20 μg/g. As these studies were mostly cross-sectional in design and were conducted 
February 2012 5-253 Draft – Do Not Cite or Quote

1 in adults whose concurrent blood Pb levels are influenced both by current Pb exposures 

2 and past Pb exposures mobilized from bone, uncertainty exists over the Pb exposure 

3 conditions that contributed to the associations observed between concurrent blood Pb 

4 level with increased BP and hypertension (Sections 4.3 and 4.7.3). 

5 A recent study in an ethnically diverse community-based cohort of women and men aged 

6 50-70 years of age found associations of both blood and tibia Pb levels with BP (Martin 

7 et al., 2006). This study also affirmed findings from other studies by demonstrating that 

8 with each increase of 1 μg/dL concurrent blood Pb level, systolic BP increased 1 mmHg 

9 and diastolic BP increased 0.5 mmHg and strengthened the evidence for an independent 

10 association with blood Pb level through the extensive examination of potential 

11 confounding. Additionally, recent epidemiologic studies provided evidence for 

12 associations in an adult cohort between blood Pb and BP and hypertension with relatively 

13 low blood Pb levels; a positive relationship was found in the NHANES adult data (1999-

14 2002) with a geometric mean blood Pb level of 1.64 μg/dL (Muntner et al., 2005).

15 However, as noted above, in adults, uncertainty exists regarding the magnitude, timing, 

16 frequency, and duration of Pb exposure that contribute to the associations observed with 

17 concurrent blood Pb levels. A new prospective study in Pb workers found independent 

18 associations of both baseline blood Pb level and subsequent changes in blood Pb over 

19 follow-up with changes in BP over follow-up and bone Pb level with hypertension

20 (Glenn et al., 2006). Although these Pb workers had higher current Pb exposure 

21 compared with nonoccupationally-exposed adults, the results indicated that different 

22 mechanisms may mediate shorter-term Pb-associated increases in BP and longer-term Pb23 associated development of hypertension.

24 In concordance with epidemiologic evidence, collectively, the animal toxicological 

25 studies providing blood Pb level and BP measurements reported higher BP with higher 

26 blood Pb levels (Figure 5-28). While the contribution of low concurrent blood Pb levels 

27 to the findings is difficult to ascertain in adults, animal toxicological studies provide 

28 support for low blood Pb level effects with increases in BP observed in groups of animals 

29 with blood Pb levels as low as 2 μg/dL (Rizzi et al., 2009; Tsao et al., 2000; Nakhoul et 

30 al., 1992). However, the majority of animal toxicological studies showing Pb-induced 

31 hypertension were conducted at higher Pb exposure levels that result in blood Pb levels > 

32 10 μg/dL. In addition, new animal evidence suggests the potential for increased BP 

33 following short-term (4 weeks) Pb treatment that included injected boluses (Fiorim et al., 

34 2011; Simões et al., 2011; Sharifi et al., 2004). New studies also demonstrated partial 

35 reversibility (not to levels in controls) of Pb-induced elevations in BP following Pb 

36 exposure cessation or chelation.
February 2012 5-254 Draft – Do Not Cite or Quote

Source: Reprinted with permission of MacMillan Press, Nawrot et al. (2002) \

Study Key: C - Caerphilly Study; HP - Welsh Heart Program; W - Whites; B - Blacks; NI - Non-immigrants; I - Immigrants; FW -

Foundry Workers; CS - Civil Servants; P - PheeCad (Public Health and Environmental Exposure to Cadmium) Study. 

Note: Circles represent individual groups and squares represent the combined association sizes. Open circles denote a 

nonsignificant association size that was assumed to be zero.

Figure 5-29 Change in systolic BP (SBP), in mmHg with 95% CI, associated 

with a doubling in the blood Pb concentration.

1 Epidemiologic studies continued to investigate the relationship between bone Pb and BP. 

2 A recently published meta-analysis (Figure 5-30) (Navas-Acien et al., 2008) included 

3 several studies (three prospective, five cross-sectional) which individually showed that 

4 bone Pb level was associated with systolic BP but not diastolic BP. In the cross-sectional 

5 studies, a pooled estimate indicated an increase in systolic BP of 0.26 mmHg (95% CI: 

6 0.02, 0.50) per 10 μg/g tibia Pb. In the longitudinal studies, a 0.33 mmHg (95% CI: -0.44, 

7 1.11) increase was estimated per 10 μg/g bone Pb. Most studies also reported associations 

8 of bone Pb with hypertension. Pooled estimates of 1.04 (95% CI: 1.01, 1.07) per 10 μg/g 
February 2012 5-255 Draft – Do Not Cite or Quote

1 increase in tibia Pb and 1.04 (95% CI: 0.96, 1.12) per 10 μg/g increase in patella Pb were 

2 reported. 

Source: Reprinted with permission of Elsevier Publishers, Navas-Acien et al. (2008)

In the Normative Aging Study, Hu et al. (1996b) reported the cross-sectional association between bone Pb levels and the 

prevalence of hypertension and Cheng et al. (2001) reported the cross-sectional association between bone Pb levels and systolic 

BP in study participants free of hypertension at baseline.

Note: The studies are ordered by increasing mean bone Pb levels. The area of each square is proportional to the inverse of the 

variance of the estimated change or log relative risk. Horizontal lines represent 95% confidence intervals. Diamonds represent 

summary estimates from inverse-variance weighted random effects models. Because of the small number of studies, summary 

estimates are presented primarily for descriptive purposes. RR indicates risk ratio.

Figure 5-30 Prospective and cross-sectional increase in systolic BP (SBP) 

and diastolic BP (DBP) and relative risk of hypertension per 

10 μg/g increase in bone Pb levels.

3 A few recent epidemiologic studies also emphasized the potential interaction between 

4 measures of long-term Pb exposure, i.e., bone Pb levels, and factors such as chronic 

5 stress and HFE genetic variants to moderate or modify the relationship of BP and 

6 hypertension with Pb. For example, among NAS men, tibia Pb level was associated with 

7 a larger risk of developing hypertension in an originally nonhypertensive group among 

8 men with higher self-reported stress (Peters et al., 2007).
February 2012 5-256 Draft – Do Not Cite or Quote

1 In addition to stress, recent epidemiologic studies investigated effect modification by 

2 race/ethnicity and genetic variants. In the NHANES 1988-1994 population of adults, the 

3 association of concurrent blood Pb with systolic BP was higher among Mexican 

4 Americans. In the same NHANES population, the association between blood Pb level and 

5 hypertension was higher among non-hispanic Blacks with the ALAD2 allele (Figure 5-25

6 and Figure 5-26 for results) (Scinicariello et al., 2010). Additionally, the association 

7 between blood Pb and PP was larger among NAS men with the HFE H63D variant 

8 (Figure 5-25). PP represents a good predictor of cardiovascular morbidity and mortality 

9 and an indicator of arterial stiffness (Zhang et al., 2010a). The aforementioned genes are 

10 related to iron metabolism and have been linked with differences in Pb distribution in 

11 blood and bone. Park et al. (2009b) provided further evidence of variants in iron 

12 metabolism genes impacting the association of bone Pb levels with QT interval changes 

13 (Table 5-19 for results). 

14 Animal toxicological evidence continued to build on the evidence characterizing the 

15 mechanisms leading to these Pb-induced cardiovascular alterations. Biological 

16 plausibility for the consistent associations observed between blood and bone Pb and 

17 cardiovascular effects is provided by enhanced understanding of Pb-induced oxidative 

stress including • 18 NO inactivation, endothelial dysfunction leading to altered vascular 

19 reactivity, activation of the RAAS, and vasomodulator imbalance.

5.4.3 Vascular Effects and Cardiotoxicity

20 Not only has Pb been shown to increase BP and alter vascular reactivity, but Pb can alter 

21 cardiac function, initiate atherosclerosis, and increase cardiovascular mortality. Past 

22 toxicological studies have reported that Pb can increase atheromatous plaque formation in 

23 pigeons, increase arterial pressure, decrease heart rate and blood flow, and alter cardiac 

24 energy metabolism and conduction (Prentice and Kopp, 1985; Revis et al., 1981). A 

25 limited number of available epidemiologic studies discussed in the 2006 Pb AQCD 

26 provided evidence of associations of blood Pb level with ischemic heart disease (IHD) 

27 and peripheral artery disease (PAD).

5.4.3.1 Effects on Vascular Cell Types

28 The endothelial layer is an important constituent of the blood vessel wall, which regulates 

29 macromolecular permeability, VSMC tone, tissue perfusion, and blood fluidity. Damage 

30 to the endothelium is an initiating step in development of atherosclerosis, thrombosis, and 

31 tissue injury. Given that epidemiologic and toxicological evidence suggests that long-
February 2012 5-257 Draft – Do Not Cite or Quote

1 term Pb exposure is associated with a number of these conditions, numerous 

2 toxicological studies have investigated and found an effect of Pb on endothelial 

3 dysfunction. A recent occupational study found that endothelial function assessed by 

4 flow-mediated dilatation was impaired in highly Pb exposed workers (mean blood Pb 

5 levels: 24.1 in workers versus 7.8 μg/dL in unexposed controls) (Poręba et al., 2010).

6 The endothelial layer makes up only a small part of the vascular anatomy; the majority of 

7 the vessel wall is composed of VSMCs, which work in concert with the endothelial cells 

8 (EC) in contraction and relaxation of the vessel, local BP regulation, and atherosclerotic 

9 plaque development. Since Pb has been shown repeatedly to result in hypertension and 

10 vascular disease in experimental animals, studies continued to investigate and find an 

11 effect of Pb on VSMCs.

12 In in vitro assays, Pb (50 μM, 2 weeks) stimulated VSMC invasiveness in isolated human 

13 arteries leading to the invasion of medial VSMC into the vessel intima and development 

14 of intimal hyperplasia, a key step in atherosclerotic progression (Zeller et al., 2010). In 

15 addition, treatment with Pb (50 μM, 12 hours) promoted VSMC elastin expression and 

16 increased arterial extracellular matrix in isolated human arteries. VSMC invasiveness was 

17 also increased in culture by treatment with supernatant of Pb-treated human EC (50 μM), 

18 suggesting that Pb-exposed ECs secrete an activating compound. This compound was 

19 confirmed to be IL-8. Pb exposure (5-50 μM) was able to, in a concentration-dependent 

20 manner, increase IL-8 synthesis and secretion in human umbilical vein EC cultures 

21 through activation of the transcription factor Nrf2. Neutralization of IL-8 could block 

22 VSMC invasion and arterial intima thickening (Zeller et al., 2010). This study provides 

23 evidence that Pb exposure stimulates ECs to secrete IL-8 in an Nrf2-dependent manner 

24 that stimulates VSMC invasion from the vessel media to intima leading to a vascular 

25 thickening and possibly atherogenesis.

26 A number of CVDs, including atherosclerosis, are characterized by increased 

27 inflammatory processes. Numerous studies have shown that Pb exposure is associated 

28 with an inflammatory environment in vascular tissues of humans and animals as indicated 

29 by higher levels of inflammatory mediators like prostaglandin E2 (PGE2). Human aortic 

30 VSMCs treated with Pb (1 μM, 1-12 hours) exhibited increased secretion of PGE2 time31 dependently through enhanced gene transcription (Chang et al., 2011). This was preceded 

32 by a Pb-induced increase in the gene expression of the rate limiting enzymes in the 

33 regulation of prostaglandins, cytosolic phospholipase A2 (cPLA2) and COX-2. The 

34 induction of these enzymes was mediated by activation of ERK1/2, MEK1, and MEK2. 

35 Further investigation into the entrance of Pb into the cell revealed that inhibition of the 

36 store-operated calcium channels (SOC) could only partially suppress cPLA2 and COX 

37 activation by Pb; however inhibition of epidermal growth factor receptor (EGFR) 
February 2012 5-258 Draft – Do Not Cite or Quote

1 attenuated Pb-induced PGE2 secretion and activation of cPLA2 and COX. A follow-up to 

2 this study found that Pb treatment (1μM) of a human epithelial cell line increased COX-2 

3 gene expression, promoter activity, and protein (Chou et al., 2011). Inhibition of NF-κB 

4 decreased the Pb-induced COX-2 activation; whereas EGFR inhibition blocked COX-2 

5 upregulation and NF-κB nuclear translocation. Overall these studies suggest that Pb can 

6 induce pro-inflammatory events in VSMC in the form of increased PGE2 secretion and 

7 cPLA2 and COX-2 expression through activation of EGFR via ERK1/2 and NF-κB 

8 pathways.

9 Damage to the endothelium is a hallmark event in the development of atherosclerosis. 

10 Past studies have shown that Pb exposure results in de-endothelialization, impaired 

11 proliferation, and inhibition of endothelium repair processes after injury (Fujiwara et al., 

12 1997; Ueda et al., 1997; Kaji et al., 1995; Kishimoto et al., 1995). However, Pb exposure 

13 was not found to lead to nonspecific cytotoxicity at low exposure levels (2-25 μM) as 

14 shown by the lack of release of lactate dehydrogenase (LDH) from Pb-treated bovine 

15 aortic EC (Shinkai et al., 2010). Instead, Pb induced specific cytotoxicity (caspase3/7 

16 activation) through endoplasmic reticulum (ER) stress that was protected against by the 

17 ER chaperones glucose-regulated protein 78 (GRP78) and glucose-regulated protein 94 

18 (GRP94). GRP78 and GRP94 play key roles in the adaptive unfolded protein response 

19 that serves as a marker of and acts to alleviate ER stress. Exposure of Pb to ECs induced 

20 GRP78 and GRP94 gene (2-25 μM) and protein (GRP78 [5-25 μM] and GRP94 [10-

21 25 μM]) expression through activation of the IRE1-JNK-AP-1 pathways (Shinkai et al., 

22 2010). This study suggests that the functional damage caused by Pb exposure to EC may 

23 be partly attributed to induction of ER stress.

5.4.3.2 Cholesterol

24 As blood cholesterol rises so does the risk of coronary heart disease. Early occupational 

25 studies (Ademuyiwa et al., 2005a; Bener et al., 2001a; Kristal-Boneh et al., 1999)

26 examining higher than current adult blood Pb levels reported higher total cholesterol 

27 levels related to Pb exposure, but mixed results for HDL, LDL, and triglycerides. More 

28 recently, Poreba et al. (2010), in an occupational study, reported no significant 

29 differences in parameters of lipid metabolism between Pb exposed workers (mean blood 

30 Pb level: 25 μg/dL) and unexposed individuals. Conversely, Kamal et al. (2011) reported 

31 that occupational Pb exposure (mean blood Pb level: >40 μg/dL) was associated with 

32 higher levels of triglycerides, total cholesterol, and LDL, and decreased HDL-C. Other 

33 Pb studies adjusted models for total cholesterol to control for this coronary heart disease 

34 risk factor. Higher mean total cholesterol with higher blood Pb levels has been reported

35 (Menke et al., 2006). In developing models to predict bone Pb levels, Park et al. (2009a)
February 2012 5-259 Draft – Do Not Cite or Quote

1 noted that total and HDL cholesterol were selected as 2 of 18 predictors for the bone Pb 

2 level model. Their findings suggested that higher Pb exposure may be associated with 

3 higher total and HDL cholesterols. A recent study reported increased LDL and decreased 

4 HDL in rats treated with Pb (20 ppm, i.p., 3 days/week, 8 weeks) (Roshan et al., 2011).

5 The major risk factor that lipids represent for heart disease make relating lipid levels to 

6 Pb exposures an interesting but challenging hypothesis to test. 

5.4.3.3 Heart Rate Variability

7 Pb has been shown not only to affect vascular contractility in animals, but also is 

8 associated with cardiac contractility. Eum et al. (2011) and Park et al. (2009b) followed 

9 up a previous NAS report (Cheng et al., 1998), which found increasing duration of 

10 corrected QT interval (QTc) with increasing bone Pb levels in men <65 years, but not in 

11 men ≥ 65 years. Eum et al. (2011) prospectively examined the association between blood 

12 and bone Pb levels and the development of electrocardiographic (ECG) conduction 

13 abnormalities among 600 men who were free of ECG abnormalities at the baseline 

14 assessment. A second ECG was obtained for 496 men 8.1 (SD = 3.1) years later on 

15 average. Baseline Pb concentrations in blood (mean [SD]: 5.8 [3.6] μg/dL), patella bone 

16 (mean [SD]: 30.3 [17.7] μg/g), and tibia bone (mean [SD]: 21.6 [12.0] μg/g) were similar 

17 to those found in other samples from the general U.S. adult population and much lower 

18 than those reported in occupationally exposed groups. Higher tibia Pb was associated 

19 with increases in QTc interval and QRSc duration. Compared with those in the lowest 

20 tertile of baseline tibia Pb (< 16 μg/g), participants in the highest tertile (> 23 μg/g) had a 

21 7.94 msec (95% CI: 1.42, 14.45) greater increase in QTc interval and a 5.94 msec (95% 

22 CI: 1.66, 10.22) greater increase in QRSc duration over 8 years after adjusting for 

23 covariates. There were no statistically significant associations with patella or blood Pb 

24 levels. These associations were observed in men with relatively low blood and bone Pb 

25 concentrations who were free of cardiac conduction abnormalities at baseline and were 

26 examined prospectively. Thus, they indicate that long-term cumulative Pb exposure may 

27 increase the risk of developing cardiac abnormalities. Uncertainty exists as to the specific 

28 Pb exposure level, timing, frequency, and duration contributing these associations 

29 observed for tibia Pb levels. A recent occupational study reported lower HRV and 

30 abnormal parameters of heart rate turbulence in Pb-exposed workers (mean blood Pb 

31 levels: ~25 μg/dL) compared to control subjects (Poręba et al., 2011a).

32 Park et al. (2009b) examined whether polymorphisms in genes known to alter iron 

33 metabolism (HFE, transferrin [TF] C2, heme oxygenase-1 [HMOX-1]) modify the 

34 association between Pb biomarker levels and the QT interval. Investigators examined 

35 associations in data stratified on polymorphisms in the three genes. They also analyzed 
February 2012 5-260 Draft – Do Not Cite or Quote

1 interaction models with cross-product terms for genotype and the Pb biomarker. The 

2 distributions of all genotypes but the HFE variant, H63D, were in Hardy-Weinberg 

3 equilibrium. Subjects homozygous for the other HFE variant, C282Y, had higher bone Pb 

4 levels and those homozygous for H63D and heterozygous for both C282Y and H63D had 

5 lower bone Pb levels. The HMOX-1 L variant (longer repeats of GT, associated with 

6 lower enzyme inducibility) alone, compared to the wild type, showed a statistically 

7 significant interaction with tibia Pb (11.35 msec longer QTc interval for each 13 μg/g 

8 increase in bone Pb in L-allele variants). No other gene variant alone showed different 

9 Pb-associated QTc intervals from those in wild types, either for tibia and patella Pb or for 

10 (linear) concurrent blood Pb. Lengthening of QTc with higher tibia and blood Pb was 

11 more pronounced with an increase in the total number of gene variants, driven by a joint 

12 effect between HFE variant and HMOX-1 L allele. There was a trend observed with 

13 blood and tibia Pb-associated QTc interval increasing with increasing number of gene 

14 variants from 0 to 3. This study provided further evidence of gene variants modifying 

15 associations of Pb biomarkers with cardiovascular effects. 

16 The interaction of key markers of the metabolic syndrome with bone Pb levels in 

17 affecting HRV was investigated in a group of 413 older adults with patella Pb 

18 measurements in the NAS (Park et al., 2006). Metabolic syndrome was defined to include 

19 three or more of the following: waist circumference >102 cm, hypertriglyceridemia 

20 (>150 mg/dL), low HDL cholesterol (<40 mg/dL in men), high BP >130/85 mmHg, and 

21 high fasting glucose (>110 mg/dL). Men using antihypertensive medication or diabetes 

22 medications were counted as high BP or high fasting glucose, respectively. The strongest 

23 relationships between patella Pb levels and lower HRV were observed among those with 

24 three or more metabolic abnormalities. A trend was observed for larger patella Pb25 associated decreases in HRV with increasing number of metabolic abnormalities. These 

26 results suggest multiplicative effects of cumulative Pb exposure and metabolic 

27 abnormalities on key predictors of CVD. Park et al. (2006) also reported the penalized 

28 spline fits to bone Pb in models assessing only main effects of bone Pb. The optimal 

29 degree of smoothing determined by the generalized cross-validation criterion for all HRV 

30 measures was 1, which indicated that the associations were nearly linear. The spline fits 

31 and associated statistics showed that the bone Pb main effects on HRV measures were 

32 linear. However, the relationship with LF/HF was linear with log(LF/HF).

33 Increased incidence of arrhythmia and atrioventricular conduction block was found in 

34 rats after 12 weeks of Pb exposure (100 ppm; mean blood Pb level 26.8 μg/dL) (Reza et 

35 al., 2008). Also, Pb exposure for 8 weeks increased heart rate and systolic BP. These 

36 increases corresponded with increased cardiac contractile force and prolonged ST 

37 interval, without alteration in QRS duration or coronary flow. In contrast, another study 

38 found that Pb (100 μM) exposure, in a concentration-dependent manner, reduced 
February 2012 5-261 Draft – Do Not Cite or Quote

myocardial contraction using rat right ventricular strips by reducing sarcolemmal Ca2+ 1

2 influx and myosin ATPase activity (Vassallo et al., 2008). This study also found that Pb 

3 exposure changed the response to inotropic agents and blunted the force produced during 

4 contraction. Conversely, past studies have found that Pb exposure increases intracellular 

Ca2+ 5 content (Lal et al., 1991; Favalli et al., 1977; Piccinini et al., 1977), which could 

6 result in increased cardiac output and hypertension.

5.4.3.4 Peripheral Artery Disease 

7 Peripheral artery disease (PAD) is an indicator of atherosclerosis and measured by the 

8 ankle brachial index, which is the ratio of BP between the posterior tibia artery and the 

9 brachial artery. PAD is typically defined as an ankle brachial index of less than 0.9. 

10 Muntner et al. (2005), whose results describing the association of blood Pb and 

11 hypertension in the NHANES 1999-2002 data set for adults were discussed previously, 

12 also examined the association of blood Pb with PAD. The authors observed an increasing 

13 trend in the odds of PAD with increasing concurrent blood Pb level. The OR for PAD 

14 comparing the fourth quartile of blood Pb (> 2.47 μg/dL) to the first quartile of blood Pb 

15 (< 1.06 μg/dL) was 1.92 (95% CI: 1.02, 3.61). These results are consistent with those 

16 from a previous NHANES analysis conducted by Navas-Acien et al. (2004).

17 Navas-Acien et al. (2004) reported a trend of increasing OR for PAD with increasing 

18 quartile of concurrent blood Pb or Cd in adults who were 40 years of age in the 1999-

19 2000 NHANES population. These authors tested both Pb and Cd in separate models, 

20 tested the metals simultaneously, and tested the interaction between the metals. The 

21 correlation coefficient between natural log Pb and natural log Cd was 0.32 (p <0.001). 

22 Although the interaction was not statistically significant, when blood Pb and blood Cd 

23 were in the same model, the ORs were diminished slightly but both showed statistically 

24 significant trends of increasing OR with increasing quartile of the metal. These results 

25 indicate that blood Cd levels did not confound the association between blood Pb level and 

26 PAD. In a subsequent analysis, Navas-Acien et al. (2005) used the same 1999-2000 

27 NHANES dataset, but constructed PAD models using a suite of urine metal 

28 concentrations. Power was reduced in this study because only 659-736 subjects 

29 (compared to 2,125) had spot urine metal tests in the data set. Urinary Cd, but not urinary 

30 Pb, was consistently associated with PAD in all models. Associations also were observed 

31 with urinary antimony and tungsten. Spot urine Pb measurements are less reliable 

32 compared to blood Pb measurements. In Navas-Acien et al. (2005), the urinary Pb level 

33 association with PAD was sensitive to adjustment for urinary creatinine, indicating that 

34 spot urine Pb measurements are affected by differences in urine dilution. This finding 
February 2012 5-262 Draft – Do Not Cite or Quote

1 illustrates the limited reliability of spot urine Pb measurements compared to blood Pb 

2 measurements. 

5.4.3.5 Ischemic Heart Disease

3 A few studies discussed in the 2006 Pb AQCD indicated associations between Pb 

4 biomarker levels and increased risk of cardiovascular outcomes associated with IHD, 

5 including left ventricular hypertrophy (Schwartz, 1991) and myocardial infarction

6 (Gustavsson et al., 2001). Recently, Jain et al. (2007) reported on the incidence of IHD 

7 (physician confirmed MI, angina pectoris) among older adult males enrolled in the NAS

8 and followed during the period of September, 1991 to December, 2001. All subjects had 

9 blood Pb and bone Pb measurements with no IHD at enrollment. Fatal and nonfatal cases 

10 were combined for analysis. Baseline blood, tibia, and patella Pb levels were log11 transformed. Blood Pb level and patella Pb level were associated with increased risk of 

12 IHD over the 10-year follow-up period. When blood Pb and patella Pb were included 

13 simultaneously in the model, each of their HRs was only moderately attenuated (HR: 

14 1.24 [95% CI: 0.80, 1.93] per SD increase in blood Pb and HR: 2.62 [95% CI: 0.99, 6.93] 

15 per SD increase in patella Pb). When blood Pb and tibia Pb were included simultaneously 

16 in the model, their risk estimates were only moderately attenuated (HR: 1.38 [95% CI: 

17 0.89, 2.13] per SD increase in blood Pb and HR: 1.55 [95% CI: 0.44, 5.53] per SD 

18 increase in tibia Pb). These findings indicate that both blood and bone Pb levels 

19 contribute independently to IHD incidence. 

20 IHD, characterized by reduced blood supply to the heart, may result from increased 

21 thrombosis. A recent animal study suggested that Pb exposure promotes a procoagulant 

22 state that could contribute to thrombus formation (Shin et al., 2007). In a rat model of 

23 venous thrombosis, Pb treatment (i.v. 25 mg/kg) resulted in increased thrombus 

24 formation. Additionally, Pb treatment to human erythrocytes (red blood cells, RBCs) 

25 increased coagulation at a dose of 5 μM and thrombin generation in a concentration26 dependent manner at doses from 2-5μM. This enhanced procoagulant activity in Pb27 treated RBCs was the result of increased outer cell membrane phosphatidylserine (PS) 

28 surfacing (human RBCs: 2-5 μM; rat RBCs: 5 μM). Similar to these in vitro results, PS 

29 externalization on erythrocytes was increased in Pb-treated rats (i.v. 50-100 mg/kg, not 

30 25 mg/kg). Increased PS externalization was likely the result of increased intracellular 

31 calcium (5 μM Pb), enhanced scramblase activity (5-10 μM Pb), inhibited flippase 

32 activity (5-10 μM Pb), and ATP depletion (1-5 μM Pb) after Pb exposure (Shin et al., 

33 2007).
February 2012 5-263 Draft – Do Not Cite or Quote

5.4.3.6 Atherosclerosis

1 Epidemiologic and toxicological studies provide evidence for increased atherosclerosis 

2 and intimal medial thickening (IMT) due to Pb exposure. The association between stroke 

3 subtypes and severity of cerebral atherosclerosis was examined in relation to a single 

4 concurrent blood Pb level and total 72-hour urinary Pb level (body Pb store-EDTA 

5 mobilization test) in a cross-sectional study of 153 patients (mean age 63.7 years) 

6 receiving digital subtraction angiography in Chang Gung Memorial Hospital in Taiwan 

7 from 2002 to 2005 (Lee et al., 2009). In an analysis adjusted for age, sex, hypertension, 

8 diabetes, triglyceride, uric acid, smoking, and alcohol consumption, a 1 μg increase in 

9 urine Pb was associated with ≥ 50% stenosis in the intracranial carotid system with an

10 OR (95% CI) of 1.02 (1.00, 1.03). Urine Pb was associated with greater stenosis in the 

11 extracranial or vertebrobasilar systems. Blood Pb level was not associated with greater 

12 stenosis in any region. As the development of atherosclerosis is a lifelong process, body 

13 Pb stores, analyzed by total 72-hour urine Pb amount, may more strongly be associated 

14 with atherosclerosis than are single blood Pb measurements. 

15 A recent study correlated greater carotid artery IMT with higher concurrent serum Pb 

16 levels (mean [SD] 0.41 [0.38] μg/dL) in hemodialysis patients (Ari et al.). A few 

17 available recent occupational studies also presented evidence for increased measures of 

18 atherosclerosis in highly Pb exposed adult populations with mean blood Pb levels around 

19 25 μg/dL. Poręba et al. (2011b) reported increased local arterial stiffness and more 

20 frequent left ventricular diastolic dysfunction in Pb-exposed workers with hypertension 

21 compared to nonexposed controls with hypertension. Occupational exposure to Pb (mean 

22 blood Pb levels: 24 μg/dL in workers, 8.3 μg/dL in nonexposed group) was also 

23 associated with greater IMT and atherosclerotic plaque presentation, analyzed by Doppler 

24 ultrasound (Poreba et al., 2011).

25 Zeller et al. (2010) examined human radial and internal mammary arteries exposed to Pb 

26 in culture and reported a concentration-dependent increase in arterial intimal thickness 

27 (statistically nonsignificant at 5 μM Pb, significant at 50 μM Pb, 2 week treatment) and 

28 intimal extracellular matrix accumulation (50 μM). Also, Pb promoted EC proliferation 

29 (5 and 50 μM, 72 hours) and VSMC elastin expression (50 μM, 12 hours), as discussed 

30 above (Section 5.4.3.1) (Zeller et al., 2010). Another study showed that Pb exposure 

31 (100 ppm in drinking water for 10 months; mean blood Pb level 28.4 μg/dL) of rats also 

32 increased the aortic media thickness, media-lumen ratio, and medial collagen content

33 (Zhang et al., 2009a). These morphological changes to the vessel due to Pb exposure 

34 indicate initiation of arteriosclerosis and could be the cause of decreased contractile 

35 response of the vessel due to altered visco-dynamic vessel properties. Alternatively, these 

36 vascular changes could be an effect of Pb-induced hypertension. 
February 2012 5-264 Draft – Do Not Cite or Quote

Table 5-19 Characteristics and quantitative data for associations of blood and 

bone Pb with other CVD measures in epidemiologic studies ordered 

as they appear in the text

Study

Population/

Location Parameter Pb Data Statistical Analysis Effect Estimate (95% CI)a

Heart rate variability

Eum et al. (2011) 600 men free of 

electrographic 

abnormalities at the 

time of baseline ECG 

from NAS in Greater 

Boston, MA area (496 

with follow-up ECG 8 

years later)

ECG 

conductionb

(QTc, QRSc, 

JTc, QT 

prolongation, 

JT 

prolongation, 

IVCDc

, AVCD, 

Arrhythmia)

Baseline Blood Pb:

Mean (SD): 5.8 (3.6) μg/dL

Baseline Patella Pb:

Mean (SD): 30.3 (17.7) μg/g

Baseline Tibia Pb:

Mean (SD): 21.6 (12.0) μg/g

Q1: <16 μg/g (n = 191)

Q2: 16.0 - 23 μg/g (n = 208)

Q3: >23 μg/g (n = 195)

Repeated measures linear 

regression adjusted for age, 

education, smoking, BMI, 

albumin-adjusted serum 

calcium, and diabetes status at 

baseline, and years between 

ECG tests and QT-prolongation 

drugs at the time of ECG 

measurement.

Tibia Pb:

Adjusted 8-year change (95% 

CI):

QTc:

Q2 vs. Q1 (reference): 7.49 

(1.22, 13.75) msec,

Q3 vs. Q1: 7.94 (1.42, 14.45) 

msec

p for trend = 0.03

QRSc:

Q2 vs. Q1: 0.52 (-3.60, 4.65) 

msec

Q3 vs. Q1: 5.94 (1.66, 10.22) 

msec

p for trend = 0.005

No associations with patella or 

blood Pb

Park et al. (2009b) 613 men from NAS in 

Greater Boston, MA 

area (8/1991 -

12/1995)

QTcb interval Baseline Blood Pb:

Median (IQR): 5 (4-7) μg/dL

Baseline Patella Pb:

Median (IQR): 26 (18-37) μg/g

Baseline Tibia Pb:

Median (IQR): 19 (14-27) μg/g

Linear regression models 

adjusted for age, BMI, smoking 

status, serum calcium, and 

diabetes.

Per IQR (3 μg/dL) increase in 

blood Pb

1.3 (-0.76, 3.36) msec after 8-

year follow up

Per IQR (19 μg/g) increase in 

patella Pb

2.64 (0.13, 5.15) msec

Per IQR (13 μg/g) increase in 

tibia Pb

2.85 (0.29, 5.40) msec

Park et al. (2006) 413 men from NAS in 

Greater Boston, MA 

area (11/14/2000 -

12/22/2004)

HRV

(SDNN, HF, 

HFnorm, LF, 

LFnorm, 

LF/HF)

Baseline Patella Pb (measured 

within 6 mo of HRV:

Median (IQR): 23.0 (15-34) μg/g

Estimateda

: Median (IQR): 

16.3 (10.4-25.8) μg/g

Baseline Tibia Pb:

Median (IQR): 19.0 (11-28) μg/g

Log linear regression models 

adjusted for age, cigarette 

smoking, alcohol consumption, 

room temperature, season 

(model 2) BMI, fasting blood 

glucose, HDL cholesterol, 

triglyceride, use of β-blockers, 

calcium channel blockers, 

and/or ACE inhibitors

Tibia Pb: Model 2

Change (95%CI)

HF: –0.9 (–3.8, 2.1) normalized 

units (nu) LF: 0.9 (–2.0, 3.9) nu

Log LF/HF: 3.3 (–10.7, 19.5) 

(%)

Per 17 μg/g tibia Pb

Patella Pb: Model 2 Change 

(95%CI)

HF: –0.6 (–3.1, 1.9) nu

LF: 0.6 (–1.9, 3.1) nu

Log LF/HF: 3.0 (–8.7, 16.2) (%)

Per 15.4 μg/g patella Pb

Effect estimates were more 

pronounced among those with 

metabolic syndrome.

Peripheral artery disease
February 2012 5-265 Draft – Do Not Cite or Quote

Study

Population/

Location Parameter Pb Data Statistical Analysis Effect Estimate (95% CI)a

Muntner et al. (2005) 9,961 NHANES (1999- 2002) participantsPAD Range Concurrent Blood Pb:

Q1: <1.06 μg/dL, 

Q2: 1.06-1.63 μg/dL 

Q3: 1.63-2.47 μg/dL 

Q4: >2.47 μg/dL

Logistic regression models 

adjusted for age, race/ethnicity, 

sex, diabetes mellitus, BMI, 

cigarette smoking, alcohol 

consumption, high school 

education, health insurance 

status

OR (95% CI): 

Q1: 1.00 (Reference), 

Q2: 1.00 (0.45, 2.22), 

Q3: 1.21 (0.66, 2.23), 

Q4: 1.92 (1.02, 3.61)

Navas-Acien et al.

(2005)

790 participants, age ≥ 

40 yr, from NHANES 

(1999-2000)

PAD Concurrent urinary Pb:

Mean (10th-90th %): 0.79 μg/L 

(0.2-2.3)

Logistic regression adjusted for 

the following:

Model 1: age, sex, race, and 

education

Model 2: covariates above plus 

smoking status

Model 3:covariates above plus 

urinary creatinine

Model 1: OR: 1.17 (0.81, 1.69) 

Model 2: OR: 1.17 (0.78, 1.76) 

Model 3: OR: 0.89 (0.45, 1.78)

Per IQR increase in urinary Pb

Array of metals in urine also 

evaluated.

Ischemic Heart Disease

Jain et al. (2007) 837 men from NAS in 

Greater Boston, MA 

area (1991-2001)

IHD

(MI or angina 

pectoris)

Baseline Blood Pb Mean (SD): 

Non-cases 6.2 (4.3) μg/dL; 

Cases 7.0 (3.8) μg/dL

Baseline Patella Pb Mean (SD): 

Non-cases 30.6 (19.7) μg/dL; 

Cases 36.8 (20.8) μg/dL

Baseline Tibia Pb Mean (SD): 

Non-Cases 21.4 (13.6) μg/g; 

Cases 24.2 (15.9) μg/g

Cases:

Blood Pb range: 1.0 to 20.0 μg/dL

Patella Pb range: 5.0 to 101 μg/g

Tibia Pb range: -5 to 75 μg/g

Cox proportional hazards 

models adjusted for age, BMI, 

education, race, smoking 

status, pack-years smoked, 

alcohol intake, history of 

diabetes mellitus and 

hypertension, family history of 

hypertension, DBP, SBP, serum 

triglycerides, serum HDL, and 

total serum cholesterol

Blood Pb level ≥ 5 μg/dL

OR over 10-year follow-up: 1.73 

(1.05, 2.87)

Ln blood Pb OR: 1.45 (1.01, 

2.06)

Ln patella Pb level OR: 2.64 

(1.09, 6.37)

Ln tibia Pb level OR: 1.84 (0.57, 

5.90)

Per 1 SD increase in Pb 

biomarker

a

Estimated patella Pb accounts for declining trend in patella Pb levels between analysis of bone Pb and HRV.

b

Heart-rate-corrected QT interval calculated by Bazett’s formula

c

IVCD, intraventricular conduction defect; AVCD, atrioventricular conduction defect

5.4.3.7 Summary of Vascular Effects and Cardiotoxicity

1 There are a limited number of studies that investigate the associations between Pb 

2 biomarkers and cardiovascular effects other than BP or hypertension (Table 5-19). As 

3 presented in Table 5-19, these studies demonstrated associations between various 

4 biomarkers of Pb exposure and clinical cardiovascular outcomes such as atherosclerosis, 

5 IHD, PAD, and HRV occurrence in adult populations after adjusting for potential 

6 confounding by variables such as age, sex, BMI, smoking, alcohol consumption, and 

7 diabetes. In a limited body of studies, mixed evidence of association between 

8 occupational exposure to Pb and altered cholesterol was reported. Studies of Pb-induced 

9 endothelial dysfunction, VMSC invasiveness, and inflammation in isolated vascular 

10 tissues and cells provide mechanistic evidence to support the biological plausibility of 

11 these other vascular effects and cardiotoxicity. A recent study provided evidence for the 
February 2012 5-266 Draft – Do Not Cite or Quote

1 interaction between biomarkers of Pb exposure and the HFE C282Y and HMOX-1 L 

2 variant on the prolonged QT interval in nonoccupationally-exposed older men (Park et 

3 al., 2009b). Also, in the NAS population, bone Pb levels were associated with larger 

4 decreases in HRV parameters among subjects identified as having metabolic 

5 abnormalities (Park et al., 2006). These metabolic abnormalities, abdominal obesity, 

6 hypertriglyceridemia, low HDL cholesterol, high BP/medication use, or high fasting 

7 glucose, have been shown to be associated with increased risk of cardiovascular events.

8 A prospective NAS study reported that higher baseline tibia Pb was associated with 

9 increases in QTc interval and QRSc duration over an 8-year follow-up period (Eum et al., 

10 2011). Concurrent blood Pb levels (population means >2.5 μg/dL) were associated with 

11 greater odds of PAD in adults in NHANES analyses (Muntner et al., 2005; Navas-Acien 

12 et al., 2004). In addition, in the NAS cohort of older adult men, blood Pb (≥ 5 μg/dL) and 

13 patella Pb levels were associated with increased morbidity from IHD (Jain et al., 2007).

14 A recent study involving both human and toxicological studies elucidated mechanisms 

15 for observed Pb-mediated arterial IMT, an early event in Pb-induced atherogenesis

16 (Zeller et al., 2010). Studies in isolated tissues and cells found that Pb stimulated the 

17 synthesis and secretion of IL-8 in ECs, which was responsible for stimulating VSMC 

18 invasion into the vessel intimal layer. Pb treatment also increased extracellular matrix and 

19 elastin, primary sites for lipid deposition in the vessel wall. Overall, the relatively 

20 available few studies provide support for associations between Pb biomarkers and other 

21 cardiovascular conditions, yet further research is warranted to understand these 

22 relationships.

5.4.4 Cardiovascular Function and Blood Pressure in Children

5.4.4.1 Introduction

23 The study of cardiovascular function effects in relation to blood Pb levels in children

24 potentially offers unique information on several topics. First, by examining endpoints 

25 predictive of future cardiovascular pathology, these studies may offer information on the 

26 potential cardiovascular effects of Pb exposure in an understudied population. Second, 

27 examination of cardiovascular changes that are antecedent to increased BP and changes 

28 in other CVD-related endpoints at later lifestages may inform uncertainties in regards to 

29 the time course of cardiovascular changes associated with Pb exposure. Finally, these 

30 studies address gaps in knowledge regarding Pb exposure effects in populations with 

31 mean blood Pb levels in the range of < 5 to 10 μg/dL.
February 2012 5-267 Draft – Do Not Cite or Quote

1 An advantage of examining the literature about the association between cardiovascular 

2 effects and blood Pb levels in children is that the blood Pb levels of children in new 

3 studies may better reflect their relatively recent Pb exposure and its effect on CVD than 

4 blood Pb levels do in adults because of the uncertainties related to the effects of earlier 

5 higher Pb exposure levels. A possible disadvantage of studying the associations of blood 

6 Pb levels and cardiovascular effects in children is the lower prevalence of such conditions 

7 in children. For example, the prevalence for hypertension in children (9 to 10 years old) 

8 ranges from to 2 to 5 percent (Daniels, 2011; Steinthorsdottir et al., 2011). The 

9 prevalence of hypertension increases with age. More than half of people aged 60 to 69 

10 years have hypertension (Chobanian et al., 2003). Further, compensatory mechanisms in 

11 children may be more active than in adults, and the cardiovascular tissue of the young 

12 may be less susceptible to damage than that of adults. The lower disease prevalence may 

13 make it more difficult to find a relationship between blood Pb level and cardiovascular 

14 effects in child studies. Although studies in children may be limited with regard to study 

15 power in assessing associations with present CVD pathology, they provide for the 

16 examination of early function changes that are associated with subsequent CVD.

17 The limited numbers of studies published on children examined endpoints such as total 

18 peripheral resistance (TPR), BP, and autonomic nervous system activation. These new 

19 and earlier studies are presented in Table 5-20. Multiple studies in New York State 

20 evaluated two child cohorts born in the 1990s after Pb was removed from gasoline in the 

21 U.S. with mean blood Pb levels of 4.62 and 1.01 μg/dL (Gump et al., 2011; Gump et al., 

22 2009; Gump et al., 2007; Gump et al., 2005). Zhang et al. (In Press) examined children in 

23 Mexico City born from 1994 to 2003, when Pb was being taken out of gasoline in 

24 Mexico (Martinez et al., 2007). The geometric mean cord and concurrent blood Pb levels 

25 of the children in the Mexico City cohort were 4.67 and 2.56 μg/dL.
February 2012 5-268 Draft – Do Not Cite or Quote

Table 5-20 Studies of Children Cardiovascular Endpoints and Pb Biomarkers 

Ordered as They Appear in the Text

Study

Population/

Location Parameters Blood Pb Dataa Statistical Analysis Effect Estimates/Results

Gump et al.

(2005)

122 children age 

9.5 yr in Oswego, 

NY (born at a single 

hospital in New 

York from 1991-94)

SBP, TPR (total 

peripheral 

vascular 

resistance)

Cord blood Pb:

GM (GSD): 

2.56 μg/dL (1.16) 

Childhood (mean age of 

measurement: 2.6 yr)

blood Pb: GM (GSD): 

4.06 μg/dL (1.14) 

Multivariate linear regression 

models examined the 

relationship of blood Pb with 

change in z-score for outcome 

(post- and pre-stress). 

Potential confounders 

considered: HOME score, 

SES, birth weight, child BMI, 

child sex.

Per 1 μg/dL increase in childhood blood 

Pb level, 0.088 (95% CI: 0.023, 0.153) 

dyne-s/cm5 change in TPR

Per 1 μg/dL increase in cord blood Pb 

level, 12.16 (95% CI: 2.44, 21.88) 

mmHg higher SBP

Gump et al.

(2007)

122 children age 

9.5 yr in 

Oswego, NY

SBP, TPR Childhood (mean age of 

measurement: 2.6 yr) 

blood Pb:

GM (GSD): 

4.06 μg/dL (1.14) 

Linear regression models 

adjusting for the same 

covariates as in Gump et al.

(2005). Separate models 

testing whether Pb is a 

mediator of SES 

associations,(Sobel test) and 

whether Pb moderates SES 

associations (Pb-SES 

interaction).

Blood Pb was a mediator of the SESTPR relationship

SES alone: -0.62 dyne-s/cm5 (p <0.05) 

SES with Blood Pb: -0.40 dyne-s/cm5

(p >0.10), change in R2 attributable to 

SES: -55.3%

Blood Pb was a potential moderator of 

the SES-TPR relationship. Blood Pb × SES interaction: p = 0.07 .

Blood Pb was a moderator of SES-SBP 

relationship

Pb × SES interaction: p = 0.007

At blood Pb levels > 4 μg/dL, SES not 

significantly associated with SBP 

Gump et al.

(2009)

122 children age 

9.5 yr in 

Oswego, NY

Salivary cortisol Cord blood Pb:

GM (GSD): 

2.56 μg/dL (1.16) 

Childhood (mean age of 

measurement: 2.6 yr) 

blood Pb: GM (GSD): 

4.06 μg/dL (1.14) 

Linear regression to examine 

whether blood Pb level 

mediates or moderates the 

relationship between SES and 

salivary cortisol as in Gump et 

al. (2007)

Blood Pb was a mediator of the SEScortisol association. SES was no longer 

significantly associated with cortisol 

after adjusting for blood Pb level. R2 for 

SES decreased by 40, 33, 50% for 

cortisol measured at 21, 40, and 60 min.

Blood Pb was not a significant 

moderator of SES-cortisol association. 

Blood Pb × SES interaction term was 

not statistically significant

Gump et al.

(2011)

140 children 

ages 9-11 yr

SBP, TPR, HRV 

(heart rate 

variability) in 

response to acute 

stress (mirror 

tracing task)

Concurrent blood Pb:

GM: 1.01 μg/dL

Quartiles:

Q1: 0.14-0.68 μg/dL

Q2: 0.69-0.93 μg/dL

Q3: 0.94-1.20 μg/dL 

Q4: 1.21-3.76 μg/dL

Outcomes were analyzed as 

continuous variables for the 

pre-stress values or the 

change post- and pre-stress. 

Regression models were 

adjusted for sex, SES, BMI, 

and age.

Blood Pb levels associated with 

autonomic and cardiovascular 

dysregulation in response to stress – greater vascular resistance, reduced 

stroke volume, and cardiac output

Change in SBP (mmHg) across 

quartiles: Q1: 5.30, Q2: 7.33, Q3: 7.07, 

Q4: 7.23, p for trend = 0.31

Change in TPR (%) across quartiles: 

Q1: 2.91, Q2: 8.18, Q3: 9.55, Q4: 9.51, 

p for trend = 0.03

Change in Stroke Volume (%) across 

quartiles: Q1: 2.23, Q2: 0.91, Q3: -3.47, 

Q4: -0.89, p for trend = 0.04
February 2012 5-269 Draft – Do Not Cite or Quote

Study

Population/

Location Parameters Blood Pb Dataa Statistical Analysis Effect Estimates/Results

Zhang et al. (In 

Press)

457 mother child 

pairs in a birth 

cohort, born 1994 to 

2003 in Mexico City. 

Children were 

evaluated 

2008-2010 at ages 

7-15 yr

SBP Cord blood Pb: 

GM (GSD): 4.67 μg/dL 

(1.18) (N=323)

Concurrent blood Pb: 

GM (GSD): 2.56 μg/dL (1.16) (N=367)

Maternal post-partum 

bone Pb: 

Median (IQR): 

Tibia Pb: 9.3 (3.3, 

16.1) μg/g

Patella Pb: 11.6 (4.5, 

19.9) μg/g

Multiple regression models 

and generalized estimating 

equations (log linear for cord 

blood, linear for concurrent 

blood and maternal bone). The 

base model considered 

maternal education, birth 

weight, BMI, sex, and child 

concurrent age as covariates.

Prenatal Pb exposure may be 

associated with higher BP in female 

offspring. 

Among girls, an IQR (13 μg/g) increase 

in maternal tibia Pb was associated with 

a 2.11 (95% CI: 0.69, 3.52) mmHg 

increase in SBP

IQR (16 μg/g) increase in maternal 

patella Pb was associated with a 0.87 

(95% CI: -0.75, 2.49) mmHg increase in 

SBP

IQR (4 μg/dL) increase in cord blood Pb 

was associated with a 0.75 (95% CI: -

1.13, 2.63) mmHg increase in SBP

Factor-Litvak et 

al. (1999; 1996)

260 children ages 

5.5 years old in K. 

Mitrovica and 

Pristina, Yugoslavia

SBP Concurrent blood Pb range: 

4.1 to 76.4 μg/dL

Linear regression analysis. 

Potential confounders 

considered: gender, maternal 

education, birth weight, HOME 

score, and BMI.

Per 1 μg/dL increase in concurrent blood 

Pb level, 0.05 (95% CI: -0.02, 0.13) 

mmHg higher SBP

Blood Pb level at birth and cumulative 

blood Pb level were not as strongly 

associated with SBP at age 5.5 yr.

Gerr et al. (2002) Young adults age 

19-29 years, born 

1965-1975, male 

and female; half of 

the subjects had 

grown up around an 

active Pb smelter in 

Silver Valley, Idaho

BP While the concurrent mean 

blood Pb level was 3.15 

μg/dL for the highest bone 

Pb category (> 10 μg/g), 

early childhood mean blood 

Pb levels in this group were 

substantially elevated for all 

bone Pb level categories 

and were highest among 

participants in the highest 

bone Pb level category. The 

mean blood Pb level was 

65 μg/dL among 

participants with bone Pb 

level >10 μg/g.

Multiple linear regression 

models always included age, 

sex, height, BMI, current 

smoking status, frequency of 

alcohol consumption, current 

use of birth-control medication, 

hemoglobin level, serum 

albumin, and income, 

regardless of significance 

levels. Both blood Pb (as a 

linear term) and bone Pb (a 

four category ordinal variable 

from <1 μg/g to >10 μg/g) 

were tested together.

Group in highest quartile of tibia Pb level 

(> 10 μg/g) had 4.26 (95% CI: 1.36, 

7.16) mmHg higher SBP and 2.80 (95% 

CI: 0.35, 5.25) mmHg higher DBP 

compared to the lowest tibia Pb group (< 

1 μg/g).

a

Blood Pb data are estimates of geometric mean (GM) and geometric standard deviation (GSD) using the arithmetic mean and SD. 

5.4.4.2 Cardiovascular Functioning in Children

1 The relationship between cardiovascular functioning (TPR, BP, stroke volume, and 

2 cardiac output,) and blood Pb levels was examined by Gump et al. (2007; 2005) in a 

3 cohort born at a single New York hospital. Higher early childhood Pb levels (average age 

4 2.6 years) were associated with greater TPR response to acute stress induced by mirror 

5 tracing on a computer at age 9.5 years as shown in Figure 5-31. Testing blood Pb with 

6 linear, quadratic, and cubic terms did not produce significantly different Pb-TPR 

7 associations which the authors suggested showed effects that were concentration 

8 dependent and, notably were not emergent at a specific exposure threshold. TPR 

9 increased with increasing quartile of blood Pb level. A mediational analysis indicated that 

10 Pb was a significant mediator of the SES-TPR reactivity association; some evidence also 

11 suggested moderation, whereby the inclusion of blood Pb into the model reduced the 

12 effect estimate for SES. Biological plausibility for these observations in children is 
February 2012 5-270 Draft – Do Not Cite or Quote

1 provided by observations that Pb exposures increases TRP in toxicological studies and by 

2 mechanistic evidence indicating that Pb-induced changes in SNS activity may mediate 

3 such effects (Section 5.4.2.3). Additionally, higher blood Pb level measured at age 2.6 

4 years was associated with a smaller stroke volume and cardiac output responses to acute 

5 stress at age 9.5 years. In a further analysis in this cohort, Gump et al. (2009) examined 

6 the possibility that Pb may mediate an association between SES and cortical responses to 

7 acute stress. Elevated cortisol has been associated with hypertension (Whitworth et al., 

8 2000). Gump et al. (2009) found that lower family income was associated with greater 

9 cortisol levels following an acute stress task and that blood Pb was a mediator of this 

10 association.

Source: Reprinted with permission of Elsevier (Gump et al., 2005)

Figure 5-31 Children’s adjusted total peripheral resistance (dyn-s/cm5) 

responses to acute stress tasks, as a function of childhood Pb 

levels.

11 In a different cohort of 140 children 9 to 11 years of age recruited from local pediatrician 

12 offices and from mailings to homes with children in this age group, Gump et al. (2011)

13 used a similar acute stress-producing paradigm to that used in the previous studies to 

14 examine the associations of concurrent blood Pb with cardiovascular responses. TPR 

15 significantly increased in a concentration-dependent relationship with blood Pb, with 

16 most of the increase occurring between the first quartile blood Pb (0.14-0.68 μg/dL) and 

17 the second quartile blood Pb (0.69-0.93 μg/dL). This result is consistent with those of 

18 Gump et al. (2005). Also, these new findings provided evidence of associations with 

19 concurrent blood Pb levels and with lower blood Pb levels (Gump et al., 2011) than were 

20 previously examined in Gump et al. (2005).
February 2012 5-271 Draft – Do Not Cite or Quote

1 Studies examining HRV in adults and animal toxicology are discussed in Section 5.4.3.3. 

2 In Gump et al. (2011), cardiac autonomic regulation decreased in a 

3 concentration-dependent manner with increasing concurrent blood Pb quartile, with the 

4 largest change relative to the first quartile (0.14-0.68 μg/dL) measured in the highest 

5 blood Pb quartile (1.21-3.76 μg/dL). Also, high frequency HRV, decreased more with 

6 acute stress in the highest Pb quartile group (1.21-3.76 μg/dL). In the earlier cohort, early 

7 childhood (mean age at collection: 2.6 years) blood Pb level was associated with reduced 

8 stroke volume and cardiac output (Gump et al., 2007; Gump et al., 2005). In this new 

9 study, Gump et al. (2011) found the same but for concurrent blood Pb level and at lower 

10 blood Pb levels.

5.4.4.3 Blood Pressure in Children

11 Zhang et al. (In Press) conducted a longitudinal study that examined changes in BP in 

12 323 girls and boys aged 7 to 15 years old in a Mexico City cohort for a relationship with 

13 maternal bone Pb measured one month post-partum (a measure of cumulative exposure 

14 that could expose fetuses to Pb through accelerated mobilization of bone Pb during 

15 pregnancy), and cord blood Pb at delivery. This was the first study to examine the 

16 association of maternal bone Pb, as a marker of prenatal exposure, with offspring BP. 

17 The model including both girls and boys (without adjustment for concurrent blood Pb) 

18 showed no statistically significant association overall of any Pb biomarker with child BP. 

19 A significant interaction was found between maternal tibia Pb and sex, and in models 

20 stratified by sex, maternal tibia Pb was associated with adjusted systolic and diastolic BP 

21 in females, but not males. Maternal post-partum median tibia Pb was 9.3 μg/g (IQR: 3.3, 

22 16.1 μg/g) with no significant differences between mothers of male and female offspring. 

23 Suboptimal growth in utero is associated with accelerated weight gain in offspring during 

24 childhood and greater risk of later hypertension (Barker and Bagby, 2005; te Velde et al., 

25 2004; Barker et al., 1989). These may represent biologically plausible mechanisms by 

26 which prenatal Pb exposure may result in increased BP later in childhood as was 

27 demonstrated in female offspring. The relationship between birth weight and Pb 

28 biomarkers is discussed in Section 5.8.7.

29 Gump et al. (2011; 2005) examined the relationship of blood Pb level with BP in their 

30 two cohorts of contemporary children around age 10 years in New York State. Gump et 

31 al. (2005) reported an association of cord blood levels with systolic BP (12.16 mmHg 

32 [95% CI: 2.44, 21.88] increase per 1 μg/dL increase in cord blood Pb level). Gump et al.

33 (2011) found that with acute stress, children in higher quartiles of concurrent blood Pb 

34 level (>0.69 μg/dL) had larger increases in systolic BP. For example, children with blood 

35 Pb levels between 1.21 and 3.76 μg/dL had a 7.23 mmHg change, and children with 
February 2012 5-272 Draft – Do Not Cite or Quote

1 blood Pb levels between 0.14 and 0.68 μg/dL had a 5.30 mmHg change. A linear trend 

2 was not observed across quartiles. An interaction between long-term perceived stress and 

3 bone Pb levels in association with BP and hypertension also was reported in a study of 

4 adults (Peters et al., 2007) (described in Section 5.4.2.1). An earlier study (Factor-Litvak 

5 et al., 1999; Factor-Litvak et al., 1996), of children with higher blood Pb levels ranging 

6 from 4.1 to 76.4 μg/dL found that a 1 μg/dL increase in concurrent blood Pb was 

7 associated with a 0.05 (95% CI: -0.02, 0.13) mmHg increase in systolic BP. An 

8 additional study (Gerr et al., 2002) reported that young adults (ages 19-29 years) in the 

9 highest category of bone Pb levels (greater than 10 μg/g), in whom the mean concurrent 

10 blood Pb level was 65 μg/dL, systolic BP was 4.26 mmHg higher compared with young 

11 adults with bone Pb levels < 1 μg/dL.

12 The pathogenesis of CVD has been hypothesized to begin in childhood (Kapuku et al., 

13 2006). Early markers observable in youth include increased blood pressure during stress, 

14 reduced heart rate variability, increased IMT, and vascular endothelium dysfunction. 

15 Kapuku et al. (2006) state that endothelial dysfunction is the center of the CVD 

16 paradigm. The factors measured in childhood or as a cumulative burden since childhood 

17 are predictors of outcomes in young adults who are still too young to experience coronary 

18 events (Li et al., 2003), and early-life exposures may induce changes in arteries that 

19 contribute to the development of atherosclerosis (Raitakari et al., 2003). Berenson et al.

20 (2002) observed that the effects of multiple risk factors on coronary atherosclerosis 

21 support evaluation of cardiovascular risk in young people. Thus, evidence relating levels 

22 of biomarkers of Pb exposure of children to cardiovascular function in the groups of 

23 studies presented in the preceding text when combined with the evidence for the potential 

24 pathogenesis of CVD starting in childhood that yield effects in adulthood provides 

25 coherence with evidence in adults supporting the effects of long-term, cumulative Pb 

26 exposures in the development of cardiovascular effects.

27 Few animal studies have examined the effect of Pb exposure during pregnancy and 

28 lactation on BP in offspring and those that have used high levels of exposure. Recently, 

29 pups of Pb-exposed dams (1,000 ppm through pregnancy and lactation) exhibited 

30 increased blood Pb level (mean blood Pb level: 58.7 μg/dL) and increased arterial 

31 systolic BP after weaning (Grizzo and Cordellini, 2008) suggesting a role for childhood 

32 Pb exposure leading to adult disease. 

5.4.4.4 Summary of Child Cardiovascular Studies

33 Studies have reported antecedent cardiovascular changes such as TPR responses to acute 

34 stress tasks as a function of childhood blood Pb levels. Also, a study reported associations 
February 2012 5-273 Draft – Do Not Cite or Quote

1 with acute stress-induced autonomic and cardiovascular dysregulation responses. 

2 Biomarkers of prenatal Pb exposure (maternal post-partum patella and tibia) were related 

3 to later higher BP. This may be related to intrauterine growth restriction and subsequent 

4 accelerated weight gain in childhood and may indicate greater risk of hypertension later 

5 in life. The results are not uniform with respect to the important lifestages of Pb exposure 

6 and can differ by sex and other factors. Uncertainties in these studies may be related to 

7 sample size, single measures of BP, variation of age of onset of puberty, and cross8 sectional design. Some of these uncertainties result in random error, however, and result 

9 in the attenuation of observed associations rather than the generation of spurious 

10 associations. These study findings indicate that in children with mean blood Pb levels in 

11 the range of < 5 to 10 μg/dL, increasing blood Pb level may be associated with small 

12 increases in BP and changes in the cardiovascular system that may be related to later 

13 development of CVD.

14 Factors may limit the ability of studies to detect statistically significant Pb-associated 

15 changes with BP. The relatively young age of the subjects may have limited the ability of 

16 these studies to detect significant BP effects (as opposed to early function effects) if 

17 longer duration Pb exposure is necessary to produce the cardiovascular changes 

18 considering the lower prevalence and strength of compensatory mechanisms in children. 

19 There is uncertainty in the shape of the concentration-response relationship to 

20 cardiovascular endpoints at lower blood Pb levels since most studies modeled a linear 

21 relationship. Several studies that compared linear and nonlinear relationships between 

22 blood Pb level and decrements in cognitive function found a better fit for the nonlinear 

23 relationship (See Section 5.3.10).

24 Cardiovascular endpoints other than baseline BP may be more sensitive outcomes by 

25 which to measure Pb-associated cardiovascular effects in very young children. The series 

26 of studies by Gump et al. (2011; 2009; 2007; 2005) evaluating much smaller samples 

27 than did the adult studies, was able to demonstrate statistically significant relationships of 

28 blood Pb levels with cardiovascular outcomes such as TPR related to acute stress. It 

29 suggests that the stress paradigm may be useful to detect associations of blood Pb levels 

30 with effects on the cardiovascular system of children. Selection of the appropriate 

31 cardiovascular outcome in children is an important factor to consider in the design of 

32 studies. Rather than using indicators of already present cardiovascular problems, such as 

33 BP, informative evaluation of cardiovascular changes that are antecedent to increased BP 

34 and changes in other CVD-related endpoints at later lifestages may reduce uncertainties 

35 regarding the time course of cardiovascular changes associated with early Pb exposure 

36 where the relevance and persistence of these endpoints has been shown to be associated 

37 with future pathology.
February 2012 5-274 Draft – Do Not Cite or Quote

1 Overall this body of evidence, based on different cohorts, locations, and study designs,

2 begins to form a literature base that indicates there is a relationship between biomarkers 

3 of Pb exposure and cardiovascular effects in children. One longitudinal study ties in 

4 maternal bone Pb level, and cord and concurrent blood Pb level for the children. 

5 Limitations exist in the studies. While blood pressure increases are more prevalent in 

6 older adults than in children, BP increases have been related to higher blood Pb level in 

7 older studies of children and young adults (Gerr et al., 2002; Factor-Litvak et al., 1999; 

8 Factor-Litvak et al., 1996). The newer children studies provide information in 

9 populations with mean blood Pb levels in the range of < 5 to 10 μg/dL for BP and 

10 antecedents for CVD such as increases in TPR and changes in cardiac autonomic 

11 regulation.

5.4.5 Mortality

12 The 2006 Pb AQCD stated that collectively the then available evidence suggested an 

13 effect of Pb on cardiovascular mortality in the general U.S. population but cautioned that 

14 these findings should be replicated before these estimates for Pb-induced cardiovascular 

15 mortality could be used for quantitative risk assessment purposes (U.S. EPA, 2006b).

16 Previous results involved NHANES II and III analyses that examined the association of 

17 adult concurrent blood Pb with all cause and cause-specific mortality (Schober et al., 

18 2006; Lustberg and Silbergeld, 2002). As blood Pb levels in adults reflect contributions 

19 from both recent Pb exposure and mobilization of historic Pb from bone, it is unclear to 

20 what extent recent, past, or cumulative Pb exposures contributed to the observed 

21 associations. Given the decline in ambient air Pb concentrations and population blood Pb 

22 levels, it is likely that study subjects had a much higher Pb exposure in their past than 

23 during the study period. Using NHANES II (1976-1980) data, Lustberg and Silbergeld

24 (2002) found significant increases in all-cause, circulatory and cancer mortality, 

25 comparing adults with blood Pb levels (measured 12-16 years before ascertainment of 

26 vital status) of 20-29 μg/dL to those with blood Pb levels less than 10 μg/dL. Using 

27 NHANES III data, Schober et al. (2006) found significant increased all-cause mortality, 

28 cardiovascular, and cancer mortality comparing adults with blood Pb levels (measured a 

29 median of 8.8 years before ascertainment of vital status) from 5-9 μg/dL and above 

30 10 μg/dL to those with blood Pb levels less than 5 μg/dL.

31 Several new studies substantially strengthen the evidence base for Pb-associated 

32 mortality. A further analysis of the NHANES III database by a different research group 

33 using different methods addressed uncertainties from other earlier analyses by 

34 considering a more extensive number of potential confounding factors and by 

35 characterizing concentration-response relationships. A few longitudinal prospective 
February 2012 5-275 Draft – Do Not Cite or Quote

1 studies in different cohorts conducted by different researchers with different methods in 

2 different parts of the U.S. provide coherence with the evidence base for blood Pb and add 

3 new evidence for mortality associated with bone Pb levels. 

4 Menke et al. (2006) examined all-cause and cause-specific mortality using NHANES III 

5 data. Subjects at least 18 years of age were followed up to 12 years after their blood Pb 

6 was measured, and 1,661 deaths were identified. Those with baseline blood Pb levels 

7 from 3.63 to 10 μg/dL had significantly higher risks of all-cause (HR: 1.25 [95% CI: 

8 1.04, 1.51]), cardiovascular (HR: 1.55 [95% CI: 1.08, 2.24]), MI (HR: 1.89 [95% CI: 

9 1.04, 3.43]), and stroke (HR: 2.51 [95% CI: 1.20, 2.26]) mortality compared to those with 

10 baseline blood Pb levels less than 1.93 μg/dL and increased risk of cancer mortality (HR: 

11 1.10 [95% CI: 0.82, 1.47]). Effect estimates adjusted for demographic characteristics 

12 were robust to the additional adjustment for factors such as smoking, alcohol 

13 consumption, diabetes, BMI, hypertension, and level of kidney function. The consistency 

14 of HRs across models with a varying number of control variables indicated little residual 

15 confounding. Hazard ratios were not higher comparing adults with blood Pb levels from 

16 1.94 to 3.62 μg/dL to those with blood Pb levels <1.93 μg/dL. However, tests for linear 

17 trend were statistically significant for all mortality outcomes except for cancer mortality. 

18 Menke et al. (2006) evaluated several of the model covariates (e.g., diabetes, 

19 hypertension, and glomerular filtration rate [GFR]) in a subgroup analysis (Figure 5-32). 

20 The authors reported that there were no interactions between blood Pb and other adjusted 

21 variables. In the previous NHANES III analysis of the association of blood Pb with 

22 mortality, Schober et al. (2006) included participants greater than 40 years of age (N = 

23 9686) and adjusted for covariates including age, sex, ethnicity, and smoking rather than 

24 the full suite of covariates evaluated by Menke et al. (2006). Schober et al. (2006)

25 reported increased HRs comparing adults with blood Pb levels ≥ 10 μg/dL to those with 

26 blood Pb levels <5 μg/dL for all-cause (HR: 1.59 [95% CI: 1.28, 1.98]), CVD (HR: 1.55 

27 [95% CI: 1.16, 2.07]), and cancer (HR: 1.69 [95% CI: 1.14, 2.52]) mortality and 

28 generally statistically nonsignificant higher HRs comparing adults with blood Pb levels 

29 from 5-9 μg/dL to those with blood Pb levels <5 μg/dL. The median follow-up time 

30 between measurement of blood Pb and death ascertainment was 8.55 years.

31
February 2012 5-276 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Lippincott Williams & Wilkins, Menke et al. (2006)

Note: Hazard ratios were calculated for a 3.4-fold increase in blood Pb level with log-blood Pb as a continuous variable. This 

increase corresponds to the difference between the 80th and 20th percentiles of the blood Pb distribution (4.92 μg/dL versus 

1.46 μg/dL, respectively).

Figure 5-32 Multivariate adjusted relative hazards of all-cause and 

cardiovascular mortality.

1 Both Menke et al. (2006) and Schober et al. (2006) presented mortality curves that plot 

2 the HRs against blood Pb level. Figure 5-33 shows the mortality hazard ratio curves (not 

3 absolute cases of mortality) for both stroke and MI reported by Menke et al. (2006).

4 Nonlinear associations were modeled. The curves were fitted using predetermined 

5 restricted quadratic splines with knots at the 10th percentile (1.00 μg/dL), the 50th 

6 percentile (2.67 μg/dL), and the 80th percentile (5.98 μg/dL) blood Pb levels. The 

7 authors did not explain the shape of the blood Pb-mortality curves in detail; however, the 

8 knots corresponded with the inflection points in the curve. In the tails of the blood Pb 

9 distribution, hazard ratios decreased with increasing blood Pb level. However, hazard 

10 ratios remained above 1 over most of the blood Pb distribution, and in the most heavily 

11 populated portion of the blood Pb distribution, hazard ratios increased with increasing 

12 blood Pb level. Hazard ratios peaked for all outcomes at a blood Pb level of 

13 approximately 6 μg/dL. Lower concentration-response functions at higher blood Pb 
February 2012 5-277 Draft – Do Not Cite or Quote

1 levels also have been found for blood Pb-cognitive function relationships in children 

2 (Section 5.3.10).

Source: Reprinted with permission of Lippincott Williams & Wilkins, Menke et al. (2006)

Note: A histogram of blood Pb levels is superimposed in the background and displayed on the right axis.

Figure 5-33 Multivariate-adjusted relative hazard (left axis) of mortality 

associated with blood Pb level between 1 μg/dL and 10 μg/dL.

3 Schober et al. (2006) examined proportional hazard assumptions, tested for linear trend 

4 across blood Pb tertiles, and evaluated log-transformed continuous blood Pb level as 

5 a 5-knot cubic spline (position of knots not reported). A statistically significant increasing 

6 linear trend for mortality was observed across blood Pb tertiles. The results of the spline 

7 fit of the continuous blood Pb level term to relative hazard of all cardiovascular diseases 

8 reported by Schober et al. (2006) are shown in Figure 5-34. In contrast to the curve 

9 presented by Menke et al. (2006), Schober et al. (2006) found the relative hazard axis and 

10 the blood Pb axis largely to be linear (solid line). Dashed lines are 95% CIs. The hazard 

11 ratio was fixed at 1.0 for the referent blood Pb level of 1.5 μg/dL. In this study, hazard 

12 ratios were less than one in the lower range of blood Pb levels. Despite differences in the 
February 2012 5-278 Draft – Do Not Cite or Quote

1 age groups included, follow-up time, categorization of blood Pb levels, and differences in 

2 hazard ratio across the blood Pb range, results reported by Menke et al. (2006) and 

3 Schober et al. (2006) are consistent in finding associations between higher blood Pb and 

4 increased CVD mortality. 

Source: Schober et al. (2006)

Note: The solid line shows the fitted five-knot spline relationship; the dashed lines are the point-wise upper and lower 95% CIs.

Figure 5-34 Relative risk of all cause mortality for different blood Pb levels 

compared with referent level of 1.5 μg/dL (12.5th percentile).

5 In addition to the NHANES analyses described above, studies of older adult males

6 (Weisskopf et al., 2009) and older adult females (Khalil, 2010; Khalil et al., 2009a) were 

7 conducted recently. Weisskopf et al. (2009) used data from the NAS to determine the 

8 associations of blood, tibia, and patella Pb with mortality. The authors identified 241 

9 deaths over an average observation period of 8.9 years (7,673 person-years). The 

10 strongest associations were observed between mortality and baseline patella Pb 

11 concentration. Baseline tibia Pb levels were more weakly associated with CVD mortality. 

12 Tibia bone Pb level is thought to reflect a longer cumulative exposure period than is 

13 patella bone Pb level because the residence time of Pb in trabecular bone is shorter than 

14 that in cortical bone. IHD contributed most to the relationship between patella Pb and all 

15 CVD death with an individual HR of 2.69 (95% CI: 1.42, 5.08). Although there was high 

16 correlation between tibia and patella Pb (Pearson r = 0.77), compared with cortical bone 

17 Pb, trabecular bone Pb may have more influence on circulating blood Pb level, and thus 

18 local organ concentration of Pb, because of its shorter residence time in bone. In contrast 
February 2012 5-279 Draft – Do Not Cite or Quote

1 to the NHANES analyses, baseline blood Pb was not significantly related to 

2 cardiovascular mortality in the NAS study. This discrepancy may be related to 

3 differences in sample size and resulting power, modeling strategies (e.g., linear versus 

4 log-linear blood Pb level terms), or age range of the study populations. The duration of 

5 follow-up was similar across studies. The youngest subjects at baseline in the Weisskopf 

6 et al. (2009) study were approximately 50-55 years old, compared to the youngest in the 

7 Menke et al. (2006) and Schober et al. (2006) studies, who were 18 and 40 years, 

8 respectively. Further, the blood Pb tertile analysis of Weisskopf et al. (2009) could have 

9 been affected if the majority of a hypothesized nonlinear association was contained 

10 largely in the lowest (reference) blood Pb tertile. 

11 Weisskopf et al. (2009) also conducted a concentration-response analysis. A linear trend 

12 was observed for increasing HR across tertiles of both tibia and patella Pb levels. The 

13 linear relationship using tertile patella Pb was confirmed in other models in which 

14 continuous patella Pb and nonlinear penalized spline terms (higher order terms) were not 

15 statistically significant. The number of knots and their placement within the Pb variable, 

16 which can influence these results, were determined by an iterative best fit procedure. 

17 Concentration-response relationships shown in Figure 5-35 were approximately linear for 

18 patella Pb on the log HR scale for all CVD, but appeared nonlinear for IHD (p <0.10). 

19 The peak HR is shown around 60 μg/g, beyond which the HR tends to decrease. It is 

20 important to note the wide confidence limits, which increase uncertainty at the lower and 

21 upper bounds of patella Pb levels. 
February 2012 5-280 Draft – Do Not Cite or Quote

Source: Reprinted with permission of Lippincott Williams & Wilkins, Weisskopf et al. (2009)

Note: The reference logHR = 0 at the mean of patella Pb concentration. The estimates are indicated by the solid line and the 95% 

pointwise CIs by the dashed lines. The P values for significance of the nonlinear component for all-cause, cardiovascular, and 

ischemic heart disease mortality were 0.42, 0.80 and 0.10 respectively. Patella Pb concentrations of all individual participants are 

indicated by short vertical lines on the abscissa. Adjusted for age, education, smoking status, and pack-years of smoking among 

participants without ischemic heart disease at baseline.

Figure 5-35 Associations between patella bone Pb level and the log of HR 

(logHR) for all-cause, cardiovascular, and ischemic heart disease.

1 The association of adult blood Pb with mortality has also been examined among women 

2 enrolled in the Study of Osteoporotic Fractures (SOF) (Khalil et al., 2009a). This 

3 prospective cohort (N = 533) enrolled female volunteers from two U.S. locations, 

4 Baltimore, MD and Monongahela Valley, PA and followed women for an average of 12 

5 years after blood Pb measurement. All-cause mortality comparing women with blood Pb 

6 levels >8 μg/dL to those with blood Pb levels <8 μg/dL was significantly increased (HR: 

7 1.59 [95% CI: 1.02, 2.49]). Combined cardiovascular disease mortality (HR: 1.78 [95% 

8 CI: 0.92, 3.45]), coronary heart disease mortality (HR: 3.08 [95% CI: 1.23, 7.70]), but not 

9 stroke mortality (HR: 1.13 [95% CI: 0.34, 3.81]) HR was increased among the women 

10 enrolled in this study with blood Pb levels >8 μg/dL. In addition, analyses of blood Pb 

11 tertiles and quintiles indicated that blood Pb-mortality HRs were consistently elevated in 

12 groups with blood Pb levels > 7 μg/dL (Khalil, 2010). The findings for elevated mortality 

13 HRs with the highest blood Pb levels are reinforced by the results displayed in Figure 

14 5-36. The HR curve for all-cause mortality is relatively flat over most of population 

15 blood Pb distribution (represented by the blue dots) and increases only in the upper tail of 

16 the blood Pb distribution where there are relatively few subjects (i.e., fewer dots).
February 2012 5-281 Draft – Do Not Cite or Quote

Source: Khalil et al. (2010)

Figure 5-36 Multivariate adjusted relative hazard (left axis) of mortality as a 

function of blood Pb levels between 1 μg/dL and 15 μg/dL.

5.4.5.1 Summary of Mortality

1 The mortality results in this review supported and expanded upon findings from the 2006 

2 Pb AQCD (U.S. EPA, 2006b), which included a few NHANES mortality studies

3 (Schober et al., 2006; Lustberg and Silbergeld, 2002). The recent NHANES mortality 

4 study discussed above (Menke et al., 2006) addressed many of the limitations of the these 

5 earlier studies, including control for a wider range of potential confounders, testing for 

6 interactions with Pb, consideration of concentration-response relationships, extensive 

7 model evaluations, and examination of mortality from specific CVDs. Further, an 

8 association with increased mortality was observed at lower mean population blood Pb 

9 levels. The mean blood Pb level of the NHANES III population was 2.58 μg/dL. The Pb 

10 attributable risk of increased cardiovascular mortality in the NHANES III analysis of 

11 Menke et al. (2006) increased with increasing blood Pb level over the most heavily 

12 population portion of the blood Pb distribution and reached its maximum at blood Pb 

13 levels between of 6 and 7 μg/dL. It is important to note that the relative contributions of 
February 2012 5-282 Draft – Do Not Cite or Quote

1 recent, past, and cumulative Pb exposure to associations observed with the baseline blood 

2 Pb levels is uncertain. In addition, the first evidence that bone Pb, a metric of cumulative 

3 Pb exposure, is associated with increased mortality was reported recently among NAS 

4 men (Weisskopf et al., 2009).

5 Quantitative differences in Pb-associated hazard for death between studies may be 

6 influenced by age range of the study groups, follow up time to death, variation in model 

7 adjustment, central tendency and range of the Pb biomarker levels, assumptions of 

8 linearity in relationship with Pb biomarkers, and choice of Pb biomarker. Quantitative 

9 differences in Pb-associated mortality across NHANES II and NHANES III studies or 

10 between different NHANES III analyses may be explained by the use of continuous or 

11 ordered blood Pb terms and different data selection strategies. Further, studies using 

12 ordered categories of blood Pb level may obtain different results, as the range of blood Pb 

13 level represented in the reference category will affect the calculated coefficients of the 

14 remaining percentiles or groups. 

15 Specifically, Menke et al. (2006) is the strongest study presently published for estimating 

16 the effects of Pb on cardiovascular disease-related mortality. The study uses the 

17 nationally representative NHANES III (1988-1994) sample of men and women. The 

18 results provide confirmation of earlier published NHANES studies but address some of 

19 the key weaknesses noted in those studies. For example, Menke et al. (2006) examined 

20 potential confounding by a large number of factors, including hypertension and kidney 

21 function. Weisskopf et al. (2009) is the first published mortality study using bone Pb as 

22 an exposure index. The study is a prospective study with nearly 100% successful follow23 up of deaths. This rigorous study found increased cardiovascular disease mortality in 

24 association with patella bone Pb. The Khalil et al. (2010; 2009a) SOF provides 

25 supporting results in a different study cohort consisting of white females aged 65-87 

26 years. Further, a number of prior studies have found association between accumulated Pb 

27 reflected in bone Pb measurements and higher CVD morbidity (Sections 5.4.2.1 and 

28 5.4.3). This evidence base is augmented with new findings indicating that biomarkers of 

29 longer-term cumulative Pb exposure increases CVD mortality. The NAS and SOF 

30 examine only men and women, respectively. However, the consistency of findings 

31 between the two studies indicates that the results of either study may be applicable 

32 widely. Despite the differences in design and methods across studies, with few exceptions 

33 associations between higher levels of Pb biomarkers and higher risk of mortality were 

34 consistently observed (Figure 5-37 and Table 5-21). In studies that examined mortality 

35 from specific CVD causes, MI, stroke, and IHD mortality, which are causes related to 

36 higher BP and hypertension were all significantly elevated with higher Pb biomarker 

37 levels.
February 2012 5-283 Draft – Do Not Cite or Quote

Note: Studies are presented in order of strength of study design and follow the order of discussion in the preceding text. Hazard 

ratios represent the hazard in the higher blood or bone Pb group relative to that in the lowest blood or bone Pb group (reference). 

Figure 5-37 Hazard ratios for associations of blood Pb (closed markers) and 

bone Pb (open markers) with all-cause mortality (black diamonds) 

and cardiovascular mortality (blue circles).
February 2012 5-284 Draft – Do Not Cite or Quote

Table 5-21 Additional characteristics and quantitative data for associations of 

blood and bone Pb with CVD mortality for studies presented in 

Figure 5-37

Study

Population/

Location Parameter Pb Data Statistical Analysis

Hazard Ratio or SMR 

(95% CI)

Menke et al.

(2006)

13,946 adult 

participants of 

NHANES III , 

≥ 17 yr (1988-1994)

All cause and cause-specific 

mortality 

(through 2000)

CVD:ICD-9 390-434; ICD-10 

I00-I99), MI (ICD-9 410-414 and 

429.2; ICD-10 I20-I25), stroke 

(ICD-9 430-434 and 436-438; 

ICD-10 I60-I69). 

Baseline Blood Pb 

(measured an 

average of 12 yr 

before mortality):

Mean: 2.58 μg/dL

Tertiles:

<1.93 μg/dL,

1.94-3.62 μg/dL, 

≥ 3.63 μg/dL

Survey-design adjusted Cox 

proportional hazard regression 

analysis (up to 12 yr follow-up) 

adjusted for Model 1: age, 

race/ethnicity, sex, Model 2: urban 

residence, cigarette smoking, alcohol 

consumption, education, physical 

activity, household income, 

menopausal status, BMI, CRP, total 

cholesterol, diabetes mellitus, Model 

3: hypertension, GFR category

All-cause (3rd vs. 1st 

tertile): 

1.25 (1.04, 1.51)

CVD (3rd vs. 1st):

1.55 (1.08, 2.24)

MI (3rd vs. 1st):

1.89 (1.04, 3.43)

Stroke (3rd vs. 1st): 

2.51 (1.20, 5.26)

Cancer (3rd vs. 1st): 

1.10 (0.82, 1.47)

Schober et al.

(2006)

9,686 adult 

participants of 

NHANES III, 

≥ 40 yr 

All cause and cause-specific 

mortality 

Ordered categorical 

blood Pb level, 

measured a median 

of 8.55 yr prior to 

death

<5 μg/dL

5-9 μg/dL

≥ 10 μg/dL

Survey-design adjusted Cox 

proportional hazard adjusted for sex, 

age, race/ethnicity, smoking, education 

level

All-cause (2nd vs. 1st): 

1.24 (1.05, 1.48) 

All-cause (3rd vs. 1st): 

1.59 (1.28, 1.98)

CVD (2nd vs. 1st): 

1.20 (0.93, 1.55)

CVD (3rd vs. 1st): 

1.55 (1.16, 2.07)

Cancer (2nd vs. 1st): 

1.44 (1.12, 1.86) 

Cancer (3rd vs. 1st): 

1.69 (1.14, 2.52)

Lustberg and 

Silbergeld (2002)

4,190 adult 

participants of 

NHANES III, yr 

(1976-1980)

All cause and cause-specific 

mortality

Categorical blood 

Pb level

Mean: 14.0 (5.1)

Median: 13 μg/dL

<10 μg/dL 

(Reference)

10-19μg/dL

20-29 μg/dL

Crude mortality RRs adjusted for age, 

sex, location, education, race, income, 

smoking, BMI, exercise

All-cause (2nd vs. 1st):

1.40 (1.16-1.69)

All-cause (3rd vs. 1st): 

2.02 (1.62-2.52)

Circulatory (2nd vs. 1st): 

1.27 (0.97-1.57)

Circulatory (3rd vs. 1st): 

1.74 (1.25-2.40)

Cancer (2nd vs. 1st): 

1.95 (1.28-2.98)

Cancer (3rd vs. 1st): 

2.89 (1.79-4.64)
February 2012 5-285 Draft – Do Not Cite or Quote

Study

Population/

Location Parameter Pb Data Statistical Analysis

Hazard Ratio or SMR 

(95% CI)

Weisskopf et al.

(2009)

868 men, >55 yr, 

95% white, from 

NAS in Greater 

Boston area, MA

All cause and cause-specific 

mortality

Pb biomarkers 

collected an 

average of 8.9 

years before death

Blood Pb:

Mean (SD): 5.6 

(3.4) μg/dL

Patella Pb:

Mean (SD): 31.2 

(19.4) μg/g

Tertiles:

<22 μg/g,

22-35 μg/g,

>35 μg/g

Tibia Pb:

Mean (SD): 21.8 

(13.6) μg/g

Cox proportional hazard regression 

analysis adjusted for age, smoking, 

education. Additional models adjusted 

for alcohol intake, physical activity, 

BMI, total cholesterol, serum HDL, 

diabetes mellitus, race, and 

hypertension

All-cause (3rd vs. 1st 

patella Pb tertile):

1.76 (0.95, 3.26)

All CVD (3rd vs. 1st 

tertile):

2.45 (1.07, 5.60)

IHD (3rd vs. 1st): 

8.37 (1.29, 54.4)

Cancer (3rd vs. 1st):

0.59 (0.21, 1.67)

After excluding 154 

subjects with CVD and 

stroke at baseline:

All-cause (3rd vs. 1st): 

2.52 (1.17-5.41)

All CVD (3rd vs. 1st): 

5.63 (1.73, 18.3)

All-cause (3rd vs. 1st 

blood Pb tertile):

0.93 (0.59, 1.45)

All CVD (3rd vs. 1st):

0.99 (0.55, 1.78)

IHD (3rd vs. 1st):

1.30 (0.54, 3.17)

Khalil et al.

(2009a)

533 women, 65-87 

yr, from Study of 

Osteoporotic 

Fractures cohort in 

Baltimore, MD and 

Monongahela 

Valley, PA

All cause and cause-specific 

mortality

Blood Pb measured 

an average 12 (SD; 

3) yr before death:

Mean (SD; range): 

5.3 (2.3; 1-21) 

μg/dL

Cox proportional hazards regression 

analysis adjusted for age, clinic, BMI, 

education, smoking, alcohol intake, 

estrogen use, hypertension, total hip 

BMD, walking for exercise, and 

diabetes

≥ 8 μg/dL vs. <8 μg/dL

All cause: 

1.59 (1.02, 2.49)

CVD: 1.78 (0.92, 3.45)

Coronary Heart 

Disease:

3.08 (1.23, 7.70)

Stroke: 1.13 (0.34, 3.81)

Cancer: 1.64 (0.73, 3.71)
February 2012 5-286 Draft – Do Not Cite or Quote

Study

Population/

Location Parameter Pb Data Statistical Analysis

Hazard Ratio or SMR 

(95% CI)

a

Neuberger et al.

(2009)

Residents at or near 

Tar Creek 

Superfund site, 

Ottawa County, OK 

(exposed pop. 

5,852, unexposed 

pop. 16,210) 

Cause-specific mortality Not reported Standardized mortality ratio (SMR) 

based on 2000 U.S. Census data

Heart disease:

Both sexes: 

114.1 (113.1, 115.2)

Men: 

118 (116.4, 119.6)

Women: 

111 (109.5, 112.5)

Stroke:

Both sexes: 

121.6 (119.2, 123.9)

Men: 

146.7 (107.4, 195.7)

Women: 

106.5 (80.2, 138.6)

a

Cocco et al.

(2007)

933 male Pb 

smelter workers 

from Sardinia, Italy 

(1973-2003)

All cause and cause-specific 

mortality

Not reported SMR All cause: 56 (46, 68)

CVD: 37 (25, 55)

a

These references not included in Figure 5-37 because they reported standardized mortality ratios.

5.4.6 Air Lead-Particulate Matter Studies

5.4.6.1 Cardiovascular Morbidity

1 A relatively small number of studies used Pb measured in PM10 and PM2.5 ambient air 

2 samples to represent Pb exposures. However, given that size distribution data for Pb-PM 

3 are fairly limited, it is difficult to assess the representativeness of these concentrations to 

4 population exposure (Section 3.5.3). Moreover, data illustrating the relationships of Pb5 PM10 and Pb-PM2.5 with blood Pb levels are lacking. A few available studies exposed 

6 rats, dogs, or humans to concentrated ambient particles (CAPS) in which Pb and several 

7 other components were measured. Consistent with epidemiologic studies of blood and 

8 bone Pb and with studies of animals exposed to Pb, exposure to Pb-containing CAPS 

9 resulted in various changes related to increased vasoconstriction (Urch et al., 2004; 

10 Wellenius et al., 2003; Batalha et al., 2002). Whereas studies of Pb biomarkers primarily 

11 found cardiovascular effects with indicators of long-term Pb exposure, studies of Pb12 containing CAPS provide evidence for cardiovascular effects with short-term exposure 

13 (2-6 hours over multiple days) It is important to note that Urch et al. (2004) estimated the 

14 Pb effect on brachial artery diameter based only on the ambient concentrations of Pb but 

15 did not directly expose their young (mean age: 35 years, SD: 10), healthy adult 

16 participants to Pb isolated from CAPS.
February 2012 5-287 Draft – Do Not Cite or Quote

1 A U.S. time-series study of almost 3 million pregnant women found that increases in 

2 ambient Pb-TSP concentrations were associated with increased odds of PIH assessed at 

3 delivery (Chen et al., 2006c). In contrast, epidemiologic studies provide weak evidence 

4 for an association between short-term changes (daily average) in ambient air 

5 concentrations of Pb- PM2.5 and cardiovascular morbidity in adults. Some of these time6 series studies analyzed Pb individually, whereas others applied source apportionment 

7 techniques to analyze Pb as part of a group of correlated components. In a time-series 

8 study of 106 U.S. counties, Bell et al. (2009) found that an increase in lag 0 Pb- PM2.5

9 was associated with an increased risk of cardiovascular hospital admissions among adults 

10 ages 65 years and older. Quantitative results were not presented; however, the 95% CI 

11 was wide and included the null value. In this study, statistically significant associations 

12 were observed for other PM metal components such as nickel, vanadium, and zinc. In the 

13 absence of detailed data on correlations among components or results adjusted for 

14 copollutants, it is difficult to exclude confounding by ambient air exposures to these other 

15 components or copollutants. 

16 To address correlations among PM chemical components, some studies applied source 

17 apportionment techniques to group components into common source categories. In these 

18 source-factor studies, it is not possible to attribute the observed association (Sarnat et al., 

19 2008) or lack of association (Andersen et al., 2007) specifically to Pb. 

5.4.6.2 Mortality

20 Time-series epidemiologic studies of ambient air Pb- PM2.5 reported positive associations 

21 with mortality. Although limited in number, these studies indicated associations in 

22 multiple cities across the U.S. In the Harvard Six Cities Study, Laden et al. (2000) found 

a 1.16% (95% CI: 0.20, 2.9%) increased risk in all-cause mortality per 461.4 ng/m3 23 (5th24 95th percentile) increase in Pb- PM2.5. In six California counties, Ostro et al. (2007)

found that a 5 ng/m3 25 (interquartile range) increase in Pb- PM2.5 was associated with a 

26 1.89% (95% CI: -0.57, 4.40%) increased risk of cardiovascular mortality and a 1.74% 

27 (95% CI: 0.24, 3.26%) increased risk of all-cause mortality during the cool season. The 

28 limitations of air-Pb studies were described in Section 5.4.6.1 above and also are relevant 

29 to the interpretation of these findings for mortality.

5.4.7 Summary and Causal Determination

30 The 2006 Pb AQCD concluded that there was a relationship between higher blood Pb and 

31 bone Pb and cardiovascular effects in adults, in particular increased BP and increased 
February 2012 5-288 Draft – Do Not Cite or Quote

1 incidence of hypertension (U.S. EPA, 2006b). This conclusion was substantiated by the 

2 coherence observed between epidemiologic and toxicological findings. The large 

3 evidence base comprising epidemiologic studies conducted in diverse populations using 

4 different designs clearly demonstrated a positive association between blood Pb level and 

5 BP. Meta-analysis of these studies found that each doubling of concurrent blood Pb level 

6 (between 1 and >40 μg/dL) was associated with a 1 mmHg increase in systolic BP and a 

7 0.6 mmHg increase in diastolic BP (Nawrot et al., 2002). In addition, most of the 

8 reviewed studies using bone Pb levels also showed an association with BP, pointing to an 

9 effect of cumulative Pb exposure. Similarly, toxicological studies provided evidence that 

10 long-term exposure to Pb (> 4 weeks) results in increased BP in experimental animals 

11 that persists long after the cessation of Pb exposure. Also, animal toxicological studies 

12 provided mechanistic evidence to support the biological plausibility of Pb-induced 

13 hypertension, including Pb-induced oxidative stress, activation of RAAS, altered 

14 sympathetic activity, and vasomodulator imbalance. Studies in the 2006 Pb AQCD also 

15 found associations between Pb biomarkers and other cardiovascular diseases such as 

16 IHD, cerebrovascular disease, peripheral vascular disease, and cardiovascular disease 

17 related mortality; however, the available evidence was limited. 

18 Building on the strong body of evidence presented in the 2006 Pb AQCD, recent 

19 epidemiologic and toxicological studies strengthened the evidence that long-term Pb 

20 exposure is associated with cardiovascular effects in adults with the largest body of 

21 evidence demonstrating associations of Pb with increased BP and hypertension. Recent 

22 epidemiologic studies addressed past uncertainties, including the potential for 

23 confounding. A recent study in an ethnically diverse community-based cohort of women 

24 and men aged 50-70 years found hypertension risk to be associated with blood and tibia 

25 Pb levels (Martin et al., 2006). These findings are consistent with those of other studies, 

26 demonstrating that with each increase of 1 μg/dL concurrent blood Pb level, systolic BP 

27 increased 1 mmHg and diastolic BP increased 0.5 mmHg. Recent epidemiologic studies 

28 in adults found associations with hypertension in populations with relatively low mean 

29 blood Pb levels. For example, a positive relationship was found in the nationally 

30 representative NHANES III (1988-1994) where the geometric mean blood Pb level of the 

31 population was 1.64 μg/dL (Muntner et al., 2005). Despite the extensive evidence for 

32 relatively low concurrent blood Pb levels, as these cardiovascular outcomes were most 

33 often examined in adults that have been exposed to higher levels of Pb earlier in life, 

34 uncertainty remains concerning the Pb exposure level, timing, frequency, and duration 

35 contributing to the observed associations. A new prospective study in Pb workers found 

36 independent associations of both baseline blood Pb level and subsequent changes in 

37 blood Pb over follow-up with changes in BP over follow-up and bone Pb level with 

38 hypertension (Glenn et al., 2006). The results indicated that different mechanisms may 
February 2012 5-289 Draft – Do Not Cite or Quote

1 mediate shorter-term Pb-associated increases in BP and longer-term Pb-associated 

2 development of hypertension.

3 Collectively, all animal toxicological studies providing blood Pb level and BP 

4 measurements reported increases in BP with increasing blood Pb level (Figure 5-29). 

5 Importantly, an animal toxicological study provided support for increased BP following 

6 long-term Pb exposure resulting in low blood Pb levels with mean blood Pb levels as low 

7 as 2 μg/dL (Tsao et al., 2000). However, a majority of studies examined Pb exposures 

8 that resulted in mean blood Pb levels > 10 μg/dL. Thus, the weight of the evidence 

9 demonstrated such effects in animals with blood Pb levels >10 μg/dL. New studies also 

10 demonstrated only partial reversibility of Pb-induced increased BP following Pb exposure 

11 cessation or chelation and the possibility for short-term Pb exposure-induced increases in 

12 BP.

13 The epidemiologic evidence for a relationship between Pb exposure and cardiovascular 

14 disease is strengthened by observations of cardiovascular effects in association with both 

15 blood and bone Pb level after adjusting for multiple potential confounding factors, 

16 including age, sex, BMI, antihypertensive medication, SES, race/ethnicity, alcohol 

17 consumption, serum total cholesterol, smoking, educational level, diabetes, and measures 

18 of renal function. The epidemiologic evidence also was substantiated by several available 

19 prospective studies that found associations between biomarkers of Pb and cardiovascular 

20 health endpoints. These studies inform the temporality of these relationship between 

21 biomarkers of Pb exposure and cardiovascular morbidity (e.g., HRV, IHD, BP, 

22 hypertension) (Eum et al., 2011; Jain et al., 2007; Peters et al., 2007; Glenn et al., 2006)

23 and mortality (Khalil et al., 2009a; Weisskopf et al., 2009; Menke et al., 2006; Schober et 

24 al., 2006).

25 Epidemiologic studies continued to demonstrate a relationship between bone Pb, which is 

26 a metric of cumulative Pb exposure, and BP in adults. Studies that examined both blood 

27 and bone Pb levels did not conclusively demonstrate a stronger association for either 

28 blood or bone Pb (Perlstein et al., 2007; Glenn et al., 2006; Martin et al., 2006). In a 

29 recent meta-analysis, Navas-Acien et al. (2008) found that studies passing the detailed 

30 inclusion criteria all showed a relationship between higher bone Pb levels and higher BP. 

31 Also, all but one that characterized hypertension showed higher relative risks or odds 

32 ratios associated with higher bone Pb levels. Recent epidemiologic studies also 

33 emphasized the interaction between bone Pb levels and factors that modify the 

34 association with BP or hypertension, such as race/ethnicity, chronic stress and metabolic 

35 syndrome. Bone Pb coupled with high perceived stress was associated with an increased 

36 risk of developing hypertension in an originally nonhypertensive group of adults (Peters 

37 et al., 2007). Also, bone Pb level was associated with larger decreases in HRV (which has 
February 2012 5-290 Draft – Do Not Cite or Quote

1 been associated with increased cardiovascular events) among adults with metabolic 

2 syndrome, also associated with increased risk of cardiovascular events (Park et al., 2006).

3 The extensive epidemiologic evidence for bone Pb levels in concert with the animal 

4 toxicological evidence for long-term Pb exposure, support an effect of long-term, 

5 cumulative Pb exposure on cardiovascular morbidity.

6 Recent epidemiologic studies of adults investigated the interaction of Pb biomarkers with 

7 genetic variants in associations with cardiovascular effects. Evidence was presented for a 

8 larger blood Pb-associated increase in BP in carriers of the ALAD2 allele, which is 

9 associated with greater binding affinity for Pb in the bloodstream (Figure 5-26 for results)

10 (Scinicariello et al., 2010). Additionally, bone Pb concentration was associated with 

11 larger increases in PP, which represents as a good predictor of cardiovascular morbidity 

12 and mortality and an indicator of arterial stiffness, among adults with the HFE H63D 

13 and/or C282Y variant (Zhang et al., 2010a) (Figure 5-26 for results). Park et al. (2009b)

14 provided further evidence of HFE and transferrin gene variants, related to iron 

15 metabolism, impacting the associations of bone Pb levels with cardiovascular effects, 

16 evaluated by QT interval changes.

17 Epidemiologic and toxicological evidence indicates that Pb exposure not only increases 

18 BP and hypertension, but can contribute to the development of other cardiovascular 

19 diseases in adults. However, fewer studies have been published compared to studies of 

20 BP and hypertension. Both recent epidemiologic and toxicological studies provide 

21 evidence in adults for blood Pb-associated increased atherosclerosis, thrombosis, IHD, 

22 PAD, arrhythmia, and cardiac contractility in populations with mean blood Pb levels 

23 >2.5 μg/dL (Table 5-19). Further, animal toxicological evidence continued to build on the 

24 evidence supporting the mechanisms leading to these cardiovascular system responses, as 

25 well as Pb-induced changes in BP and hypertension. Enhanced understanding of Pbinduced oxidative stress including • 26 NO inactivation, endothelial dysfunction leading to 

27 altered vascular reactivity, activation of the RAAS, and vasomodulator imbalance 

28 provides biological plausibility for the consistent associations observed between higher 

29 blood and bone Pb levels and greater cardiovascular effects.

30 Several studies in children reported associations of childhood (concurrent and measured 

31 at an average age of 2.6 years) blood Pb levels with cardiovascular changes such as TPR 

32 and autonomic and cardiovascular dysregulation in response to acute stress tasks 

33 measured between ages 5 to 15 years (Gump et al., 2011; 2009; Gump et al., 2007). Also, 

34 maternal bone and cord blood Pb levels, biomarkers of prenatal Pb exposure, were related 

35 to higher BP in children (Gump et al., 2005; Zhang et al., In Press). These study findings 

36 of children add evidence that Pb exposure is associated with small increases in BP, and 
February 2012 5-291 Draft – Do Not Cite or Quote

1 that changes in the cardiovascular system precedent to later CVD are potentially 

2 impacted by early-life Pb exposure.

3 New evidence extended the potential continuum of Pb-related cardiovascular effects by 

4 demonstrating associations between Pb biomarkers and both cardiovascular and all-cause 

5 mortality with follow-up periods ranging between 8 and 12 years. All-cause mortality 

6 was increased with increasing blood Pb level. A recent analysis of the NHANES III 

7 sample reported associations of adult blood Pb level with cardiovascular mortality, with 

8 stronger associations observed with myocardial infarction and stroke mortality (Menke et 

9 al., 2006). These findings were supported by a community-based cohort of women age 

10 65-87 years, in which higher effect estimates were observed for mortality from 

11 cardiovascular disease and coronary heart disease (Khalil et al., 2009a). Weisskopf et al.

12 (2009) published the first mortality study using bone Pb as an exposure index. This 

13 prospective study found that patella bone Pb levels were associated with increased 

14 mortality from cardiovascular disease and IHD with hazard ratios of 5.6 and 8.4, 

15 respectively.

16 Changes in BP that have been associated with biomarkers of Pb exposure indicate a 

17 modest change for an individual; however, these modest changes can have a substantial 

18 implication at the population level. The reported effects represent a central tendency of 

19 Pb-induced cardiovascular effects among individuals; some individuals may differ in risk 

20 and manifest effects that are greater in magnitude. For example, a small increase in BP 

21 may shift the population distribution and result in considerable increases in the 

22 percentages of individuals with BP values that are clinically significant, i.e., an indication 

23 of hypertension and medication use. Studies in the medical literature show that increasing 

24 BP, even within the nonhypertensive range, is associated with increased rates of death 

25 and cardiovascular disease, including coronary disease, stroke, PAD, and cardiac failure

26 (Ingelsson et al., 2008; Chobanian et al., 2003; Pastor-Barriuso et al., 2003; Prospective 

27 Studies Collaboration, 2002; Kannel, 2000a, b; Neaton et al., 1995).

28 In summary, new studies evaluated in the current review supported and expanded upon 

29 the strong body of evidence presented in the 2006 Pb AQCD that Pb exposure is 

30 associated with cardiovascular health effects. The weight of both epidemiologic and 

31 toxicological evidence continues to support a consistent relationship between Pb 

32 exposure and increased BP or hypertension development in adults. The epidemiologic 

33 evidence is strengthened by several prospective studies that find associations between 

34 biomarkers of Pb and BP and hypertension and by effect estimates that are observed after

35 adjustment for multiple potential confounding factors. The weight of epidemiologic 

36 evidence supported associations in adults with mean concurrent blood Pb levels less than 

37 5 μg/dL. As these outcomes in epidemiologic studies were most often observed in adults 
February 2012 5-292 Draft – Do Not Cite or Quote

1 with likely higher past than current Pb exposures, uncertainty exists as to the Pb exposure 

2 level, timing, frequency, and duration contributing to the observed associations. Recent 

3 epidemiologic studies found that bone Pb level, a metric of cumulative exposure, is 

4 strongly related to hypertension risk in adults with mean bone Pb levels greater than 

5 20 μg/g. However, uncertainties also exist as to the specific Pb exposure conditions that 

6 contributed to the associations. The weight of animal evidence also demonstrates an 

7 increase in BP after long-term (i.e., greater than 4 weeks) exposure to Pb. Whereas the 

8 majority of studies examined and found increases in BP in animals with mean blood Pb 

9 levels greater than 10 μg/dL, a recent studies found elevated BP in animals with a mean 

blood Pb level of 2 μg/dL. By demonstrating Pb-induced oxidative stress including • 10 NO 

11 inactivation, endothelial dysfunction leading to altered vascular reactivity, activation of 

12 the RAAS, and vasomodulator imbalance, toxicological studies have characterized the 

13 modes of action of Pb and provided biological plausibility for the consistent associations 

14 observed in epidemiologic studies between blood and bone Pb and cardiovascular effects. 

15 These associations of Pb with cardiovascular morbidity observed in both epidemiologic 

16 and toxicological studies support recent epidemiologic findings of increased Pb17 associated cardiovascular mortality. Collectively, the evidence integrated across 

18 epidemiologic and toxicological studies as well as across the spectrum of other 

19 cardiovascular endpoints examined is sufficient to conclude that there is a causal 

20 relationship between Pb exposures and cardiovascular health effects. 

5.5 Renal Effects

5.5.1 Introduction

21 This section summarizes key findings with regard to effects of Pb on the kidney in animal 

22 toxicology and epidemiologic studies. Findings summarized across epidemiologic and 

23 toxicological studies indicate that chronic Pb exposure is associated with pathological 

24 changes in the renal system such as proximal tubule (PT) cytomegaly, renal cell 

25 apoptosis, mitochondrial dysfunction, aminoaciduria, increased electrolyte excretion, 

26 ATPase dysfunction, oxidant redox imbalance, altered glomerular filtration rate (GFR), 

chronic kidney disease (CKD) development, and altered • 27 NO homeostasis with ensuing 

28 elevated BP. As several of these outcomes are most often observed in adults with likely 

29 higher past Pb exposures, uncertainty exists as to the Pb exposure level, timing, 

30 frequency, and duration contributing to the associations observed with blood or bone Pb 

31 levels. 
February 2012 5-293 Draft – Do Not Cite or Quote

1 The cardiovascular and renal systems are intimately linked. Homeostatic control at the 

2 kidney level functions to regulate water and electrolyte balance via filtration, re3 absorption and excretion and is under tight hormonal control. Pb exposure damages the 

4 kidneys and its vasculature and systemic hypertension ensues with effects on the 

5 cardiovascular and renal systems (Section 5.4). Chronic increases in vascular pressure 

6 can contribute to glomerular and renal vasculature injury, which can lead to progressive 

7 renal dysfunction and kidney failure. In this manner, Pb-induced hypertension has been 

8 noted as one cause of Pb-induced renal disease. However, the relationship between BP 

9 and renal function is more complicated. Not only does hypertension contribute to renal 

10 dysfunction but damage to the kidneys can also cause increased BP. Long-term control of 

11 arterial pressure is affected by body fluid homeostasis which is regulated by the kidneys. 

12 In examining the physiological definition of BP (i.e., mean BP equates to cardiac output 

13 multiplied by total peripheral resistance [TPR]) the role of the kidneys in BP regulation is 

14 highlighted. Cardiac output is driven by left ventricular and circulating blood volume. 

15 TPR is driven by vasomodulation and electrolyte balance. Thus, it is possible to dissect 

16 the causes of hypertension from features of primary kidney disease. Increased 

17 extracellular fluid volume results in increased blood volume which enhances venous 

18 return of blood to the heart and increases cardiac output. Increased cardiac output not 

19 only directly increases BP, but also increases TPR due to a compensatory autoregulation 

20 or vessel constriction. In addition, damage to the renal vasculature will alter the intra21 renal vascular resistance thereby altering kidney function and affecting the balance 

22 between renal function and BP. The interactions between these systems can lead to 

23 further exacerbation of vascular and kidney dysfunction following Pb exposure. As 

24 kidney dysfunction can increase BP and increased BP can lead to further damage to the 

25 kidneys, Pb-induced damage to both systems may result in a cycle of further increased 

26 severity of disease.

27 In general, associations between bone Pb (particularly in the tibia) and health outcomes in 

28 adults indicate chronic effects of cumulative Pb exposure. In adults without current 

29 occupational Pb exposure, blood Pb level represents both recent and cumulative Pb 

30 exposure. In particular, blood Pb level may represent cumulative exposure in 

31 physiological circumstances of increased bone remodeling or loss (e.g., osteoporosis and 

32 pregnancy) when Pb from bone of adults contributes substantially to blood Pb 

33 concentrations. Blood Pb in children is also influenced by Pb stored in bone due to rapid 

34 growth-related bone turnover in children relative to adults. Thus, blood Pb in children is 

35 also reflective of cumulative dose. Additional details on the interpretation of Pb in blood 

36 and bone are provided in Section 4.3.5. The toxicokinetics of Pb in blood and bone are 

37 important considerations in making inferences about etiologically-relevant Pb exposures 

38 that contributed to associations observed between blood and bone Pb levels and health 

39 outcomes.
February 2012 5-294 Draft – Do Not Cite or Quote

5.5.1.1 Kidney Outcome Measures

1 The primary function of the kidneys is to filter waste from the body while maintaining 

2 appropriate levels of water and essential chemicals, such as electrolytes, in the body. 

3 Therefore, the gold standard for kidney function assessment involves measurement of the 

4 GFR through administration of an exogenous radionuclide or radiocontrast marker 

5 (e.g., 125I-iothalamate, iohexol) followed by timed sequential blood samples or, more 

6 recently, kidney imaging, to assess clearance through the kidneys. This procedure is 

7 invasive and time-consuming. Therefore, serum levels of endogenous compounds are 

8 routinely used to estimate GFR in large epidemiologic studies and clinical settings. 

9 Creatinine is the most commonly measured endogenous compound; blood urea nitrogen 

10 (BUN) has also been examined. Increased serum concentration or decreased kidney 

11 clearance of these markers both indicate decreased kidney function. The main limitation 

12 of endogenous compounds identified to date is that non-kidney factors impact their serum 

13 levels. Specifically, since creatinine is metabolized from creatine in muscle, muscle mass 

14 and diet affect serum levels resulting in variation in different population subgroups

15 (e.g., women and children compared to men), that are unrelated to kidney function. 

16 Measured creatinine clearance, involving measurement and comparison of creatinine in 

17 both serum and urine, can address this problem. However, measured creatinine clearance 

18 utilizes timed urine collections, traditionally over a 24-hour period, and the challenge of 

19 complete urine collection over an extended time period makes compliance difficult. 

20 Therefore equations to estimate kidney filtration that utilize serum creatinine but also 

21 incorporate age, sex, race, and, in some, weight (in an attempt to adjust for differences in 

22 muscle mass), have been developed. Although these are imperfect surrogates for muscle 

23 mass, such equations are currently the preferred outcome assessment method. 

24 Traditionally, the Cockcroft-Gault equation (Cockcroft and Gault, 1976), which estimates 

25 creatinine clearance, a GFR surrogate, has been used. In the last decade, the abbreviated 

26 Modification of Diet in Kidney Disease (MDRD) Study equation (Levey et al., 2000; 

27 Levey et al., 1999), which estimates GFR, has become the standard in the kidney 

28 epidemiologic and clinical communities. With widespread use of the MDRD equation, it 

29 became clear that the equation underestimates GFR at levels in the normal range. 

30 Therefore, the CKD-Epidemiology Collaboration (CKD-EPI) equation was recently 

31 developed to be more accurate in this range (Levey et al., 2009). This is a decided 

32 advantage in nephrotoxicant research since most participants in occupational and many 

33 even in general population studies have GFRs in a range that is underestimated by the 

34 MDRD equation.

35 Both the MDRD and CKD-EPI equations use serum creatinine. Due to the inability to 

36 adjust serum creatinine levels for muscle mass, alternative serum biomarkers have been 
February 2012 5-295 Draft – Do Not Cite or Quote

1 evaluated such as cystatin C, a cysteine protease inhibitor that is filtered, reabsorbed, and 

2 catabolized in the kidney (Fried, 2009). It is produced and secreted by all nucleated cells 

3 thus avoiding the muscle mass confounding that exists with serum creatinine (Fried, 

4 2009). However, recent research indicates that serum cystatin C varies by age, sex, and 

5 race (Kottgen et al., 2008). Thus, a cystatin C-based eGFR equation was recently 

6 developed that includes age, sex, and race (Stevens et al., 2008).

7 Most of the kidney outcome measures discussed above were developed for use in the 

8 clinical setting. Unfortunately, they are insensitive for detection of early kidney damage, 

9 as evidenced by the fact that serum creatinine remains normal after kidney donation. 

10 Therefore, in the last two decades, the utility of early biological effect (EBE) markers as 

11 indicators of preclinical kidney damage has been of interest. These can be categorized as 

12 markers of function (i.e., low molecular weight proteins that should be reabsorbed in the 

13 PT such as β2-microglobulin and retinol-binding protein [RBP]); biochemical alteration 

14 (i.e., urinary eicosanoids such as prostaglandin E2, prostaglandin F2 alpha, 6-keto15 prostaglandin F1 alpha, and thromboxane B2); and cytotoxicity (e.g., N-acetyl-β-D16 glucosaminidase [NAG]) (Cardenas et al., 1993). Elevated levels may indicate an 

17 increased risk for subsequent kidney dysfunction. However, most of these markers are 

18 research tools only, and their prognostic value remains uncertain since prospective 

19 studies of most of these markers in nephrotoxicant-exposed populations are quite limited 

20 to date. Recently, microalbuminuria has been identified as a PT marker, not just 

21 glomerular as previously thought (Comper and Russo, 2009). Kidney EBE markers are a 

22 major recent focus for research in patients with acute kidney injury (AKI) and markers 

23 such as neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 

24 (Kim-1), developed in AKI research, may prove useful for chronic nephrotoxicant work 

25 as well (Ferguson et al., 2008; Devarajan, 2007).

5.5.2 Nephrotoxicity and Renal Pathology

5.5.2.1 Toxicology

26 Figure 5-38 and Table 5-22 presents the recent animal toxicological data for studies 

27 investigating the effects of Pb (as blood Pb level) on various measures of kidney health 

28 and function. In animals, Pb has been found to induce changes in a wide range of 

29 indicators of renal function. Most studies examined Pb exposure concentrations that 

30 resulted in higher blood Pb levels (> 20 μg/dL) than those in the current U.S. general 

31 population. As indicated in Figure 5-38 and Table 5-22, toxicological information on 

32 renal dysfunction with blood Pb levels < 10 μg/dL) generally is not available. 
February 2012 5-296 Draft – Do Not Cite or Quote

1 Dysfunction in kidney function measures, including urinary flow, ALP, microalbumin, 

2 and NAG, was observed at blood Pb concentrations above 20 μg/dL (Wang et al., 

3 2010d).

Figure 5-38 Concentration-response representation of the effect of Pb on 

renal outcomes in animal toxicology studies.

1 10 100
February 2012 5-297 Draft – Do Not Cite or Quote

Table 5-22 Additional characteristics for results of toxicological studies 

presented in Figure 5-38

Reference

Species; 

Lifestage; 

Sex

Pb Dose; Exposure 

Duration

Blood Pb Level 

with Response 

(μg/dL) Outcome

Wang et al.

(2010d)

Rat; Adult; 

Female

300 ppm Pb-acetate in drinking 

water; 8 weeks 20 Biomarker - Aberrant NAG, GGT, β2-microgobulin expression

Roncal et al. (2007)Rat; Adult; 

Male

150 ppm Pb-acetate in drinking 

water; 16 weeks with remnant 

kidney surgery at week 4

26

Inflammation - Elevation in number of macrophages & marker MCP-1 

in Pb exposed kidneys with remnant kidney surgery.

Navarro-Moreno 

et al. (2009)

Rat; Adult; 

Male

500 ppm Pb-acetate in drinking 

water; 28 weeks 43 Oxidative stress - Increased kidney lipid peroxidation (i.e., TBARS) 

Wang et al.

(2010d)

Rat; Adult; 

Female

300 ppm Pb-acetate in drinking 

water; 8 weeks 20

Oxidative Stress - Pb caused increased lipid peroxidation (i.e., MDA 

production), elevated kidney antioxidant enzymes (SOD, GPx, CAT), 

and depleted GSH

Massó-González 

et al. (2009)

Rat; 

Weanling 

pups

300 ppm Pb-acetate in drinking 

water; GD1 to PND21 23

Oxidative stress - Elevated TBARS and catalase activity 

Roncal et al.

(2007)

Rat; Adult; 

Male

150 ppm Pb-acetate in drinking 

water; 16 weeks with remnant 

kidney surgery at week 4

26

Morphology - Pb induced pre-glomerular vascular disease of kidney 

(i.e., sclerosis, fibrosis, peritubular capillary loss)

Navarro-Moreno 

et al. (2009)

Rat; Adult; 

Male

500 ppm Pb-acetate in drinking 

water; 28 weeks 43 Morphology - Electron micrography showed lumen reduction, microvilli loss, brush border loss, and mitochondrial damage

Wang et al.

(2010d)

Rat; Adult; 

Female

300 ppm Pb-acetate in drinking 

water; 8 weeks 20

Morphology - Electron micrography showed Pb damages 

mitochondria, basement membrane, and brush border in kidney 

tissue. Some focal tubal necrosis observed.

Massó-González 

et al. (2009)

Rat; 

Weanling 

pups

300 ppm Pb-acetate in drinking 

water; GD1 to PND21 23

Morphology - Pb elevated relative kidney weight at PND21

Navarro-Moreno 

et al. (2009)

Rat; Adult; 

Male

500 ppm Pb-acetate in drinking 

water; 28 weeks 43 Kidney function - Pb exposed males had elevated urinary pH and protein, and glucose and blood in the urine.

Roncal et al.

(2007)

Rat; Adult; 

Male

150 ppm Pb-acetate in drinking 

water; 16 weeks with remnant 

kidney surgery at week 4

26

Kidney function - Remnant kidney surgery and Pb exposure induced 

decreased creatinine clearance and proteinuria.

Wang et al.

(2010d)

Rat; Adult; 

Female

300 ppm Pb-acetate in drinking 

water; 8 weeks 20 Kidney function - Elevated urinary total protein, urinary albumin, and serum urea nitrogen.

Ademuyiwa et al.

(2009)

Rat; Adult 200, 300, and 400 ppm 

Pb-acetate in drinking water; 

12 weeks

39 and 61

Kidney Function - Renal phospholipidosis and depletion of renal 

cholesterol after gestational Pb exposure.

Renal Function and Interstitial Fibrosis

1 Past studies have shown that chronic continuous or repeated Pb exposure can result in 

2 interstitial nephritis and focal or tubular atrophy. After an initial 3 months of Pb exposure 

3 (in a longitudinal 12-month exposure study to either 100 ppm [lower dose] or 5,000 ppm

4 [higher dose] Pb-acetate in drinking water, male rats), elevated GFR, consistent with 

5 hyperfiltration, and renal hypertrophy were observed; high dose animals also had 

6 increased NAG and GST (Khalil-Manesh et al., 1993a; Khalil-Manesh et al., 1992a; 

7 Khalil-Manesh et al., 1992b). At 6 months of exposure, GFR decreased in the high dose 

8 animals, albuminuria was present, and pathology ensued with focal tubular atrophy and 
February 2012 5-298 Draft – Do Not Cite or Quote

1 interstitial fibrosis formation. This pathology was persistent out to 12 months, and at 12 

2 months glomeruli developed focal and segmental sclerosis. Similarly, GFR remained 

3 decreased after 12 months of exposure in the high dose group. The toxicological evidence 

4 for differences in GFR according to duration of Pb exposure, i.e., hyperfiltration with 3-

5 month exposure versus decreased GFR with 6- or 12-month exposure, provide biological 

6 plausibility for epidemiological studies that observed a similar phenomenon by age in 

7 adults in association with Pb biomarker levels. These duration-dependent dichotomous 

8 changes in GFR are consistent between the toxicological and epidemiologic literature. 

9 Biomarkers of Pb-induced renal toxicity have been developed including the enzymes 

10 lysosomal NAG, GST, brush border antigens (BB50, BBA, HF5), and Tamm-Horsfall 

11 protein. GST functions as a renal biomarker since renal ALAD is protected by the kidney 

12 antioxidant GSH. Urinary NAG and GST levels were found to increase in rats after 3 

13 months of high dose Pb exposure (resulting in blood Pb level of 125 μg/dL) in rats

14 (Khalil-Manesh et al., 1992a), whereas only urinary NAG was increased following low 

15 dose Pb exposure (resulting in blood Pb level of 29 μg/dL) in rats (Khalil-Manesh et al., 

16 1993a). An in vitro study also found an increase in NAG with high-level Pb exposure 

17 (50 μg/dL) (Dehpour et al., 1999). Occupational studies found that urinary NAG 

18 correlated best with recent blood Pb changes. 

19 The renal effects of chronic Pb exposure as detailed above were partially rescued in rats 

20 with chelation therapy such as DMSA (Khalil-Manesh et al., 1992b). Improvements 

21 include increased GFR, decreased albuminuria, and decreased inclusion body numbers 

22 but little change in tubulointerstitial scarring. Administration of an Indian herb to Pb23 exposed mice, as is discussed in greater detail in the antioxidant section (Section 5.5.5), 

24 produced similar findings. There was a functional rescue however Pb-induced pathology 

25 remained (Jayakumar et al., 2009). Thus, administration of various compounds 

26 (chelators, antioxidants) to Pb-exposed animals produced hemodynamic rescue. 

27 Recent studies have corroborated the previously observed increase in serum creatinine 

28 following Pb exposure in rats. Abdel Moneim et al. (2011b) reported Pb-induced (i.p. 

29 20 ppm, 5 days) increased serum creatinine accompanied by histological alterations in rat 

30 kidneys. Berrahal et al. (2011) reported on the effects of age-dependent exposure to Pb on 

31 nephrotoxicity in male rats. Pups were exposed to Pb lactationally (as a result of dams 

32 consuming water containing 50 ppm Pb-acetate) until weaning. Thereafter, the offspring 

33 were exposed to the same solution from weaning (day 21) until sacrifice. Male pups were 

34 sacrificed at age 40 days (puberty; blood Pb level 12.7 μg/dL) and at age 65 days (post35 puberty; blood Pb level 7.5 μg/dL). Serum creatinine was elevated at both 40 days and 65 

36 days (0.54 and 0.60 mg/dL compared to control values of 0.45 mg/dL 

37 [p <0.001]).Various parameters of Pb-induced renal dysfunction are listed in Table 5-23
February 2012 5-299 Draft – Do Not Cite or Quote

1 below. The elevated serum creatinine in the Pb-exposed animals compared to controls 

2 suggests that animals exposed to low dosage (i.e., 50 ppm) Pb from birth may develop 

3 renal abnormalities. However, the lack of measurements of GFR or renal pathology 

4 weakens the conclusions. Other investigators have also shown that chronic Pb exposure 

5 has detrimental effects on renal function. Pb-exposed male rats (500 ppm Pb-acetate in 

6 drinking water for 7 months) had elevated urinary pH, proteinuria, as well as glucose and 

7 blood in the urine (Navarro-Moreno et al., 2009).

Table 5-23 Indicators of renal damage in male rats exposed to 50 ppm Pb for 40 

and 65 days, starting at parturition

Biomarker (Mean±SD) PND40

Control

PND40

Pb

PND65

Control

PND65

Pb

Blood Pb level

(μg/dL)

1.8±0.33 12.7±1.7 2.1±0.35 7.5±0.78

Plasma Creatinine

(mg/L)

4.5±0.21 5.35±0.25a 4.55±0.27 6.04±0.29a

Plasma Urea

(mg/L)

0.37±0.019 0.47±0.021a 0.29±0.009 0.29±0.009

Plasma Uric Acid

(mg/L)

7.51±0.44 7.65±0.32 9.39±0.82 5.91±0.53a

a

p <0.001

Source: Modified with permission of John Wiley & Sons, Berrahal et al. (2011)

8 Qiao et al. (2006) measured the effect of Pb on the expression of the renal fibrosis-related 

9 nuclear factor-kappa B (NF-κB), transforming growth factor (TGF-β) and fibronectin in 

10 Sprague-Dawley rat kidney. Pb was administered at a dose of 0.5% Pb-acetate, 

11 continuously for either one, two or three months. All growth factors increased by the end 

12 of three months of treatment but only NF-κB increased progressively at each time period. 

13 These changes were hypothetically related to the development of Pb-induced renal 

14 fibrosis in rats, but no histology was performed.

15 Roncal et al. (2007) found that Pb accelerated arteriolopathy and tubulointerstitial injury 

16 in non-Pb-related CKD. Sprague-Dawley rats were administered Pb-acetate at 150 ppm

17 for 4 weeks, then subjected to remnant kidney surgery (left kidney mass reduced by 2/3 

18 and right kidney removed), and subsequently exposed to Pb for an additional 12 weeks. 

19 Pb-treated rats had higher systolic BP, lower creatinine clearance, and higher proteinuria 

20 than did controls. Most striking was development of worse arteriolar disease, peritubular 

21 capillary loss, tubulointerstitial damage, and macrophage infiltration. Pb treatment was 

22 associated with significant worsening of pre-glomerular vascular disease, as characterized 

23 by an increase in the media-to-lumen ratio. There was also a higher percentage of 
February 2012 5-300 Draft – Do Not Cite or Quote

1 segmental sclerosis within glomeruli and a tendency for a higher number of sclerotic 

2 glomeruli. Additionally, a loss of peritubular capillaries, as reflected by a reduction in 

3 thrombomodulin staining, was observed. This was associated with worse tubular injury 

4 (osteopontin staining) due to more interstitial fibrosis (type III collagen staining) and a 

5 greater macrophage infiltration in the interstitium. The increase in macrophages was 

6 associated with higher renal MCP-1 mRNA. As a whole, these findings indicate that Pb 

7 exposure concomitant with existing renal insufficiency due to surgical kidney resection 

8 accelerated vascular disease and glomerular pathology. These findings are consistent with 

9 the previous work of Bagchi and Preuss (2005) also showing that Pb-exposed animals 

10 with non-Pb-related CKD (remnant surgery) had kidney dysfunction including 

11 impairment of the renin-angiotensin system (Losartan challenge), elevated systolic BP, 

and alterations in renal excretion of Pb, K+, and Na+ 12 . Thus, this model shows that low Pb 

13 exposure may exacerbate pre-existing underlying kidney disease.

Histological Changes 

14 Historical studies discussed in previous Pb AQCDs have identified Pb-related renal

15 damage by the presence of dense intranuclear inclusion bodies, which are capable of 

16 sequestering Pb (Goyer et al., 1970a). Chelators such as CaNa2EDTA removed these 

17 inclusion bodies from affected nuclei (Goyer et al., 1978). Multiple endpoints indicate 

18 dysfunction in the PT after Pb exposure. Pb-induced formation of intranuclear inclusion 

19 bodies in the PT is protective; Pb is sequestered such that it is not in its bioavailable, free, 

20 toxicologically active form. Intranuclear inclusion bodies are found in the kidney with 

21 acute (i.e., < 4 weeks) Pb exposures but present to a lesser degree with chronic exposures 

22 (See Section 5.2.3 for further discussion). Other PT ultrastructural changes in Pb-induced 

23 nephropathy include changes to the PT epithelium, endoplasmic reticulum dilation, 

24 nuclear membrane blebbing, and autophagosome enlargement (Fowler et al., 1980; Goyer 

25 et al., 1970b). Symptoms similar to the PT transport-associated Fanconi syndrome appear 

26 with Pb exposure, albeit often at high doses of Pb, i.e., Pb-poisoning. These symptoms, 

27 which include increased urinary electrolyte excretion (zinc), decreased Na-K-ATPase 

28 activity, mitochondrial aberrations, and aminoaciduria, also have been associated with 

29 blood Pb levels in children. 

30 New studies since the 2006 Pb AQCD are consistent with the historical findings and 

31 build upon the literature base by including the role of antioxidants. Jabeen et al. (2010)

32 exposed pregnant albino BALB/c mice to a daily oral dose of Pb-acetate (10 mg/kg body 

33 weight, daily throughout pregnancy) until GD18, at which point the fetal kidneys were 

34 processed for histological examination. Histology revealed Pb exposure induced 

35 decreased kidney cortical thickness, decreased diameter of renal corpuscles, and 

36 increased renal tubular atrophy with desquamated epithelium and degenerated nuclei in 
February 2012 5-301 Draft – Do Not Cite or Quote

1 the distal and proximal tubules. Blood Pb levels were not reported in this study. 

2 Nonetheless, these data show that in utero Pb exposure had significant histological effects 

3 on the fetal kidney, which could contribute to altered renal function including clearance 

4 of waste products, electrolyte balance, and vasoregulation. Massanyi et al. (2007)

5 reported on Pb-induced alterations in male Wistar rat kidneys after single i.p. doses of 

6 Pb-acetate (50, 25, and 12.5 mg/kg); kidneys were removed and analyzed 48 hours after 

7 Pb administration. Qualitative microscopic analysis detected dilated Bowman’s capsules 

8 and dilated blood vessels in the interstitium with evident hemorrhagic alterations. 

9 Quantitative histomorphometric analysis revealed increased relative volume of 

10 interstitium and increased relative volume of tubules in the experimental groups. The 

11 diameter of renal corpuscles and the diameter of glomeruli and Bowman’s capsule were 

12 significantly increased. Measurement of tubular diameter showed dilatation of the tubule 

13 with a significant decrease of the height of tubular epithelium compatible with 

14 degenerative renal alterations. These findings extend the observations of Fowler et al.

15 (1980) and Khalil-Manesh et al. (1992a; 1992b); in particular, the enlarged glomeruli are 

16 consistent with the early hyperfiltration caused by Pb. 

17 Abdel Moneim et al. (2011b) reported histological evidence of inflammation after Pb 

18 treatment in rats (i.p. 20 ppm, 5 days). This evidence included increased inflammatory 

19 cellular infiltrations, cytoplasmic vacuolation, and dilatation of some kidney tubules. 

20 Inflammation was accompanied by an increase in apoptotic cells and increased oxidative 

21 stress.

22 A recent study has also reported inclusion body formation in the nuclei, cytoplasm, and 

23 mitochondria of PT cells of Pb-treated rats (50 mg Pb/kg bw i.p., every 48 hours for 14 

24 days) (Navarro-Moreno et al., 2009). These inclusion bodies were not observed in 

25 chronically Pb-exposed rats (500 ppm Pb in drinking water, 7 months). However, chronic 

26 Pb exposure resulted in morphological alterations including loss of PT apical membrane 

27 brush border, collapse and closure of the PT lumen, and formation of abnormal 

28 intercellular junctions.

29 Vogetseder et al. (2008) examined the proliferative capacity of the renal PT (particularly 

30 the S3 segment) following i.v. administration of Pb to juvenile and adult male Wistar 

31 rats. Proliferation induction was examined by detection of Bromo-2’-deoxyuridine 

32 (BrdU), Ki-67 (labels S, G2, and M phase cells), and cyclin D1 (an essential cell cycle 

33 progression protein). The cycling marker Ki-67 revealed a much higher proliferation rate 

34 in the S3 segment in control juvenile rats (4.8 ± 0.3%) compared with control adult rats 

35 (0.4 ± 0.1%). Pb administration (3.8 mg/100 g bw) increased the proportion of Ki-67-

36 positive cells to 26.1 ± 0.3% in juvenile rats and 31.9 ± 0.3% in adult rats. Thus, the 

37 increased proliferation caused by Pb was age independent. The proliferation induction 
February 2012 5-302 Draft – Do Not Cite or Quote

caused by Pb administration may be a result of reduced cell cycle inhibition by p27kip-1 1 . 

2 Acute Pb treatment increased the incidence of cyclin D1 labeling in the BrdU-positive 

3 cells suggesting Pb was able to accelerate re-entry of cells into the cell cycle and cause 

4 proliferation in the PT. Pb-induced cellular proliferation has also been reported in the 

5 retina with gestational and early postnatal rodent Pb exposure (Giddabasappa et al., 

6 2011).

7 Ademuyiwa et al. (2009) examined Pb-induced phospholipidosis and cholesterogenesis in 

8 rat tissues. Sprague-Dawley rats were exposed to 200, 300 and 400 ppm Pb-acetate for 12 

9 weeks. The Pb exposure resulted in induction of phospholipidosis in kidney tissue, 

10 accompanied by depletion of renal cholesterol. The authors suggested that induction of 

11 cholesterogenesis and phospholipidosis in kidney may be responsible for some of the 

12 subtle and insidious cellular effects found with Pb-mediated nephrotoxicity. Drug13 induced PT phospholipidosis is seen clinically with use of the potentially nephrotoxic 

14 aminoglycoside drugs, including gentamicin (Baronas et al., 2007).

15 Various antioxidants have been shown to attenuate Pb-induced histopathological changes 

16 to the kidney. Ozsoy et al. (2010) found L-carnitine to be protective in a model of 

17 experimental Pb toxicity in female rats. Markers of histopathological change in the 

18 kidney, including tubule dilatation, degeneration, necrosis, and interstitial inflammation 

19 were rescued by L-carnitine treatment in females. Male rats exposed to Pb (0.2% for 6 

20 weeks) also displayed tubular damage, whereas concomitant treatment with Pb and an 

21 extract of Achyranthes aspera ameliorated the observed damage (Jayakumar et al., 2009).

22 El-Nekeety et al. (2009) found an extract of the folk medicine plant Aquilegia vulgaris to 

23 be protective against Pb-acetate-induced kidney injury in Sprague-Dawley rats. Rats were 

24 treated with Pb (20 ppm; 2 weeks) and extract (administered before, during, or after Pb). 

25 Pb treatment resulted in tubular dilatation, vacuolar and cloudy epithelial cell lining, 

26 interstitial inflammatory cell infiltration, hemorrhage, cellular debris, and glomerulus 

27 hypercellularity. Concomitant exposure to Pb and extract produced histology 

28 indiscernible from that in controls. Post treatment with extract partially rescued the Pb29 induced histopathology. El-Neweshy and El-Sayed (2011) studied the influence of 

30 vitamin C supplementation (20 mg/kg pretreatment every other day) on histopathological 

31 alterations in Pb-exposed male rats (20 mg/kg by intragastric feeding once daily for 60 

32 days). Control rats showed normal histology, while Pb-treated rats exhibited karyomegaly 

33 with eosinophilic intranuclear inclusion bodies in the epithelial cells of the proximal 

34 tubules. Glomerular damage and tubular necrosis with invading inflammatory cells were 

35 also found. Rats treated with Pb-acetate plus vitamin C exhibited relatively mild or no 

36 karyomegaly with eosinophilic intranuclear inclusion bodies in the proximal tubules. 

37 Normal glomeruli were noted in animals exposed to Pb and vitamin C. These findings are 

38 presented in more detail in Section 5.5.5 but they consistently show that some 
February 2012 5-303 Draft – Do Not Cite or Quote

1 antioxidants are capable of preventing or rescuing Pb-induced renal histopathological 

2 changes.

Alteration of Renal Vasculature and Reactivity

3 As discussed in Section 5.5.1, changes in renal vasculature function or induction of 

4 hypertension can contribute to further renal dysfunction. Pb can increase BP through the 

5 promotion of oxidative stress and altered vascular reactivity. Also, Pb has been shown to 

6 act on known vasomodulating systems in the kidney. In the kidney, two vascular tone 

mediators, • 7 NO and ET-1, are found to be affected by Pb exposure. Antioxidants 

8 attenuated Pb-related oxidative/nitrosative stress in the kidney and abrogated the Pb9 induced increased BP (Vaziri et al., 1999b). Administration of the vasoconstrictor 

10 endothelin-1 (ET-1) affected mean arterial pressure (MAP) and decreased GFR (Novak 

11 and Banks, 1995). Acute high-dose Pb exposure (24 nmol/min for 15 or 30 minutes) 

12 completely blocked this ET-1-mediated GFR decrease but had no effect on MAP. 

13 Depletion of the endogenous antioxidant glutathione using the drug buthionine 

14 sulfoximine, a GSH synthase inhibitor, increased BP and increased kidney nitrotyrosine 

15 formation without Pb exposure, demonstrating the importance of GSH in maintenance of 

16 BP (Vaziri et al., 2000). Multiple studies have shown that Pb exposure depletes GSH 

17 stores. Catecholamines are vascular moderators that are also affected by Pb exposure

18 (Carmignani et al., 2000). The effect on BP with Pb exposure is especially relevant to the 

19 kidney because it is both a target of Pb deposition and a mitigator of BP. These historic 

20 data detail the interaction of known modulators of vascular tone with Pb.

21 Recently, Vargas-Robles et al. (2007) examined the effect of Pb exposure (100 ppm

22 Pb-acetate for 12 weeks) on BP and angiotensin II vasoconstriction in isolated perfused 

23 kidney and interlobar arteries. Vascular reactivity was evaluated in the presence and 

24 absence of the nitric oxide synthase inhibitor L-NAME in both Pb-treated and control 

25 animals. Pb exposure significantly increased BP (134 ± 3 versus 100 ± 6 mmHg), eNOS 

26 protein expression, oxidative stress, and vascular reactivity to angiotensin II. L-NAME 

27 potentiated the vascular response to angiotensin II in the control group, but had no effect 

28 on the Pb-treated group. Conversely, passive microvessel distensibility, measured after 

29 deactivation of myogenic tone by papaverine, was significantly lower in the arteries of 

30 Pb-exposed rats. Nitrites released from the kidney under the influence of angiotensin II in 

31 the Pb group were lower as compared to the control group whereas 3-nitrotyrosine was 

32 higher in the Pb group. The authors concluded that Pb exposure increases vascular tone 

33 through nitric oxide-dependent and -independent mechanisms, increasing renal vascular 

34 sensitivity to vasoconstrictors.
February 2012 5-304 Draft – Do Not Cite or Quote

1 Simões et al. (2011) reported that acute Pb treatment (Pb-acetate i.v. bolus dose of 

2 320 μg/kg bw, blood Pb of 37 μg/dL at 120 minutes after Pb administration) in adult 

3 male Wistar rats increased systolic arterial pressure 60 minutes after treatment without 

4 affecting diastolic arterial pressure or heart rate. With this single injection, serum 

5 angiotensin converting enzyme (ACE) activity was significantly elevated. The Pb6 induced altered systolic BP was under the control of the renin-angiotensin system as 

7 evidenced by attenuation of the effects of Pb in Losartan (Ang II receptor blocker) or 

8 Enalapril (ACE inhibitor) co-treated animals (Simões et al., 2011). These data agree with 

9 earlier reports of Pb-related increases in ACE activity in young rats exposed to Pb for 2-8 

10 weeks (Sharifi et al., 2004) and adult rats exposed to Pb for 10 months (Carmignani et al., 

11 1999).

Apoptosis and/or Ischemic Necrosis of Tubules and Glomeruli

12 Apoptosis or programmed cell death in excess can cause cell atrophy while an 

13 insufficiency can lead to uncontrolled cell proliferation, such as cancer. Pb exposure has 

14 been shown to cause morphological changes to the kidney structure. Some of these Pb15 induced changes are a result of cellular apoptosis or necrosis. Past studies have shown 

16 Pb-induced necrosis in proximal tubule cells (Fowler et al., 1980). Pb-induced apoptosis 

17 is known to act through the mitochondria (Rana, 2008). Pb-induced calcium overload 

18 may depolarize the mitochondria, resulting in cytochrome c release, caspase activation, 

19 and apoptosis. The apoptosis is mediated by Bax translocation to the mitochondria and 

20 can be blocked by overexpression of Bcl-xl. Also, Pb-induced ALA accumulation can 

21 generate ROS, which may damage DNA leading to apoptosis. 

22 Mitochondria are targets of Pb toxicity and often involved in apoptosis. Pb can induce 

23 uncoupling of oxidative phosphorylation, decreased substrate utilization, and 

24 modification of mitochondrial ion transport. ATP energetics are affected when ATP-Pb 

25 chelates are formed and ATPase activity is decreased. ROS formation can contribute to 

26 these mitochondrial changes and to other changes within the kidney. Antioxidant 

27 supplementation after Pb exposure can remedy some changes. All of these outcomes, in 

28 conjunction with Pb-related depletion of antioxidants (e.g., GSH) and elevation of lipid 

29 peroxidation point to possible susceptibility of the kidney to apoptosis or necrosis. 

30 Rodriguez-Iturbe et al. (2005) reported that chronic exposure to low doses of Pb 

31 (100 ppm in drinking water for 14 weeks) results in renal infiltration of immune cells, 

32 apoptosis, NF-κB activation and overexpression of tubulointerstitial Ang(II). Similarly, 

33 higher level Pb treatment in rats (i.p. 20 mg/kg, 5 days) induced inflammatory cellular 

34 infiltrations and an increase in apoptotic cells, accompanied by more pronounced BAX 

35 staining in kidney tubule epithelial cells (Abdel Moneim et al., 2011b). Pb treatment (0.5-
February 2012 5-305 Draft – Do Not Cite or Quote

1 1 μM) of isolated rat proximal tubular cells increased cell death by apoptosis and necrosis 

2 in a concentration- and time-dependent manner (Wang et al., 2011b). This was 

3 accompanied by increased morphological changes typical of apoptosis such as 

4 fragmented chromatin, condensed chromatin, and shrunken nuclei. These cells also 

5 exhibited decreased mitochondrial membrane potential, decreased intracellular pH, 

6 inhibition of Na+-K+ ATPase and Ca2+-ATPase activity, and increased intracellular 

7 Ca2+ following Pb treatment.

8 Navarro-Moreno et al. (2009) examined the effect of 500 ppm Pb in drinking water over 

9 7 months on the structure (including intercellular junctions), function, and biochemical 

10 properties of PT cells of Wistar rats. Pb effects in epithelial cells consisted of an early 

11 loss of the apical microvilli, followed by a decrement of the luminal space and the 

12 respective apposition and proximity of apical membranes, resulting in the formation of 

13 atypical intercellular contacts and adhesion structures. Inclusion bodies were found in 

14 nuclei, cytoplasm, and mitochondria. Lipid peroxidation (TBARS measurement) was 

15 increased in the Pb-treated animals as compared to controls. Calcium uptake was 

16 diminished and neither proline nor serine incorporation that was present in controls was 

17 noted in the PT of Pb-exposed animals. The authors speculated that Pb may compete with 

18 calcium in the establishment and maintenance of intercellular junctions.

19 Tubular necrosis was also observed in rats treated with Pb-acetate (100 ppm s.c.) for 30 

20 days (El-Sokkary et al., 2005). Histological sections of kidneys from Pb-treated rats 

21 showed tubular degeneration with some necrotic cells. Similarly, El-Neweshy and El22 Sayed reported glomerular damage and tubular necrosis with invading inflammatory 

23 cells after Pb treatment (20 mg/kg by intragastric feeding once daily for 60 days) to male 

24 rats. The incidence of necrosis was decreased in both of these studies by pretreatment 

25 with either melatonin or vitamin C. Pretreatment with melatonin (10 mg/kg), an 

26 efficacious free radical scavenger and indirect antioxidant, resulted in a near normal 

27 tubular structure. The authors concluded that melatonin protected the liver and kidneys 

28 from the damaging effects of exposure to Pb through inhibition of lipid peroxidation and 

29 stimulation of endogenous antioxidative defense systems (El-Sokkary et al., 2005).

30 Vitamin C supplementation (20 mg/kg pretreatment every other day) protected the renal 

31 architecture and histology (El-Neweshy and El-Sayed, 2011).

32 Wang et al. (2009c) examined the effect of Pb-acetate (0.25, 0.5 and 1 μM) on cell death 

33 in cultured rat primary PT cells. A progressive loss in cell viability, due to both apoptosis 

34 and necrosis, was observed in cells exposed to Pb. Apoptosis predominated and could be 

35 ameliorated with concomitant N-acetylcysteine exposure, whereas necrosis was 

36 unaffected. Elevation of ROS levels and intercellular calcium, depletion of mitochondrial 

37 membrane potential, and intracellular glutathione levels was observed during Pb 
February 2012 5-306 Draft – Do Not Cite or Quote

1 exposure. Pb-induced apoptosis was demonstrated morphologically (Hoechst 33258 

2 staining) with condensed/fragmented chromatin and apoptotic body formation. CAT and 

3 SOD activities were significantly elevated, reflecting the response to accumulation of 

4 ROS.

5 Table 5-24 presents the acute and chronic renal effects of Pb exposure observed in recent 

6 and past animal toxicology studies.

Table 5-24 Effects of Pb on the kidney/renal system related to exposure 

duration– evidence from animal toxicology studies

Effects with less than 3 months of exposure Effects with 6 or 12 months of exposure

Mitochondrial dysfunction

Renal cell apoptosis 

Nuclear Inclusion Body Formation

Proximal Tubule Cytomegaly

Glomerular Hypertrophy

Increased GFR

Mitochondrial dysfunction

Renal cell apoptosis

Oxidant redox imbalance

Altered NO homeostasis

ATPase dysfunction

Aminoaciduria

Increased electrolyte excretion 

Elevated blood pressure

Decreased GFR

5.5.2.2 Epidemiology in Adults

7 A number of advances in research on the impact of Pb on the kidney in the 20 years 

8 following the 1986 Pb AQCD (U.S. EPA, 1986a) were noted in the 2006 Pb AQCD (U.S. 

9 EPA, 2006b). These included research in general and CKD patient populations at much 

10 lower blood Pb levels (5-10 μg/dL) at the time of evaluation than were previously 

11 studied. These advances contributed to the understanding of the effects of Pb exposure on 

12 kidney dysfunction overall in the population. Pb, at much lower doses than those causing 

13 chronic Pb nephropathy, may act as a cofactor with other more established kidney risks to 

14 increase the risk for CKD and disease progression in susceptible patients. Maric and Hall

15 (2011) note that data from basic and clinical studies suggest that obesity, hypertension, 

16 hyperglycemia, hyperlipedemia, and other elements of the metabolic syndrome are highly 

17 interrelated and contribute to the development and progression of diabetic nephropathy 

18 and thus represent populations potentially at increased risk for kidney dysfunction.

19 In the 2006 Pb AQCD (U.S. EPA, 2006b), several key issues could not be completely 

20 resolved based on the Pb-kidney literature published to date. These included 

21 characterizing the lowest Pb dose at which altered kidney function effects occur, the 
February 2012 5-307 Draft – Do Not Cite or Quote

1 impact of higher past exposures on associations with current Pb biomarker levels, the 

2 impacts of Pb on the kidney in children, the use of paradoxical Pb-kidney associations on 

3 risk assessment in the occupational setting, and the impact of co-exposure to other 

4 environmental nephrotoxicants, such as cadmium. In the intervening five years, relevant 

5 data addressing several of these challenges have been published. 

General Population Studies

6 The 2006 Pb AQCD reported studies that examined associations between indicators of Pb 

7 exposure and kidney function in general populations. This was a new approach to Pb8 kidney research in the two decade time period covered by the 2006 Pb AQCD. As 

9 illustrated in Figure 5-39 and Table 5-25, studies consistently demonstrate associations 

10 between higher blood Pb level and lower renal function in adults. The studies in this 

11 category provided critical evidence that the effects of Pb on the kidney occur at much 

12 lower doses than previously appreciated based on occupational exposure data. However, 

13 because blood Pb level in nonoccupationally-exposed adults reflects both recent and past 

14 Pb exposures, the magnitude, timing, frequency, and duration of Pb exposure 

15 contributing to the observed associations was uncertain. The evidence of Pb-associated 

16 renal effects in general population studies was substantiated by results that were adjusted 

17 for multiple potential confounding factors including age, race, sex, education, household 

18 income, smoking, alcohol use, cadmium exposure, and various health indicators such as 

19 diabetes, SBP, BMI, and history of cardiovascular disease.

20 The landmark Cadmibel Study was the first large environmental study of this type that 

21 adjusted for multiple kidney risk factors (Staessen et al., 1992). It included 965 men and 

22 1,016 women recruited from cadmium exposed and control areas in Belgium. Mean 

23 concurrent blood Pb was 11.4 μg/dL (range 2.3-72.5) and 7.5 μg/dL (range 1.7-60.3) in 

24 men and women, respectively. After adjustment, log transformed blood Pb was 

25 negatively associated with measured creatinine clearance. A 10-fold increase in blood Pb 

26 was associated with a decrease in creatinine clearance of 10 and 13 mL/min in men and 

27 women, respectively. Blood Pb was also negatively associated with estimated creatinine 

28 clearance. 

29 Multiple analyses assessing the kidney impact of Pb exposure have been conducted in the 

30 NAS population (Tsaih et al., 2004; Wu et al., 2003a; Kim et al., 1996; Payton et al., 

31 1994). Participants in this study were originally recruited in the 1960s in the Greater 

32 Boston area. Inclusion criteria included male sex, age 21 to 80 years, and absence of 

33 chronic medical conditions. Longitudinal data contained in NAS publications remain 

34 essential, particularly in light of the dearth of prospective data on the kidney effects of 

35 Pb. The first of these included 459 men whose blood Pb levels from periodic 
February 2012 5-308 Draft – Do Not Cite or Quote

1 examinations, conducted every 3 to 5 years during 1979-1994, were estimated based on 

2 measurements in stored packed red blood cell samples adjusted for hematocrit level (Kim 

3 et al., 1996). Participants were randomly selected to be representative of the entire NAS 

4 population in terms of age and follow-up. Kidney function was assessed with serum 

5 creatinine. Data from four evaluations were available for the majority of participants. At 

6 baseline, mean (SD) age, blood Pb level, and serum creatinine, at baseline, were 56.9 

7 (8.3) years, 9.9 (6.1) μg/dL, and 1.2 (0.2) mg/dL, respectively. In the longitudinal 

8 analysis, using random-effects modeling with repeated measures, ln-transformed blood 

9 Pb was associated with an increase in serum creatinine from the previous to current 

10 follow-up period in the 428 participants whose highest blood Pb level was ≤ 25 μg/dL 

11 (β= 0.027 mg/dL [95% CI: 0.0, 0.054] per unit increase in ln blood Pb); effect estimates 

12 in the entire group and subsets with different peak blood Pb levels (≤ 10 or 40 μg/dL) 

13 also were positive (and larger for blood Pb levels ≤ 10 μg/dL) but had p-values between 

14 0.07 and 0.13. 

15 This study made two other key contributions. In order to address the question of whether 

16 nephrotoxicity observed at current blood Pb levels is due to higher blood Pb levels from 

17 past exposure, these authors performed a sensitivity analysis in participants whose peak 

18 blood Pb levels, dating back to 1979, were ≤ 10 μg/dL. A statistically significant positive 

19 association between blood Pb and concurrent serum creatinine remained in a cross20 sectional analysis. These authors also addressed reverse causality, which attributes 

21 increased blood Pb levels to lack of kidney excretion rather than as a causative factor for 

22 CKD, by showing in adjusted plots that the association between blood Pb and serum 

23 creatinine occurred over the entire serum creatinine range (0.7-2.1 mg/dL), including the 

24 normal range where reverse causality would not be expected.

25 Cortical and trabecular bone Pb measurements were obtained in addition to whole blood 

26 Pb in evaluations performed in the NAS between 1991 and 1995. Associations between 

27 baseline blood, tibia, and patella Pb and change in serum creatinine over an average of 6 

28 years in 448 men were reported in a subsequent NAS publication (Tsaih et al., 2004). At 

29 baseline 6 and 26% of subjects had diabetes and hypertension, respectively. Mean blood 

30 Pb levels and serum creatinine decreased significantly over the follow-up period in the 

31 group. Baseline blood Pb level was not associated with change in creatinine in all 

32 participants. However, diabetes was observed to be an effect modifier of the relations of 

33 blood and tibia Pb with change in serum creatinine. Per unit increase in ln blood Pb, the 

34 increase in serum creatinine between follow-up periods was substantially stronger in 

35 diabetics (β = 0.076 mg/dL [95% CI: 0.031, 0.121]) compared to non-diabetics (β = 

36 0.006 mg/dL [95% CI: -0.004, 0.016]). A similar relationship was observed for tibia Pb. 

37 An interaction was also observed between tibia Pb and hypertension, although it is 

38 possible that many of the 26 diabetics were also included in the hypertensive group and 
February 2012 5-309 Draft – Do Not Cite or Quote

1 were influential there as well. Reverse causality was addressed in a sensitivity analysis of 

2 participants whose serum creatinine was <1.5 mg/dL; the authors reported that 

3 longitudinal associations did not materially change.
February 2012 5-310 Draft – Do Not Cite or Quote

a

Blood Pb data are presented as median and (IQR) in μg/dL for blood Pb. For uniform presentation, median and IQR were estimated 

from the given distributional statistics by assuming normal distributions.

b

The cross product of logged blood Pb and ranked urine Hg was included in the regression to model the interaction between these 

two variates. The significant hyperfilrative effect to these children could be due to a biphasic time course sometimes seen in early 

exposure.

Note: Results are presented first for kidney function tests where an increase is considered impaired function (black circles) then for 

tests where a decrease is considered impaired function (blue circles, outlined in box). Within a category, results are presented first 

for longitudinal analyses followed by cross-sectional analyses. To compare results for linear and nonlinear modeling, effect 

estimates were standardized to a 1 μg/dL increase in blood Pb level within the 10th-90th percentile interval. Magnitudes of the effect 

should not be compared among different kidney metrics.

Figure 5-39 Kidney metric slopes for blood Pb or bone Pb.
February 2012 5-311 Draft – Do Not Cite or Quote

1 The impact of Pb on the kidney has been examined in multiple NHANES datasets 

2 obtained over the last few decades (Figure 5-40 and Table 5-25). NHANES data analyses 

3 benefit from a number of strengths including large sample size, ability to adjust for 

4 numerous Pb risk factors, and the fact that the study population is representative of the 

5 U.S. non-institutionalized, civilian population. The results, covering different time 

6 frames, have been consistent in providing support for Pb as a CKD risk factor, including 

7 NHANES III, conducted from 1988-1994, in which adults with hypertension and diabetes 

8 were observed to be susceptible populations (Muntner et al., 2003) and NHANES 1999-

9 2002 (Muntner et al., 2005). However, because the various NHANES analyses were 

10 cross-sectional in design, examining associations between concurrent measures of kidney 

11 function and blood Pb levels, a common limitation is the uncertainty regarding the 

12 magnitude, timing, frequency, and duration of Pb exposure that contributed to the 

13 observed associations.

14 A recent publication examined NHANES data collected from 1999 through 2006 (Navas15 Acien et al., 2009). The geometric mean concurrent blood Pb level was 1.58 μg/dL in 

16 14,778 adults aged ≥ 20 years. After adjustment for survey year, sociodemographic 

17 factors, CKD risk factors, and blood cadmium, the odds ratios for albuminuria (≥ 

30 mg/g creatinine), reduced eGFR (<60 mL/min/1.73 m2 18 ), and both albuminuria and 

19 reduced eGFR were 1.19 (95% CI: 0.96, 1.47), 1.56 (95% CI: 1.17, 2.08), and 2.39 (95% 

20 CI: 1.31, 4.37), respectively, comparing the highest (> 2.4 μg/dL) to the lowest (≤ 1.1 

21 μg/dL) blood Pb quartiles. Thus, in the subset of the population with the most severe 

22 kidney disease (both reduced eGFR and albuminuria), the magnitude of association with 

23 concurrent blood Pb was greater. When blood cadmium was included as a covariate, 

24 blood Pb remained significantly associated. In fact, the most important contribution of 

25 this recent NHANES analysis was the evaluation of joint Pb and cadmium exposure 

26 (discussed in Section 5.5.4.1).

27 An important contribution of all NHANES publications is that they provide evidence that 

blood Pb remains associated with reduced kidney function (<60 mL/min/1.73 m2 28 as 

29 estimated with the MDRD equation cross-sectionally) despite steadily declining blood Pb 

30 levels during the time periods covered. Additional studies in this category have also 

31 reported worse kidney function related to blood Pb levels (Lai et al., 2008a; Hernandez32 Serrato et al., 2006; Goswami et al., 2005).
February 2012 5-312 Draft – Do Not Cite or Quote

Note: These articles reported ORs of kidney function measures by grouping the population into quartiles of blood Pb and then 

comparing each group to the quartile with the lowest blood Pb (reference group). The blood Pb distribution of the examined group is 

shaded black and the reference group is shaded gray. To express these odds ratios in terms of blood Pb concentration, a log 

normal distribution was fit to the statistics presented and then the medians of each group were determined. The adjusted OR was 

the exponentiated quantity (log(OR) divided by the difference in the medians of the groups compared). The resulting odds ratio is 

presented in terms of percent change=100*(OR-1). 

Figure 5-40 Percent change for kidney outcomes associated with blood Pb.

 

 

 

 

 

5

10

2 

-40 -10 10 50

 

Study Quartiles of Blood Pb Distribution Used

Muntner et al. (2003)

Elevated Serum Creatinine

Hypertensive

Normotensive

Navas-Acien et al. (2009)

Albuminuria >30 mg/g creatinine

eGFR <60 mL/min/1.73m2

μg/dL Blood Pb % Change per μg/dL Blood Pb
February 2012 5-313 Draft – Do Not Cite or Quote

Table 5-25 Additional characteristics and quantitative data for associations of 

blood and bone Pb with kidney outcomes for results presented in 

Figure 5-39 and Figure 5-40

Reference Population

Study 

Location; 

Time 

Period

N Pb Biomarker 

Data Outcome Statistical Analysis Effect Estimate (95% CI)

FIGURE 5-39: Positive Effect Estimates Indicate Poorer Function

Per 1 μg/dL increase in blood 

Pb within the 10th-90th 

percentile interval

Longitudinal Results

Kim et al.

(1996)

Adult males Boston, MA; 

Multiple 

examination

s 1979-

1994

459 Median baseline 

blood = 8.6 μg/dL

10th-90th 

percentile: 4.0-17.5

Change in 

serum creatinine 

between visits x 

10 (mg/dL)

Random-effects modeling 

adjusted for baseline age, 

time since initial visit, BMI, 

smoking status, alcohol 

ingestion, education level, 

hypertension, baseline 

serum creatinine, and time 

between visits

Peak blood Pb ≤ 40 μg/dL: 

0.012 (-0.0001, 0.025)

Peak blood Pb ≤ 25 μg/dL: 

0.015 (0.0002, 0.03)

Peak blood Pb ≤ 10 μg/dL: 

0.021 (-0.005, 0.048)

Tsaih et al.

(2004)

Adult males Boston, MA; 

8/1991-

1995 with 

mean 

6 year 

follow-up

448 Mean (SD) 

Baseline

Blood Pb = 6.5 (4.2) 

μg/dL

10th-90th 

percentile: 2.1-7.6

Tibia Pb = 21.5 

(13.5) μg/g

Patella Pb = 32.4 

(20.5) μg/g

Change in 

serum creatinine 

per year 

x 10 (mg/dL)

Log linear regression 

adjusted for age, age 

squared, BMI, 

hypertension, diabetes, 

smoking status, alcohol 

consumption, analgesic 

use, baseline serum 

creatinine, serum creatinine 

squared

With diabetes: 0.18 (0.07, 0.29)

Without diabetes: 0.014 (-0.009, 

0.037)

With hypertension: 0.019 (-

0.027, 0.065)

Without hypertension: 0.021 

(-0.007, 0.049)

Per unit increase in lntransformed tibia Pb

With diabetes: 0.082 (0.03, 

0.14)

Without diabetes: 0.005 (-0.01, 

0.02)

With hypertension: 0.023 

(0.003, 0.04)

Without hypertension: 0.0004 

(-0.01, 0.01)

Cross-Sectional Results

Kim et al.

(1996)

Adult males Boston, MA; 

Multiple 

examination

s 1979-

1994

459 Median baseline 

blood = 8.6 μg/dL

10th-90th 

percentile: 4.0-17.5

Serum 

creatinine 

(mg/dL)

Random-effects modeling 

adjusted for baseline age, 

time since initial visit, BMI, 

smoking status, alcohol 

ingestion, education level, 

hypertension.

Peak blood Pb ≤ 40 μg/dL: 

0.0017 (0.0005, 0.003)

Peak blood Pb ≤ 25 μg/dL: 

0.0021 (0.0007, 0.0035)

Peak blood Pb ≤ 10 μg/dL: 

0.0033 (0.0012, 0.0053)
February 2012 5-314 Draft – Do Not Cite or Quote

Reference Population

Study 

Location; 

Time 

Period

N Pb Biomarker 

Data Outcome Statistical Analysis Effect Estimate (95% CI)

Tsaih et al.

(2004)

Adult males Boston, MA; 

8/1991-

1995 with 

mean 6 yr 

follow-up

448 Mean (SD) baseline

Blood Pb = 

6.5 (4.2) μg/dL

10th-90th 

percentile: 2.6-11.5

Repeated 

measures 10th-90th 

percentile: 2.1-7.6

Tibia Pb = 21.5 

(13.5) μg/g

Patella Pb = 32.4 

(20.5) μg/g

Serum 

creatinine 

(mg/dL)

Log linear regression 

adjusted for age, age 

squared, BMI, 

hypertension, diabetes, 

smoking status, alcohol 

consumption, analgesic use

Baseline blood Pb

With diabetes: -0.009 (-0.038, 

0.020) 

Without diabetes: -0.004 

(-0.010, 0.003)

With hypertension: 0 (-0.013, 

0.013)

Without hypertension: -0.005

(-0.011, 0.002)

Follow-up blood Pb

With diabetes: 0.053 (-0.032, 

0.138)

Without diabetes: 0.034 (0.007, 

0.061)

With hypertension: 0.083 

(0.038, 0.128)

Without hypertension: 0.014 (-

0.016, 0.044)

De Burbure et 

al. (2006)

Children, mean 

age = 10 years, 

age range = 

8.5-12.3 years

France, 

Czech 

Republic, 

and Poland; 

dates not 

provided

804 Concurrent Blood 

Pb Median (IQR) = 

3.9 (2.6, 5.7) μg/dL

10th-90th 

percentile: 1.8-8.1

Log-transformed 

serum 

creatinine, 

cystatin C, and 

β2-microglobulin

Log linear regression 

adjusted for cadmium, 

urinary creatinine, urinary 

mercury

Log serum creatinine (mg/L): 

-0.062 (-0.106, -0.017)a

Log Cystatin C: -1.3 (-2.4, 

-0.21)a

Log β2-microglobulin: -2.2 

(-4.0, -0.54)a

FIGURE 5-39: Negative Effect Estimates Indicate Poorer Function 

Per 1 μg/dL increase in blood 

Pb within the 10th-90th 

percentile interval

Longitudinal Results:

Yu et al. (2004) Adult CKD 

patients

Taipei, 

Taiwan; 48 

month 

longitudinal 

study period

121 Mean (SD) 

Baseline blood = 

4.2 (2.2) μg/dL

10th-90th 

percentile: 2.0-5.1

Change in MDRD 

eGFR over 4 

yr/100 

(mL/min/1.73 m2 

body surface 

area)

Generalized estimating 

equations adjusted for 

age, sex, BMI, 

hyperlipidemia, 

hypertension, smoking, 

use of ACE inhibitor, 

baseline serum creatinine, 

daily protein excretion, 

daily protein intake, 

underlying kidney disease

-0.040 (-0.072, -0.008)a

Cross-Sectional Results: 

Akesson et al.

(2005)

WHILA, 

adult women

Sweden; 

6/1999-

1/2000

820 Median (5-95%) 

concurrent blood = 

2.2 (1.1, 4.6) μg/dL

10th-90th 

percentile: 1.3-3.8

Creatinine 

clearance/100 

(mL/min) 

Linear regression 

adjusted for age, BMI, 

diabetes, hypertension, 

regular use of nephrotoxic 

drug, smoking status

-0.018 (-0.03, -0.006)

Cystatin C-based 

eGFR (Larsson et 

al. 2004)/100 

(mL/min)

-0.02 (-0.03, 0.007)
February 2012 5-315 Draft – Do Not Cite or Quote

Reference Population

Study 

Location; 

Time 

Period

N Pb Biomarker 

Data Outcome Statistical Analysis Effect Estimate (95% CI)

Staessen et al.

(1992)

Adults Belgium; 

1985-1989

1,981 Concurrent Blood 

Pb Mean (SD)

Males: 11.4 μg/dL

Females: 7.5 μg/dL

10th-90th 

percentile: 3.7-15.1

Creatinine 

clearance/100 

(mL/min)

Log linear regression 

adjusted for age, age 

squared, sex, BMI, BP, 

ferritin level, smoking 

status, alcohol ingestion, 

rural/urban residence, 

analgesic and diuretic 

use, blood and urinary 

cadmium, diabetes, 

occupational exposure to 

heavy metals, and gamma 

glutamyl transpeptidase

Females: -0.067 

(-0.108, -0.027)a

Males: -0.051 (-0.097, -0.047)a

Payton et al.

(1994)

Adult males Boston, MA; 

1988-1991

744 Mean (SD) 

concurrent blood = 

8.1 (3.9) μg/dL

10th-90th 

percentile: 4.1-12.9

Log-transformed 

creatinine 

clearance 

(mL/min)

Log linear regression 

adjusted for age, BMI, 

analgesic and diuretic 

use, alcohol consumption, 

smoking status, SBP, DBP

-0.040 (-0.079, -0.0015)

Fadrowski et al.

(2010)

NHANES, 

adolescents

U.S.; 

1988 1994

769 Median concurrent 

blood = 1.5 μg/dL

10th-90th 

percentile: 0.4-5.4

Q1: <1.0

Q2: 1.0 to 1.5

Q3: 1.6 to 2.9

Q4: >2.9

Cystatin C-based 

eGFR/100

(mL/min/1.73 m2

; 

calculated using 

the Filler and 

Lepage equation)

Log linear regression 

adjusted for age, sex, 

race/ethnicity, urban/rural 

residence, smoking, 

obesity, household 

income, education level of 

family reference person, 

BP, lipid levels, glucose

levels

-0.022 (-0.038, -0.0054)

Q1: Referent

Q2: -1.4 (-7.4, 4.5)

Q3: -2.6 (-7.3, 2.2)

Q4: -6.6 (-12.6, -0.07)

FIGURE 5-40: Analysis of Blood Pb Quartiles: %change in kidney 

outcome

Muntner et al.

(2003)

NHANES III, 

adults 

U.S.; 

1988-1994

4813 Mean (SD) 

concurrent blood 

Pb

With Hypertension: 

4.2 (0.14) μg/dL

Q1: 0.7 to 2.4

Q2: 2.5 to 3.8

Q3: 3.9 to 5.9

Q4: 6.0 to 56.0 

Without 

Hypertension: 3.3 

(0.10) μg/dL

Q1: 0.7 to 1.6

Q2: 1.7 to 2.8

Q3: 2.9 to 4.6

Q4: 4.7 to 52.9

Elevated Serum 

Creatinine 

(99th percentile of 

each race-sex 

specific 

distribution for 

healthy young 

adults)

CKD

Logistic regression 

adjusted for age, race, 

sex, diabetes, SBP, 

smoking, history of CVD, 

BMI, alcohol consumption, 

household income, 

education level, marital 

status, health insurance

Q1: Referent

With hypertension

Q2: 47% (3, 110)

Q3: 80% (34, 142)

Q4: 141% (46, 297)

Without hypertension

Q2: 11% (-44, 121)

Q3: 19% (-38, 125)

Q4: 9% (-47, 122)

With hypertension

Q2: 44% (0, 109)

Q3: 85% (32, 159)

Q4: 160% (52, 345)

Without hypertension

Q2: -10% (-63, 116)

Q3: 0% (-55, 122)

Q4: 9% (-59, 189)

Navas-Acien et 

al. (2009)

NHANES III, 

adults

U.S.; 

1999-2006

14,778 Geometric 

concurrent blood 

mean = 1.58 μg/dL

Q1: ≤ 1.1

Q2: 1.2 to 1.6

Q3: 1.7 to 2.4

Q4: >2.4

eGFR <60 

mL/minute/1.73 

m2

Logistic regression 

adjusted for survey year, 

age, sex, race/ethnicity, 

BMI, education, smoking, 

cotinine, alcohol intake, 

hypertension, diabetes, 

menopausal status

Q1: Referent

Q2: 10% (-20, 51)

Q3: 36% (-1, 85)

Q4: 56% (17, 108)

Albuminuria and 

eGFR <60 

mL/minute/1.73 

m2

Q2: 53% (-15, 177)

Q3: 57% (-17, 198)

Q4: 139% (31, 337)

a

95% CI estimated from given p-value.
February 2012 5-316 Draft – Do Not Cite or Quote

Patient Population Studies

1 CKD as defined by the National Kidney Foundation - Kidney Disease Outcomes Quality 

2 Initiative (NKF-K/DOQI) workgroup (National Kidney Foundation, 2002) is the presence 

of markers of kidney damage or GFR <60 mL/min/1.73 m2 3 for ≥ 3 months. The MDRD 

4 equation is the most common one used in the eGFR determination for this definition. 

5 Notably, decreased GFR is not required for the first criterion and markers of kidney 

6 damage are not required for the second criterion.

7 Several key studies in CKD patients provide prospective data to evaluate the impact of 

8 baseline blood Pb level on CKD progression (kidney function decline) in patient 

9 populations (Table 5-26). Yu et al. (2004), discussed in the 2006 Pb AQCD, followed 

10 121 patients over a four year period. Eligibility required well-controlled CKD with serum 

11 creatinine between 1.5 and 3.9 mg/dL. Importantly, EDTA-chelatable Pb <600 μg/72 h, a 

12 level below that traditionally thought to indicate risk for Pb-related nephrotoxicity, was 

13 required at baseline. Patients with potentially unstable kidney disease were excluded 

14 (i.e., due to systemic diseases such as diabetes). Mean blood Pb and EDTA-chelatable Pb 

15 levels were 4.2 μg/dL and 99.1 μg/72 hours, respectively. In a Cox multivariate 

16 regression analysis, chelatable Pb was significantly associated with overall risk for the 

17 primary endpoint (doubling of serum creatinine over the 4-year study period or need for 

18 hemodialysis). When the group was dichotomized by EDTA chelatable Pb level, Kaplan19 Meier analysis demonstrated that significantly more patients (15/63) in the high-normal 

20 group (EDTA chelatable Pb level ≥ 80 but <600 μg/72 hours) reached the primary end 

21 point than did those in the lower EDTA chelatable Pb levels (<80 μg Pb/72 hours) group 

22 (2/58). Associations between baseline chelatable or blood Pb level and change in serial 

23 measurements of eGFR (estimated by the MDRD equation (Levey et al., 1999) were 

24 modeled separately using generalized estimating equations. Based on these models, a 10 

25 μg higher chelatable Pb level or 1 μg/dL higher blood Pb level reduced the GFR by 1.3 

and 4.0 mL/min/1.73 m2 26 , respectively, during the 4-year study period. Recent studies 

27 expanded the CKD patient populations in which this effect was observed to those with 

28 diabetic nephropathy (Lin et al., 2006a) and with the lowest blood Pb levels studied to 

29 date (Lin et al., 2006b). Results of these observational studies have been summarized

30 (Weaver and Jaar, 2010).
February 2012 5-317 Draft – Do Not Cite or Quote

Table 5-26 Patient population studies: kidney function decline

Study n

Baseline 

mean (SD) 

blood Pb 

(μg/dL)

Baseline mean 

(SD) chelatable 

Pb (μg/72 hours)

Baseline mean 

(SD) eGFR 

(mL/min/1.73 m2)

Years of 

follow-up

Decline in eGFR 

per 1 SD higher 

Pb dose at 

baseline per year

Comments

Lin et al.

(2003)

202 5.3 (2.9) 104.5 (106.3) 41.6 (14.4) 2 0.16 Largest study to date

Yu et al.

(2004)

121 4.2 (2.2) 99.1 (83.4) 36.0 (9.8) 4 2.7 (chelatable) 

2.2 (blood Pb) 

Longest follow-up; 1 μg/dL 

higher blood Pb, at baseline, 

associated with 4.0 

mL/min/1.73 m2 reduction in 

eGFR over 4 years

Lin et al.

(2006a)

87 6.5 (3.4) 108.5 (53.8) 35.1 (9.0) 1 3.87 Type II diabetics with 

nephropathy

Lin et al.

(2006b)

108 2.9 (1.4)a 40.2 (21.2) 

(all <80)

47.6 (9.8) 2 1.1 Lowest Pb exposed CKD 

patients

a

Notably, mean blood Pb level in this study was below that observed in a recent large general population study of 50- to 70-year olds in Baltimore, MD

(Martin et al., 2006).

Source: Reprinted with permission of UpToDate.com, Weaver and Jaar (2010)

1 A recent population-based case-control study examined occupational Pb exposure as a 

2 risk factor for severe CKD (Evans et al., 2010). The study included 926 cases with first 

3 time elevations of serum creatinine >3.4 mg/dL for men and >2.8 mg/dL for women and 

4 998 population-based controls. Occupational Pb exposure was assessed using an expert 

5 rating method based on job histories. Eighty-one cases and 95 controls were judged to 

6 have had past occupational Pb exposure. Of those, 23 cases and 32 controls were thought 

to have been exposed to Pb levels ≥ 30 μg/m3 (the current U.S. OSHA limit is 50 μg/m3 7 ). 

8 In multivariable logistic regression modeling, the OR for CKD (adjusted for age, sex, 

9 smoking, alcohol consumption, diabetes, education, and BMI) was 0.97 (95% CI: 0.68, 

10 1.38) in Pb-exposed compared to non-exposed participants. In analyses comparing low 

11 (≥ 3 to <10% of occupational exposure limit), medium (≥ 10 to < 30% of limit), and high 

12 (≥ 30% of limit) exposure groups, although ORs were elevated (statistically 

13 nonsignificant) in the medium exposure group compared to the never exposed group for 

14 average and lifetime cumulative exposure metrics, a monotonic increase in OR across 

15 exposure groups was not observed. In the low and high exposure groups, odds of CKD 

16 tended to be lower compared with the never exposed group. In addition, the CKD patients 

17 were followed prospectively for a mean of 2.5 years for the 70 Pb-exposed patients and 

18 2.4 years for the 731 patients without past occupational Pb exposure. Mean eGFRs (using 

the MDRD equation) were 16.0 and 16.6 mL/min/1.73 m2 19 in exposed and non-exposed 

20 patients, respectively, indicating severe disease in both groups. Using mixed-effects 

multivariable models, eGFRs declined by 4.27 and 3.39 mL/min/1.73 m2 21 /y in ever and 

most Pb-exposed CKD patients, respectively, compared with 4.55 mL/min/1.73 m2 22 /y in 

23 patients without occupational Pb exposure. Thus, the results overall did not provide 

24 strong evidence that Pb exposure was associated with renal effects.
February 2012 5-318 Draft – Do Not Cite or Quote

1 Strengths noted by the authors included virtually complete case ascertainment and 

2 minimal loss to follow-up. Exposure assessment was listed as both a strength and a 

3 limitation. Expert rating methods are commonly used when biological monitoring is not 

4 an option and in case-control studies where many occupational exposures are considered. 

5 In Pb-kidney research, this approach is uncommon except in the case-control setting. 

6 However, given the challenges of interpreting blood Pb in dialysis patients (discussed 

7 below), this approach may have advantages in this study of such severe CKD. Other case8 control studies examining occupational risk factors for CKD found Pb exposure to be a 

9 risk factor (Nuyts et al., 1995; Steenland et al., 1990). Nuyts et al. (1995) found adults 

10 with history of occupational Pb exposure to have elevated odds of CKD (OR for ever11 versus never-exposed: 2.11 [95% CI: 1.23, 4.36]). The association was weaker in 

12 Steenland et al. (1990) (OR for ever- versus never-exposed: 1.73 [95% CI: 0.82, 3.65]). 

13 Regular moonshine consumption, also a potential source of Pb exposure, was a stronger 

14 risk factor for CKD (OR: 2.42 [95% CI: 1.10, 5.36]).

15 The prospective observational aspect of Evans et al. (2010) is similar in design to the 

16 work of Lin and colleagues but differs in several important respects. In Evans et al.

17 (2010), only occupational Pb exposure was considered whereas the work in Taiwan 

18 excluded occupational exposure and used blood and chelatable Pb measures. In the past 

19 in developed countries, environmental exposures were substantial. For example, mean 

20 tibia Pb levels were 21.5 and 16.7 μg/g bone mineral, in environmentally-exposed 50- to 

21 70-year-old African-Americans and whites, respectively, in Baltimore (Martin et al., 

22 2006). In Korean Pb workers, mean baseline tibia Pb level was only twofold higher (35.0 

23 μg/g) (Weaver et al., 2003a) which illustrates the substantial body burden in middle- and 

24 older-aged Americans from lifetime Pb exposure. Declines in blood Pb levels in Sweden 

25 have been reported and attributed to the leaded gasoline phase-out (Strömberg et al., 

26 1995; Elinder et al., 1986), although blood Pb levels were lower than those noted during 

27 the U.S. phase-out. Finally, the severe degree of CKD among subjects in Evans et al.

28 (2010) creates a survivor bias at enrollment and limits the eGFR decline possible during 

29 follow-up, thus limiting the ability to identify factors that influence that decline. 

ESRD Patient Studies

30 End stage renal disease (ESRD) is a well-established public health concern, and is 

31 characterized by the use of dialysis to perform the normal functions of the kidney. 

32 Incidence and prevalence in the U.S. continue to increase resulting in rates that are the 

33 third highest among nations reporting such data (U.S. Renal Data System, 2009). Studies 

34 in patients with CKD requiring chronic hemodialysis (ESRD) have also been published in 

35 the past five years. A study of 271 adult patients on regular thrice weekly dialysis 
February 2012 5-319 Draft – Do Not Cite or Quote

1 reported much higher blood Pb levels than had been appreciated by the treating clinicians

2 (Davenport et al., 2009). Blood Pb levels ranged from 3 to 36.9 μg/dL; 25.5% had levels 

3 >20 μg/dL, 59% had values of 10-20 μg/dL, and 15.5% were <10 μg/dL. Few details on 

4 the statistical analysis were provided which complicates interpretation of the findings. 

5 However, blood Pb was positively correlated with hemodialysis vintage (months on 

6 dialysis; Spearman r = 0.38, p <0.001); negatively correlated with urine output (r = -0.44, 

7 p <0.001) and higher in patients using single carbon filter and reverse osmosis water 

8 purification devices. Another recent publication reported higher Pb in dialysate than in 

9 the tap water used in its preparation (Chen et al., 2009a). A systematic review of a wide 

10 range of trace elements in hemodialysis patients reported higher Pb levels in patients 

11 compared to controls although the difference was not large (Tonelli et al., 2009). These 

12 data suggest that blood Pb monitoring in dialysis patients may be useful.

13 Interpretation of blood and bone Pb in patients on dialysis is challenging for several 

14 reasons. First, renal osteodystrophy, the bone disease related to kidney disease, may 

15 result in increased release of Pb from bone stores. Thus, interpretation of blood and even 

16 bone Pb levels may require adjustment with one or more of a range of osteoporosis 

17 variables. Secondly, as observed above (Davenport et al., 2009), residual kidney function 

18 may have a substantial impact on blood Pb levels in populations with such minimal 

19 excretion. Third, as illustrated in the studies cited above (Chen et al., 2009a; Davenport et 

20 al., 2009), water and concentrates used in dialysis may be variable sources of Pb. A 

21 recent study reported decreased blood Pb in post-dialysis compared to pre-dialysis 

22 samples (Kazi et al., 2008). Thus, substantial fluctuations in blood Pb are possible while 

23 on dialysis. Finally, anemia is common in CKD and Pb is stored in red blood cells. Thus, 

24 measurement of blood Pb in anemia may require adjustment for hemoglobin; no 

25 standardized approach to this currently exists.

26 Given these caveats, a pilot study observed higher median blood Pb levels in 55 African27 American dialysis patients compared to 53 age- and sex-matched controls (6 and 3 μg/dL 

28 respectively; p <0.001) (Muntner et al., 2007). This study was unique in that tibia Pb 

29 levels were assessed. Median tibia Pb was higher in ESRD patients although the 

30 difference did not reach statistical significance (17 and 13 μg/g bone mineral, 

31 respectively [p = 0.13]). In order to determine the potential impact of renal 

32 osteodystrophy, median blood and tibia Pb levels in the dialysis patients were compared 

33 by levels of serum parathyroid hormone, calcium, phosphorus, and albumin and were not 

34 found to be significantly different (Ghosh-Narang et al., 2007). A study of 211 diabetic 

35 patients on hemodialysis (Lin et al., 2008) found parathyroid hormone and serum 

36 creatinine to be associated with blood Pb level in crude but not adjusted associations. In 

37 contrast, a study of 315 patients on chronic peritoneal dialysis observed parathyroid 

38 hormone to be positively correlated and residual renal function to be negatively 
February 2012 5-320 Draft – Do Not Cite or Quote

1 correlated with logarithmic-transformed blood Pb levels after adjustment (Lin et al., 

2 2010). In the prospective portion of this study, blood Pb levels at baseline were 

3 categorized by tertile (range of 0.1 to 29.9 μg/dL with cut points of 5.62 and 8.66 μg/dL). 

4 Cox multivariate analysis, after adjustment for parathyroid hormone level, residual renal 

5 function, and 20 other variables, showed increased all-cause mortality in the middle 

6 (5.62-8.66 μg/dL) and highest (> 8.66 μg/dL) compared to the lowest (< 5.62 μg/dL) 

7 tertiles after 18 months of follow-up (hazard ratio= 2.1 [95% CI: 2.0, 2.2] and 3.3 [95% 

8 CI: 1.3, 13.5], respectively). A recent publication of an 18-month follow-up of 927 

9 patients on maintenance hemodialysis also reported increased hazard ratios for all-cause 

10 (4.7 [95% CI: 1.9, 11.5]), cardiovascular-cause (9.7 [95% CI: 2.1, 23.3]), and infection11 cause (5.4 [95% CI: 1.4, 20.8]) 18-month mortality in the highest (> 12.64 μg/dL) 

12 compared to the lowest tertile (< 8.51 μg/dL) of baseline blood Pb level, after adjustment 

13 for sex, urban residence, hemodialysis vintage, hemoglobin, serum albumin, and ferritin

14 (Lin et al., 2011). Given other recent publications in hemodialysis patients by this group, 

15 it would be valuable to examine these risks after adjustment for hemoglobin A1C (Lin16 Tan et al., 2007a), and blood cadmium (Yen et al., 2011; Hsu et al., 2009a).

Clinical Trials in Chronic Kidney Disease Patients

17 Randomized chelation trials in CKD patients, uncommon in nephrotoxicant research, 

18 provide unique information on the kidney impact of Pb. These studies have been 

19 performed by Lin and colleagues in Taiwan and involve similar study designs. Initially, 

20 patients are observed in order to compare CKD progression prior to chelation. Then, 

21 CKD patients whose diagnostic EDTA chelatable Pb levels are within certain ranges 

22 (generally 60-600 μg/72 hours and thus below the level commonly considered for 

23 chelation) are randomized. The treated group receives weekly chelation with 1 g EDTA 

24 intravenously for up to 3 months. The control group receives placebo infusions. In the 

25 follow-up period, chelation is repeated for defined indications such as increased serum 

26 creatinine or chelatable Pb levels above specified cut-offs. Placebo infusions are repeated 

27 in the controls as well. The results of the most recent of these trials are summarized in 

28 Table 5-27 below.
February 2012 5-321 Draft – Do Not Cite or Quote

Table 5-27 Clinical randomized chelation trials in chronic kidney disease 

patients

Reference Group n

Baseline 

mean 

(SD) 

blood Pb 

(μg/dL)

Baseline mean (SD) 

chelatable Pb (μg/72 

hr)

Baseline mean (SD) 

eGFR (mL/min/1.73 

m2)

Months of 

treatment/ 

follow-up

Change in 

eGFR per yr

(mL/min/1.73 

m2)

Comments

Lin et al.

(2003)

Chelated 32 6.1 (2.5) 150.9 (62.4) 32.0 (12.1) 27 + 1.07

Control 32 5.9 (3.0) 144.5 (87.9) 31.5 (9.0) - 2.7

Lin et al.

(2006a)

Chelated 15 7.5 (4.6) 148.0 (88.6) 22.4 (4.4) 15 -3.5 Subjects with 

Type II 

diabetes and 

nephropathy

Control 15 5.9 (2.2) 131.4 (77.4) 26.3 (6.2) -10.6

Lin et al.

(2006b)

Chelated 16 2.6 (1.0)a 43.1 (13.7) 41.2 (11.2) 27 +3.0 Lowest Pb 

exposed and 

treated range 

Body Lead 

Burden (72 h 

urinary Pb 

excretion) ≥ 20- <80 μg

Control 16 3.0 (1.1) 47.1 (15.8) 42.6 (9.7) -2.0

Lin-Tan et al.

(2007b)

Chelated 58 5.0 (2.2) 164.1 (111.1) 36.8 (12.7) 51 -0.3 Subjects 

without 

diabetes

Control 58 5.1 (2.6) 151.5 (92.6) 36.0 (11.2) -2.9

a

Notably, mean blood Pb level in this study was below that observed in a recent large general population study of 50- to 70-year olds in Baltimore, MD

(Martin et al., 2006).

1 This study design requires replication in larger populations at multiple clinical centers. If 

2 confirmed, the effect may be due to removal of Pb. However, chelation may also have a 

3 direct beneficial effect on kidney function, regardless of Pb exposure. Antioxidant effects 

4 of CaNa2EDTA which may improve kidney function directly via improved blood flow to 

5 the kidneys have been reported (Saxena and Flora, 2004; Jacobsen et al., 2001). EDTA 

6 benefits in a Pb rodent model appeared to occur via reduced oxidation (Saxena and Flora, 

7 2004). EDTA administration reduced kidney damage in a rat model of acute renal failure 

8 induced by ischemia (Foglieni et al., 2006). Similarly DMSA has been reported to 

9 prevent renal damage when co-administered during induction of nephrosclerosis in a 

10 nonPb-exposed rat model (Gonick et al., 1996). Benefits from chelation reported in 

11 rodent models of Pb-related nephrotoxicity (Sanchez-Fructuoso et al., 2002a; Sanchez12 Fructuoso et al., 2002b; Khalil-Manesh et al., 1992b) did not appear to occur via reversal 

13 of structural damage (Khalil-Manesh et al., 1992b); again suggesting that improved 

14 hemodynamics from reduction of reactive oxidant species, which could be due to reduced 

15 Pb and/or directly to the chelating agent, may be a mechanism (Gonick et al., 1996).

16 However, the most parsimonious explanation for the combination of the observational 
February 2012 5-322 Draft – Do Not Cite or Quote

1 and experimental chelation work of Lin and colleagues is that reduced Pb is the 

2 underlying reason. 

3 The unique body of work in patient populations by Lin and co-workers, both 

4 observational and experimental, has numerous strengths including prospective study 

5 design, randomization, Pb assessment that includes estimates of the bioavailable dose, 

6 longitudinal statistical analysis, and control for multiple kidney risk factors. However, the 

7 generalizability of the results to broader populations is unknown. In addition, the 

8 association observed between Pb dose and decline in GFR has been variable; the annual 

9 decline in eGFR per standard deviation (SD) higher Pb dose at baseline was much lower 

10 in the 2003 study than in subsequent publications (Table 5-27 above). Small sample sizes 

11 and differences in renal diagnoses between groups may be factors in this variability. 

12 Additional research in large populations at multiple centers with assessment of 

13 neuropsychological as well as kidney outcomes is needed. 

Occupational Studies

14 The vast majority of studies in the literature on the impact of Pb on the kidney have been 

15 conducted in the occupational setting. In general, study size and extent of statistical 

16 analysis are much more limited than those in general population studies. Publications in 

17 few populations have reported adjusted results in occupationally exposed workers in the 

18 five years since the 2006 Pb AQCD. In a two-year prospective cohort study, generalized 

19 estimating equations were used to model change in kidney function between each 

20 evaluation in relation to tibia Pb and concurrent change in blood Pb in 537 current and 

21 former Pb workers (Weaver et al., 2009). Tibia Pb was evaluated at the beginning of each 

22 follow-up period (yearly on average) and Pb biomarker levels were adjusted for baseline 

23 levels and other covariates. In males, serum creatinine decreased and calculated 

24 creatinine clearance increased over the course of the study; these changes were largest in 

25 participants whose blood Pb declined concurrently or whose tibia Pb was lower at the 

26 beginning of the follow-up interval. In females, decreasing serum creatinine was 

27 associated with declining blood Pb (as in males); however, increasing blood Pb was 

28 associated with a concurrent increase in serum creatinine. Women (25.9% of the study 

29 population) were older and more likely to be former Pb workers than were men which 

30 may have been important factors in the effect modification observed by sex. 

31 Chia and colleagues observed a significant, positive association between concurrent 

32 blood Pb and urine NAG in linear regression models after adjustment for age, sex, race, 

33 exposure duration, ALAD G177C polymorphism and the interaction between ALAD 

34 genotype and blood Pb (Chia et al., 2006). Similar positive associations were observed 

35 between blood Pb and a wider range of EBE markers in models that adjusted for age, sex, 
February 2012 5-323 Draft – Do Not Cite or Quote

1 race, exposure duration, and the HpyCH4 ALAD polymorphism (Chia et al., 2005).

2 Other studies published in the last 5 years also focused on ALAD polymorphisms but did 

3 not find effect modification to be in a consistent direction (Gao et al., 2010a; Wang et al., 

4 2009a; Weaver et al., 2006; Weaver et al., 2005b). In adults with the ALAD2 genotype, 

5 Pb has been associated with better and poorer renal function in separate cohorts of Pb 

6 workers. 

7 A study of 155 male workers reported significant, positive correlations between blood 

8 and urine Pb and urine NAG and albumin after controlling for age and job duration (Sun 

9 et al., 2008a). An important additional study that analyzed occupational Pb exposure is 

10 discussed below under patient population studies (Evans et al., 2010).

11 A few studies of occupationally-exposed adults have performed benchmark dose 

12 calculations for the effect of Pb on the kidney. Both used only EBE markers and found 

13 NAG to be the most sensitive outcome; reported lower confidence limits on the 

14 benchmark doses were 10.1 μg/dL (Sun et al., 2008a), and 25.3 μg/dL (Lin and Tai-yi, 

15 2007).

16 A number of other publications in the five years since the 2006 Pb AQCD have reported 

17 significantly worse kidney outcomes in unadjusted analyses in occupationally-exposed 

18 adults compared to unexposed controls (Onuegbu et al., 2011; Patil et al., 2007) and/or 

19 significant correlations between higher levels of Pb biomarkers and worse kidney 

20 function (Alasia et al., 2010; Khan et al., 2008; Sun et al., 2008a; Garcon et al., 2007; Lin 

21 and Tai-yi, 2007; Alinovi et al., 2005). One small study found no significant differences

22 (Orisakwe et al., 2007). In a study of 108 Pb workers with mean blood Pb level of 

23 36.2 μg/dL, no significant correlations were observed between blood Pb concentration 

24 and GFR, creatinine clearance, uric acid clearance or uric acid excretion fraction

25 (Karimooy et al., 2010). However, interpretation of this study is limited by the fact that 

26 "only 30 subjects had a correct 24 hours urine volume" and no methods are described for 

27 kidney outcome measurement or analysis.

28 Overall, the occupational literature published in the last five years on the kidney impact 

29 of Pb exposure has been more consistent in reporting statistically significant associations 

30 than were data reviewed for the 2006 Pb AQCD. This may reflect increased reliance on 

31 EBE markers as more sensitive outcome measures, publication bias, or multiple 

32 comparisons due to a greater number of outcomes assessed. 

33 A small number of publications that include concentration-response information provides 

34 evidence of Pb-related nephrotoxicity in the occupational setting across the blood Pb 

35 ranges analyzed (Weaver et al., 2003a; Ehrlich et al., 1998). Data in 267 Korean Pb 

36 workers in the oldest age tertile (mean age = 52 years) did not provide evidence of a 
February 2012 5-324 Draft – Do Not Cite or Quote

1 threshold for a Pb effect on serum creatinine levels (added variable plot shown in Figure 

2 5-41) (Weaver et al., 2003a). It is important to note the uncertainty regarding whether the 

3 concentration-response information provided in these studies applies to lower blood Pb 

4 levels or to populations with lower current environmental Pb exposures.

Source: Reprinted with permission of the BMJ Publishing Group, Weaver et al. (2003a)

Note: Both the adjusted regression line (straight line) and the line estimated by the smoothing method of the S-PLUS statistical 

software function lowess (line with curves) are displayed. Both have been adjusted for covariates. For ease of interpretation, axes 

have been scaled, so that the plotted residuals are centered on the means, rather than zero.

Figure 5-41 Added variable plot of association between serum creatinine and 

blood Pb in 267 Korean Pb workers in the oldest age tertile.

5 A major challenge in interpretation of the occupational literature is the potential for Pb6 related hyperfiltration. Hyperfiltration involves an initial increase in glomerular 

7 hypertension which results in increased GFR. If persistent, the risk for subsequent CKD 

8 increases. This pattern has been observed in diabetes, hypertension, and obesity (Nenov 

9 et al., 2000). As discussed in the 2006 Pb AQCD, findings consistent with hyperfiltration 

10 have been observed in occupational populations (Weaver et al., 2003a; Hsiao et al., 2001; 

11 Roels et al., 1994), a study of adults who were Pb poisoned as children (Hu, 1991), and a 

12 study in European children (De Burbure et al., 2006). Longitudinal data in Pb-exposed 

13 rodents provide evidence of a hyperfiltration pattern of increased, followed by decreased 

14 GFR, associated with Pb exposure and are critical in interpretation of the human Pb15 kidney literature (Khalil-Manesh et al., 1992a). Pb could induce glomerular hypertension 

16 resulting in hyperfiltration by several mechanisms including increased ROS, changes in 

Adjusted blood Pb level (μg/dL)

Adjusted serum creatinine (mg/dL)
February 2012 5-325 Draft – Do Not Cite or Quote

1 eicosanoid levels, and/or an impact on the renin-angiotensin system (Vaziri, 2008b; Roels 

2 et al., 1994). Whether hyperfiltration contributes to pathology in humans is unclear; 

3 longitudinal studies are needed. 

4 Regardless, significant findings could be obscured if opposite direction associations are 

5 present in different segments of the study population and interaction models are not 

6 performed to address this. In the Korean Pb workers (Weaver et al., 2003a; Weaver et al., 

7 2003b), significant associations in opposite directions were observed only when relevant 

8 effect modifiers such as age or genetic variants in ALAD, VDR, and NOS were included 

9 in the model. This is a valid concern for risk assessment, since the factors involved in 

10 these inverse associations in Pb-exposed workers are not well defined at present.

5.5.2.3 Epidemiology in Children

Lead Nephrotoxicity in Children

11 Both the 2006 and 1986 Pb AQCDs noted that the degree of kidney pathology observed 

12 in adult survivors of untreated childhood Pb poisoning in the Queensland, Australia 

13 epidemic (Inglis et al., 1978) has not been observed in other studies of childhood Pb 

14 poisoning. Recent publications remain consistent with that conclusion; a recent study 

15 observed an impact of childhood Pb poisoning on IQ but not kidney outcomes (Coria et 

16 al., 2009). Chelation was raised as a potential explanation for this discrepancy in the 2006 

17 Pb AQCD. 

18 With declining Pb exposure levels, recent work has focused on studies in children with 

19 much lower blood Pb levels. However, insensitivity of the clinical kidney outcome 

20 (i.e., GFR) measures for early kidney damage is a particular problem in children who do 

21 not have many of the other kidney risk factors that adults do, such as hypertension and 

22 diabetes. As a result, such studies have utilized EBE markers. However, data to 

23 determine the predictive value of such biomarkers for subsequent kidney function decline 

24 in Pb exposed populations are extremely limited (Coratelli et al., 1988) and may pose 

25 particular challenges in children due to puberty-related biomarker changes (Sarasua et al., 

26 2003). The few studies included the 2006 Pb AQCD that analyzed clinical kidney 

27 outcomes in children found associations with indicators of Pb exposure that were 

28 inconsistent in direction. Fels et al. (1998) found no difference in mean serum creatinine 

29 between 62 children living near Pb-producing factories and 50 control children living in 

30 communities without Pb emission sources. In a study of 200 Belgian adolescents aged 17 

31 years, higher concurrent blood Pb level was associated with higher serum cystatin-C in 

32 200 (De Burbure et al., 2006); however, among 300-600 European children (n varied by 
February 2012 5-326 Draft – Do Not Cite or Quote

1 outcome), higher concurrent blood Pb level was associated with lower serum creatinine 

2 and cystatin C (Staessen et al., 2001).

3 Recent studies of children with elevated Pb exposure did not consistently indicate that Pb 

4 exposure was associated with reduced kidney function. A study in 123 children of 

5 workers in Pakistani Pb smelters and battery recycling plants and 123 control children, 

6 ages 1-6 years, reported elevated blood Pb levels, serum creatinine and urea in children of 

7 Pb-exposed workers compared to controls (medians: 8.1 versus 6.7 μg/dL; 56 versus 

8 52 μM; and 4,500 versus 4,300 μM, respectively (p ≤ 0.01 for all) in unadjusted analyses

9 (Khan et al., 2010a). Blood Pb levels were correlated with serum creatinine (Spearman r 

10 = 0.13; p = 0.05). However, a study of 77 participants, ages 10-25 years, who were 

11 previously Pb poisoned through contaminated flour and chelated, reported no difference 

12 in renal effects between children with blood Pb levels > 48 μg/dL and < 43 μg/dL 

13 although lower IQ was observed in the subset who were exposed before the age of six 

14 years (Coria et al., 2009).

15 One of the key gaps identified in the 2006 Pb AQCD was limited data in children and 

16 adolescents particularly with respect to GFR measures and in populations without the 

17 elevated Pb exposure associated with Pb poisoning, living near a Pb source, or having 

18 parents with occupational Pb exposures. A recently published NHANES analysis in 

19 adolescents begins to fill this gap (Fadrowski et al., 2010). Associations between 

20 concurrent blood Pb and kidney function were investigated in 769 adolescents aged 12-20 

21 years in the U.S. NHANES III, conducted 1988-1994. Kidney function was assessed with 

22 two eGFR equations. One utilized serum cystatin C and the other used the more 

23 traditional marker, serum creatinine. Median concurrent blood Pb and cystatin C-based 

eGFR levels were 1.5 μg/dL and 112.9 mL/min/1.73 m2 24 , respectively. Cystatin C-based 

eGFR was lower (-6.6 mL/min/1.73 m2 25 [95% CI: -0.7, -12.6]) in participants with blood 

26 Pb levels in the highest quartile (≥ 3.0 μg/dL) compared with those in the lowest (<1 

μg/dL). A doubling of blood Pb level was associated with a -2.9 mL/min/1.73 m2 27 (95% 

28 CI: -0.7, -5.0) lower eGFR. In contrast, the association between blood Pb and creatinine29 based eGFR, although in the same direction, was not statistically significant. As these 

30 children were born between 1968 and 1982, some likely had higher Pb exposures in 

31 earlier childhood, although notably, not as high or as long in duration as did older adults 

32 examined in aforementioned studies. Nonetheless, in this study of NHANES adolescents, 

33 there also is uncertainty regarding the magnitude, timing, frequency, and duration of Pb 

34 exposure that contributed to the observed associations. Additional research in children is 

35 warranted, in particular studies with longitudinal follow-up, multiple outcome assessment 

36 methods, and examination of children born after Pb was banned from gasoline. 
February 2012 5-327 Draft – Do Not Cite or Quote

5.5.2.4 Associations between Lead Dose and New Kidney 

Outcome Measures

1 As noted above, in an effort to more accurately estimate kidney outcomes, new equations 

2 to estimate GFR based on serum creatinine have been developed, and the utility of other 

3 biomarkers, such as cystatin C, as well as equations based on them, are being studied. 

4 However, few publications have utilized these state-of-the-art techniques when 

5 evaluating associations between Pb or cadmium dose and renal function. In addition to 

6 the study in NHANES adolescents discussed above (Fadrowski et al., 2010), a cross7 sectional study of Swedish women reported that higher concurrent blood Pb (median: 

8 2.2 μg/dL) and cadmium (median: 0.38 μg/L) levels were associated with lower eGFR 

9 based on serum cystatin C alone (without age, sex, and race) after adjustment for socio10 demographic and CKD risk factors (Akesson et al., 2005). Associations were comparable 

11 to those using creatinine clearance as the kidney outcome for Pb; however associations of 

12 cadmium dose measures were stronger for the cystatin C based outcome. Staessen et al.

13 (2001) found a statistically significant association between concurrent blood Pb level and 

14 serum cystatin C in a cross-sectional study of adolescents; creatinine-based measures 

15 were not reported. However, in a cross-sectional study of European children, higher 

16 concurrent blood Pb levels were associated with lower serum cystatin C and creatinine; 

17 these inverse associations were attributed to hyperfiltration (De Burbure et al., 2006). A 

18 very recent publication compared associations of blood Pb and eGFR using the traditional 

19 MDRD equation to those with four new equations: CKD-EPI, and cystatin C single 

20 variable, multivariable, and combined creatinine/cystatin C, in 3,941 adults who 

21 participated in the 1999-2002 NHANES cystatin C subsample (Spector et al., 2011).

22 Similar to the NHANES adolescent analysis, associations with the cystatin C outcomes 

23 were stronger. After multivariable adjustment, differences in mean eGFR for a doubling 

24 blood Pb were -1.9 (95% CI: -3.2, -0.7), -1.7 (95% CI: -3.0, -0.5), and -1.4 (95% CI: -2.3, 

-0.5) mL/min/1.73 m2 25 , using the cystatin C single variable, multivariable and combined 

26 creatinine/cystatin C equations, respectively, reflecting lower eGFR with increased blood 

27 Pb. The corresponding differences were -0.9 (95% CI: -1.9, 0.02) and -0.9 (95% CI: -1.8, 

28 0.01) using the creatinine-based CKD-EPI and MDRD equations, respectively.

5.5.2.5 Reverse Causality

29 As discussed briefly above, reverse causality has been considered as an alternative 

30 hypothesis to explain associations observed between indicators of Pb exposure and renal 

31 dysfunction in adults. The reverse causality hypothesis attributes increased blood and 

32 bone Pb levels to reduced Pb excretion from nonPb-related causes rather then implicating 

33 Pb-related renal dysfunction as a contributing factor to CKD. The 2006 Pb AQCD 
February 2012 5-328 Draft – Do Not Cite or Quote

1 concluded that "reverse causality is not likely to be a major explanatory factor accounting 

2 for observed associations between Pb and renal dysfunction" (U.S. EPA, 2006b). The 

3 strongest evidence against reverse causality was provided by longitudinal studies in 

4 which associations between baseline measurements of Pb biomarkers and subsequent 

5 changes in renal function were demonstrated. In the NAS, baseline blood Pb levels were 

6 associated with subsequent declines in renal function over follow-up periods ranging 

7 from 3 to 6 years after adjustment for baseline renal function (Tsaih et al., 2004; Kim et 

8 al., 1996). Similar results were reported in studies of patients with renal disease (Yu et 

9 al., 2004; Lin et al., 2003). The longitudinal analyses in NAS studies also indicated that 

10 associations between blood Pb level and serum creatinine persisted in the lower range of 

11 serum creatinine levels, including those in the normal range (Tsaih et al., 2004; Kim et 

12 al., 1996). Thus, the association was not limited to the segment of the population with 

13 potentially clinically significant renal dysfunction in whom reduced Pb excretion would 

14 be more likely.

15 Additional support against reverse causality was provided by findings in Swedish women 

16 that both higher blood and urinary Pb were associated with lower creatinine clearance

17 (Akesson et al., 2006; Akesson et al., 2005). If reverse causality were the more likely 

18 hypothesis for these associations, lower creatinine clearance would be associated with 

19 lower urinary Pb, which it is not. Among adults with chronic kidney disease, renal failure 

20 was not associated with increases in blood or bone Pb levels or chelatable Pb levels (Van 

21 De Vyver et al., 1988). Batuman et al. (1983) found that chelatable Pb levels were similar 

22 in adults with renal disease of unknown and known nonPb-related causes. Study had bone 

23 Pb levels (group means: 18 and 19 μg/g) in the range of those measured in recent 

24 epidemiologic studies. In summary, evidence that higher blood Pb levels are associated 

25 with subsequent declines in renal function from baseline levels, that associations persist 

26 among adults with normal renal function, and that renal failure does not increase Pb 

27 biomarker levels collectively do not support reverse causality and thus demonstrate that 

28 reverse causality does not provide a more suitable alternative hypothesis to explain the 

29 associations consistently observed between Pb biomarker levels and renal dysfunction.

5.5.3 Modes of Action for Lead-Induced Nephrotoxicity

5.5.3.1 Altered Uric Acid

30 Higher occupational Pb exposure or blood Pb levels have been linked to increased risk 

31 for both gout and kidney disease (Shadick et al., 2000; Batuman, 1993). Pb is thought to 

32 increase serum uric acid by decreasing its kidney excretion (Emmerson and Ravenscroft, 
February 2012 5-329 Draft – Do Not Cite or Quote

1 1975; Ball and Sorensen, 1969; Emmerson, 1965). Research during the past decade 

2 indicates that uric acid is nephrotoxic at lower levels than previously recognized (Johnson 

3 et al., 2003). Therefore, the 2006 Pb AQCD reviewed literature implicating increased uric 

4 acid as a mechanism for Pb-related nephrotoxicity (Weaver et al., 2005a; Shadick et al., 

5 2000). However, this does not appear to be the only mechanism, since associations 

6 between blood Pb and serum creatinine have remained significant even after adjustment 

7 for uric acid (Weaver et al., 2005a). These mechanistic relations have more than just 

8 theoretical importance. Clinically relevant therapies may be possible since EDTA 

9 chelation has been reported to improve both kidney function and urate clearance in 

10 patients with kidney insufficiency and gout, even when EDTA-chelatable Pb body 

11 burdens were low (Lin et al., 2001).

12 Alterations in serum uric acid have been studied in animal models exposed to Pb. 

13 Conterato et al. (2007) followed various parameters of kidney function after single or

14 multiple Pb injections in rats. The single dosing regimen consisted of a single i.p. 

15 injection of 25 or 50 mg/kg Pb-acetate, while the multiple injections involved once daily 

16 i.p. injection of either vehicle or Pb-acetate (5 or 25 mg/kg) for 30 days. Single and 

17 multiple injections at both dose levels increased plasma uric acid levels. Similarly, Abdel 

18 Moneim et al. (2011b) reported increased serum uric acid and urea levels after 5 days of 

19 Pb-acetate treatment (i.p. 20 mg/kg). In male rats exposed to Pb in drinking water from 

20 lactation to puberty (40 days) or post-puberty (65 days), Berrahal et al. (2011) found that 

21 plasma urea levels increased after 40 days of exposure (puberty blood Pb level of 12.7 

22 μg/dL) but decreased after 65 days of Pb exposure (post-puberty blood Pb level of 7.5 

23 μg/dL) (Table 5-23).

5.5.3.2 Oxidative Damage

24 A role for ROS in the pathogenesis of experimental Pb-induced hypertension and renal 

25 disease has been well characterized (Vaziri, 2008a, b; Vaziri and Khan, 2007). The 

26 production of oxidative stress following Pb exposure is detailed in respect to modes of 

27 action of Pb (Section 5.2.4). Past studies have shown that Pb treatment (single or three 

28 daily i.p. injections) can elevate kidney GST levels, affecting glutathione metabolism

29 (Daggett et al., 1998; Moser et al., 1995; Oberley et al., 1995).

30 Animal studies continue to provide evidence for increased oxidative stress playing a role 

31 in the pathogenesis of Pb-induced renal toxicity. Increased ROS, serum NO, and renal 

32 NO were observed after Pb injections in rats (i.p. 20 mg/kg, 5 days) (Abdel Moneim et 

33 al., 2011b). Pb exposure to rat proximal tubular cells (0.5-1 μM) also increased ROS 

34 production, in a concentration-dependent manner (Wang et al., 2011b). Oxidative stress 
February 2012 5-330 Draft – Do Not Cite or Quote

1 was also demonstrated by increased lipid peroxidation (i.e., MDA) in serum and renal 

2 tissue after Pb exposure (Abdel Moneim et al., 2011b; Lodi et al., 2011; Wang et al., 

3 2011b). Berrahal et al. (2011) reported increased MDA in Pb-exposed (50 ppm

4 Pb-acetate pre- and post-natally) rat kidney relative to controls at both 40 (puberty; blood 

5 Pb 12.7 μg/dL) and 65 (post-puberty; blood Pb 7.5 μg/dL) days of age. In addition, total 

6 sulfhydryl groups were significantly decreased at 65 days. These increases in oxidative 

7 stress were accompanied by age-dependent Pb nephrotoxicity in male rats (Table 5-24). 

8 Alterations in endogenous antioxidants and antioxidant enzymes that may lead to 

9 oxidative stress have also been reported after Pb exposure. Pb treatment decreased the 

10 activity of the renal antioxidant enzymes, CAT, SOD, GST, GPx, and GR (Abdel 

11 Moneim et al., 2011b) and protein levels of CAT and GSH (Lodi et al., 2011).

12 Additionally, proteomic analysis of high-level Pb treated (1,500 ppm, 5 weeks; resulting 

13 in blood Pb level of 53.4 μg/dL) rat kidney identified decreased abundance of a rate14 limiting enzyme in the synthesis of GSH (glutamate cysteine ligase) (Chen et al., 2011).

15 Conterato et al. (2007) examined the effect of Pb-acetate on the cytosolic thioredoxin 

16 reductase activity and oxidative stress parameters in rat kidneys. A single injection of 

17 Pb-acetate consisted of a single i.p. injection of 25 or 50 mg/kg Pb-acetate, while 

18 repeated injections consisted of one daily i.p. injection of Pb-acetate (5 or 25 mg/kg) for 

19 30 days. Measured were thioredoxin reductase-1, a selenoprotein involved in many 

20 cellular redox processes, SOD, δ-ALAD, GST, GPx, non protein thiol groups (NPSH), 

21 CAT, as well as plasma creatinine, uric acid, and inorganic phosphate levels. The single 

22 injection at the 25 mg Pb dose level resulted in increased SOD and thioredoxin reductase23 1 activity, while the 50 mg dose level increased CAT activity and inhibited δ-ALAD 

24 activity in the kidney. Repeated injections at the 5 mg dose level of Pb inhibited δ-ALAD 

25 and increased GST, NPSH, CAT, and thioredoxin reductase-1. Repeated injections at the 

26 25-mg dose level reduced δ-ALAD but increased GST, NPSH, and plasma uric acid 

27 levels. No changes were observed in TBARS, GPx, creatinine or inorganic phosphate 

28 levels after either single or repeated injection dosing. As both dosing regimens increased 

29 thioredoxin reductase-1 activity, the authors suggest that this enzyme may be a sensitive

30 indicator of renal changes with low dose Pb treatment.

31 Jurczuk et al. (2006) published a study of the involvement of some low molecular weight 

32 thiols in the peroxidative mechanisms of action of Pb in the rat kidney. Wistar rats were 

33 fed a diet containing 500 ppm Pb-acetate for a period of 12 weeks and were compared to 

34 a control group receiving distilled water for the same time period. GSH, metallothionein 

35 (MT), total and nonprotein SH groups (TSH and NPSH) were measured, as were the 

36 blood activity and urinary concentration of δ-ALA. The concentrations of GSH and 

37 NPSH were decreased by Pb administration, while MT concentration was unchanged. δ-
February 2012 5-331 Draft – Do Not Cite or Quote

1 ALAD in blood was decreased, whereas urinary δ-ALA was increased by Pb 

2 administration. Negative correlations were found between the kidney GSH concentrations 

3 and previously reported concentrations of Pb and MDA in kidneys of these rats. It is 

4 apparent from graphical presentation of the data that GSH was reduced by more than 

5 50% following Pb administration, while TSH was reduced by approximately 15%. No 

6 values for either blood or kidney Pb levels or kidney MDA were reported in this article. 

7 In 2007, the same authors (Jurczuk et al., 2007) reported on the renal concentrations of 

8 the antioxidants, vitamins C and E, in the kidneys of the same Pb-treated and control rats. 

9 Exposure to Pb significantly decreased vitamin E concentration by 13% and vitamin C 

10 concentration by 26%. The kidney concentration of vitamin C negatively correlated with 

11 MDA concentration. The authors concluded that vitamins E and C were involved in the 

12 mechanism of peroxidative action of Pb in the kidney, and their protective effect may be 

13 related to scavenging of free radicals.

14 Studies have used antioxidant compounds to investigate the role of oxidative stress in Pb15 induced nephrotoxicity. Abdel Moneim et al. (2011b) reported that flaxseed oil treatment 

16 protected rats from Pb-induced (i.p. 20 mg/kg, 5 days) oxidative stress, inflammation, 

17 and apoptosis. However, the flaxseed oil also decreased the accumulation of Pb in renal 

18 tissue making it difficult to ascertain whether the protection was due to decreased 

19 oxidative stress or to altered Pb uptake kinetics.

20 El-Neweshy and El-Sayed (2011) studied the influence of vitamin C supplementation on 

21 Pb-induced histopathological alterations in male rats. Rats were given Pb-acetate, 

22 20 mg/kg by intragastric feeding once daily for 60 days. Control rats were given 15 mg of 

23 sodium acetate per kg once daily, and an additional group was given Pb-acetate plus 

24 vitamin C (20 mg/kg every other day) 30 minutes before Pb feeding. Control rats showed 

25 normal histology, while Pb-treated rats exhibited karyomegaly with eosinophilic 

26 intranuclear inclusion bodies in the epithelial cells of the proximal tubules. Glomerular 

27 damage and tubular necrosis with invading inflammatory cells were also seen in Pb28 treated animals. Among rats treated with Pb-acetate plus vitamin C, five exhibited 

29 relatively mild karyomegaly and eosinophilic intranuclear inclusion bodies of proximal 

30 tubules and an additional five rats were normal. Normal glomeruli were noted in all. 

31 Thus, vitamin C was shown to ameliorate the renal histopathological effects of Pb 

32 intoxication, however no measures of Pb accumulation were provided to clarify the 

33 mechanism of action of vitamin C. 

34 Masso-Gonzalez and Antonio-Garcia (2009) studied the protective effect of natural 

35 antioxidants (zinc, vitamin A, vitamin C, vitamin E, and vitamin B6) against Pb-induced 

36 damage during pregnancy and lactation in rat pups. At weaning, pups were sacrificed and 

37 kidneys were analyzed. Pb-exposed pups had decreased body weights. Blood Pb levels 
February 2012 5-332 Draft – Do Not Cite or Quote

1 were 1.43 μg/dL in the control group, 22.8 μg/dL in the Pb group, 21.2 μg/dL in the Pb 

2 plus zinc plus vitamins group, and 0.98 μg/dL in the zinc plus vitamin group. The kidney 

3 TBARS were significantly elevated in Pb exposed pups, while treatment with vitamins 

4 and zinc returned TBARS to control levels. Kidney CAT activity was significantly 

5 increased above control with Pb treatment; however supplementation with zinc and 

6 vitamins reduced CAT activity toward normal. Pb exposure inhibited kidney Mn7 dependent SOD but not Cu-Zn-dependent SOD activity. Thus, supplementation with zinc 

8 and vitamins during gestation and lactation was effective in attenuating the redox 

9 imbalance induced by developmental, chronic low-level Pb exposure, likely through the 

10 alteration of Pb accumulation. 

11 Bravo et al. (2007) reported further that mycophenolate mofetil (an immunosuppressive 

12 agent used in renal transplantation which inhibits T and B cell proliferation) 

13 administration reduces renal inflammation, oxidative stress and hypertension in Pb14 exposed rats. Thus, an inflammatory immune and oxidative stress component can be seen 

15 as contributing to Pb-induced renal effects and hypertension.

16 Although the majority of studies of the effects of Pb exposure have been conducted in 

17 male rats, a couple of studies have compared the response of male rats with female rats

18 (Sobekova et al., 2009; Alghazal et al., 2008a). Sobekova et al. (2009) contrasted the 

19 activity response to Pb on the antioxidant enzymes, GPx and GR, and on TBARS in both 

20 male and female Wistar rats of equal age. Males weighing 412 ± 47 g and females 

21 weighing 290 ± 19 g were fed diets containing either 100 ppm or 1,000 ppm Pb-acetate

22 for 18 weeks. In the male rats, kidney Pb content increased by 492% on the 100 ppm Pb 

23 diet and by 7,000% on the 1,000 ppm Pb diet. In the female rats, kidney Pb content 

24 increased by 410% on the 100 ppm Pb diet and by 23,000% on the 1,000 ppm Pb diet. 

25 There was virtually no change in GPx in the kidney of male rats given the 100 ppm Pb 

26 diet but there was a significant reduction in GPx in the female rats on both the 100 ppm

27 diet and 1,000 ppm diet. In male rats, GR was increased from 182 units/gram of protein 

28 in control kidneys to 220 units on the 100 ppm Pb diet and 350 units on the 1,000 ppm

29 diet. In female rats, kidney GR decreased from 242 units in control animals to 164 units 

30 in animals on the 100 ppm Pb diet and 190 units in animals on the 1,000 ppm diet. In 

31 male rats, kidney TBARS content increased from 7.5 units/gram protein to 10.0 units 

32 (1,000 ppm Pb diet group). In female rats, there was a reduction in TBARS from 14.4 

33 units per gram protein to 10.0 units in rats on the 100 ppm Pb diet and to 11 units in rats 

34 on the 1,000 ppm Pb diet. 

35 Alghazal et al. (2008a) compared the activity responses of the antioxidant enzyme, SOD 

36 and the detoxifying enzyme, GST, of the same rats exposed to 100 ppm or 1,000 ppm

37 Pb-acetate for 18 weeks. Similar to the previous study, kidney TBARS were increased 
February 2012 5-333 Draft – Do Not Cite or Quote

1 only in male rats given the higher dose of Pb. Kidney SOD activity, on the other hand, 

2 was increased in both males and females at the higher dose of Pb, while GST activity was

3 increased in kidney of males at the higher dose of Pb and decreased at the lower dose, but 

4 was decreased at both doses of Pb in females. Thus there were significant differences in 

5 the responses of male and female rats to Pb exposure. Differences may be accounted for 

6 in part due to the greater deposition of Pb in female rat kidneys. Another explanation, 

7 offered by the authors, is that male rats are known to metabolize some foreign 

8 compounds faster than do females, so the biological half-life of xenobiotics in the 

9 females may be longer.

5.5.3.3 Renal Gangliosides

10 Gangliosides are constituents of the plasma membrane that are important for control of 

11 renal GFR because they can act as receptors for various molecules and have been shown 

12 to take part in cell-cell interactions, cell adhesion, recognition and signal transduction. 

13 Aguilar et al. (2008) studied changes in renal gangliosides following Pb exposure 

14 (600 ppm Pb-acetate in drinking water for 4 months) in adult male Wistar rats. Pb 

15 exposure caused an increase in blood Pb from 2.1 to 35.9 μg/dL. There was no change in 

16 serum creatinine or in hemoglobin, but there was an increase in urinary δ-ALA. The 

17 following renal gangliosides were measured by immunohistochemistry and by thin layer 

18 chromatography: GM1, GM2, GM4, and 9-O-acetylated modified form of the GD3 

19 ganglioside (9-O-Ac-GD3). The ganglioside pattern was mainly characterized by a 

20 decrease in the GM1 ganglioside as well as by a mild increase in GM4 and GM2 

21 gangliosides, while the strongest alteration was observed in the 9-O-Ac-GD3, which was 

22 overexpressed. The latter was observed only in the glomerular zone. This was associated 

23 with a decrease in apoptotic glomerular cells, as assessed by the TUNEL assay. The 

24 authors hypothesized that the increase in GD3-O-acetylation could represent a strategy to 

25 attenuate the normal renal apoptotic process and therefore contribute to cell survival 

26 during Pb exposure.

5.5.3.4 Role of Metallothionein

27 Yu et al. (2009) described dichotomous effects of Pb-acetate on the expression of MT in 

28 the liver and kidney of mice. Male mice were i.p. injected with Pb-acetate in doses of 

29 100, 200, and 300 μmol/kg and sacrificed 4, 8, and 24 hours after Pb treatment. 

30 Administration of Pb increased the levels of MT-1 mRNA in the liver and kidneys but 

31 increased MT protein only in the liver. Treatment of mouse PT cells in vitro with Pb also 

32 resulted in an increase in MT mRNA but little increase in MT protein. Thus, Pb appears 
February 2012 5-334 Draft – Do Not Cite or Quote

1 to exert a dual effect on MT expression in the kidney: enhancement of MT gene 

2 transcription but suppression of MT mRNA translation.

3 Zuo et al. (2009) examined the potential role of α-Synuclein (Scna) and MT in Pb4 induced inclusion body formation. They used MT-I/II double knockout (MT-null) and 

5 parental wild type (WT) cell lines to explore the formation process of Pb-induced 

6 inclusion bodies. Unlike WT cells, MT-null cells did not form inclusion bodies after Pb 

7 exposure. Western blot of the cytosol showed that soluble MT protein in WT cells was 

8 lost during Pb exposure as inclusion bodies formed. However, transfection of MT-1 into 

9 MT-null cells allowed inclusion body formation after Pb exposure. As Scna is a protein 

10 with a natural tendency to aggregate into oligomers, Scna was measured in WT cells and 

11 MT-null cells after Pb exposure. Scna protein showed poor basal expression in MT-null 

12 cells, and Pb exposure increased Scna expression only in WT cells. MT transfection 

13 increased Scna transcript to WT levels. In both of these cell lines Pb-induced Scna 

14 expression rapidly increased and then decreased over 48 hours as Pb-induced inclusion 

15 bodies were formed. A direct interaction between Scna and MT was confirmed ex vivo 

16 by an antibody pull down assay, where the proteins co-precipitated with an antibody to 

17 MT. Pb exposure caused increased colocalization of MT and Scna proteins. In archival 

18 kidney samples of renal cortex from WT mice chronically treated with Pb, MT was 

19 localized to the surface of inclusion bodies. Thus, Scna may be a component of Pb20 induced inclusion bodies and, with MT, may play a role in inclusion body formation.

5.5.4 Effects of Exposure to Lead Mixtures

21 The effect of Pb on other cations, specifically calcium, is well established in the kidney 

22 literature. Calcium-mediated processes involving receptors, transport proteins, and 

23 second messenger signaling among other endpoints have been shown to be significantly 

24 affected by Pb exposure. The disposition of Pb in the soft tissues (kidney and spleen) can 

25 change with exposure to Pb and other compounds. Pb plus Cd exposure changed Pb 

26 disposition with increased blood Pb (versus Pb alone group) and decreased metal 

27 concentration in the kidney and liver (versus Pb alone). An iron deficient diet 

28 significantly increased Pb deposition in adult animals (Hashmi et al., 1989), pregnant 

29 dams, and maternally-exposed fetuses (Singh et al., 1991). Dietary thiamine plus zinc 

30 slightly reduced blood and kidney Pb in exposed animals (Flora et al., 1989). Selenium, a 

31 cofactor for GPx, attenuated Pb-induced lipid peroxidation and abrogated the Pb-induced 

32 attenuation of GR and SOD. Concomitant exposure to the cations aluminum and Pb 

33 protected animals from ensuing nephropathy (Shakoor et al., 2000). In summary, Pb has 

34 been shown to affect processes mediated by endogenous divalent cations. In addition, 
February 2012 5-335 Draft – Do Not Cite or Quote

1 exposure to other metals or divalent cations can modulate Pb disposition and its effects in 

2 the body. 

5.5.4.1 Lead and Cadmium

3 Cd shares many similarities with Pb; it has been shown to be a ubiquitous PT 

4 nephrotoxicant and accumulates in the body. Despite the similarities, few studies have 

5 evaluated associations between Cd exposure and CKD or the impact of joint exposure of 

6 Pb and Cd or other metals on CKD. As discussed in the 2006 Pb AQCD, environmental 

7 exposure to Cd, at levels common in the U.S. and other developed countries, has been 

8 shown to impact substantially associations between indicators of Pb exposure and the 

9 kidney EBE marker, NAG, even in the presence of occupational level Pb exposure. In an 

10 occupational study, mean NAG, although higher in the Pb-exposed worker group 

11 compared to controls, was correlated with urine Cd but not blood or tibia Pb (Roels et al., 

12 1994). In another occupational population where both metals were significantly 

13 associated with NAG, a 0.5 μg/g creatinine increase in Cd had the same effect on NAG as 

14 did a 66.9 μg/g bone mineral increase in tibia Pb (Weaver et al., 2003a).

15 The 2006 Pb AQCD noted that data examining the concentration-response relation 

16 between environmental Cd and the kidney were too scarce to determine the impact of Cd 

17 exposure on relations between Pb exposure and other kidney outcomes. A recent 

18 publication in NHANES data collected from 1999 through 2006 addresses this need; 

19 (results pertaining solely to Pb were discussed in Section 5.5.2.2) (Navas-Acien et al., 

20 2009). Geometric mean concurrent blood Cd level was 0.41 μg/L in 14,778 adults aged ≥ 

21 20 years. After adjustment for survey year, sociodemographic factors, CKD risk factors, 

22 and blood Pb, the ORs for albuminuria (≥ 30 mg/g creatinine), reduced eGFR (<60 

mL/min/1.73 m2 23 ), and both albuminuria and reduced eGFR were 1.92 (95% CI: 1.53, 

24 2.43), 1.32 (95% CI: 1.04, 1.68), and 2.91 (95% CI: 1.76, 4.81), respectively, comparing 

25 the highest with the lowest blood Cd quartiles. Both Pb and Cd remained significantly 

26 associated after adjustment for the other. Effect modification was not observed; however, 

27 ORs were higher for adults in the highest quartiles of both metals compared with the ORs 

28 for the highest quartiles of concurrent blood Cd or Pb alone (Table 5-25). Compared with 

29 adults with blood Cd levels ≤ 0.2 μg/L and blood Pb levels ≤ 1.1 μg/dL, adults with blood 

30 Cd levels > 0.6 μg/L and blood Pb levels > 2.4 μg/dL had ORs (95% CIs) of 2.34 (95% 

31 CI: 1.72, 3.18) for albuminuria, 1.98 (95% CI: 1.27, 3.10) for reduced eGFR, and 4.10 

32 (95% CI: 1.58, 10.65) for albuminuria and reduced eGFR together. These findings are 

33 consistent with other recent publications (Akesson et al., 2005; Hellstrom et al., 2001),

34 support consideration of both metals as independent CKD risk factors in the general 
February 2012 5-336 Draft – Do Not Cite or Quote

1 population, and provide novel evidence of increased risk in those with higher 

2 environmental exposure to both metals.

3 However, a very recent study suggests that interpretation of Cd associations with GFR 

4 measures may be much more complex. Conducted in Pb workers to address the fact that 

5 few studies have examined the impact of environmental Cd exposure in workers who are 

6 occupationally exposed to other nephrotoxicants such as Pb, the study assessed Cd dose 

7 with urine Cd, which is widely considered the optimal dose metric of cumulative Cd 

8 exposure. In 712 Pb workers, mean (SD) blood and tibia Pb, urine Cd, and eGFR using 

9 the MDRD equation were 23.1 (14.1) μg/dL, 26.6 (28.9) μg/g, 1.15 (0.66) μg/g 

creatinine, and 97.4 (19.2) mL/min/1.73m2 10 , respectively (Weaver et al., 2011). After 

11 adjustment for age, sex, BMI, urine creatinine, smoking, alcohol use, education, annual 

12 income, diastolic BP, current or former Pb worker job status, new or returning study 

13 participant, and blood and tibia Pb, higher urine Cd was associated with higher calculated 

creatinine clearance, eGFR (β = 8.7 mL/min/1.73 m2 14 [95% CI: 5.4, 12.1] per unit 

15 increase in ln-transformed urine Cd) and ln-NAG, but lower serum creatinine. These 

16 unexpected paradoxical associations have been reported in a few other publications

17 (De Burbure et al., 2006; Hotz et al., 1999) and have been observed in other populations. 

18 Potential explanations for these paradoxical results included a normal physiologic 

19 response in which urine Cd levels reflect renal filtration; the impact of adjustment for 

20 urine dilution with creatinine in models of kidney outcomes; and Cd-related 

21 hyperfiltration. 

22 Wang et al. (2009c) studied the effects of Pb and/or Cd on oxidative damage to rat kidney 

23 cortex mitochondria. In this study young female Sprague Dawley rats were fed for 8 

24 weeks with either Pb-acetate (300 ppm), Cd chloride (50 ppm), or Pb and Cd together in 

25 the same dosage. Lipid peroxidation was assessed as MDA content. Renal cortex pieces 

26 were also processed for ultrastructural analysis and for quantitative rtPCR to identify the 

27 mitochondrial damage and to quantify the relative expression levels of cytochrome 

28 oxidase subunits (COX-I/II/III). Cytochrome oxidase is the marker enzyme of 

29 mitochondrial function, and COX-I, II, and III are the three largest mitochondrially30 encoded subunits which constitute the catalytic functional core of the COX holoenzyme. 

31 Mitochondria were altered by either Pb or Cd administration, but more strikingly by Pb 

32 plus Cd administration, as indicated by disruption and loss of mitochondrion cristae. 

33 Kidney cortex MDA levels were increased significantly by either Pb or Cd, given 

34 individually, but more so by Pb plus Cd. COX-I/II/III were all reduced by either Pb or Cd 

35 administration, but more prominently by Pb plus Cd administration. This study adds to 

36 knowledge of the synergistic effects of Pb and Cd on kidney mitochondria.
February 2012 5-337 Draft – Do Not Cite or Quote

5.5.4.2 Lead, Cadmium, and Arsenic

1 Wang and Fowler (2008) present a general review of the roles of biomarkers in 

2 evaluating interactions among mixtures of Pb, Cd, and arsenic. Past studies have found 

3 that addition of Cd to treatment of rats with Pb or Pb and As significantly reduced the 

4 histological signs of renal toxicity from each element alone; on the other hand, animals 

5 exposed to Cd in addition to Pb or Pb and As showed an additive increase in the urinary 

6 excretion of porphyrins, indicating that, although measured tissue burdens of Pb were 

7 reduced, the biologically available fraction of Pb is actually increased (Mahaffey et al., 

8 1981; Mahaffey and Fowler, 1977).

9 Stress proteins were examined after exposure to mixtures of Pb and other metals. 

10 Induction of MT was strongest in groups with Cd treatment. However, co-exposure to Pb 

11 and As induced higher levels of MT protein than did either Pb or As exposure alone in 

12 kidney tubule cells. Heat shock proteins (Hsps) are commonly altered with exposure to 

13 metal mixtures. A study found in vitro (low dose) and in vivo that Pb induced Hsps in a 

14 metal/metalloid-, concentration- and time-specific manner (Wang et al., 2005). Additive 

15 or more than additive interactions occurred among Pb, Cd and As under combined 

16 exposure conditions.

5.5.4.3 Lead and Zinc

17 Zinc has been investigated as a protective compound against the effects of Pb. Pb 

18 treatment (35 mg/kg i.p. for 3 days) caused a significant fall in hemoglobin content, 

19 significant increases in lipid peroxidation and decreased level of reduced glutathione in 

20 liver, together with diminished total protein content in liver and kidney. Co-treatment of 

21 Pb with zinc (10 mg/kg i.p.) or ascorbic acid (10, 20 and 30 mg/kg i.p.) showed a 

22 moderate therapeutic effect when administered individually, but more pronounced 

23 protective effects after combined therapy (Upadhyay et al., 2009).

24 Jamieson et al. (2008) studied the effect of dietary zinc content on renal Pb deposition. 

25 Weanling Sprague Dawley rats were assigned to marginal zinc (MZ, 8 mg Zn/kg diet), 

26 zinc adequate control (CT, 30 mg Zn/kg), zinc-adequate diet-restricted (30 mg Zn/kg), or 

27 supplemental zinc (SZn, 300 mg Zn/kg) groups, with or without Pb-acetate (200 ppm for 

28 3 weeks). Pb exposure did not result in nephromegaly or histological alterations. The MZ 

29 rats had higher renal Pb (35%) and lower renal zinc (16%) concentrations than did CT 

30 rats. On the other hand, SZn was more protective than the CT diet was against renal Pb 

31 accumulation (33% lower). Standard procedures for indirect immunoperoxidase staining 

32 were used to determine MT localization in the kidney. Pb had no effect on MT staining 
February 2012 5-338 Draft – Do Not Cite or Quote

1 intensity, distribution, or relative protein amounts. Western blot analysis confirmed that 

2 MT levels were responsive to dietary zinc but not to Pb exposure.

5.5.4.4 Lead and Mercury

3 Stacchiotti et al. (2009) studied stress proteins and oxidative damage in a renal-derived 

4 cell line exposed to inorganic mercury and Pb. The time course of the expression of 

5 several Hsps, glucose-regulating proteins and MTs in a rat proximal tubular cell line 

6 (NRK-52E) exposed to subcytotoxic doses of inorganic mercury (HgCl2, 1-40 μM) and 

7 Pb (PbCl2, 2-500 μM) were analyzed. ROS and reactive nitrogen species (RNS) were 

8 detected by flow cytometric analysis. Endogenous total GSH content and the enzymatic 

9 activity of GST were determined in cell homogenates. Western blot analysis and 

10 immunohistochemistry were used for quantification of hsps and MTs. Reverse 

11 transcription PCR was used for quantification of metallothionein. The higher doses of 

12 mercury (20 μM and 40 μM) were shown to markedly inhibit growth of the cell line 

13 while the higher doses of Pb (60 μM to 500 μM) inhibited cell growth to a lesser degree. 

14 After 24 hours of exposure at 20 μM mercury, the cells presented abnormal size and 

15 pyknotic nuclei, swollen mitochondria and both apoptosis and overt necrosis. In the 

16 presence of 60 or 300 μM Pb, the cells lost cell-cell and cell-matrix contacts, showed a 

17 round size, irregular nuclear contour and often mitotic arrest, but no apoptosis or overt 

18 necrosis at 24 hours. Mercury induced a significant increase in both ROS and RNS, 

19 maximal RNS at 24 hours, and maximal ROS at 48 hours. Pb (60 or 300 μM) did not 

20 cause an increase in ROS or RNS beyond the levels measured in control cells. Total GSH 

21 significantly increased in cells grown in the presence of Pb; the effect was concentration22 dependent and GSH reached its maximal value at a dose of 300 μM Pb. The effect of 

23 mercury was biphasic: 10 μM significantly enhanced GSH by 600%, while the amount of 

24 GSH detected after 20 μM mercury only increased by 50% compared to control levels. 

25 GST activity was enhanced by both Pb and mercury. Hsp25 and Hsp72 were up-regulated 

26 by mercury but there was no effect on Grp78 as compared to control. On the contrary, Pb 

27 treatment only upregulated Grp78. Mercury induced a time-dependent effect on MT 

28 mRNA expression, which reached its maximal value 3 hours after beginning treatment 

29 and reverted to control values at 24 hours. With Pb, on the other hand, mRNA 

30 transcription was concentration- and time-dependent. The transcripts remained 

31 overexpressed compared to controls up to 72 hours. The results of this study with regard 

32 to the Pb effect on MT synthesis clearly differ from those of Jamieson et al. (2008),

33 which found no increase in MT following Pb exposure. This discrepancy remains to be 

34 clarified.
February 2012 5-339 Draft – Do Not Cite or Quote

5.5.5 Impact of Treatment with Antioxidants on Renal Lead Accumulation 

and Pathology

5.5.5.1 Treatment with Antioxidants

1 Wang et al. (2010d) assessed the protective effect of N-acetylcysteine (NAC) on 

2 experimental chronic Pb nephrotoxicity in immature female rats. NAC is a potent oxygen 

3 free radical scavenger, a metal chelator, and the precursor to the antioxidant glutathione. 

4 Sprague-Dawley rats received Pb-acetate (300 ppm in drinking water) and/or NAC 

5 (100 mg/kg/day, by i.p. injection) for 8 weeks to investigate the protective effect of NAC 

6 on Pb-induced renal damage and oxidative stress. Serum and renal cortical Pb levels were 

7 markedly increased in the Pb-treated animals, but reduced in the Pb plus NAC treated 

8 animals. There were time-related increases in urinary alkaline phosphatase, urinary GGT, 

9 urinary NAG, urinary total protein, urinary β-2 microglobulin, and urinary microalbumin, 

10 which were all decreased by NAC. Serum urea nitrogen was significantly increased by Pb 

11 administration and reduced toward normal by Pb plus NAC. Alterations in proximal 

12 tubular structures were observed in most kidney samples from Pb-treated rats, but 

13 animals treated with combination Pb plus NAC showed well-preserved cell structures and 

14 organelles. Indices of oxidative stress (MDA, SOD, GSH, GPx, and CAT) were altered 

15 by Pb treatment and restored to or toward normal by Pb plus NAC treatment (MDA 

16 increased and the remainder decreased). Thus NAC was shown to have both an anti17 oxidative and a chelator effect on Pb intoxication.

18 Saxena et al. (2005) investigated the beneficial role of monoesters of meso-2, 3-

19 dimercaptosuccinic acid in the mobilization of Pb and recovery of tissue oxidative injury 

20 in rats. Dimercaptosuccinic acid (DMSA) is known as a Pb chelator and as an antioxidant 

21 by virtue of its possession of thiol groups. In this study, DMSA, and two of its analogues, 

22 monomethyl dimercaptosuccinic acid (MmDMSA) and mono-cyclohexyl 

23 dimercaptosuccinic acid (MchDMSA) were assessed for their capability to reduce Pb 

24 concentration in blood and soft tissues and to recover Pb-induced oxidative stress. Male 

25 Wistar rats were exposed to Pb-acetate (0.1% in drinking water) for 20 weeks. Rats were 

26 then treated orally once daily for five days with DMSA or its two analogues at doses up 

27 to 100 mg/kg. Exposure to Pb caused a rise in blood Pb levels to approximately 25 μg/dL. 

28 Exposure to Pb also caused a significant decrease in blood ALAD activity and GSH 

29 levels, accompanied by inhibition of kidney ALAD and an increase in δ-aminolevulinic 

30 acid synthetase (ALAS) activity in liver and kidneys. Pb exposure also resulted in 

31 increased blood and soft tissue (brain, liver, and kidney) Pb and TBARS levels and 

32 decreased GSH levels. These were restored by treatment with DMSA and its analogues, 

33 particularly MchDMSA. 
February 2012 5-340 Draft – Do Not Cite or Quote

1 Abdallah et al. (2010) examined the effect of Pb on coenzyme Q levels in rat tissues. 

2 Coenzyme Q acts as an electron and proton carrier in mitochondria and functions as an 

3 antioxidant in its reduced form (ubiquinol). Both coenzyme Q9 and coenzyme Q10 were 

4 measured in rat tissues as coenzyme Q9 is the predominant form found in the rat. Male 

5 albino rats were injected i.p. with Pb-acetate at a dose of 5 mg/kg daily for 6 weeks. No 

6 blood Pb levels were reported. TBARS were elevated above levels in controls in serum, 

7 liver, kidney and brain while non-protein sulfhydryl groups (indicative of GSH) were 

8 decreased in serum and kidney. Both oxidized and reduced coenzyme Q9 levels were 

9 significantly reduced in kidneys from Pb-treated rats as contrasted to controls (mean 

10 [SE]: respectively, 48.6 [5.6] versus 95.5 [10.1] nmol/g tissue for oxidized form and 35.4 

11 [3.0] versus 61.4 [5.1] nmol/g tissue for reduced form). On the other hand, levels of 

12 oxidized and reduced coenzyme Q10 were unchanged. Thus, the reduced levels of 

13 coenzyme Q attributable to Pb exposure may be a factor in the diminished antioxidant 

14 defense mechanism. 

15 El-Sokkary et al. (2005) evaluated the effect of melatonin against Pb-induced hepatic and 

16 renal toxicity in male rats. Melatonin is known to be efficacious as a free radical 

17 scavenger and indirect antioxidant. Three groups of animals were used: control, 

18 Pb-acetate-treated (100 ppm) and Pb-acetate and melatonin (10 mg/kg) given 

19 subcutaneously for 30 days. Lipid peroxidation was measured as the sum of MDA and 4-

20 hydroxyalkenals (4-HAD). Pb increased kidney lipid peroxidation products, but these 

21 were reduced toward normal with melatonin co-treatment. Both SOD and GSH levels 

22 were reduced by Pb and were increased by melatonin. Histological section of kidneys of 

23 Pb-treated rats showed tubular degeneration with some apparently necrotic cells, while 

24 melatonin-treated rats demonstrated a near normal structure. The authors concluded that 

25 melatonin protected the liver and kidneys from the damaging effects of exposure to Pb 

26 through inhibition of lipid peroxidation and stimulation of endogenous antioxidative 

27 defense systems.

28 Ozsoy et al. (2010) studied the protective effects of L-carnitine on experimental Pb 

29 toxicity in rats. Female two month-old rats were fed 0.5 mg/kg Pb-acetate alone or with 

30 daily injections of 0.5 mg/kg L-carnitine for 60 days. Control animals were injected with 

31 physiological saline. Pb caused an increase in serum creatinine and histopathological 

32 changes in the kidney, consisting of tubule dilatation, degeneration and necrosis and 

33 interstitial inflammation. In the Pb plus L-carnitine group, serum creatinine was reduced 

34 to control values and the histopathological changes were reversed. Immunological 

35 staining indicated Cu/Zn-SOD stimulation by Pb feeding alone and reduction by L36 carnitine co-treatment. The authors attributed the beneficial effects of L-carnitine to its 

37 antioxidant effect.
February 2012 5-341 Draft – Do Not Cite or Quote

1 Reddy et al. (2010) treated Sprague-Dawley rats with 10 mg/kg/day of Pb-acetate and/or 

2 thiamine (25 mg/kg/day) for 7 weeks. Thiamine treatment normalized the Pb-induced 

3 alterations in blood ALAD activity and urinary NAG activity.

4 The effectiveness of various plant or bacterial extracts as antioxidants in the kidney was 

5 examined in a few studies. Kharoubi et al. (2008a) described the prophylactic effects of 

6 Wormwood (Artemisia absinthium L.) plant extracts on kidney function in Pb-exposed 

7 animals. Male Wistar rats were exposed to Pb-acetate (750 ppm in drinking water) for 11 

8 weeks, and then received Wormwood extract (200 mg/kg) for 4 weeks. Significant 

9 differences in blood and urinary Pb concentration were observed between the Pb group 

10 and the Wormwood group (e.g., 55.6 μg/dL blood Pb versus 22.3 μg/dL, respectively).

11 Pb induced lipid peroxidation (TBARS and protein carbonyls in the kidney), but these 

12 levels were reduced by Wormwood extract. Wormwood extract also attenuated the 

13 effects of Pb on renal function. These results indicated that Wormwood extract had 

14 significant antioxidant activity and protected the kidney from Pb-induced toxicity.

15 Jayakumar et al. (2009) evaluated the effect of a methanolic extract of the Indian herb, 

16 Achyranthes aspera, in preventing Pb-induced nephrotoxicity in rats. Male albino Wistar 

17 rats, received Pb-acetate (0.2% for 6 weeks) or Pb-acetate plus A. aspera (200 mg/kg for 

18 6 weeks) simultaneously. A. aspera partially prevented the increases in kidney weight, 

19 BUN, serum uric acid, and serum creatinine caused by Pb administration. The levels of 

20 urinary marker enzymes, GGT, β-glucuronidase, NAG, Cathepsin D, and LDH, which 

21 were reduced by Pb administration, were increased to or toward normal by A. aspera. 

22 Kidney histology revealed that Pb-treated animals showed tubular damage, whereas the 

23 Pb plus A. aspera-treated animals showed a reduction in tubular damage. 

24 El-Nekeety et al. (2009) evaluated the protective effect of an extract of the folk medicine 

25 plant Aquilegia vulgaris against Pb-acetate-induced oxidative stress in Sprague-Dawley 

26 rats. The experimental group was treated with 200 ppm Pb-acetate and/or 100 ppm of an 

27 extract of A. vulgaris for 2 weeks prior to Pb-acetate. Pb-acetate increased serum urea 

28 and decreased serum total protein and albumin. These changes were reversed by 

29 treatment with the extract. Histological examination of kidneys of rats treated with Pb 

30 showed tubular dilatation, interstitial inflammatory cells, hemorrhage, cellular debris, and 

31 hypercellularity in the glomerulus, with apoptotic nuclei in renal tubular epithelial cells. 

32 The rats treated simultaneously with Pb and the extract showed essentially normal renal 

33 tubules and glomeruli while rats treated with Pb and then the extract showed 

34 improvement in tubular structure, but interstitial fibrosis was still present. This 

35 experiment indicated that exposure to Pb generates free radicals, and that an extract of A. 

36 vulgaris resulted in restoration of the different parameters tested. The second experiment 

37 in this group was by Ponce-Canchihuaman et al. (2010) who evaluated the antioxidant 
February 2012 5-342 Draft – Do Not Cite or Quote

1 activity of the cyanobacterium Spirulina maxima against Pb-acetate-induced 

2 hyperlipidemia and oxidative damage in the liver and kidney of male rats. Male Wistar 

3 rats were treated with Pb-acetate by i.p. injection (25 mg/rat on a weekly basis for 3 

4 weeks), and a 5% supplement of Spirulina was given in food. The findings in the kidney 

5 were similar to those in the liver (see Section 5.9.1). Thus, Pb-induced oxidative stress 

6 and renal damage can be attenuated by treatment with Spirulina extract.

7 Finally, there is a need to examine whether the chelator, CaNa2EDTA, acts also as an 

8 antioxidant and promotes increased vasodilatation and subsequent increased renal blood 

9 flow by enhancing the delivery of NO. This question arose because of the observations of 

10 Lin et al. (2006b) that repeated injections of CaNa2EDTA lead to improvement in kidney 

11 function in patients with chronic renal failure, even in individuals with very low body Pb 

12 stores as indicated by EDTA mobilization tests (i.e., < 80 μg 72 hour urinary Pb 

13 excretion). Jacobsen et al. (2001) examined the anti-oxidative effects of Gallic acid, 

14 EDTA, and an emulsifier in mayonnaise enriched with 16% fish oil. EDTA was shown to 

15 be an efficient antioxidant in the fish oil enriched mayonnaise as it strongly inhibited the 

16 formation of free radicals and volatile oxidation compounds. The authors suggested that 

17 the antioxidative effect appears to be due to its ability to chelate free iron in egg yolk at 

18 the oil-water interface. 

5.5.5.2 Treatment with Antioxidants plus Chelators 

19 Santos et al. (2006a) assessed the potentiating effects of chelators (2,3-

20 dimercaptopropanol [BAL], 2,3-dimercaptopropane-l-sulfonic acid [DMPS], and meso21 2,3-dimercaptosuccinic acid [DMSA]) given simultaneously with Pb-acetate on δ-ALAD 

22 activity, both in vivo and ex vivo. Ex vivo, human blood was pre-incubated with BAL or 

23 DMSA (10 μM) or DMPS (1 μM) then Pb-acetate added to the reaction mixture. In vivo, 

24 mice were given daily injections of 50 mg/kg Pb-acetate for 15 days and then injected 

25 with 1/3 of LD50 of the chelating agents. In human blood, the inhibitory effect of 

26 Pb-acetate (1 and 100 μM) on δ-ALAD activity was markedly increased in the presence 

27 of BAL and DMPS, whereas DMSA ameliorated the enzyme inhibition caused by 1 μM 

28 Pb-acetate. In vivo, Pb-acetate inhibited δ-ALAD activity by 42%. Parallel to the ex vivo 

29 results, BAL and DMPS, but not DMSA, increased the inhibitory potency of Pb in blood. 

30 In the kidney, BAL and DMSA but not DMPS increased inhibitory activity. The authors 

31 conjectured that the chelators may deplete the cells of zinc, an essential element for δ32 ALAD activity. These observed effects with chelators were supported by Bradberry and 

33 Vale (2009), Hamidinia et al. (2006), and Aslani et al. (2010) who found decreased 

34 kidney Pb content post-chelation.
February 2012 5-343 Draft – Do Not Cite or Quote

5.5.6 Summary and Causal Determination

1 The 2006 Pb AQCD concluded that “in the general population, both circulating and 

2 cumulative Pb was found to be associated with a longitudinal decline in renal function,” 

3 evidenced by increased serum creatinine and decreased creatinine clearance or eGFR 

4 over follow-up of 4 to 15 years in association with higher baseline blood and bone Pb 

5 levels (U.S. EPA, 2006b). Data in general and patient populations of adults provided 

6 consistent evidence of Pb-associated lower renal function in populations with mean 

7 concurrent or baseline blood Pb levels of 2-10 μg/dL (Akesson et al., 2005; Tsaih et al., 

8 2004; Yu et al., 2004; Kim et al., 1996); associations with lower eGFR were observed in 

9 adults with hypertension with a mean concurrent blood Pb level of 4.2 μg/dL (Muntner et 

10 al., 2003). The conclusion from the 2006 Pb AQCD was substantiated by the coherence 

11 of effects observed across epidemiologic and toxicological studies. Both human and 

12 animal studies observed Pb-associated hyperfiltration. In animals during the first 3 

13 months after Pb exposure, effects were characterized by increased GFR and increased 

14 kidney weight due to glomerular hypertrophy. However, exposure for 6 or 12 months 

15 resulted in decreased GFR, interstitial fibrosis, and kidney dysfunction. Additionally, 

16 toxicological studies found that early effects of Pb on tubular cells were generally 

17 reversible, but continued exposure resulted in chronic irreversible damage. Toxicological 

18 studies provided mechanistic evidence to support the biological plausibility of Pbinduced renal effects, including oxidative stress leading to • 19 NO inactivation. Despite the 

20 strong body of evidence presented in the 2006 Pb AQCD, uncertainty remained on the 

21 contribution of past Pb exposures to associations observed in adults, the impact in 

22 children, and the implication of hyperfiltration. 

23 Recent epidemiologic studies in adult general and patient populations continue to support 

24 Pb-related nephrotoxicity with consistently observed associations of blood and bone Pb 

25 levels with worse kidney function. These studies benefit from a number of strengths that 

26 vary by study but include comprehensive assessment of Pb dose with measurements of 

27 blood Pb, bioavailable Pb, and bone Pb as a biomarker of cumulative exposure; 

28 prospective study design; and statistical approaches that utilize a range of exposure and 

29 outcome measures, while adjusting for numerous potential confounding factors including 

30 age, race, sex, education, household income, smoking, alcohol use, cadmium exposure, 

31 and various health indicators such as diabetes, SBP, BMI, and history of cardiovascular 

32 disease. Large sample sizes provide strength to the general population studies. Re33 examination of a study from the 2006 Pb AQCD provided data to conclude that a 10-fold 

34 increase in concurrent blood Pb (e.g., from 1 to 10 μg/dL) was associated with an 18 

35 mL/min decrease in estimated creatinine clearance or a 25% decrease from the mean, and 

36 that an increase in blood Pb from the 5th to the 95th percentile (3.5 μg/dL) had the same 

negative impact on eGFR as did an increase of 4.7 years in age or 7 kg/m2 37 in body mass 
February 2012 5-344 Draft – Do Not Cite or Quote

1 index (Akesson et al., 2005). At current blood Pb levels in the U.S. adult population, a 

2 downward shift in kidney function of the entire population due to Pb may not result in 

3 CKD in identifiable individuals; however, that segment of the population with the lowest 

4 kidney reserve may be at increased risk for CKD when Pb is combined with other kidney 

5 risk factors. For example, in adults with mean (concurrent or baseline measured 4-6 years 

6 before kidney function tests) blood Pb levels that are comparable to that of the general 

7 U.S. population (1.6 to 4.2 μg/dL), higher blood Pb level was found to be associated with 

clinically-relevant effects (e.g., eGFR < 60 mL/min/1.73 m2 8 , doubling of serum 

9 creatinine) (Fadrowski et al., 2010; Yu et al., 2004) and larger magnitudes of effect in 

10 potentially at-risk populations with cormorbidities for CKD such as diabetes mellitus

11 (Tsaih et al., 2004) and hypertension (Tsaih et al., 2004; Muntner et al., 2003) or higher 

12 co-exposure to other environmental nephrotoxicants such as cadmium (Navas-Acien et 

13 al., 2009).

14 Recent NHANES analyses added to the evidence for Pb-associated lower renal function 

15 in populations with low concurrent mean blood Pb levels (< 2 μg/dL) (Fadrowski et al., 

16 2010; Navas-Acien et al., 2009). However, because of uncertainties concerning the 

17 magnitude, timing, frequency, and duration of Pb exposure that contributed to the 

18 observed associations, it is difficult to assess whether a threshold exists for Pb-related 

19 renal effects.

20 Research in the occupational setting has traditionally been far less consistent than that in 

21 environmentally exposed populations (Section 5.5.2.2). A number of explanatory factors 

22 for this inconsistency, all due to limitations of the occupational literature, were discussed 

23 in the 2006 Pb AQCD. The observation of paradoxical or inverse associations (higher Pb 

24 dose with lower serum creatinine, and/or higher eGFR or calculated or measured 

25 creatinine clearance) in several of these studies reflects limitations inherent in the study 

26 design. Irrespective of the mechanism, these associations have risk assessment 

27 implications. If associations are in opposite directions in different subgroups of the 

28 population and the relevant effect modifier is not considered, null associations will be 

29 observed. For these reasons, nonsignificant associations or paradoxical associations in the 

30 occupational setting cannot be used as a rationale for discounting Pb-related 

31 nephrotoxicity at lower environmental levels.

32 Important data on the effects of Pb on the kidney in children were reported in a recent 

33 NHANES analysis in adolescents, ages 12-20 years, which observed an association 

34 between higher concurrent blood Pb (mean: 1.5 μg/dL) and lower cystatin C-based eGFR

35 (Fadrowski et al., 2010). These findings are consistent with results from a rodent model 

36 study in which a low dose of Pb (50 ppm) administered from birth resulted in renal 

37 impairment (elevated serum creatinine as compared to control rats), but these 
February 2012 5-345 Draft – Do Not Cite or Quote

1 observations require confirmation by measurement of GFR and renal pathology (Berrahal 

2 et al., 2011). This recent epidemiologic study along with several previous studies that 

3 included children with higher Pb exposures (due to residence near sources, Pb poisoning, 

4 or parental occupational exposure) provide evidence that renal function in children may 

5 be affected by Pb exposure; however, additional research is warranted. It is important to 

6 recognize that the NHANES adolescents from Fadrowski et al. (2010) likely had higher 

7 Pb exposures earlier in childhood, thus, the magnitude, timing, frequency, and duration of 

8 Pb exposure contributing to the observed association is uncertain.

9 CKD results in substantial morbidity and mortality, and, even at earlier stages than those 

10 requiring kidney dialysis or transplantation, is an important risk factor for cardiac disease. 

11 As kidney dysfunction can increase BP and increased BP can lead to further damage to 

12 the kidneys, Pb-induced damage to either or both renal or cardiovascular systems may 

13 result in a cycle of further increased severity of disease. Pb exposure has been causally 

14 linked to both increased BP and other cardiovascular effects (Section 5.4). Interestingly, 

15 animal studies have shown Pb-induced vascular injury in the kidney was associated with 

16 increased glomerular sclerosis, tubulointerstitial injury, increased collagen staining, and 

17 an increase in macrophages associated with higher levels of MCP-1 mRNA. It is possible 

18 that the cardiovascular and renal effects of Pb observed are mechanistically linked and 

19 are contributing to the progression of the diseases. 

20 Recently available animal toxicological studies strengthen the evidence regarding the 

21 modes of action for Pb exposure leading to renal alterations, including the influence of 

22 Pb-induced oxidative stress. The mode of action of Pb in the kidneys has been extended 

23 to the field of immunology, evidenced by observations that Pb exposure resulted in 

24 infiltration of lymphocytes and macrophages associated with increased expression of NF25 κB in proximal tubules and infiltrating cells (Roncal et al., 2007). Additionally, recent 

26 findings expand on the evidence of acute effects of Pb, including mitochondrial 

27 dysfunction, renal cell apoptosis, and glomerular hypertrophy. These mechanisms are 

28 useful in understanding the occurrence of acute hyperfiltration followed by chronic 

29 kidney dysfunction. As indicated in Figure 5-38 and Table 5-22, studies found 

30 dysfunction in various kidney function measures, including urinary flow, ALP, 

31 microalbumin, and NAG in animals with blood Pb levels > 20 μg/dL (Wang et al., 

32 2010d). Lower concentration Pb exposures and lower blood Pb levels in animals have not 

33 been examined widely.

34 In summary, new epidemiologic and toxicological studies evaluated in the current review 

35 support or expand upon the strong body of evidence presented in the 2006 Pb AQCD 

36 indicating that Pb exposure is associated with renal effects. The weight of epidemiologic 

37 evidence demonstrates consistently a relationship between higher blood Pb level and 
February 2012 5-346 Draft – Do Not Cite or Quote

1 kidney dysfunction (e.g., lower creatinine clearance, higher serum creatinine, and lower 

2 GFR) in nonoccupationally-exposed adults with mean concurrent or baseline blood Pb 

3 levels of 2-10 μg/dL. A few analyses find higher blood Pb levels to be associated with a 

4 greater longitudinal decrease in kidney function over time (4-15 years), suggesting that 

5 past Pb exposures may contribute to ongoing renal effects. The epidemiologic evidence is 

6 strengthened by associations between Pb biomarker levels and renal function that were 

7 observed after adjustment for multiple potential confounding factors such as age, sex, 

8 comorbid cardiovascular conditions, BMI, smoking, and alcohol use. Similar findings 

9 among adults with CKD indicate that in populations with pre-existing renal disease, Pb 

10 exposure may be associated with greater progression of disease. Because blood Pb level 

11 in nonoccupationally-exposed adults reflects both recent and past Pb exposures, the 

12 magnitude, timing, frequency, and duration of Pb exposure contributing to the observed 

13 associations is uncertain. Coherence for epidemiologic findings is provided by 

14 observations in animal models that Pb exposure for greater than 6 months decreases GFR 

15 and increases serum creatinine. The weight of evidence in animal studies indicates Pb16 induced histopathological changes, including tubular atrophy and sclerosis. Overall, 

17 reduced renal function and increased kidney damage in animals are observed with 

18 chronic Pb (> 4 weeks) exposure that result in blood Pb levels > 20 μg/dL. By 

19 demonstrating Pb-induced renal oxidative stress, inflammation, mitochondrial 

20 dysfunction, apoptosis, and glomerular hypertrophy, toxicological studies provide 

21 biological plausibility for the associations observed in epidemiologic studies between 

22 blood Pb levels and kidney dysfunction. Collectively, the evidence integrated across 

23 epidemiologic and toxicological studies as well as across the spectrum of renal outcomes 

24 is sufficient to conclude that there is a causal relationship between Pb exposures and renal 

25 health effects.

5.6 Immune System Effects

5.6.1 Introduction

26 With respect to studies conducted in laboratory animal and in vitro models, the immune 

27 effects of Pb exposure have been extensively examined over several decades. Animal 

28 studies of the effects of Pb exposure on host resistance date back to the 1960s while those 

29 focusing on Pb-induced immune functional alterations, including developmental 

30 immunotoxicity, were first conducted during the 1970s. Despite this long history of 

31 research, Pb-associated immune effects in animals with blood Pb levels in the range of 

32 current U.S. population levels (i.e., <10 μg/dL), particularly early in life, have been 

33 observed only relatively recently within the last 10-15 years (Dietert and McCabe, 2007).
February 2012 5-347 Draft – Do Not Cite or Quote

1 Over the last 10-15 years, advances in the understanding of Pb-associated changes in 

2 immunological parameters in humans have substantiated the immune effects of Pb. 

3 The 2006 Pb AQCD presented consistent evidence for immune system effects associated 

4 with Pb exposure (U.S. EPA, 2006b). Rather than producing overt cytotoxicity or 

5 pathology, Pb exposure was found to be associated with alterations in several subclinical 

6 parameters related to cellular and humoral immunity (Figure 5-42). These conclusions 

7 were based most heavily on toxicological evidence in animals and in vitro models for the 

8 direct effects of Pb exposure in inducing changes in a wide spectrum of immune 

9 outcomes, but principally, a shift away from T-derived lymphocyte helper (Th)1 

10 cytokines toward Th2 cytokines, suppression of Th1-dependent delayed type 

11 hypersensitivity (DTH), elevation in Th2-driven immunoglobulin E (IgE), and 

12 modulation of macrophages into a hyperinflammatory phenotype. Relatively fewer 

13 studies were available in humans and for fewer immune-related endpoints. A majority of 

14 studies in humans were conducted in Pb-exposed male workers, and while a larger 

15 number of immune outcomes were examined, the most consistent evidence comprised 

16 effects on neutrophil functionality. In a smaller body of studies of humans without 

17 occupational exposures, the weight of evidence indicated associations of higher blood Pb 

18 levels with lower abundance of T lymphocyte cells and, in concordance with 

19 toxicological studies, higher IgE levels in children. Due to limited examination, the 

20 immune effects of Pb exposure in adults without occupational exposures were not well 

21 characterized.
February 2012 5-348 Draft – Do Not Cite or Quote

Figure 5-42 Immunological pathways by which Pb exposure may increase risk 

of immune-related diseases.

1 The pathways by which Pb exposure may alter immune cell function and consequently 

2 increase the risk of immune-related diseases are presented in Figure 5-42. Both 

3 toxicological and epidemiologic studies of children indicated Pb exposure effects on T 

4 cells. A large body of toxicological evidence demonstrated Pb-induced effects on 

5 macrophages. Neutrophil functionality was found to be reduced with higher Pb exposure, 

6 based on studies of Pb-exposed workers with mean blood Pb levels > 40 μg/dL. 

7 Alterations in immune cells can lead to changes in cell-to-cell interactions, multiple 

8 signaling pathways, and inflammation that affect both innate and acquired immunity, that 

9 in turn, influence the risk of developing infectious, allergic and autoimmune diseases as 

10 well as exacerbating inflammatory responses in other organ systems. Studies conducted 

11 in animal and in vitro models provided consistent evidence for Pb exposure inducing 

12 effects on the range of immune effects presented in this continuum. Among the hallmarks 

13 reported for Pb-induced changes in functional pathways were: (1) a suppression of T14 derived lymphocyte helper (Th)1-driven cell-mediated immunity (as measured by a DTH 

15 response); (2) an increase in Th2-driven IgE antibody and Th2 cytokine production; and 

Increased risk of atopy

and allergic disease

Pb

Macrophages and 

Other Innate Immune Cells

Reduced:

Phagocytosis

Nitric oxide production

Peroxynitrite production

Lysosomal activity

Reduced host resistance 

to bacterial infection

Elevated TNF-α

Overproduction of ROS

Depleted antioxidant 

defenses

Damaged epithelia 

and mucosal barriers

Tissue damage 

and de novo 

antigen 

appearance

Removal of normal 

myelomonocytic

suppression

Inappropriate 

T cell proliferation 

activation

Increased risk of 

autoimmunity

Increased tissue 

inflammation 

(e.g. lung, gut, skin)

Dendritic Cells

Elevated IL-10

Suppressed IL-12

T Cells

Elevated IL-4 and IL-5

Suppressed INF-γ

Skewed 

Th2-biased

responses

Increased IgE

production

Increased risk of 

later-life cancer

Suppressed 

Th1- mediated 

anti-tumor 

host defense

Increased lipid 

and DNA oxidation 

in tissues

Increased tumor 

cell formation

B Cells

Increased risk of tissue 

inflammatory diseases
February 2012 5-349 Draft – Do Not Cite or Quote

1 (3) a pro-inflammatory shift in macrophage function. The latter was characterized by 

2 increased production of reactive oxygen species (ROS), prostaglandin E2 (PGE2), and 

3 inflammatory cytokines such as tumor necrosis factor-αlpha (TNF-α) and interleukin 

4 (IL)-6 and decreased production of IL-12 and nitric oxide (NO). While epidemiologic 

5 studies did not provide evidence for this full spectrum of immune effects, a shift to a Th2 

6 phenotype was indicated by associations observed between higher blood Pb level and 

7 higher serum IgE levels in children. 

8 Reflecting an inhibition of Th1 activity, toxicological evidence presented in the 2006 Pb 

9 AQCD linked Pb exposure of animals to impaired host resistance to bacteria (U.S. EPA, 

10 2006b). Indicating a hyperinflammatory state and local tissue damage, a few available 

11 toxicological studies found Pb exposure-induced generation of autoantibodies, suggesting 

12 an elevated risk of autoimmune reactions. Additionally, the shift toward a Th2 response 

13 suggested that Pb could elevate the risk of atopy and allergic responses. While 

14 toxicological evidence for Pb-induced Th2 activity, elevated IgE, and inflammation 

15 supported the biological plausibility of such effects, epidemiologic evidence was too 

16 sparse to draw conclusions regarding associations between blood Pb levels and these 

17 broader indicators of immune dysfunction in humans.

18 Changes in the spectrum of immune endpoints were associated with a wide range of 

19 blood Pb levels. Several toxicological studies found blood Pb levels in the range of 7-

20 100 μg/dL to be associated with juvenile and/or adult immune effects (e.g., suppressed 

21 DTH, elevated IgE, changes in cytokine levels). Most epidemiologic studies examined 

22 and found lower T cell abundance and higher serum IgE levels in association with 

23 population mean (or quantiles) concurrent blood Pb levels > 10 μg/dL.

24 With respect to critical lifestages of Pb exposure, animal studies provided strong 

25 evidence for immune effects induced by prenatal Pb exposures and by postnatal 

26 exposures in adult animals. There was uncertainty regarding critical lifestages of Pb 

27 exposure in humans as epidemiologic studies of children primarily were cross-sectional 

28 in design and examined concurrent blood Pb levels. Several other limitations of 

29 epidemiologic studies were noted, including small sample sizes; inconsistent adjustment 

30 for potential confounders such as age, sex, smoking, and comorbid conditions; and 

31 reliance on comparisons of immune endpoints among groups with different blood Pb 

32 levels, which provided limited information on the concentration-response function.

33 Collectively, the small numbers of toxicological and epidemiologic studies published 

34 since the 2006 Pb AQCD supported the previous findings of Pb-associated immune 

35 effects. Epidemiologic studies supported previous findings in children and provided new 

36 evidence for effects in nonoccupationally-exposed adults. Recent studies also expanded 

37 on the array of immunological parameters affected by Pb exposure as presented in Figure 
February 2012 5-350 Draft – Do Not Cite or Quote

1 5-42. For example, a new toxicological study indicated that Pb may modulate the 

2 function of dendritic cells. Results from new toxicological and epidemiologic studies 

3 strengthened the link between Pb-associated effects on immune cells and immune- and 

4 inflammatory-based diseases by providing evidence for changes in intermediary signaling 

5 and inflammatory pathways (Figure 5-42). Several new epidemiologic studies examined 

6 signaling molecules such as pro-inflammatory cytokines and NO to produce findings 

7 parallel with toxicological studies. New toxicological studies expanded knowledge of the 

8 broader role of Pb-associated immune modulation in mediating Pb effects in 

9 nonlymphoid tissues (e.g., in the nervous, reproductive, and respiratory systems). 

10 Although primarily cross-sectional in design, recent epidemiologic studies improved on 

11 the design of earlier studies through greater examination of children and adults with 

12 blood Pb levels more comparable to contemporary levels in the U.S. population and 

13 greater consideration of confounding by age, sex, smoking, comorbid conditions, and 

14 SES-related variables.

5.6.2 Cell-Mediated Immunity

5.6.2.1 T Cells

15 A majority of the evidence indicating effects of Pb exposure on T cells was provided by 

16 older toxicological and epidemiologic studies reviewed in the 2006 Pb AQCD (U.S. 

17 EPA, 2006b). Toxicological studies demonstrated Pb exposure-induced shifts in the 

18 partitioning of CD4+ (T helper) cell populations to favor Th2 cells (10-100 μM in vitro) 

19 and the production of Th2 cytokines and to suppress production of Th1 cytokines (wide 

20 range of Pb exposures) (Section 5.6.5.4). Previous epidemiologic findings were limited 

21 largely to associations between higher concurrent blood Pb level and lower T cell 

22 abundance in children.

23 Consistent with previous epidemiologic findings, in a recent study of 7 week-old Wistar 

24 rats administered 200 ppm Pb-acetate, the percentages of CD4+ and CD8+ cytotoxic T 

25 cells were decreased (with CD4-CD8- cells elevated) in the submaxillary lymph nodes, 

26 but only with i.p. Pb dosing (p <0.05) and not oral exposure (200 ppm Pb-acetate both 

27 routes of administration) (Teijon et al., 2010). Limited recent toxicological investigation 

28 added mechanistic information by indicating that Pb may induce a shift to Th2 responses 

29 via T cell-dependent and -independent pathways. Previously, in cultures of human CD4+ 

30 T cells, Pb (1 μM, 30 minutes) was shown to activate transcription factor NF-κB 

31 (regulates T cell activation) (Pyatt et al., 1996) and to increase, in a concentration32 dependent manner (10 and 50 μM PbCl2, 24 hours), the expression of MHC class II 
February 2012 5-351 Draft – Do Not Cite or Quote

1 surface antigens (e.g., HLA-DR), which mediate the CD4+ response to exogenous 

2 antigens (Guo et al., 1996b). Recent studies examined higher Pb exposure concentrations 

3 than did previous studies. Heo et al. (2007) recently showed that Pb (25 μM) blocked 

4 production of the Th1 cytokine interferon-γ (IFN-γ) in cultures of stimulated mouse T 

5 cells not by affecting gene expression but by suppressing translation of the protein. This 

6 blockage was rescued with the addition of IL-12, which promotes Th1 activity. These 

7 results demonstrated a T cell-dependent pathway to skewing toward Th2 responses. 

8 However, Pb was able to decrease the IFN-γ to IL-4 ratio (indicating a shift to Th2) in the 

9 absence of STAT6, the preferential signaling pathway for T cells. Thus, Pb also was 

10 found to skew toward Th2 responses via a T cell-independent pathway. Similar 

11 observations were made in vivo by Kasten-Jolly et al. (2010). In this study, 

12 developmental Pb exposure of mice (100 μM Pb-acetate in drinking water of dams from 

13 GD8 to PND21, resulting in pup blood Pb levels of 10-30 μg/dL) induced gene 

14 expression of IL-4 and suppressed production of IFN-γ in splenic cells. These changes 

15 occurred in the absence of STAT4 or STAT6 and occurred with concomitant increases in 

16 adenylate cyclase 8 and phosphatidylinositol 3-kinase, adding to the evidence that Pb 

17 may promote Th2 activity via T cell-independent pathways.

18 While a few available recent epidemiologic studies found associations of blood Pb levels 

19 with lower levels of Th1 cytokines and higher levels of Th2 cytokines in humans 

20 (Section 5.6.5.4), the extant evidence for effects on T cells in humans is derived largely 

21 from older studies describing differences in the abundance of several T cell subtypes that 

22 mediate acquired immunity responses to antigens. In most studies of children, higher 

23 blood Pb levels were associated with lower T cell abundance, primarily CD3+ cells. Such 

24 associations were observed in studies that adjusted for potential confounding factors (as 

25 described below) and studies that compared mean cell abundances among groups with 

26 different blood Pb levels. Blood Pb level was less consistently associated with lower 

27 abundance of other T cell subtypes such as CD4+ and CD8+.

28 The weight of evidence supported lower T cell abundance in association with concurrent 

29 blood Pb level and in groups of children with levels > 10 μg/dL (Zhao et al., 2004; 

30 Sarasua et al., 2000; Lutz et al., 1999). Associations were less consistent in comparisons 

31 of children with lower blood Pb levels. In analyses of 331 children in Germany, Karmaus 

32 et al. (2005) found that children (ages 7-10 years) with concurrent blood Pb levels 2.2-

33 2.8 μg/dL (2nd quartile) had a 9 to 11% lower abundance of several T cell subtypes (p < 

34 0.05, t-test) compared with children with blood Pb levels < 2.2 μg/dL (lowest quartile). 

35 Unlike other studies in children, Karmaus et al. (2005) considered potential confounding 

36 by adjusting for sex, age, number of infections in the past 12 months, passive smoke 

37 exposure, serum lipids, and serum organochlorine levels. Other studies that examined 
February 2012 5-352 Draft – Do Not Cite or Quote

1 blood Pb levels < 10 μg/dL (population mean < 2 μg/dL or quantile 5-9 μg/dL) did not 

2 find associations with T cell abundance (Hegazy et al., 2011; Belles-Isles et al., 2002).

3 Another study of U.S. (multiple unspecified locations) children that considered 

4 confounding also found an association between higher concurrent blood Pb level and 

5 lower T cell abundance; however, it was limited to the youngest subjects (Sarasua et al., 

6 2000). Among 241 children 6-35 months in age, a 1 μg/dL higher blood Pb level was 

7 associated with a 0.18% (95% CI: -0.34, -0.02) lower CD3+ cell count, a 0.10% (95% CI: 

8 -0.24, 0.04) lower CD4+ cell count, and a 0.04% (95% CI: -0.15, 0.07) lower CD8+ cell 

9 count, adjusting for city of residence, age, and sex. In older age groups (36-71 months, 6-

10 15 years), many effect estimates were positive. Analysis of blood Pb level categories 

11 indicated that associations were driven by lower T cell abundance (3-6%) among children 

12 6-35 months in age with blood Pb levels > 15 μg/dL. It is important to note that 76% of 

13 subjects lived near a Pb smelting operation. These subjects living near Pb sources likely 

14 had higher blood Pb levels and may have driven the observed associations. Neither 

15 Karmaus et al. (2005) nor Sarasua et al. (2000) found a monotonic decrease in T cell 

16 abundance across blood Pb level groups. Neither of these studies adjusted for SES, which 

17 has been associated with blood Pb levels and immune-related conditions such as asthma, 

18 allergy, and viral infections. However, it is difficult to assess the potential for 

19 confounding by SES as neither study reported the SES characteristics of the study 

20 population.

21 In the limited investigation of nonoccupationally-exposed adults, higher concurrent blood 

22 Pb levels were associated with higher T cell abundance (Boscolo et al., 2000; Sarasua et 

23 al., 2000; Boscolo et al., 1999); however, occupational studies conducted in the U.S. and 

24 Asia did not find Pb-exposed workers consistently to have lower or higher abundance of 

25 T cells (Mishra et al., 2010; Pinkerton et al., 1998; Yucesoy et al., 1997b; Undeger et al., 

26 1996; Fischbein et al., 1993). Some studies found that compared with unexposed 

27 controls, Pb-exposed workers had a lower ratio of CD4+/CD8+ cells (Mishra et al., 2010; 

28 Fischbein et al., 1993). In particular, Fischbein et al. (1993) found this lower ratio in New 

29 York area firearms instructors with a relatively lower mean blood Pb level of 14.6 μg/dL 

30 and after adjusting for age and smoking. Changes in the CD4+/CD8+ ratio have not been 

31 examined in populations with lower blood Pb levels.

32 Although several epidemiologic studies have found lower T cell abundance in association 

33 with higher blood Pb levels, after adjusting for a range of potential confounding factors, it 

34 is not clear what effects these small magnitudes of change may have on the cell-to-cell 

35 interactions that mediate downstream acquired immune responses. In children, groups 

36 with higher blood Pb level had lower CD3+ cell abundance that ranged between 1 and 

37 9%. Larger decreases (20-35%) were observed in studies of occupationally-exposed 
February 2012 5-353 Draft – Do Not Cite or Quote

1 males with higher blood Pb levels than those found in the general U.S. population (means 

2 15 and 75 μg/dL) (Undeger et al., 1996; Fischbein et al., 1993).

3 In summary, a majority of the evidence indicating effects of Pb exosure on T cells was 

4 provided by previous toxicological and epidemiologic studies reviewed in the 2006 Pb 

5 AQCD. Previous toxicological studies demonstrated Pb-induced expansion of Th2 cells 

6 and increased Th2 cytokine production. A small number of new toxicological studies 

7 expanded the evidence by describing that Pb-induced Th2 skewing may occur via T cell8 dependent pathways (Heo et al., 2007) and -independent pathways (Kasten-Jolly et al., 

9 2010; Heo et al., 2007). Previous studies did not provide consistent evidence that higher 

10 blood Pb levels or Pb exposure was associated with lower T cell abundance in 

11 nonoccupationally-exposed or occupationally-exposed adults, respectively. A new study 

12 of children did not find children with higher blood Pb level to have lower T cell 

13 abundance (Hegazy et al., 2011). Epidemiologic findings were limited largely to 

14 associations between higher concurrent blood Pb level (> 10 μg/dL) and lower T cell 

15 abundance observed in previous studies of children (Karmaus et al., 2005; Zhao et al., 

16 2004; Sarasua et al., 2000; Lutz et al., 1999).

5.6.2.2 Lymphocyte Activation

17 A majority of the evidence indicating that Pb exposure stimulates lymphocyte activation 

18 is provided by previous toxicological studies in which exposures to high concentrations 

19 of Pb (10-100 μM) induced an expansion of alloreactive B and T lymphocytes (U.S. 

20 EPA, 2006b). Lymphocyte activation occurs as a result of reversing the normal 

21 suppression that is mediated by a macrophage-like subpopulation. In the limited recent 

22 investigation of Pb-induced lymphocyte activation, toxicological studies characterized 

23 potential mechanisms underlying this effect. Gao et al. (2007) described a potential role 

24 for dendritic cells. Dendritic cells that matured in the presence of 25 μM PbCl2 promoted 

25 enhanced alloreactive T cell proliferation compared to control dendritic cells. In addition, 

26 using the local lymph node assay (LLNA), Carey et al. (2006) found that PbCl2 (injection 

27 doses 25-50 μg) was able to provide a costimulatory signal to antigens that could activate 

28 T cells in adult female mice. The exact mechanistic basis for this is not known. As 

29 discussed in Section 5.6.5.2, changes in NO production appear to be involved in Pb30 induced lymphocyte activation (Farrer et al., 2008).

31 The available epidemiologic evidence for Pb-associated lymphocyte activation was 

32 provided by a small number of previous studies in children and nonoccupationally33 exposed adults. Instead of directly measuring lymphocyte proliferation, these studies 

34 provided indirect evidence by measuring the abundance of cells that expressed HLA-DR, 
February 2012 5-354 Draft – Do Not Cite or Quote

1 an indicator of activated cells. It is important to note that HLA-DR+ cells also may 

2 indicate the presence of activated monocytes. Further, these studies did not adjust 

3 extensively for potential confounding. In the only study of children (ages 9 months-6 

4 years, Missouri), the mean percentage of HLA-DR+ cells was approximately 2-fold 

5 higher (p > 0.05, Kruskal-Wallis) in the 19 children with concurrent blood Pb levels 15-

6 19 μg/dL than in children with blood Pb levels 10-14 μg/dL (n = 61) or < 10 μg/dL 

7 (n = 178) after adjusting for age (Lutz et al., 1999). However, activated cells were not 

8 elevated in children with blood Pb levels 20-44 μg/dL. Studies of nonoccupationally9 exposed adults in Italy found that concurrent blood Pb level was correlated positively 

10 with the percentage of HLA-DR expressing cells in men with and without allergies 

11 (Spearman r = 0.51, p < 0.002, n = 17 each, median blood Pb level both groups 

12 combined: 11 μg/dL) (Boscolo et al., 1999) but only in women without allergies 

13 (Spearman r = 0.44, p < 0.05, n = 25, median blood Pb level: 5.5 μg/dL) (Boscolo et al., 

14 2000).

15 Comparisons of Pb-exposed workers and unexposed controls indicated similar levels of 

16 lymphocyte proliferation (≤ 1% difference) between groups (Queiroz et al., 1994b; 

17 Cohen et al., 1989) or lower lymphocyte proliferation among Pb-exposed workers (8-

18 25%) (Mishra et al., 2003; Fischbein et al., 1993; Alomran and Shleamoon, 1988; Kimber 

19 et al., 1986) Toxicological studies have demonstrated the selective expansion of Th2 cells 

20 and suppression of Th1 cells (U.S. EPA, 2006b). Therefore, the differential activation of 

21 specific subtypes may not be discernable in epidemiologic studies that measure overall 

22 lymphocyte proliferation. Additionally, because occupational studies did not provide 

23 concentration-response information, it is difficult to infer whether findings apply to 

24 populations with lower blood Pb levels. Toxicological studies also did not provide 

25 information on Pb-induced lymphocyte activation at exposure concentrations relevant to 

26 humans without occupational Pb exposures.

5.6.2.3 Delayed-type Hypersensitivity

27 Although recent investigation was limited, a large body of previous toxicological studies 

28 reviewed in the 2006 Pb AQCD (U.S. EPA, 2006b) and recent reviews (Mishra, 2009; 

29 Dietert and McCabe, 2007) identified a suppressed DTH response as one of the most 

30 consistently observed and well-documented immunomodulatory effects of Pb exposure in 

31 animal models. A new study indicated that this effect may be mediated by dendritic cells. 

32 The DTH assay is commonly used to assess the T cell-mediated adaptive immune 

33 response, i.e., induration and erythema resulting from the activation of T cells and 

34 recruitment of monocytes to the site of antigen deposition. The DTH response is largely 

35 Th1-dependent in that Th1 cytokines drive the production of antigen-specific T cells 
February 2012 5-355 Draft – Do Not Cite or Quote

1 directed against the antigen (sensitizing phase) and the recruitment of antigen-specific T 

2 cells and monocytes to the site of antigen deposition (elicitation phase).

3 Previous toxicological studies demonstrated suppressed DTH responses in animals after

4 gestational (Chen et al., 2004; Bunn et al., 2001a; Bunn et al., 2001b, c; Lee et al., 2001b; 

5 Chen et al., 1999; Miller et al., 1998; Faith et al., 1979) and postnatal (McCabe et al., 

6 1999; Laschi-Loquerie et al., 1984; Muller et al., 1977) Pb exposures. Blood Pb level 

7 data were not available in all studies; however, DTH was suppressed in animals with a 

8 wide range of blood Pb levels (11 to > 100 μg/dL), with the associations of DTH with 

9 lower blood Pb levels occurring as a result of gestational exposure. 

10 In some studies that examined Pb exposures at multiple stages of gestation, exposures 

11 later in gestation suppressed DTH in animals (Bunn et al., 2001c; Lee et al., 2001b).

12 These latter findings may reflect the status of thymus and T cell development. A recent 

13 study contributed to the robust evidence by indicating a role for dendritic cells in the Pb14 induced suppression of the DTH response. Gao et al. (2007) exposed bone marrow15 derived dendritic cells in vitro to PbCl2 (25 μM, 10 days) then the antigen ovalbumin 

16 (OVA) and injected the cells into naïve adult mice. Mice treated with Pb-exposed 

17 dendritic cells had a diminished OVA-specific DTH footpad response compared with 

18 mice treated with non Pb-exposed dendritic cells.

19 The capability of Pb to suppress the DTH response is strongly supported by mechanistic 

20 studies in which Pb suppresses Th1 cytokine production (Section 5.6.5.4). In some 

21 animal studies, the suppressed DTH response was accompanied by a decreased 

22 production of IFN-γ (Lee et al., 2001b; Chen et al., 1999), which is the primary cytokine 

23 that stimulates recruitment of macrophages, a key component of the DTH response. 

24 Observations of a concomitant decrease in IFN-γ further link Pb-induced inhibition of 

25 Th1 functional activities with suppression of the DTH response. Further, coherence is 

26 provided by associations observed between Pb exposure or blood Pb levels and other 

27 responses in animals related to the inhibition of Th1-driven adaptive immune responses, 

28 including decreased host resistance (Section 5.6.4.1). 

5.6.2.4 Macrophages and Monocytes

29 As reported in the 2006 Pb AQCD, based on a large body of toxicological evidence, Pb30 induced promotion of a hyperinflammatory phenotype in macrophages was considered to 

31 be a hallmark of Pb-associated immune effects (U.S. EPA, 2006b). Pb-induced 

32 hyperinflammation was indicated by the enhanced production of ROS, suppressed 

33 production of NO, enhanced production of TNF-α, excessive metabolism of arachidonic 

34 acid into immunosuppressive metabolites (e.g., PGE2), impaired growth and 
February 2012 5-356 Draft – Do Not Cite or Quote

1 differentiation of cells, and potentially altered receptor expression [e.g., toll-like 

2 receptors]). Several of these findings are described in detail in Sections 5.6.5.2 and 

3 5.6.5.4. Because macrophages are major resident populations in most tissues and organs 

4 and also are highly mobile in response to microbial signals and tissue alterations, their 

5 functional impairment in response to Pb exposure may serve as a link between Pb6 induced immune effects and impaired host defense, tissue integrity, and organ 

7 homeostasis in numerous physiological systems (Section 5.6.4.5). A small number of 

8 available recent studies continue to support previous findings. A study in Pb-treated 

9 (40 ppm by oral gavage, once daily for 40 days) adult mice reported decreased 

10 macrophage yield and viability, and phagocytic index in the kidney (Lodi et al., 2011). In 

11 addition, several studies reinforced the effect of Pb exposure to alter a broad spectrum of 

12 functional alterations in human monocytes (the blood form of macrophages) or mouse 

13 macrophages in vitro (Khan et al., 2011; Bussolaro et al., 2008; Mishra et al., 2006a). In 

14 particular, Bussolaro et al. (2008) found reduced phagocytic activity with a relatively low 

15 concentration of Pb exposure (0.2 μM Pb nitrate, 72 hours). Khan et al. (2011) found that 

16 exposure of human monocytes to 25-100 μM Pb-acetate for 3-6 hours induced increases 

17 in the pro-inflammatory cytokine TNF-α. Mishra et al. (2006a) found that 100 ppm

18 Pb-acetate suppressed lipopolysaccharide (LPS)-induced NO production.

19 Epidemiologic studies have not widely examined the effects of Pb exposure on altered 

20 macrophage functional activity in humans, and the indices of macrophage function have 

21 varied among the few available studies. Pineda-Zavaleta et al. (2004) was unique in 

22 examining the hyperinflammatory state specifically in macrophages, albeit in relation to 

23 blood Pb levels higher than those in the current U.S. population (range of blood Pb 

24 levels: 3.5-47.5 μg/dL). This study included children in Lagunera, Mexico, attending 

25 schools at varying distances from an active Pb smelter. Consistent with the large body of 

26 toxicological evidence, higher concurrent blood Pb level was associated with lower NO 

27 production and higher superoxide anion production in macrophages isolated from child 

28 sera (Section 5.6.5.2). Model covariates included sex, age, and presence of allergies.

29 Other studies in humans examined occupationally-exposed adults and did not find a clear 

30 association between concurrent measurements of blood Pb level and macrophage 

31 abundance. Adjusting for age, race, smoking, and workshift, Pinkerton et al. (1998) found 

32 a lower abundance of monocytes among 145 U.S. Pb smelter workers with a mean blood 

33 Pb level of 39 μg/dL (7.8%) than among 84 unexposed controls with a mean blood Pb 

34 level of <2 μg/dL (8.5%) (p = 0.03). Conterato et al. (In Press) examined a group of male 

35 Pb-exposed painters with blood Pb levels ranging between 1.4 and 14.0 μg/dL (mean: 

36 5.4 μg/dL), lower than those in other occupational studies. The mean level and 

37 percentage of monocytes in the 50 painters were similar to those in the 36 controls (mean 

38 blood Pb level: 1.5 μg/dL) and in the 23 battery workers with much higher blood Pb 
February 2012 5-357 Draft – Do Not Cite or Quote

1 levels (mean: 50 μg/dL). Fischbein et al. (1993) found lower (p < 0.001) abundance of 

2 HLA-DR+ cells in two groups of New York metropolitan area firearms instructors with 

3 mean blood Pb levels of 14.6 μg/dL (n = 36, 8.8%) and 31.4 μg/dL (n = 15, 8.7%) than 

4 among the 36 unexposed controls (15.2%). HLA-DR+ is an indicator of activated 

5 functional state of antigen presenting cells (APCs, e.g., macrophages, dendritic cells, T 

6 cells) and is upregulated in response to cell signaling. While this study did not provide 

7 direct evidence for Pb effects on macrophages, the investigators cited previous 

8 observations that 70% of monocytes express HLA-DR antigen compared with 15% of 

9 lymphocytes to suggest that the reduced expression of HLA-DR antigen in Pb-exposed 

10 workers was due primarily to a reduction in activated monocytes. 

11 In summary, a small body of new in vitro studies (Khan et al., 2011; Bussolaro et al., 

12 2008; Mishra et al., 2006a) adds to the extensive base toxicological evidence indicating 

13 that Pb exposure decreases functionality of macrophages and promotes a 

14 hyperinflammatory phenotype. The sparse epidemiologic data are provided primarily by 

15 older studies and are not conclusive. Whereas an association between blood Pb level and 

16 a hyperinflammatory state of macrophages was observed in a previous study of children

17 (Pineda-Zavaleta et al., 2004), studies of occupationally-exposed adults did not clearly 

18 indicate Pb exposure effects on macrophages. A new study did not find large differences 

19 in monocyte abundance between Pb-exposed workers and unexposed controls (Conterato 

20 et al., In Press).

5.6.2.5 Neutrophils

21 In the 2006 Pb AQCD, Pb exposure was not judged to have strong effects on neutrophils

22 (U.S. EPA, 2006b). This conclusion was based on the relatively limited available 

23 toxicological evidence as compared with that for effects on other immune cells. However, 

24 the modulation of neutrophil activity may have important consequences on the 

25 dysregulation of inflammation and ability of organisms to respond to infectious agents. 

26 Studies of cultured human polymorphonuclear cells (PMNs) (Governa et al., 1987) and 

27 occupationally-exposed adults (Queiroz et al., 1994a; Queiroz et al., 1993; Valentino et 

28 al., 1991; Bergeret et al., 1990) found Pb-associated reductions in PMN functionality, as 

29 indicated by reduced chemotactic response, phagocytic activity, respiratory oxidative 

30 burst activity, or reduced ability to kill ingested antigen. Important limitations to applying 

31 epidemiologic findings broadly include male-only study populations, the relatively high 

32 blood Pb levels of workers (range of mean levels: 33.1-71 μg/dL), lack of concentration33 response information for associations between blood Pb level and neutrophil function, 

34 and lack of consideration of potential confounding factors.
February 2012 5-358 Draft – Do Not Cite or Quote

1 Instead of examining neutrophil functional activities, a few available recent studies of 

2 animals and occupationally-exposed adults examined the effects of Pb exposure on 

3 neutrophil counts, an increase in which has been interpreted by some investigators to be a 

4 compensatory response to Pb-induced impairment in neutrophil chemotactic activity and 

5 a hyperinflammatory response. In male rats with Pb spheres implanted in their brains, 

6 neutrophils were a major responding cell (Kibayashi et al., 2010). Compared with rats 

7 implanted with glass spheres, rats with Pb spheres had greater neutrophil infiltration with 

8 inflammatory-related damage that included apoptosis and indications of 

9 neurodegeneration. It is important to acknowledge the uncertain relevance of these 

10 findings to those expected from typical routes of Pb exposure in humans.

11 In a group of 68 ceramic, Pb recycling, or bullet manufacturing workers and 50 controls 

12 selected among food plant workers, DiLorenzo et al. (2006) provided information on the 

13 concentration-response relationship and adjusted for potential confounding variables. 

14 Among all subjects, a 1 μg/dL higher concurrent blood Pb level was associated with a 

15 21.8 cells/μL (95% CI: 11.2, 32.4 cells/μL) higher absolute neutrophil count (ANC) 

16 adjusted for age, BMI, and smoking status. The geometric mean (range) of concurrent 

17 blood Pb levels was 20.5 μg/dL (range: 3.2 to 120 μg/dL) among workers and 3.5 μg/dL 

18 (range: 1 to 11 μg/dL) among controls. Eight workers described to have medium to high 

19 Pb exposures (exact blood Pb levels not reported), but no controls had neutrophilia 

(n >7,500 cells/mm3 20 ), suggesting that chronic, higher-level Pb exposures can lead to a 

21 biologically meaningful excess of circulating neutrophils. Additionally, in analyses 

22 comparing three blood Pb level groups, controls, workers with blood Pb levels ≤ 

23 30 μg/dL, and workers with blood Pb levels >30 μg/dL, ANC was observed to increase 

24 monotonically across increasing blood Pb groups, supporting a blood Pb concentration25 dependent relationship. When the three blood Pb groups were further stratified by current 

26 smoking, two-way ANOVA indicated an interaction between concurrent blood Pb level 

27 and current smoking. Higher blood Pb level was associated with higher ANC only in 

28 current smokers. Among nonsmokers, ANCs were similar across blood Pb groups. In 

29 contrast, Conterato et al. (In Press) found lower neutrophil concentrations among two 

30 groups of Pb-exposed male workers with mean concurrent blood Pb levels of 50.0 and 

31 5.4 μg/dL than among controls with a mean blood Pb level of 1.5 μg/dL. Pb-exposed 

32 workers did not consistently have higher levels of other immune cells such as 

33 eosinophils, basophils, monocytes, or total lymphocytes either.

34 Additional evidence for the effects of Pb exposure on neutrophils is provided by findings 

35 that blood Pb level is associated with mediators of neutrophil proliferation, survival, 

36 maturation, and functional activation. These mediators include cytokines such as TNF-α 

37 (Section 5.6.5.4) and complement. The complement system is a component of the innate 

38 immune system that controls various cell-mediated immune responses such as 
February 2012 5-359 Draft – Do Not Cite or Quote

1 chemotaxis of macrophages and neutrophils and phagocytosis of antigens. The effects of 

2 Pb exposure on complement have not been widely examined; however the limited data 

3 suggest Pb may suppress complement activity. Both Ewers et al. (1982) and Undeger et 

4 al. (1996) measured lower serum complement C3 protein among Pb-exposed workers 

5 compared with unexposed controls, with Ewers et al. (1982) additionally observing an 

6 association between higher concurrent blood Pb level and lower C3 in a regression 

7 analysis combining Pb-exposed workers and controls. However, the broad applicability 

8 of these findings may be limited due to the high blood Pb levels in these occupationally9 exposed groups (range of blood Pb levels: 18.6-85.2 μg/dL and 38-100 μg/dL, 

10 respectively) and by the lack of adjustment for potential confounding variables.

11 In summary, a majority of evidence indicating the effects of Pb exposure on neutrophils 

12 was provided by previous studies that found that compared with unexposed controls, Pb13 exposed workers had lower neutrophil functionality (Queiroz et al., 1994a; Queiroz et al., 

14 1993; Valentino et al., 1991; Bergeret et al., 1990) and lower complement, which is a 

15 mediator of phagocyte functionality (Undeger et al., 1996; Ewers et al., 1982). In the 

16 limited new epidemiologic investigation that examined neutrophil abundance, studies did 

17 not conclusively find Pb-exposed workers to have higher or lower neutrophil abundance. 

18 Overall, the epidemiologic evidence is limited by the high blood Pb levels (range of mean 

19 levels: 33.1 to 71 μg/dL) with which neutrophil functionality was observed to be 

20 decreased and the lack of consideration of potential confounding.

5.6.2.6 Dendritic Cells

21 Since the publication of the 2006 Pb AQCD, new results (from both an ex vivo and in 

22 vitro models) suggest that the effects of Pb exposure on suppressing Th1 activity and 

23 promoting Th2 activity may be a consequence of the direct action of Pb on the function 

24 of dendritic cells (a major APC). Prior research on the effects of Pb in favoring Th2 over 

25 Th1 activity emphasized the direct measurement of Th1 versus Th2 T-cell populations 

26 and cytokine profiles. But new research techniques have been developed (Gao and 

27 Lawrence, 2010) that provide an opportunity to look upstream at how dendritic cells may 

28 be involved in mediating the effects of Pb on acquired immunity. Gao et al. (2007) used 

29 bone marrow cultures exposed to Pb to examine the impact of Pb on dendritic cell 

30 maturation and function. PbCl2 (25 μM, 10 days) was found to alter the course of 

31 dendritic cell maturation by changing the ratio of cell surface markers, such as the 

32 CD86/CD80 ratio, that promote Th2 cell development. Additionally, upon activation with 

33 LPS, Pb-matured dendritic cells produced less IL-6, TNF-α, and IL-12 (stimulates growth 

34 and differentiation of T cells) than did control cells but the same amount of IL-10 

35 (inhibits production of Th1 cytokines). The effect of Pb in altering the cytokine 
February 2012 5-360 Draft – Do Not Cite or Quote

1 expression profile of dendritic cells, in particular, the lower IL-12/IL-10 ratio, may serve 

2 as an important signal to shift naïve T cell populations toward a Th2 phenotype. 

3 Strengthening the role of dendritic cells in mediating Pb immune effects were ex vivo 

4 results from the same study which showed that Pb-naïve adult BALB/c mice implanted 

5 with Pb-exposed dendritic cells were skewed toward Th2 activity as indicated by 

6 inhibited DTH (Section 5.6.2.3) and IgG2a antibody (Section 5.6.3) responses (Gao et al., 

7 2007).

5.6.2.7 Natural Killer Cells

8 Based mostly on epidemiologic studies, previous and recent evidence does not clearly 

9 indicate that the innate immune natural killer (NK) cells are affected to a large extent by 

10 Pb exposure. In previous studies of nonoccupationally-exposed adults in Italy, concurrent 

11 blood Pb levels were correlated with NK cell abundance among men with and without 

12 allergy (Spearman r = 0.49, p ~ 0.05, n = 17 each, median blood Pb level both groups 

13 combined: 11 μg/dL) (Boscolo et al., 1999) and women without allergies (Spearman r = 

14 0.44, p < 0.05) (Boscolo et al., 2000) In a study of children from St. Lawrence River 

15 communities in Quebec, Canada, cord blood Pb level was not significantly associated 

16 with cord blood NK cell abundance (quantitative results not reported (Belles-Isles et al., 

17 2002). Similar means of NK cell abundance or functional activity were observed in Pb18 exposed workers and unexposed controls (García-Lestón et al., 2011; Mishra et al., 2003; 

19 Pinkerton et al., 1998; Yucesoy et al., 1997b; Undeger et al., 1996; Fischbein et al., 1993; 

20 Kimber et al., 1986). The lack of evidence for a strong effect on NK cells is underscored 

21 by epidemiologic observations within individual studies that blood Pb levels are 

22 associated with T cell abundance but are not associated with NK cell abundance or their 

23 level of functional activity (Karmaus et al., 2005; Sarasua et al., 2000; Pinkerton et al., 

24 1998). Consistent with the epidemiologic evidence, toxicological evidence is not 

25 conclusive. In a recent in vitro study comparing the toxicities of metals for different 

26 populations of immune cells, Fortier et al. (2008) found that PbCl2 (7.5-20.7 μg/dL) did 

27 not affect NK cytotoxicity compared with the DMSO vehicle. However, PbCl2 was not 

28 found to affect other immune parameters (e.g., monocyte phagocytic activity or 

29 lymphocyte proliferation) either. A recent study did show a decrease in NK cell activity 

30 in mice albeit with high-level Pb exposure (1,300 ppm Pb-acetate in drinking water, 10 

31 days, blood Pb level ~100 μg/dL) (Queiroz et al., 2011).
February 2012 5-361 Draft – Do Not Cite or Quote

5.6.3 Humoral Immunity

1 The 2006 Pb AQCD described another hallmark effect of Pb on the immune system to be 

2 an enhanced humoral immune response as characterized by increased production of IgE 

3 antibodies (U.S. EPA, 2006b). Several previous toxicological and epidemiologic studies 

4 (Table 5-28) demonstrated Pb-associated increases in IgE production, which is strongly 

5 implicated in mediating allergic responses and inflammation in allergic asthma. In studies 

6 of juvenile rodents, Pb exposures (Pb-acetate) through drinking water from gestation 

7 through lactation induced concomitant increases in IgE and IL-4 production (a Th2 

8 cytokine) by T cells (Snyder et al., 2000; Chen et al., 1999), consistent with the 

9 hypothesis that Th2-mediated mechanisms can induce class switching of B cells to 

10 produce IgE. Evidence from both earlier toxicological and epidemiologic studies (Table 

11 5-28) did not consistently indicate that Pb exposure was associated with changes in other 

12 classes of Igs including IgG, IgM, and IgA. Previous epidemiologic studies of children 

13 (Table 5-28) and adults [Table 5-28 with group comparisons and (Boscolo et al., 2000; 

14 Boscolo et al., 1999) with correlation analyses] did not find a consistent association 

15 between blood Pb level and the abundance of B cells, which produce IgE and other 

16 antibodies.

17 Although previous evidence for effects on Ig classes other than IgE was inconsistent, a 

18 small number of available new toxicological studies found Pb-induced increases in IgG 

19 and pointed to a role for T cell-mediated mechanisms in Pb-induced activation of B cells 

20 and production of Ig antibodies. Important limitations of this evidence include high Pb 

21 exposures tested and use of injection routes of exposure that may have little relevance to 

22 human routes of exposure. Fernandez-Cabezudo et al. (2007) reported evidence for a 

23 subtle shift toward a Th2 immune response following Salmonella infection in mice 

24 exposed to high concentrations of Pb-acetate (5,000-10,000 μM, 16 weeks, resulting in 

25 blood Pb levels of 20.5-106 μg/dL). Serum levels of Salmonella-specific IgG1 antibodies 

26 were increased in the Pb-exposed mice compared to controls, whereas IgG2a levels were 

27 increased in control but not Pb-exposed mice following infection. The impaired Th1 and 

28 enhanced Th2 response also was evident by the decreased secretion of IL-12 and 

29 increased production of IL-4 by spleen cells taken from Pb-exposed (blood Pb levels 

30 > 20.5 μg/dL) mice (Fernandez-Cabezudo et al., 2007).

31 In a highly-specialized strain of knockout adult mice lacking the ability to produce IFN-γ, 

32 i.p. injection with 50 μg PbCl2 increased the IgG2a/IgG1 ratio (Gao et al., 2006). This 

33 result was surprising given evidence that IFN-γ usually directs secretion of IgG2a; 

34 however, the authors suggested that in these knockout mice, Pb may initiate a Th1 

35 response via an IFN-γ independent pathway to enhance IgG2a production. Other animal 

36 studies that administered Pb via the i.p., found Pb-induced humoral responses to 
February 2012 5-362 Draft – Do Not Cite or Quote

1 mediated preferentially via Th2 mechanisms. Carey et al. (2006) treated BALB/c mice 

2 with subsensitizing doses of a T cell-independent (Trinitrophenyl-Ficoll [TNP-Ficoll]) or 

3 T cell-dependent (TNP-ovalbumin [TNP-OVA]) hapten-protein conjugate with or 

4 without co-exposure to PbCl2. Seven days later, investigators examined the effects of Pb 

5 on the LLNA response to TNP-Ficoll or TNP-OVA. A bolus injection of PbCl2

6 (25-50 μg) increased the numbers of T and B cells in the lymph node against both TNP7 Ficoll and TNP-OVA. Further, in a concentration-dependent manner, Pb induced 

8 statistically significant elevations of IgM-, IgG2a-, and IgG1-producing cells in the 

9 lymph node. While the increase in IgM-producing cells against TNP-Ficoll indicated a T10 cell independent mechanism, the increases in IgG2a- and IgG1-producing cells against 

11 both antigens indicated a Th1- and Th2-mediated mechanism, respectively. Despite 

12 finding increases in both IgG1- and IgG2a-producing cells, the authors concluded that Pb 

13 skewed the response toward Th2 and had considerable potential for promoting allergic 

14 sensitization against T-dependent antigens.

15 In a recent microarray study in BALB/c mice, Kasten-Jolly et al. (2010) found that early16 life Pb exposure (100 μM Pb-acetate in drinking water of dams from GD8 to PND21, 

17 resulting in pup blood Pb levels 10-30 μg/dL) produced statistically significant increases 

18 in the expression of genes encoding Ig antibodies or those involved in B lymphocyte 

19 function and activation. These genes included those for the heavy chain of IgM, IL-4, IL20 7 and IL-7 receptor, IL-21, RAG-2, CD antigen 27, B-cell leukemia/lymphoma 6, RNA 

21 binding motif protein 24, Histocompatibility class II antigen A (beta 1), Notch gene 

22 homolog 2, and histone deacetylase 7A.

23 In epidemiologic studies, several of which are new, associations between biomarkers of 

24 Pb exposure and serum IgE level were demonstrated primarily in children, although a 

25 monotonic dose-dependent increase was not consistently observed (Hegazy et al., 2011; 

26 Hon et al., 2010; Hon et al., 2009; Karmaus et al., 2005; Annesi-Maesano et al., 2003; 

27 Sun et al., 2003; Lutz et al., 1999) (Table 5-28). Karmaus et al. (2005) had the most 

28 extensive adjustment for potential confounding factors and examined differences with 

29 lower blood Pb levels. Compared with children with concurrent blood Pb level 

30 <2.2 μg/dL (quartile 1), children with blood Pb levels 2.84-3.41 μg/dL (quartile 3) and 

31 >3.4 μg/dL (quartile 4) had 28% higher serum IgE levels (p = 0.03, F-test). These 

32 differences were observed after the adjustment for potential confounding by age, 

33 biomarkers of various organochlorine exposures, number of infections in the previous 12 

34 months, serum lipids, and passive smoke exposure. Similar differences in IgE count on 

35 basophils were not observed among the blood Pb quartiles; however, it is important to 

36 note that although serum IgE and basophil-bound IgE have been correlated in adults

37 (Malveaux et al., 1978; Conroy et al., 1977), little data are available in children (Dehlink 

38 et al., 2010). A recent study in children found that serum IgE levels were not correlated 
February 2012 5-363 Draft – Do Not Cite or Quote

1 with basophil-bound IgE but correlated (Spearman r = -0.003) with other IgE receptor2 expressing cells such as dendritic cells and monocytes (Spearman r = 0.43 to 0.65, p < 

3 0.05) (Dehlink et al., 2010). The number of IgE-bound basophils also has been found to 

4 be highly variable across individuals, particularly children (Hausmann et al., 2011; 

5 Dehlink et al., 2010). Thus, it is not unexpected that higher blood Pb level was associated 

6 with higher serum IgE but not basophil-bound IgE counts in Karmaus et al. (2005). In 

7 this study, blood Pb level was not associated with serum levels of IgG, IgA, and IgM or B 

8 cell abundance. Lutz et al. (1999) found higher serum IgE in children after adjusting for 

9 age, albeit with concurrent blood Pb levels > 10 μg/dL.

10 Recent studies in children also reported associations between concurrent blood Pb level 

11 and elevated serum IgE but did not adjust for potential confounding variables (Hegazy et 

12 al., 2011; Hon et al., 2010; Hon et al., 2009). The studies by Hon and colleagues (2010; 

13 2009) demonstrated associations in children with low blood Pb levels (range: 1.4-

14 6.0 μg/dL) and found that blood Pb level was correlated with both serum IgE and atopic 

15 dermatitis, a condition commonly characterized by elevated IgE levels. 
February 2012 5-364 Draft – Do Not Cite or Quote

Table 5-28 Comparison of serum immunoglobulin levels and B cell abundance 

among various blood Pb groups

Study Population/Study Details

Blood Pb 

Level 

Mean or 

Range 

(μg/dL) IgEa IgGb IgMb IgAb B cellsc

Children

Karmaus et al.

(2005)

331 children, ages 7-10 yr, Hesse, Germany

Results were adjusted for age, sex, passive smoke 

exposure, number of infections in the previous 12 

months, serum lipid concentration, and organochlorine 

exposures.

<2.2

2.21 - 2.83

2.84 - 3.41

>3.41

46 (1.0)

30 (0.65)

59 (1.28)

59 (1.28)d

1210

1214

1241

1201

150

143

153

148

123

121

133

136

418e (1.0)

353 (0.84)

389 (0.93)

393 (0.94)

Hegazy et al.

(2011)

318 children, ages 6 mo-7 yr, Egypt < 5

5-9

10 - 14

15 - 19

20 - 44

45 - 69

13.0 (1.0)

12.0 (0.92)

20.8 (1.60)

14.9 (1.15)

20.4 (1.57)

10.2 (0.78)d

Sarasua et al.

(2000)

382 children, ages 6-30 mo, Multiple U.S. locations

Results were adjusted for age, sex, and study location.

0.6 - 4.9

5 - 9.9

10 - 14.9

≥ 15

609 

666d

680d

630

103

108

105

124d

50.1

55.0

58.2

61.4d

19.1 (1.0)

20 (1.05)

20.4 (1.07)

22.2 (1.16)

Sarasua et al.

(2000)

562 children, ages 36-71 mo, Multiple U.S. locations

Results were adjusted for age, sex, and study location.

0.6 - 4.9

5 - 9.9

10 - 14.9

≥ 15

817 

813

856

835

120

116

125

121

88.6

90.9

96.3

94.1

18.4 (1.0)

17.6 (0.96)

19.2 (1.04)

18.6 (1.01)

Sarasua et al.

(2000)

675 children ages 5-16 yr, Multiple U.S. locations

Results were adjusted for age, sex, and study location.

0.6 - 4.9

5 - 9.9

10 - 14.9

≥ 15

1,031 

1,094d

1,048

1,221

128

131

136

106

140

143

140

108

16.1 (1.0)

15.8 (0.98)

15.3 (0.95)

20.1 (1.25)

Lutz et al.

(1999)

279 children, ages 9 mo-6 yr, Springfield, MO

Results were adjusted for age.

<10

10 - 14

15 - 19

20 - 44

51.8 (1.0)

74.0 (1.43)

210.7 (4.07)

63.7 (1.23)d

13.4 (1.0)

12.6 (0.94)

16.9 (1.26)

11.1 (0.83)

Zhao et al.

(2004)

75 children, ages 3-6 yr, Zhejiang Province, China <10

≥ 10

16.58 (1.0)

16.82 (1.01)

Adults without Occupational Pb Exposures

Sarasua et al.

(2000)

433 children and adults, ages 16-75 yr, Multiple U.S. 

locations

Results were adjusted for age, sex, study location, and 

smoking.

0.6-4.9

5-9.9

10-14.9

≥ 15

1,099 

1,085

1,231

1,169

175

175

262d

139

252

242

283

193

13.9 (1.0)

13.0 (0.94)

12.4 (0.89)

14.8 (1.06)

Adults with Occupational Pb Exposures

Fischbein et al.

(1993)

36 unexposed controls, mean age 47 yr

36 firearms instructors, mean age 49 yr

15 firearms instructors, mean age 48 yr

New York metropolitan area

NR

14.6

31.4

8.6 (1.0)

10.5 (1.22)

11.2 (1.30)d

Kimber et al.

(1986)

21 unexposed male controls, ages 20-60 yr

39 male tetraethyl Pb plant workers, ages 25-61 yr

U.K.

11.8

38.4

1062

1018

1294

1040

2235

2425
February 2012 5-365 Draft – Do Not Cite or Quote

Study Population/Study Details

Blood Pb 

Level 

Mean or 

Range 

(μg/dL) IgEa IgGb IgMb IgAb B cellsc

Pinkerton et al.

(1998)

84 unexposed controls, mean age 30 yr

145 Pb smelter workers, mean age 33 yr

U.S., exact location NR

Results were adjusted for age, race, current smoking 

status, and workshift.

<2

39

1,090 

1,110

94.5 

106.2

180

202

14.6 (1.0)

13.2 (0.90)

Heo et al.

(2004)

606 Pb battery plant workers

Korea

<10

10-29

≥ 30

112.5 (1.0)

223.3 (1.99)

535.8 

(4.76)d

Anetor and 

Adeniyi (1998)

50 male controls, ages 22-58 yr

80 male Pb-exposed workers, ages 21-66 yr, Nigeria

30.4

56.3

1,997

1,187d

215

191

188

144d

Ewers et al.

(1982)

53 male unexposed controls, ages 21-54 yr

72 male Pb battery/smelter workers, ages 16-58 yr, 

Germany

11.7

59.0

193f

171

161f

127

140f

128

Undeger et al.

(1996)

25 unexposed male controls, ages 22-56 yr

25 male Pb battery plant workers, ages 22-55 yr, 

Turkey

16.7

74.8

1,202.1

854.6d

140.4

93.3d

210.3

168.1

545.5e (1.0)

635.9 (1.17)

Alomran and 

Shleamoon

(1988)

18 unexposed age-matched controls

39 Pb battery workers, mean age 35 yr, Iraq

NR

NR

1713

1610

183

170

545.5e (1.0)

635.9 (1.17)

a

IgE data are presented as IU/mL. (In parentheses are the ratio of IgE in the higher blood Pb group to IgE in the lowest blood Pb group.) b

Other Ig data are presented as mg/dL unless otherwise specified.

c

B cell data are presented as the percentage of B cells among all lymphocytes unless otherwise specified. (In parentheses are the ratio of B cells in the 

higher blood Pb group to B cells in the lowest blood Pb group.) d

p <0.05 for group differences. e

Data represent the number of cells/μL serum. f

Data are presented as IU/mL.

1 Sarasua et al. (2000) found associations of higher concurrent blood Pb level with higher 

2 IgA, IgG, and IgM in U.S. children. Blood Pb level was associated with higher levels of 

3 all three Igs (0.8 [95% CI: 0.2, 1.4], 4.8 [95% CI: 1.2, 8.4], and 1.0 [95% CI: 0.1, 

4 1.9] mg/dL higher IgA, IgG, and IgM, respectively, per 1 μg/dL higher blood Pb level, 

5 adjusted for age, sex, and location) in the youngest age group (6-35 months) but not in 

6 older age groups (36-71 months, 6-15 years, 16-75 years), suggesting elevated risk in 

7 very young children. Among infants aged 6-35 months with concurrent blood Pb levels ≥ 

8 15 μg/dL, serum levels of all three examined Igs were elevated over levels measured in 

9 infants with blood Pb levels < 5 μg/dL. In this study, IgE was not examined. 

10 While most epidemiologic studies examined concurrent blood Pb levels, some studies 

11 indicated that prenatal Pb exposure may impact Ig levels in newborns (Annesi-Maesano 

12 et al., 2003; Belles-Isles et al., 2002). These studies also pointed to an increased risk in 

13 infants; however, important limitations of these studies are the lack of extensive 

14 consideration of potential confounding variables. Belles-Isles (2002) examined newborns 

15 in St. Lawrence River communities in Quebec, Canada and found an association between 
February 2012 5-366 Draft – Do Not Cite or Quote

1 higher cord blood Pb level and higher cord serum IgG in a regression analysis adjusted 

2 for prenatal smoke exposure. Annesi-Maesano et al. (2003) found that infant hair Pb level 

3 but not cord or placental Pb level was associated with cord serum IgE in newborns in 

4 Paris. From these findings, the authors inferred a stronger effect of Pb exposure 

5 integrated over the entire gestational period compared to exposures closer to birth. 

6 Cotinine was not associated significantly with Pb biomarker levels or with IgE. The 

7 magnitude of association was larger in the subgroup of infants with mothers without 

8 allergies (Spearman r = 0.21, p < 0.01 in infants with mothers without allergies versus r = 

9 0.12 in infants with mothers with allergies, pointing to the possible masking of a blood 

10 Pb association with IgE by the stronger association of family history of allergy. 

11 Associations between blood Pb level and IgE also were reported in studies of adults, 

12 without (Pizent et al., 2008) and with occupational Pb exposure (Heo et al., 2004). In a 

13 study of urban adults aged 19 to 67 years and of similar SES (i.e., white-collar office 

14 workers) in Zagreb, Croatia, a statistically significant association between higher 

15 concurrent blood Pb level and higher IgE was found in women but not men (Pizent et al., 

16 2008). Several covariates were considered in a stepwise multiple regression, including 

17 age, smoking intensity, and alcohol consumption. Among women not on hormone 

18 replacement therapy or oral contraceptives, a 1 μg/dL higher blood Pb level was 

19 associated with a 0.60 higher log of IgE (95% CI: 0.58, 1.18). Concurrent blood Pb levels 

20 were low in these women who were aged 19-67 years (mean: 2.16 μg/dL, range 0.56-

21 7.35 μg/dL); however, due to the cross-sectional nature of this study, it is difficult to 

22 characterize the timing, level, frequency, and duration of Pb exposure that contributed to 

23 the observed association. Because of likely higher past Pb exposures of adults and the 

24 mobilization of Pb from bone to blood, the associations may reflect effects of higher past 

25 Pb exposures. Investigators did not report an effect estimate in men because it did not 

26 attain statistical significance. Without quantitative results, it was difficult to ascertain 

27 whether there was suggestion of association in men but insufficient power to indicate 

28 statistical significance due to the smaller number of men examined (50 men versus 166 

29 females). Another study of 34 men in Italy also did not report quantitative results but 

30 indicated a lack of statistically significant correlation between blood Pb level and IgE in 

31 men (Boscolo et al., 1999).

32 A majority of the epidemiologic evidence for the effects of Pb on IgA, IgG, and IgM 

33 levels is provided by previous studies of Pb-exposed workers (Anetor and Adeniyi, 1998; 

34 Pinkerton et al., 1998; Undeger et al., 1996; Queiroz et al., 1994b; Alomran and 

35 Shleamoon, 1988; Kimber et al., 1986; Ewers et al., 1982). Consistent with the collective 

36 body of toxicological findings for these other Ig classes, epidemiologic evidence is 

37 mixed, with studies reporting higher, lower, and similar Ig levels in Pb-exposed workers 

38 compared with unexposed controls. Some studies reporting lower Ig levels in Pb-exposed 
February 2012 5-367 Draft – Do Not Cite or Quote

1 workers included workers with the highest mean blood Pb levels (> 50 μg/dL) (Anetor 

2 and Adeniyi, 1998; Undeger et al., 1996). The lack of rigorous statistical analyses in 

3 occupational studies precludes characterization of the factors that may contribute to 

4 inconsistent associations. 

5 In summary, a majority of the evidence for Pb exposure affecting humoral immunity is 

6 provided by previous toxicological and epidemiologic studies that found Pb-associated 

7 increases in IgE. Previous toxicological studies provided evidence for the independent 

8 effects of Pb exposure increasing IgE levels. A few available recent epidemiologic 

9 studies add to the large body of extant evidence supporting associations between higher 

10 concurrent blood Pb level and higher IgE levels in children; however, they did not adjust 

11 for potential confounding variables (Hegazy et al., 2011; Hon et al., 2010; Hon et al., 

12 2009). Collectively, epidemiologic evidence indicates higher IgE in children with 

13 concurrent blood Pb levels > 10 μg/dL. The epidemiologic evidence in children equally 

14 comprises studies that did not and did adjust for potential confounding. Studies that 

15 adjusted for potential confounding varied in the number and the specific variables 

16 included as model covariates (Table 5-28) but were consistent in finding an association 

17 between blood Pb and IgE. None of the studies in children adjusted for SES or allergen 

18 exposure. Lower SES, which may indicate poorer housing conditions, is associated with 

19 higher exposures to Pb as well as cockroach, rodent, and other allergens. Allergen 

20 exposure and lower SES are associated with higher IgE and IgE-related conditions such 

21 as allergies and asthma (Bryant-Stephens, 2009; Dowd and Aiello, 2009; Aligne et al., 

22 2000). Studies generally did not provide detailed demographic or residential information 

23 to assess whether SES and/or allergen exposure potentially confounded the observed 

24 associations between blood Pb level and IgE. Thus, uncertainty remains as to the extent 

25 to which blood Pb-IgE associations in children are confounded by unmeasured SES 

26 and/or allergen exposure. Evidence in adults is limited and thus, inconclusive. A new 

27 study of environmentally-exposed adults found an association between concurrent blood 

28 Pb level and IgE in women (Pizent et al., 2008). Evidence integrated across toxicological 

29 and epidemiologic studies does not consistently indicate that Pb exposure affects other 

30 classes of Igs; however, a few new toxicological studies found Pb-induced increases in 

31 IgG (Fernandez-Cabezudo et al., 2007; Gao et al., 2006) or increases in the expression of 

32 genes encoding Ig antibodies (Kasten-Jolly et al., 2010).
February 2012 5-368 Draft – Do Not Cite or Quote

5.6.4 Immune-based Diseases

5.6.4.1 Host Resistance

1 The capability of Pb to reduce host resistance of animals to bacteria has been known for 

2 almost 40 years and was supported by several animal studies described in the 2006 Pb 

3 AQCD (U.S. EPA, 2006b). A recent animal study provided supporting evidence for such 

4 an effect. Biological plausibility for these findings has been provided by observations that 

5 Pb affects mechanisms underlying diminished host resistance, e.g., suppressed 

6 Th1-driven acquired immune responses and increased inflammatory responses in target 

7 tissue resulting in further damage to host protective barriers. Host resistance to bacteria 

8 such as Listeria requires effective Th1-driven responses including the production of IL9 12 and IFN-γ (Lara-Tejero and Pamer, 2004) and these have been found to be inhibited 

10 by Pb exposure. The lack of IFN-γ can inhibit appropriate and timely macrophage 

11 activation. Nitric oxide is produced by macrophages during cellular activation and plays a 

12 role in host defense against bacterial infection (U.S. EPA, 2006b), and NO production 

13 also has been found to be suppressed by Pb exposure. A recent animal study 

14 characterized a potential mechanism by which Pb may impact both innate immune cells 

15 and natural host defense barriers. Kasten-Jolly et al. (2010) showed that developmental 

16 exposure of mice to Pb (100 μM Pb-acetate in drinking water of dams from GD8 to 

17 PND21, resulting in pup blood Pb levels of 10-30 μg/dL) resulted in an upregulation of 

18 splenic RNA of caspase-12, a cysteine protease that inhibits the clearance of bacteria both 

19 systemically and in the gut mucosa (Saleh et al., 2006).

20 In the few available epidemiologic studies, a range of indicators of Pb exposure (i.e., cord 

21 blood Pb, concurrent blood Pb, Pb content in total deposition samples, Pb content in 

22 lichen) were associated with viral and bacterial infections in children. Collectively, study 

23 limitations, including lack of consideration for potential confounding variables (Karmaus 

24 et al., 2005; Rabinowitz et al., 1990), lack of statistical analysis (Karmaus et al., 2005),

25 and ecological design (Carreras et al., 2009), limit the ability to draw inferences 

26 regarding the effects of Pb exposure on viral or bacterial infections in children. Similar to 

27 these studies in children, an occupational study finding higher frequency of self-reported 

28 colds or influenza among Pb battery or smelter plant workers with higher blood Pb levels 

29 than among unexposed controls is limited by a lack of statistical analysis (Ewers et al., 

30 1982). Studies that address the aforementioned limitations are needed to characterize the 

31 relationship between Pb exposure and resistance to bacterial and viral infection in 

32 humans.
February 2012 5-369 Draft – Do Not Cite or Quote

1 With limited investigation, the effect of Pb on host resistance to parasitic agents is 

2 unclear. The 2006 Pb ACQD described a study in which high-level Pb-exposed (≥ 

3 10 μM) mouse macrophages had diminished ability to kill Leishmania enrietti parasites

4 (Mauel et al., 1989). However, given the well-characterized effect of Pb in promoting 

5 Th2 activity, it is plausible for Pb to enhance host resistance to parasites that require 

6 robust Th2 responses (e.g., helminths) (U.S. EPA, 2006b). In a recent study, high-level 

7 Pb exposure (≥ 10 μM) enhanced host resistance to malaria (Koka et al., 2007). However, 

8 this was attributed to the capability of Pb to induce eryptosis and the rapid removal of 

9 malaria-infected erythrocytes and not to Pb-induced alterations in immune function. 

10 Nriagu et al. (2008) also reported that higher blood Pb level was associated with lower 

11 malaria prevalence among children (ages 2-9 years) from three Nigerian cities. A 

12 majority of children (75%) had blood Pb levels below 10 μg/dL. The association 

13 persisted after adjusting for age, sex, number of siblings, and other comorbidities such as 

14 headaches, depressed mood, and irritability.

5.6.4.2 Asthma and Allergy

15 Toxicological evidence and to a relatively limited extent, epidemiologic evidence, have 

16 supported the effects of Pb exposure on multiple immune parameters, including elevated 

17 production of Th2 cytokines such as IL-4, increased IgE antibody production (Table 

18 5-24), and increased inflammation. These are well-recognized pathways in the 

19 development and exacerbation of allergy and allergic disease, including asthma. This 

20 mechanistic evidence is coherent with a small body of epidemiologic studies that found 

21 associations of blood Pb levels with asthma or allergy in children (Figure 5-43 and Table 

22 5-29). Children examined in these studies encompassed a wide range of ages (i.e., less 

23 than 1 year to 12 years) and across studies, blood Pb was measured during different 

24 lifestages. This body of evidence in children included large studies with multivariate 

25 analyses and studies with prospective follow-up of subjects in which disease occurrence 

26 was ascertained after the measurement of blood Pb levels.
February 2012 5-370 Draft – Do Not Cite or Quote

Note: Results are organized by endpoint and for asthma in order of increasing blood Pb level. For analyses with blood Pb level as a 

continuous variable, odds ratios are standardized to a 1 μg/dL increase in blood Pb level. SPT = skin prick test, BR = bronchial 

responsiveness. Black diamond represents associations with concurrent blood Pb levels, green triangles represent associations with 

prenatal (cord) blood Pb levels, and blue circles represent associations with blood Pb levels measured in childhood up to 12 months 

prior to outcome assessment.

Figure 5-43 Associations of blood Pb levels with asthma- and allergy-related 

conditions in children.

Study

Rabinowitz et al. (1990)

Jedrychowski et al. (2011)

Rabinowitz et al. (1990)

Pugh Smith et al. (2011)

Joseph et al. (2005)

Population

Children

Children

Children

Children

Children, Caucasian

Children, African American

Children, African American

Outcome

Eczema

Positive SPT

Asthma

Asthma

Incident Asthma

Requiring Medical Care

Blood Pb Level Mean

or Group (μg/dL)

≥10 vs. <10

prenatal: 1.16

concurrent: 2.02

≥10 vs. <10

≥10 vs. <10

≥5 vs. <5

≥5 vs. <5

≥10 vs. <5

►

0 1 2 3 4 5 6 7 8

Odds ratio (95% CI)
February 2012 5-371 Draft – Do Not Cite or Quote

Table 5-29 Additional characteristics and quantitative results for studies 

presented in Figure 5-43

Study Population/Location Blood Pb Level Data (μg/dL) Statistical Analysis Outcome

Odds Ratio 

(95% CI)a

Rabinowitz et 

al. (1990)

1,768 children followed 

from birth to 

unspecified age 

Boston area, MA

Prenatal (cord) ≥ 10 vs. <10a Logistic regression with no 

additional covariates included in 

model

Eczema 1.0 (0.6, 1.6)

Jedrychowski 

et al. (2011)

224 children followed 

prenatally to age 5 yr

Krakow, Poland

Prenatal (cord): GM: 1.16 (95% CI: 1.12, 1.22)

Concurrent: GM: 2.02 (95% CI: 1.95, 2.12)

Logistic regression adjusted for 

sex, parity, maternal age, 

maternal education, maternal 

atopy, maternal smoking

Positive SPT 2.3 (1.1, 4.6)b

1.1 (0.7, 1.6)b

Joseph et al.

(2005)

4,634 children, ages 1-3 

yr followed prospectively 

for 12 months

Southeastern MI

Measured up to 12 mos before outcome

Caucasian ≥ 5 vs. Caucasian <5c

African American ≥ 5 vs. African American <5c

African American ≥ 10 vs. African American <5c

Logistic regression adjusted for 

sex, birth weight, and annual 

income

Incident asthma 

requiring 

medical care 2.7 (0.9, 8.1)

1.1 (0.8, 1.7)

1.3 (0.6, 2.6)

Rabinowitz et 

al. (1990)

1,768 children followed 

from birth to 

unspecified age 

Boston area, MA

Prenatal (cord blood)

≥ 10 vs. <10a

Logistic regression with no 

additional covariates included in 

model

Prevalent 

asthma

1.3 (0.8, 2.0)

Pugh Smith 

and Nriagu

(2011)

356 children, 

ages 0-12 yr

Saginaw, MI

Levels ascertained from statewide database, 

timing unreported but varied among subjects

≥ 10 vs. <10a

Logistic regression adjusted for 

age, sex, number of stories in 

unit, cat in home, dog in home, 

cockroach problem, number of 

persons in home, household 

smoking, clutter, 

candles/incense, type of cooking 

stove, main heating source, 

months of residency, housing 

tenure, type of air conditioning, 

peeling paint, ceiling/wall 

damage, age of housing, water 

dampness/mold/mildew

Prevalent 

asthma

7.5 (1.3, 42.9)

GM = geometric mean, SPT = skin prick test.

a

Odds ratio in children with blood Pb level ≥ 10 μg/dL with children with blood Pb level <10 μg/dL serving as the reference group

b

Odds ratio presented per 1 μg/dL increase in blood Pb level.

c

Relative risk in each specified subgroup with children with blood Pb level <5 μg/dL serving as the reference group.

1 In a study of 4,634 children in southeastern Michigan, blood Pb levels were measured at 

2 ages 1 to 3 years, up to 12 months prior to asthma assessment. In analyses that controlled 

3 for annual income, birth weight, and sex, an elevated risk of incident asthma requiring a 

4 doctor visit or medication (indicator of severe asthma) was reported in association with 

5 blood Pb levels ≥ 5 μg/dL among Caucasian children (relative risk [RR]: 2.7 [95% CI: 

6 0.9, 8.1] compared with Caucasian children with blood Pb levels < 5 μg/dL) (Joseph et 

7 al., 2005) (Figure 5-43 and Table 5-29). In analyses restricted to African Americans, 

8 children with blood Pb levels ≥ 10 μg/dL had an elevated risk of asthma requiring 

9 medical care (RR: 1.3 [95% CI: 0.6, 2.6] compared with children with blood Pb level < 

10 5 μg/dL) (Figure 5-43 and Table 5-29). In analyses that used Caucasian children with 

11 blood Pb level <5 μg/dL as the reference group, blood Pb level was associated with 

12 increased risk of asthma requiring medical care among African American children in all 

13 blood Pb level categories. Collectively, the results indicated a stronger association with 

14 race/ethnicity than with blood Pb level. It is important to recognize the small numbers of 
February 2012 5-372 Draft – Do Not Cite or Quote

1 children with asthma requiring medical care in the high blood Pb level categories, which 

2 likely accounted for the wide 95% CIs (5 Caucasian children with blood Pb ≥ 5 μg/dL 

3 and 9 African American children with blood Pb level ≥ 10 μg/dL).

4 Consistent with Joseph et al. (2005), a recent cross-sectional study conducted in Saginaw, 

5 Michigan found a higher prevalence of asthma in children (ages ≤ 12 years) with blood 

6 Pb levels ≥ 10 μg/dL (Pugh Smith and Nriagu, 2011). Of the 356 children randomly 

7 selected from a statewide surveillance database of blood Pb measurements, 78% were 

8 African American. This study had greater power than did Joseph et al. (2005) due to the 

9 larger percentage of children with blood Pb levels ≥ 10 μg/dL (18.6%). Compared with 

10 children with initial blood Pb levels < 10 μg/dL, children with initial blood Pb levels ≥ 

11 10 μg/dL had a higher odds of having a doctor diagnosis of asthma within the past 12 

12 months (OR: 7.5 [95% CI: 1.3, 42.9]). A strength of this study was the adjustment for a 

13 large number of potential confounding variables such as age, sex, pets in the home, 

14 housing characteristics, and household smoking. However, because children were 

15 identified from a statewide database of initial blood Pb measurements collected at 

16 unspecified ages, the timing of blood Pb varied among children. In a study that did not 

17 consider potential confounding variables, Rabinowitz et al. (1990) reported an increased 

18 risk of asthma in children (age of assessment not reported) in association with cord blood 

19 Pb levels > 10 μg/dL relative to cord blood Pb levels ≤ 10 μg/dL (Figure 5-43 and Table 

20 5-29).

21 While the aforementioned studies examined blood Pb levels measured at a single point in 

22 time, a recent prospective birth cohort study compared associations of prenatal (cord and 

23 maternal) and concurrent blood Pb levels with risk of allergic sensitization at age 5 years

24 (Jedrychowski et al., 2011). Cord and prenatal maternal blood Pb level were associated 

25 with greater risk of positive skin prick test (SPT) to dust mite, dog, or cat allergen than 

26 was child concurrent blood Pb level (Figure 5-43 and Table 5-29). For prenatal Pb 

27 biomarkers, similar effect estimates were obtained before and after adjusting for sex, 

28 parity, maternal age, maternal education, maternal atopy, and environmental tobacco 

29 smoke exposure. Cord and concurrent blood Pb levels were weakly correlated (r = 0.29), 

30 providing support for an independent association for prenatal Pb biomarkers. The 

31 independent effects of Pb also were substantiated by observations that indicators of other 

32 exposures, including blood levels of mercury, polycyclic aromatic hydrocarbon DNA 

33 adducts, and residential levels of dust mite or pet allergen were associated with lower 

34 risks of SPT than was blood Pb level. While associations were observed with relatively 

35 low cord blood Pb levels (geometric mean: 1.16 μg/dL [95% CI: 0.12, 1.22]), it is 

36 uncertain the extent to which higher past Pb exposures of the mothers may have 

37 influenced their pregnancy blood Pb levels and newborn cord blood Pb levels. Among 

38 children approximately age 10 years in Hong Kong, Hon et al. (2010; 2009) found 
February 2012 5-373 Draft – Do Not Cite or Quote

1 correlations between low concurrent blood Pb levels (population means < 2 μg/dL) and 

2 severity of atopic dermatitis (Spearman r = 0.46, p <0.001 and r = 0.33, p <0.005), 

3 another inflammatory condition commonly related to elevated IgE levels. Neither 

4 analysis considered potential confounding variables.

5 Among the studies in children that found associations of blood Pb level with asthma and 

6 allergy-related outcomes, several adjusted for potential confounding by SES-related 

7 variables. Joseph et al. (2005) adjusted for annual income. Pugh Smith et al. (2011), who 

8 examined a primarily low SES population of children in Michigan, adjusted for various 

9 factors associated with SES, including multiple indices of housing condition, and 

10 presence of pets and cockroaches in the home, which are indicators of allergens in the 

11 home. Jedrychowski et al. (2011) adjusted for maternal education, and found similar 

12 magnitudes of association between cord blood Pb level (and positive SPT) as those in the 

13 unadjusted analysis. Further, residential levels of dust mite or pet allergen were 

14 associated with lower risks of SPT than was blood Pb level. Thus, while SES and/or 

15 allergen exposure has been associated with both Pb exposure and asthma and allergy in 

16 children (Bryant-Stephens, 2009; Dowd and Aiello, 2009; Aligne et al., 2000), these 

17 collective findings in different populations do not demonstrate that confounding by SES 

18 and/or allergen exposure fully accounts for the associations observed between blood Pb 

19 levels and asthma and allergy. As allergic sensitization, asthma, and elevated IgE have 

20 been correlated in children, the evidence of association of blood Pb level with asthma and 

21 allergy after adjusting for SES, housing conditions, and allergen exposures, may provide 

22 support for the associations observed between higher blood Pb and higher IgE in children 

23 (Section 5.6.3).

24 Studies in nonoccupationally-exposed adults did not find associations of biomarkers of 

25 Pb exposure with asthma or allergy (Pizent et al., 2008; Mendy et al., In Press). The 

26 largest of these studies was a U.S. NHANES 2007-2008 analysis of adults ages 20 years 

27 and older, in which urinary Pb level was not associated with an increase in asthma (OR: 

28 0.72 [95% CI: 0.46, 1.12] per 1 μg/g increase in creatinine-adjusted urine) (Mendy et al., 

29 In Press). The results did not provide strong evidence that urinary Pb level was associated 

30 with other respiratory conditions such as emphysema or chronic bronchitis either. In a 

31 study of 216 adults without occupational Pb exposures, Pizent et al. (2008) found that 

32 among women, the association between concurrent blood Pb level and serum IgE was 

33 statistically significant, whereas the association with positive SPT to common inhaled 

34 allergens was not. Among men, higher concurrent blood Pb level was associated with 

35 lower odds of positive SPT (OR: 0.92 [95% CI: 0.86, 0.98] but was not statistically 

36 significantly associated with IgE. These findings appeared to be discordant because an 

37 increase in IgE commonly mediates the acute inflammatory response to allergens. 

38 However, the interpretation of the findings is difficult because only statistically 
February 2012 5-374 Draft – Do Not Cite or Quote

1 significant effect estimates were reported; thus it is not known whether odds ratios were 

2 in the same direction for SPT and IgE. Bener et al. (2001b) found higher prevalence of 

3 asthma and allergy-related conditions such as rhinitis and dermatitis among Pb industrial 

4 workers than among control subjects; however, the blood Pb levels in both the Pb5 exposed group and the control group (geometric means: 77.5 and 19.8 μg/dL, 

6 respectively) were higher than those in the current U.S. general adult population.

7 In summary, an available recent epidemiologic study added evidence for associations 

8 between blood Pb level and asthma in children (Pugh Smith and Nriagu, 2011), and a few 

9 recent studies provided new evidence for associations with allergic conditions in children

10 (Jedrychowski et al., 2011; 2010; Hon et al., 2009). This epidemiologic evidence was 

11 strengthened by several observations that associations were robust to the adjustment of 

12 multiple potential confounding factors, in particular, SES and allergen exposures. 

13 Because of the heterogeneity in the relatively small body of evidence, it was difficult to 

14 identify whether the strength of association with asthma and allergy differed by age of 

15 children, lifestage of blood Pb measurement (prenatal, sometime in childhood prior to 

16 outcome assessment, concurrent), or level of blood Pb. The epidemiologic evidence for 

17 associations of blood Pb level with asthma and allergy is well supported by toxicological 

18 and epidemiologic evidence indicating Pb effects on increasing IgE (Section 5.6.3), Th2 

19 cytokines (Section 5.6.5.4), and inflammation (Section 5.6.5.1). In the few recent studies 

20 that investigated nonoccupationally-exposed adults, Pb biomarker levels were not 

21 associated significantly with greater asthma or allergy (Pizent et al., 2008; Mendy et al., 

22 In Press).

5.6.4.3 Other Respiratory Effects

23 The respiratory effects of Pb have been examined primarily in a small number of recent 

24 epidemiologic studies of nonoccupationally-exposed adults, and as with evidence for 

25 asthma, evidence for Pb-associated respiratory effects in adults is weak. Increased 

26 bronchial responsiveness (BR) is a characteristic feature of asthma and other respiratory 

27 diseases and can result from the activation of innate immune responses and increased 

28 airway inflammation. In a study of 525 middle-aged adults in Seoul, Korea, Min et al.

29 (2008a) found an association between concurrent blood Pb level and BR. A 1 μg/dL 

30 higher concurrent blood Pb level was associated with a higher BR index (log [% decline 

31 in forced expiratory volume in 1 second (FEV1)/log of final methacholine concentration 

32 in mg/dL]) of 0.018 (95% CI: 0.004, 0.03), adjusting for age, sex, height, smoking, lung 

33 function, and asthma diagnosis (Min et al., 2008a). The concurrent blood Pb levels in 

34 these adults were low (mean [SD]: 2.90 [1.59] μg/dL); however, it is uncertain what 

35 timing, level, frequency, and duration of Pb exposures contributed to the observed 
February 2012 5-375 Draft – Do Not Cite or Quote

1 association. In contrast to Min et al. (2008a), Pizent et al. (2008) found that higher 

2 concurrent blood Pb level was associated with lower BR in men (2.4% decrease [95% CI: 

3 -4.2, -0.52%] in percent change FEV1 post-histamine challenge per 1 μg/dL increase in 

4 blood Pb level). Although this finding was not discussed in details by investigators, it was 

5 consistent with the observation that higher blood Pb level was associated with lower odds 

6 of positive SPT among men in this study.

7 In studies of male Pb workers, Pb-associated respiratory effects were not clearly 

8 indicated. None of the studies directly examined associations between blood Pb levels 

9 and lung function. In bus drivers in Hong Kong, China, (Jones et al., 2008; Jones et al., 

10 2006), drivers of non-air conditioned buses had lower exposures to PM10, lower blood Pb 

11 levels (mean 3.7 μg/dL versus 5.0 μg/dL in air conditioned buses) but lower indices of 

12 lung function than did drivers of air conditioned buses (Jones et al., 2006). In this study, 

13 the authors attributed the slightly higher blood Pb levels of air conditioned bus drivers to 

14 the poor efficiency in the filters and higher PM10 levels measured on those buses versus 

15 the non-air conditioned buses. In a comparison of roadside vendors and adjacent 

16 shopkeepers, blood Pb levels and various lung function parameters were similar between 

17 groups (Jones et al., 2008). Pb industrial workers in the United Arab Emirates had higher 

18 prevalence of respiratory symptoms such as cough, phlegm, shortness of breath, and 

19 wheeze than did unexposed controls (Bener et al., 2001b). Blood Pb levels in both the Pb20 exposed group and the control group (geometric means: 77.5 and 19.8 μg/dL, 

21 respectively) were higher than those in the current U.S. adult general population, and the 

22 analysis did not consider potential confounding variables. 

23 Toxicological evidence for Pb-associated respiratory effects is provided by observations 

24 that exposure to Pb-containing PM induces a range of inflammatory-related effects in the 

25 airways of animals and in cultured airway cells (Section 5.6.6). Specific effects on the 

26 lung also were demonstrated in a recent study of rats injected with Pb-acetate (25 mg/kg, 

27 3 consecutive days, resulting in 2.1 μg/dL blood Pb) (Kaczynska et al., 2011). Pb-treated 

28 rats exhibited ultrastructural changes in lung tissue, including substantial pulmonary 

29 fibrosis containing numerous lipofibroblasts, collagens, and elastin filaments in the 

30 interstitium. Mast cells also were present in the interstitium following Pb exposure. 

31 Pulmonary inflammation was observed, evidenced by the increased recruitment of 

32 monocytes and thrombocytes inside the capillary vessels and increased macrophage 

33 accumulation in the alveolar space. Evidence of damaged surfactant lining and 

34 destruction of the laminae inside lamellar bodies of epithelial type II cells also was 

35 observed. While the observed pulmonary histological changes have been linked with 

36 functional pulmonary decrements in other studies (unrelated to Pb exposure), they were 

37 observed with injected Pb. It is not clear whether routes of Pb exposure more relevant to 

38 those in humans would result in similar effects.
February 2012 5-376 Draft – Do Not Cite or Quote

5.6.4.4 Autoimmunity

1 Evidence for the effects of Pb on increasing the risk of autoimmunity is provided 

2 primarily by a small number of older toxicological studies reviewed in the 2006 Pb 

3 AQCD in which Pb exposure of animals (pre- and postnatal) was associated with the 

4 generation of autoantibodies (Hudson et al., 2003; Bunn et al., 2000; El-Fawal et al., 

5 1999; Waterman et al., 1994). Whereas some evidence linked this risk of autoimmunity 

6 to a shift toward Th2 responses, other evidence pointed to a shift toward Th1 responses. 

7 While recent studies did not examine Pb-induced production of autoantibodies, some 

8 provided indirect evidence by indicating that the changes induced by Pb had broader 

9 implications for increasing risk of autoimmunity. For example, Kasten-Jolly et al. (2010)

10 examined the impact of developmental (100 μM Pb-acetate in drinking water of dams 

11 from GD8 to PND21, resulting in pup blood Pb levels 10-30 μg/dL) Pb exposure of mice 

12 on changes in gene expression in the spleen. Investigators found that Pb upregulated 

13 digestive and catabolizing enzymes that could lead to the generation of self-peptides, 

14 which in conjunction with other Pb-induced immunomodulatory effects, had the potential 

15 to induce the generation of autoantibodies. In Carey et al. (2006), the activation of 

16 neoantigen-specific T cells in PbCl2-treated adult mice (25-50 μg i.p.) also indicated the 

17 potential for autoantibody generation. Evidence of Pb-associated autoimmune responses 

18 in humans is limited to an older study of male Pb battery workers with blood Pb levels 

19 ranging from 10 to 40 μg/dL (El-Fawal et al., 1999). In this study, the Pb-exposed 

20 workers had higher levels of IgM and IgG autoantibodies to neural proteins compared 

21 with unexposed controls (blood Pb levels not reported) (El-Fawal et al., 1999). Pb also 

22 was found to modify neural proteins in rats (Waterman et al., 1994). The generation of 

23 self-peptides and modification of proteins can result in formation of neoantigens, thereby 

24 increasing the risk of autoimmune reactions.

5.6.4.5 Specialized Cells in Other Tissues

25 As discussed in Section 5.6.2.4, Pb exposure consistently has been found to alter the 

26 function of macrophages. Nonlymphoid tissues also contain specialized macrophages 

27 whose altered function can contribute to organ/tissue dysfunction, cell death, tissue 

28 pathology and tissue-specific autoimmune reactions. Among the specialized macrophages 

29 are microglia and astrocytes in the brain, Kupffer cells in the liver, alveolar macrophages 

30 in the lung, keratinocytes and Langerhans cells in the skin, osteoclasts in the bone, and 

31 preadipocytes in adipose tissue. The evidence demonstrating the effects of Pb on 

32 specialized macrophages, which was provided by recent toxicological studies, is 

33 important as it demonstrates the contribution of immune dysfunction to the effects of Pb 

34 on dysfunction in nonlymphoid tissues (Figure 5-44). Because these specialized cells are 
February 2012 5-377 Draft – Do Not Cite or Quote

1 not always recognized as macrophages, the resulting diseases and conditions are not 

2 always recognized as being linked with Pb-induced immune dysfunction. It is important 

3 to note that in this small body of available studies, several studies treated animals by Pb 

4 injection, and it is unclear whether the effects would be observed with routes of exposure 

5 that are more relevant for humans. 

Figure 5-44 Specialized macrophages in nonlymphoid tissue may serve as a 

link between Pb exposure and disease in multiple organ systems.

6 Fan et al. (2009b) reported that Kupffer cells undergo significant changes in phenotypic 

7 expression (e.g., CD68 and ferritin light chain), organization, and functional activity 

8 connected to Pb-induced apoptosis in the liver. Dosing of juvenile Wistar rats with Pb 

9 injections (15 mg/kg of Pb-acetate daily for 2 weeks, resulting in a mean blood Pb level 

10 of 30 μg/dL) during early postnatal maturation was observed to produce chronic glial 

11 activation, increase pro-inflammatory cytokines, and increase neurodegeneration, as 

12 indicated by an increase in GFAP, increase in IL-1β, TNF-α and IL-6 cytokines, and a 

13 decrease in synaptophysin (component of presynaptic vesicles), respectively, in brain 

14 tissue (Struzynska et al., 2007). In bone, osteoclasts regulate osteoblast function (Chang 

15 et al., 2008a), and osteoblasts have been shown to be affected by Pb exposure. Effects on 

16 these cell types can contribute to later life diseases such as arthritis [reviewed in Zoeger 

17 et al. (2006)]. Pb-induced elevation of TGF-β production was found to be involved in 

Pb

Osteoblastsand 

Osteoclasts

Periodontitis

and 

Chondrogenesis

Kupffer cells

Altered liver metabolism

Oxidative damage

Glutathione status

Microglia 

and Astrocytes

Neurobehavioral and 

neurodegenerative 

conditions

Alveolar 

macrophages

Bronchial hyper-reactivity

Respiratory allergy

Asthma

Susceptibility to infections

Resident bone

macrophages
February 2012 5-378 Draft – Do Not Cite or Quote

1 chondrogenesis in bone (Zuscik et al., 2007). Kaczynska et al. (2011) reported effects on 

2 alveolar macrophages after Pb treatment (i.p. 25 mg/kg, 3 days, resulting in blood Pb 

3 levels of 2.1 μg/dL) in rats. Macrophage recruitment increased, and this macrophage 

4 infiltration limited air space available to gas exchange and contained parts of 

5 phagocytized surfactant and alveolar lining.

6 Resident immune cells in reproductive organs have been shown to be affected by Pb 

7 exposure. Pace et al. (2005) reported that Pb exposure in mice contributed to poor 

8 reproductive performance that was concomitant with altered homeostasis of the testicular 

9 macrophage population in that organ. The authors proposed that increased oxidative 

10 damage and apoptosis among these macrophages and reduced potential to maintain organ 

11 homeostasis contributed to the observed pattern of male sterility.

5.6.4.6 Tumors

12 While toxicological evidence indicates that high concentration Pb exposures directly 

13 promote tumor formation or induce mutagenesis and genotoxicity (Section 5.10), 

14 evidence for involvement of the immune system is limited. Kerkvliet and Baecher15 Steppan (1982) observed that male C57Bl/6 mice exposed to 130 and 1,300 ppm of 

16 Pb-acetate in drinking water had enhanced moloney sarcoma virus-induced tumor growth 

17 compared with control animals. The findings indicated that Pb-induced 

18 immunomodulation affecting tumors likely resulted from a combination of suppressed 

19 Th1 responses and increased inflammation leading to excessive release of ROS into 

20 tissues. The promotion of cancer is a relatively common outcome in chemical-induced 

21 immunotoxicology, particularly when early life exposures are involved (Dietert, 2011).

5.6.5 Modes of Action for Lead Immune Effects

5.6.5.1 Inflammation

22 The 2006 Pb AQCD indicated that misregulated inflammation represents one of the 

23 major immune-related effects of Pb and a major mode of action for Pb effects in multiple 

24 organ systems such as the liver, kidney, and vasculature (Section 5.2.5). In the 2006 Pb 

25 AQCD, several lines of toxicological evidence demonstrated Pb-induced inflammation: 

26 Pb-induced increases in PGE2 and ROS (Section 5.6.5.2) and increases in pro27 inflammatory cytokines (Section 5.6.5.4), both of which promote a hyperinflammatory 

28 phenotype in immune cells. As described in sections that follow, these findings are 
February 2012 5-379 Draft – Do Not Cite or Quote

1 corroborated by a small number of available recent toxicological studies. A few available 

2 recent epidemiologic studies provided new evidence of Pb-associated inflammation, as 

3 indicated by associations of blood Pb levels with pro-inflammatory cytokines and other 

4 nonspecific indicators of inflammation.

5 Enhanced inflammation and tissue damage occurs through the modulation of 

6 inflammatory cell function and production of pro-inflammatory cytokines and 

7 metabolites. It is important to note that while inflammation can mediate immune-related 

8 conditions such as asthma, respiratory infections, and BR, inflammation can exacerbate 

9 disease and damage in almost any organ given the distribution of immune cells as both 

10 permanent residents and infiltrating cell populations. Among the problems presented by 

11 this immunomodulation are the overproduction of ROS and an apparent depletion of 

12 antioxidant protective enzymes and factors (e.g., selenium). For example, toxicological 

13 studies have found that Pb-induced inflammatory damage involves the depletion of 

14 antioxidants such as glutathione and catalase (Lodi et al., 2011; Chetty et al., 2005).

15 While several processes have been proposed to explain the mechanisms of Pb-induced 

16 oxidative damage, the exact combination of processes involved remains to be determined 

17 (Section 5.2.4). 

18 In epidemiologic studies, Pb-associated changes in pro-inflammatory cell function 

19 (Section 5.6.2) and cytokine production (Section 5.6.5.4) have been found in the 

20 cumulative body of evidence. A few recent epidemiologic studies added to this evidence 

21 with observations of associations between blood Pb level and other nonspecific indicators 

22 of inflammation that may be related to multisystemic effects as have been demonstrated 

23 in animal and in vitro studies. Using 1999-2004 NHANES data, Songdej et al. (2010)

24 examined the relationship between concurrent blood Pb levels and the inflammation 

25 markers, C-reactive protein (CRP), fibrinogen, and white blood cell (WBC) count in 

26 adults 40 years of age or older. Adjusting for age, sex, race/ethnicity, education, income, 

27 BMI, physical activity, smoking status, diabetes status, inflammatory disease status, and 

28 cardiovascular disease status, investigators found larger magnitudes of association 

29 between blood Pb and inflammation in men compared with women. Among women, 

30 most ORs for associations between quintiles of blood Pb level and tertiles of CRP, 

31 fibrinogen, and WBC count were less than 1.0 whereas corresponding ORs in men tended 

32 to be greater than 1.0 but were not always statistically significant. For example, compared 

33 with men with concurrent blood Pb levels less than 1.16 μg/dL, men with blood Pb levels 

34 of 1.16-<1.63 μg/dL, 1.63-<2.17, 2.17-<3.09 μg/dL, and ≥ 3.09 μg/dL had elevated odds 

35 of elevated CRP (OR [95% CI]: 2.22 [1.14, 4.32], 1.67 [0.85, 3.28], 2.12 [1.07, 4.21], and 

36 2.85 [1.49, 5.45], respectively). For all inflammation markers, although the OR was 

37 highest in the highest quintile of blood Pb level (≥ 3.09 μg/dL), monotonic concentration38 dependent increases were not observed.
February 2012 5-380 Draft – Do Not Cite or Quote

1 Consistent with these findings, among men in Incheon, Korea without occupational Pb 

2 exposures, Kim et al. (2007) reported associations of higher concurrent blood Pb level 

3 with higher levels of WBCs and IL-6. Larger effects were estimated for men in the upper 

4 two quartiles of blood Pb levels, 2.51-10.47 μg/dL than for the full range of blood Pb 

5 levels. The findings are consistent with Pb effects on promoting a Th2 phenotype. Th2 

6 cells produce IL-6 which is the primary stimulus for expression of CRP and fibrinogen

7 (Hage and Szalai, 2007; Fuller and Zhang, 2001). Despite the low concurrent blood Pb 

8 level of adults in these aforementioned studies, it is important to acknowledge that the 

9 relative contributions of recent versus past Pb exposures to the associations observed with 

10 concurrent blood Pb levels are not delineated. 

11 In a genome-wide association study that included 37 children with autism and 15 children 

12 without autism (ages 2-5 years; blood Pb level range: 0.37 to 5.2 μg/dL) in California, in 

13 models that included age, sex, and autism diagnosis, concurrent blood Pb level was 

14 associated with the expression of several genes related to immune function and 

15 inflammation, including human leukocyte antigen genes (HLA-DRB) and MHC Class II16 associated invariant chain CD74 (involved in antigen presentation) (Tian et al., 2011).

17 Although blood Pb levels were similar between children with and without autism and 

18 correlations were observed in both groups, they were in opposite directions (positive 

19 among children with autism and negative among children without autism). Pb has been 

20 shown to increase MHC molecule surface expression in mouse and human HLA antigen 

21 presenting cells (Guo et al., 1996a; McCabe and Lawrence, 1991); however, additional 

22 larger studies with a priori hypotheses regarding specific indices of inflammation are 

23 warranted to characterize Pb-associated changes in inflammation in children.

5.6.5.2 Increased Prostaglandin E2 and Decreased Nitric Oxide 

24 Consistent with the large body of evidence presented in the 2006 Pb AQCD (U.S. EPA, 

25 2006b), a small number of available recent studies continued to indicate that Pb exposure 

26 alters the levels of signaling molecules such as PGE2 and NO. Collectively, the weight of 

27 evidence was provided by toxicological studies. These signaling molecules are involved 

28 in mediating inflammation and host resistance (Figure 5-42). A recent in vitro study with 

29 human neuroblastoma cells found increases in PGE2 with lower Pb concentrations (0.01-

30 1 μM) than those previously reported (Chetty et al., 2005). A large body of evidence 

31 reviewed in the 2006 Pb AQCD demonstrated a Pb-associated decreased production of 

32 NO by macrophages [see 2006 Annex Table AX5.9.6 (U.S. EPA, 2006h)].

33 In adult animal models, decreases in NO were observed with short-term exposures (hours 

34 to days) to a wide range of Pb concentrations. Decreases in NO can impact not only 
February 2012 5-381 Draft – Do Not Cite or Quote

1 innate host defenses, but also, acquired immunity. In a recent study, Farrer et al. (2008)

2 found that high-level Pb glutamate exposure (5 μM) of lymphocyte cultures led to 

3 decreased NO production, decreased inducible NO synthase function in myeloid cells, 

4 but no change in its gene expression. Additionally, Pb abrogated the myeloid cell 

5 (CD11b+)-mediated suppression of CD4+ T cell proliferation. Together, these findings 

6 indicated that Pb may indirectly enhance T cell proliferation through its effect on 

7 decreasing NO production. Combined with the observation that Pb can alter antigen 

8 processing (Farrer et al., 2005) and, hence, the quality and magnitude of the acquired 

9 immune response signal against pathogenic challenge, evidence indicated that multiple 

10 arms of the host defense against infectious challenge can be compromised. The loss of 

11 NO production in innate immune cells such as macrophages would be expected to affect 

12 other physiological systems (e.g., neurological, cardiovascular, endocrine) that require 

13 NO signaling cascades. 

14 Relative to studies in animal and in vitro models, fewer epidemiologic studies have 

15 examined the effects of Pb on signaling molecules; however the limited data supported 

16 associations of blood Pb level with suppressed NO production (Barbosa et al., 2006a; 

17 Pineda-Zavaleta et al., 2004) and increased ROS production (Pineda-Zavaleta et al., 

18 2004) in populations living near Pb sources. In a previous study of children in Mexico, 

19 with increasing residential proximity to the Pb smelter, mean concurrent blood Pb levels 

20 increased (7.02 to 20.6 to 30.38 μg/dL) as did superoxide anion release from 

21 macrophages (directly activated by IFN-γ/LPS) isolated from children (Pineda-Zavaleta 

22 et al., 2004). NO release from macrophages (indirectly activated by phytohemagglutinin, 

23 PHA) was lower with higher blood Pb levels. After adjusting for age and sex, a 1 μg/dL 

24 higher blood Pb level was associated with a higher level of superoxide anion of 0.00389 

25 (95% CI: 0.00031, 0.00748) μmol/mg protein and a lower level of NO of 0.00089 (95% 

26 CI: -0.0017, -0.00005) nmol/μg protein. Because PHA activates macrophages indirectly 

27 through the activation of lymphocytes and IFN-γ directly activates macrophages, these 

28 results indicated that Pb suppressed T cell-mediated macrophage activation and 

29 stimulated cytokine-induced macrophage activation. Group-level comparisons indicated 

30 that associations likely were driven by changes observed in the group of children living in 

31 closest proximity to the smelter who had blood Pb levels 10.31-47.49 μg/dL. Results also 

32 demonstrated a larger magnitude of association between blood Pb levels and superoxide 

33 anion release in males. Although not described in detail, higher blood Pb level was not 

34 associated with lower NO in girls. Barbosa et al. (2006a) also observed an association 

35 between higher blood Pb level and lower plasma NO in a group of adults in Sao Paolo, 

36 Brazil residing near a closed battery plant, particularly among adults with the TC or CC 

37 eNOS genotype (r = 0.23, p = 0.048) which is associated with reduced promoter activity 

38 and potentially reduced gene expression. Quantitative results were not reported for 

39 analysis of all adults combined or adults with the TT genotype, but p-values were greater 
February 2012 5-382 Draft – Do Not Cite or Quote

1 than 0.05. Because NO was measured in plasma, it was not possible to identify immune 

2 cells as the specific sources of NO. 

3 Studies of occupationally-exposed adults provided less clear indication of associations of 

4 blood Pb level with NO and ROS. Despite large differences in blood Pb levels between 

5 30 male Pb recycling plant workers (mean: 106 μg/dL) and 27 unexposed controls (mean: 

6 4.5 μg/dL), levels of ROS released from neutrophils (indicators of respiratory burst) were 

7 similar between groups (Mishra et al., 2006a). In a study of male foundry workers (mean 

8 blood Pb level: 21.7 μg/dL), pottery workers (mean blood Pb level: 9.7 μg/dL), and 

9 unexposed workers (mean blood Pb level: 3.9 μg/dL), Valentino et al. (2007) found 

10 similar plasma NO levels in controls compared with Pb-exposed workers. Also, although 

11 quantitative results were not reported, blood Pb level was reported not to be correlated 

12 with NO.

5.6.5.3 Cellular Death (Apoptosis, Necrosis)

13 The 2006 Pb AQCD reviewed a small number of toxicological studies in which Pb 

14 exposure had contrasting effects on the apoptosis of macrophages. Since then, a few 

15 available recent toxicological studies found that Pb exposure induces apoptosis or 

16 mediators of apoptosis in immune cells. In a study in mice, Bishayi and Sengupta (2006)

17 found that Pb treatment of adult mice (10 mg/kg, i.p.) elevated DNA fragmentation in 

18 splenic macrophages. Using mouse resident peritoneal macrophages, Gargioni et al.

19 (2006) found that 20 and 40 μM Pb nitrate induced both necrosis and apoptosis in vitro. 

20 While the exact pathways involved were not determined, the authors concluded that 

21 activation of the Bax pro-apoptotic protein was not the key effect of Pb on inducing 

22 macrophage apoptosis. In an in vivo study in 3 week-old mice, Xu et al. (2008) found 

23 that a 4-week administration of Pb-acetate (50-100 mg/kg, oral) significantly elevated 

24 both ROS and malondialdehyde (an indicator of ROS-induced peroxidation) levels in 

25 peripheral blood lymphocytes. Pb also induced DNA damage (determined by the comet 

26 assay), which was accompanied by elevations in p53 and Bax expression with no change 

27 in Bcl-2 expression (creating a Bax/Bcl-2 imbalance). The authors proposed that 

28 oxidative stress was a likely route to Pb-induced apoptosis and tumorigenesis. Because of 

29 the high Pb concentrations administered and systemic administration of Pb employed, 

30 other studies are warranted to examine whether apoptosis of macrophages represent a 

31 mode of action relevant to human Pb exposures.
February 2012 5-383 Draft – Do Not Cite or Quote

5.6.5.4 Cytokine Production

1 The 2006 Pb AQCD presented a large body of toxicological evidence that clearly 

2 demonstrated Pb-induced changes in cytokine production in vivo and in vitro (U.S. EPA, 

3 2006b). Previous toxicological studies indicated that Pb affects immune cytokine 

4 production via action on T cells and macrophages. The combination of cytokine changes 

5 induced by Pb in multiple cell types can create a hyperinflammatory state among innate 

6 immune cells to skew acquired immunity responses away from Th1 responses and toward 

7 Th2 responses. As illustrated in Figure 5-42, downstream effects include altered IgE 

8 production, ROS production, and inflammation. In support of a Pb-induced skew toward 

9 Th2 responses, several studies in rats, mice, and chickens found that pre- and postnatal 

10 Pb exposure suppressed the production of Th1 cytokine IFN-γ and/or increased 

11 production of Th2 cytokines such as IL-4 [Table 5-7 of the 2006 Pb AQCD (U.S. EPA, 

12 2006g)]. Recent toxicological studies continued to find such Pb-induced changes in 

13 cytokine production, and a study provided new evidence that Th2 skewing may be 

14 mediated via effects on dendritic cells.

15 Cheng et al. (2006) found that Pb exposure affected TNF-α production in vitro in A/J 

16 mice macrophages by affecting the mitogen-activated protein kinase (MAPK) signaling 

17 pathway. Pb-acetate (10 μM) co-administered with LPS stimulated the phosphorylation 

18 of p42/44 MAPK and TNF-α expression (Cheng et al., 2006). Blocking protein kinase C 

19 or MAPK reduced TNF-α production of macrophages in vitro, which in turn, protected 

20 against Pb + LPS-induced liver injury in vivo. Thus, Pb exposure may induce local tissue 

21 damage through the modulation of immune responses. These findings were consistent 

22 with those from recent studies. Gao et al. (2007) showed that treatment of mouse 

23 dendritic cells with 25 μM PbCl2 produced an increased phosphorylation of the 

24 Erk/MAPK signaling molecule, and Khan et al. (2011) showed that monocytes treated 

25 with 25-50 μM Pb-acetate increased TNF-α through ERK1/2 and p38 signaling. 

26 In an in vivo study conducted across a lifetime (developmental through adulthood) in 

27 Swiss mice (females and males) using a broad range of dietary Pb concentrations, Iavicoli 

28 et al. (2006a) found a nonlinear hierarchical cytokine response. At the lowest dietary Pb 

29 concentration (0.11 ppm Pb-acetate, resulting in blood Pb level: 1.6 μg/dL), IL-2 and 

30 IFN-γ were decreased compared to those in the controls (0.02 ppm Pb-acetate, resulting 

31 in blood Pb level: 0.8 μg/dL), indicating a suppressed Th1 response. As dietary Pb 

32 exposure increased (resulting in blood Pb levels 12-61 μg/dL), a Th2 phenotype was 

33 observed with suppressed IFN-γ and IL-2 and elevated IL-4 production. These findings 

34 support the notion that the immune system is differentially modulated by low-level versus 

35 high-level Pb exposures. Other studies found variable Pb-induced changes in IL-2, with 

36 no change or elevated production, depending upon the protocol used. Recently, Gao et al.
February 2012 5-384 Draft – Do Not Cite or Quote

1 (2007) found that Pb-treated dendritic cells (25 μM) promoted a slight but statistically 

2 significant increase in IL-2 production (quantitative results not reported) among 

3 lymphocytes.

4 In vitro studies also reported a Pb-induced shift to production of Th2 cytokines. In 

5 conjunction with other indicators of Th2 skewing described in Sections 5.6.2.3, 5.6.2.6, 

6 and 5.6.3, Gao et al. (2007) observed that 25 μM Pb elevated the production of Th2 

7 cytokines such as IL-4, IL-5 and IL-6 in dendritic cells. In cultures of human PMNs 

8 activated with Salmonella enteritidis or with monoclonal antibodies of CD3, CD28, and 

9 CD40, Pb-acetate concentrations of 0.15 μg/dL and higher suppressed expression of Th1 

10 cytokines, IFN-γ, IL-1β, and TNF-α, and increased secretion of Th2 cytokines, IL-5, IL11 6, and IL-10 (Hemdan et al., 2005).

12 Consistent with toxicological studies, a few available epidemiologic studies also found 

13 higher concurrent blood Pb levels in children and occupationally-exposed adults to be

14 associated with a shift toward production of Th2 cytokines relative to Th1 cytokines. The 

15 evidence in children was based on comparisons of serum cytokine levels among groups 

16 with different blood Pb levels without consideration of potential confounding factors. 

17 Among children ages 9 months to 6 years in Missouri, Lutz et al. (1999) found that 

18 children with concurrent blood Pb levels 15-19 μg/dL had higher serum levels of IL-4 (p 

19 = 0.08, Kruskal Wallis) and IgE (Section 5.6.3) than did children with lower blood Pb 

20 levels. However, IL-4 levels in children with blood Pb levels 20-44 μg/dL were lower 

21 than those in children with blood Pb levels < 15 μg/dL. The elevated IL-4 and IgE in 

22 children with blood Pb levels 15-19 μg/dL were consistent with the mode of action for 

23 IL-4 to activate B cells to induce B cell class switching to IgE. In another study of 

24 children in grades 5 and 6 in Taiwan, investigators did not group children by blood Pb 

25 levels but by potential for Pb exposures due to age of home and location of residence

26 (Hsiao et al.). Concurrent blood Pb levels did not differ by residence in old versus new 

27 homes or by urban versus rural residence (means: 3.2-3.8 μg/dL) but were higher among 

28 children living near an oil refinery, in particular, among children with known respiratory 

29 allergies (mean: 8.8 μg/dL). This latter group of children also had the lowest serum levels 

30 of IFN-γ (45-fold) and highest levels of IL-4 (6-fold) (lower p < 0.05 for comparisons 

31 with any subgroup). There was no direct comparison of cytokine levels between blood Pb 

32 level groups in the population overall; however, cytokine levels were similar between 

33 healthy and allergy groups in the other Pb source groups that had similar blood Pb levels 

34 (p > 0.05 for comparisons with any subgroup). Thus, the differences in cytokine levels 

35 between healthy and allergic children living near the oil refinery may have been 

36 influenced by differences in their blood Pb levels. 
February 2012 5-385 Draft – Do Not Cite or Quote

1 Evidence of association between blood Pb levels and cytokine levels in 

2 nonoccupationally-exposed adults was equivocal. Among adult university students in 

3 Incheon, Korea, Kim et al. (2007) found associations of concurrent blood Pb level with 

4 serum levels of TNF-α and IL-6 that were larger among males in the upper two quartiles 

5 of blood Pb levels, 2.51-10.47 μg/dL. Notably, the relative contributions of recent versus 

6 past Pb exposures to these associations are not known. In models that adjusted for age, 

7 BMI, and smoking status, a 1 μg/dL higher blood Pb level was associated with a 0.75 

8 (95% CI: 0.14, 1.36) pg/mL higher TNF-α and a 0.18 (95% CI: -0.02, 0.38) pg/mL higher 

9 IL-6. The association between levels of blood Pb and plasma TNF-α was greater among 

10 men who were GSTM1 null (1.14 [95% CI: 0.20, 2.10] pg/mL higher per 1 μg/dL higher 

11 blood Pb level) than men who were GSTM1 sufficient (0.12 [95% CI: -0.43, 0.67] 

12 pg/mL). Blood Pb also was associated with a larger increase in TNF-α among men who 

13 had the TNF-α GG genotype (0.80 [95% CI: 0.15, 1.45] pg/mL higher per 1 μg/dL higher 

14 blood Pb level) than men who had the GA or AA genotype (-0.21 [95% CI: -1.1, 0.71] 

15 pg/mL). For the association between blood Pb level and plasma IL-6, the effect estimate 

16 was slightly elevated in TNF-α GG genotype but not elevated in the GSTM1 positive 

17 group. The effects of Pb on several physiological systems have been hypothesized to be 

18 mediated by the generation of ROS (Daggett et al., 1998). Thus, it is biologically 

19 plausible that the null variant of GSTM1, which is associated with reduced elimination of 

20 ROS, may increase the risk of Pb-associated immune effects. The results for the TNF-α 

21 polymorphism were difficult to interpret. The GG genotype is associated with lower 

22 expression of TNF-α, and the literature is mixed with respect to which variant increases 

23 risk of inflammation-related conditions. Among adults in Italy, concurrent blood Pb 

24 levels were not statistically significantly correlated with either Th2 or Th1 cytokine levels 

25 in men (Boscolo et al., 1999) or women (Boscolo et al., 2000) (quantitative results not 

26 reported).

27 Results from studies of occupationally-exposed adults also suggested that Pb exposure 

28 may be associated with decreases in Th1 cytokines and increases in Th2 cytokines; 

29 however, analysis were mostly limited to comparisons of mean cytokine levels among 

30 different blood Pb groups or Pb exposure groups (Di Lorenzo et al., 2007; Valentino et 

31 al., 2007; Yucesoy et al., 1997a) that did not consider potential confounding variables. 

32 The exception was a study of male foundry workers, pottery workers, and unexposed 

33 workers (Valentino et al., 2007). Multiple regression analyses were performed with age, 

34 BMI, smoking, and alcohol consumption included as covariates; however, regression 

35 coefficients describing the concentration-response functions were not reported. Pb36 exposed workers had higher IL-10 and TNF-α (ANOVA, p < 0.05). Levels of IL-2, IL-6, 

37 and IL-10 also increased from the lowest to highest blood Pb group (ANOVA, p > 0.05). 

38 In contrast with most other studies, both exposed worker groups had lower IL-4 levels 

39 compared with controls (ANOVA, p > 0.05). In a similar analysis, DiLorenzo et al.
February 2012 5-386 Draft – Do Not Cite or Quote

1 (2007) separated exposed workers into intermediate (9.1-29.4 μg/dL) and high (29.4-

2 81.1 μg/dL) blood Pb level groups, with unexposed workers comprising the low exposure 

3 group (blood Pb levels 1-11 μg/dL). Mean TNF-α levels showed a monotonic increase 

4 from the low to high blood Pb level group. Results also indicated a potential interaction 

5 between blood Pb level and smoking. Among current smokers, a 12- to 16-fold difference 

6 in TNF-α levels was observed among blood Pb groups. Among nonsmokers, the 

7 differences were less than two fold. In Yucesoy et al. (1997a), levels of the Th1 

8 cytokines, IL-1β and IFN-γ, were lower in workers than in controls.

9 In summary, a large body of previous toxicological evidence clearly demonstrated Pb10 induced increases in Th2 cytokines and decreases in Th1 cytokines. Several recent 

11 toxicological studies added to this evidence by showing Pb-induced increases in TNF-α 

12 that were mediated by MAPK signaling pathways (Khan et al., 2011; Gao et al., 2007; 

13 Cheng et al., 2006; Iavicoli et al., 2006a; Hemdan et al., 2005), and demonstrating Pb14 induced increases in cytokine production in dendritic cells (Gao et al., 2007). The 

15 collective epidemiologic evidence is sparse; however, in the few available new studies, 

16 higher concurrent blood Pb level was associated with higher levels of various pro17 inflammatory cytokines in children, nonoccupationally-exposed adults, and 

18 occupationally-exposed adults. The recent epidemiologic study of children in Taiwan 

19 found an association between higher IL-4 levels in children with higher blood Pb levels

20 (Hsiao et al.), similar to a previous study of children in Missouri (Lutz et al., 1999).

21 Neither study of children considered potential confounding variables. In contrast to 

22 previous studies, a recent study in nonoccupationally-exposed adults found associations 

23 between higher concurrent blood Pb level and higher TNF-α and IL-6 cytokine levels in 

24 men in Korea (Kim et al., 2007). Recent studies of occupationally-exposed adults added 

25 to the evidence for higher levels of several cytokines among Pb-exposed workers 

26 compared to unexposed controls (Di Lorenzo et al., 2007; Valentino et al., 2007). Due to 

27 the limited investigation, it is difficult to draw conclusions about the effects of Pb 

28 exposure on cytokine levels in any particular group in the human population.

5.6.6 Air-Lead Studies

29 Although comprising a smaller body of evidence than do immune studies of Pb 

30 biomarkers, several recent studies used Pb measured in PM10 and PM2.5 air samples to 

31 represent Pb exposures. Some studies analyzed the Pb component individually, whereas 

32 others analyzed Pb as part of a group of correlated components using source 

33 apportionment techniques or principal component analysis. In concordance with blood Pb 

34 studies, recent time-series epidemiologic studies that examined ambient air Pb-PM 

35 concentrations found associations with respiratory morbidity in children (Gent et al., 
February 2012 5-387 Draft – Do Not Cite or Quote

1 2009; Hong et al., 2007b). Adjusting for season, day of week, and date, Gent et al. (2009)

2 found that an increase in lag 0-2 average Pb-PM2.5 was associated with elevated odds of 

wheeze (OR: 1.03, p = 0.13 per 5 ng/m3 3 increase in Pb PM2.5), shortness of breath (OR = 

4 1.12, p = 0.01), and fast-acting inhaler use (OR: 1.04, p = 0.10) among children with 

5 asthma (ages 4-12 years), followed for 1 year during 2000-2003. Among children in 

6 Korea (grades 3-6), a 1 log increase in lag 1 Pb PM10 was associated with a -6.83 L/min 

7 decrease (p < 0.01) in morning peak expiratory flow and a −6.37 L/min decrease 

8 (p < 0.01) in daily average peak expiratory flow adjusting for age, sex, height, weight, 

9 smoking exposure, and meteorological factors (Hong et al., 2007b). Older and recent 

10 toxicological studies also found Pb-containing CAPs to induce pulmonary inflammation. 

11 Uzu et al. (2011) specifically tested particles emitted at a Pb recycling plant and found 

12 that these Pb-rich particles increased the release of the cytokine granulocyte-macrophage 

13 colony-stimulating factor from human epithelial cells. Other toxicological studies found 

14 pulmonary inflammation induced by CAPs in which Pb was one of numerous 

15 components (Wei et al., 2011; Duvall et al., 2008; Godleski et al., 2002; Saldiva et al., 

16 2002). As with blood Pb, epidemiologic studies did not consistently find associations of 

17 ambient air Pb-PM concentrations with respiratory-related hospitalizations or mortality in 

18 older adults. Both studies adjusted for meteorological factors and for temporal trends. 

Among adults ages 65 years and older in 6 California counties, a 4 ng/m3 19 increase in lag 

20 3 Pb-PM2.5 was associated with an increased relative risk of respiratory mortality all year 

21 of 1.01 (95% CI: 0.99, 1.03) and during summer months (quantitative results not 

22 reported) (Ostro et al., 2007). However, among adults ages 65 years and older in 106 U.S 

23 counties, Bell et al. (2009) found that an increase in lag 0 Pb-PM2.5 was associated with a 

24 decrease in respiratory hospital admissions. The 95% CI was wide, indicating lack of 

25 precision in effect estimate.

26 Although limited available recent findings suggest a relationship between respiratory 

27 effects in children and short-term (over several days) changes in ambient air Pb-PM

28 concentrations, it is important to note uncertainties that limit the ability to draw 

29 conclusions regarding airborne Pb exposure. Size distribution data for Pb-PM are 

30 relatively limited, so it is difficult to assess the representativeness of these concentrations 

31 to population exposure (Section 3.5.3). Moreover, data on the relationship between blood

32 Pb and air Pb are relatively limited (see Section 4.5.1) and do not characterize 

33 relationships of varying Pb-PM size distribution with blood Pb level. In several air-Pb 

34 studies, other PM components such as elemental carbon (EC), copper (Cu), and zinc (Zn)

35 also were associated with respiratory effects. In the absence of detailed data on 

36 correlations among all PM components, measurements on other co-occurring ambient 

37 pollutants, or results adjusted for copollutants, it is difficult to exclude confounding by 

38 ambient air exposures to other PM components or ambient pollutants. In several studies 

39 that analyzed PM component mixtures, of which Pb particles comprised one component, 
February 2012 5-388 Draft – Do Not Cite or Quote

1 it is not possible to attribute the observed associations or lack of associations specifically 

2 to Pb (Sarnat et al., 2008; Andersen et al., 2007; Veranth et al., 2006; Maciejczyk and 

3 Chen, 2005).

5.6.7 Immune Effects of Lead within Mixtures

4 Several toxicological studies published since the 2006 Pb AQCD examined immune 

5 effects resulting from exposures to metal mixtures. Some studies indicated that immune 

6 effects may be observed with lower levels of Pb exposure when they occur in conjunction 

7 with other metals. In a study of mice treated with Pb-acetate (10 mg/kg i.p. injection by 

8 weight, daily for 15 days), As (0.5 mg/kg i.p. injection by weight, daily for 15 days), or 

9 both, Bishayi and Sengupta (2006) reported a greater than additive effect of co10 administered Pb and As on macrophages in decreasing bacterial resistance, 

11 myeloperoxidase (MPO) release, and NO production. Investigators assessed the Pb-As 

12 interaction on MPO release using the multivariate ANOVA and constructing an 

13 isobologram by running an ordinary least squares regression between effects (% MPO 

14 release) and dose levels of metals (single and multimetal) in log-linear form.

15 Epidemiologic studies have not widely examined interactions between Pb and other 

16 metals. However, consistent with Bishayi and Sengupta (2006), Pineda-Zavaleta et al.

17 (2004) (Section 5.6.5.2) found interactions between Pb and As among children in Mexico 

18 aged 6-11 years. Contamination of drinking water by both Pb and As was a concern in 

19 the study area; however, urinary As levels were higher in children who had lower blood 

20 Pb levels. In multiple regression analyses, urinary As was associated with lower NO 

21 release from macrophages (similar to blood Pb). An interaction was observed between Pb 

22 and As, which indicated that high internal doses of both metals were associated with a 

23 larger decrease in NO than was either metal alone (p for interaction = 0.037). Urinary As 

24 was associated with lower superoxide anion release (opposite direction of Pb). The 

25 interaction between Pb and As indicated that higher internal doses of both Pb and As 

26 were associated with a larger increase in superoxide anion than was blood Pb level alone 

27 (p for interaction = 0.042). Due to the high blood Pb in these children (means in three 

28 groups at varying distances from a Pb smelter: 7, 20.6, 30.4 μg/dL), it is not clear 

29 whether these relationships would apply to children with lower blood Pb levels.

30 Results from Institoris et al. (2006) indicated that metal co-exposures potentiated the 

31 effects of Pb. Lymph node weight decreased with exposure of 4 week-old rats to 

32 20 mg/kg Pb-acetate by drinking water plus a second metal (Cd or Hg) but not with 

33 20 mg/kg of Pb alone. In contrast with the aforementioned studies, Fortier et al. (2008)

34 did not find Hg co-exposure to increase the effects of Pb. PbCl2-exposed (7.5-20.7 μg/dL) 
February 2012 5-389 Draft – Do Not Cite or Quote

1 human leukocytes did not have alterations in lymphocyte proliferation, monocytic 

2 phagocytic activity, or NK cell activity. The combination of 20.7 μg/dL PbCl2 plus 

3 12.0 μg/dL methylmercuric chloride (MeHgCl) decreased lymphocyte proliferation; 

4 however, these effects were attributed to MeHgCl, which had a stronger suppressive 

5 effect independently. Other toxicological studies found immune effects of multiple metal 

6 mixtures that included Pb (e.g., decreased antibody titers, increased neutrophil counts)

7 (Jadhav et al., 2007; Massadeh et al., 2007) but did not test each metal individually. Thus, 

8 it cannot be ascertained whether findings are due to interactions between Pb and other 

9 components within the mixture. Overall, results indicated that exposures to Pb-containing 

10 metal mixtures are associated with immune effects. Moreover, several studies found an 

11 interaction between Pb and metals such as As, Cd, and Hg, suggesting that a threshold for 

12 producing Pb-induced immune effects may be lower if additional metals are present.

5.6.8 Summary and Causal Determination

13 The collective body of evidence demonstrates that Pb exposure is associated with 

14 immune effects. The evidence indicates that rather than resulting in overt cytotoxicity to 

15 lymphoid tissues, Pb exposure is associated predominantly with subtle changes in a 

16 spectrum of cellular and humoral immune responses, most notably, altered function of T 

17 lymphocytes and macrophages, suppression of the DTH response and resistance to 

18 bacterial infection, increased IgE production, changes in cytokine production, and 

19 inflammation. The strength of evidence for Pb-associated immune effects is derived not 

20 only from the consistency of findings within particular endpoints but also from the 

21 coherence of findings among the spectrum of immune changes operating within the same 

22 pathway. In particular, the evidence integrated across the examined spectrum of immune 

23 outcomes clearly indicates that the prominent immune effect of Pb exposure is to shift 

24 responses from a Th1 phenotype toward a Th2 phenotype.

25 The weight of evidence is provided largely by an extensive body of animal studies 

26 characterizing effects on the broad range of immune endpoints as described above. 

27 Comprising a smaller body of evidence, epidemiologic studies in children are consistent 

28 with toxicological studies in reporting associations of blood Pb levels with indicators of 

29 increased Th2 activity, principally, higher levels of IgE, asthma, and allergy in children. 

30 Across outcomes, because most studies examined concurrent blood Pb levels, there is 

31 uncertainty regarding the critical timing, frequency, duration, and level of Pb exposures 

32 in children that contributed to observed associations. In toxicological studies, the shift to 

33 a Th2 phenotype is well characterized by observations that Pb exposures suppress the 

34 production of Th1 cytokines (e.g., IFN-γ) and increase production of Th2 cytokines 

35 (e.g., IL-4) (Section 5.6.5.4). In animal studies, the shift to Th2 cytokine production was 
February 2012 5-390 Draft – Do Not Cite or Quote

1 observed in juvenile animals with prenatal Pb exposures and in adult animals with long2 term (> 4 weeks) Pb exposures [Table 5-7 in the 2006 Pb AQCD (U.S. EPA, 2006g)]. 

3 Blood Pb levels of animals were reported infrequently, and a wide range of Pb exposures 

4 was found to induce changes in cytokine levels. In a recent study, lifetime Pb exposure 

5 that resulted in blood Pb levels between 2-3 μg/dL suppressed Th1 cytokine production 

6 in adult mice (Iavicoli et al., 2006a). A few available epidemiologic studies in children

7 (Hsiao et al., 2001; Lutz et al., 1999) found higher levels of IL-4, a Th2 cytokine, in 

8 groups of children with higher concurrent blood Pb levels (e.g., mean 8.8 μg/dL or range 

9 15-19 μg/dL, respectively); however, these studies are limited by their lack of 

10 consideration of potential confounding variables and lack of information on the 

11 concentration-response function.

12 Coherence for a Pb-associated skewed ratio of cytokine production is provided by 

13 evidence demonstrating Pb-induced activation of T cells in animals, with recent studies 

14 describing mechanisms underlying T cell activation (Sections 5.6.2.1 and 5.6.2.2). A 

15 recent toxicological study expanded the extant evidence by showing in vivo and in vitro 

16 that Pb may promote Th2 responses by directly increasing production of Th2 cytokines in 

17 dendritic cells, the major effector in antigen response (Gao et al., 2007). Pb-induced T 

18 cell activation also is indicated by a relatively small body of evidence for prenatal and 

19 postnatal Pb exposures of animals resulting in the generation of autoantibodies in 

20 response to new antigens. These findings suggest that Pb exposure may increase the risk 

21 of developing autoimmune conditions (Section 5.6.4.4). In epidemiologic studies, 

22 evidence for the effects of Pb on T cells comprised associations of concurrent blood Pb 

23 levels > 10 μg/dL in children with lower T cell abundance, in particular CD3+ cells. 

24 However, the functional relevance of these changes is unclear. 

25 In addition to T cell responses, a prominent effect of Pb exposure, as demonstrated in an 

26 extensive historical toxicological evidence base, was the induction of macrophages into a 

27 hyperinflammatory state as characterized by enhanced production of ROS, suppressed 

28 production of NO, enhanced production of TNF-α, and excessive metabolism of 

29 arachidonic acid into immunosuppressive metabolites (e.g., PGE2). Consistent with these 

30 observations, a previous epidemiologic study examined and found greater release of ROS 

31 and lower release of NO from macrophages, primarily in children with concurrent blood 

32 Pb levels 10.31-47.49 μg/dL (Pineda-Zavaleta et al., 2004). Misregulated inflammation 

33 represents one of the major modes of action for Pb-induced immune effects. 

34 Toxicological studies provide evidence for the modulation of inflammatory cell function, 

35 production of pro-inflammatory cytokines and metabolites, enhanced inflammatory 

36 chemical messengers, and pro-inflammatory signaling cascades. In addition to the 

37 associations reported with IL-4, epidemiologic evidence for Pb effects on inflammation is 

38 limited to a few recent studies in nonoccupationally-exposed adults in which concurrent 
February 2012 5-391 Draft – Do Not Cite or Quote

1 blood Pb level was associated with other indicators of inflammation such as CRP

2 (Songdej et al., 2010) and IL-6 (Kim et al., 2007). The studies commonly adjusted for 

3 potential confounding by age, sex, BMI, and smoking status. However, because only 

4 concurrent blood Pb levels were examined, there is uncertainty regarding the magnitude, 

5 timing, frequency, and duration of Pb exposures that contributed to the observed 

6 associations. 

7 The toxicological evidence for Pb-induced production of Th2 cytokines provides 

8 biological plausibility for the evidence linking Pb exposure with elevated IgE levels. An 

9 increase in IL-4 from activated Th2 cells induces differentiation of B cells into antibody10 producing cells, thereby amplifying B cell expansion to secrete IgE, IgA, and IgG. 

11 Animal studies describe Pb-induced (resulting in blood Pb levels 10-30 μg/dL) increases 

12 in IgE (Snyder et al., 2000; Miller et al., 1998; Heo et al., 1996). Additionally, 

13 epidemiologic studies in children consistently demonstrated associations between higher 

14 concurrent blood Pb levels and higher in serum IgE (Section 5.6.3). While most studies 

15 found elevated IgE in groups of children with blood Pb levels > 10 μg/dL, Karmaus et al.

16 (2005) found higher serum in IgE in children with blood Pb levels 2.8-3.4 μg/dL 

17 compared with children with lower blood Pb levels. A few studies considered potential 

18 confounding by factors such as age (Karmaus et al., 2005; Lutz et al., 1999), smoking 

19 exposure, serum lipids, organochlorine biomarkers, and number of previous infections

20 (Karmaus et al., 2005) but not SES or allergen exposure. It is important to acknowledge 

21 the extensive evidence in animals for Pb-induced increases in IgE that is not subject to 

22 confounding by SES. Additional support for effects on IgE is provided by toxicological 

23 evidence for Pb-induced increases in Th2 cytokines. Epidemiologic studies did not find 

24 blood Pb level to be associated consistently with B cell abundance.

25 The toxicological evidence for Pb-induced suppression of Th1 cytokine production is 

26 coherent with historical observations in animals that Pb exposure decreases responses to 

27 antigens and bacterial infection (U.S. EPA, 2006b). Toxicological studies provide clear, 

28 consistent evidence that prenatal and postnatal Pb exposure of animals (resulting in blood 

29 Pb levels 11->100 μg/dL) suppresses the DTH response to antigens (Section 5.6.2.3), and 

30 a recent in vitro study indicates such effects may be mediated by dendritic cells (Gao et 

31 al., 2007). Recent toxicological studies provided additional support for Pb-induced 

32 decreased host resistance by demonstrating that Pb exposure impairs phagocytic and 

33 chemotactic activity of macrophages (Lodi et al., 2011; Bussolaro et al., 2008). These 

34 observations reflect suppressed Th1 activity given the role of Th1-dependent IFN-γ in 

35 enhancing the killing capacity of macrophages. Epidemiologic evidence for Pb-associated 

36 diminished response to bacterial infection is limited to previous observations of reduced 

37 neutrophil functionality in Pb-exposed workers with mean blood Pb levels > 30 μg/dL. 

38 While a few previous epidemiologic studies in children found higher prevalence of 
February 2012 5-392 Draft – Do Not Cite or Quote

1 respiratory infections in children with higher blood Pb levels (Karmaus et al., 2005; 

2 Rabinowitz et al., 1990), the findings are limited by the weak statistical methods of 

3 studies and the lack of consideration of potential confounding variables.

4 The effects of Pb exposure on macrophages also suggest a role for the immune system in 

5 mediating Pb-associated effects in multiple other physiological systems. A small body of 

6 new toxicological studies indicated Pb-induced changes in specialized macrophages in 

7 nonlymphoid tissue such as alveolar macrophages, testicular macrophages, and brain 

8 microglia (Section 5.6.4.5); however, these studies primarily used the i.p. route to 

9 administer Pb. Thus, the relevancy of observations to those expected from typical routes 

10 of human exposure is not clear.

11 Th2-dependent increases in IgE mediate type 1 hypersensitivity resulting in various 

12 allergic conditions and asthma. Observations of Pb-associated increases in Th2 cytokines 

13 and circulating IgE levels provide biological plausibility for the small body of available 

14 epidemiologic evidence indicating associations of blood Pb levels with asthma and 

15 allergic conditions in children (Jedrychowski et al., 2011; Pugh Smith and Nriagu, 2011; 

16 Joseph et al., 2005). Several of these studies considered a larger set of potential

17 confounding variables than did studies of IgE in children. While the set of particular 

18 factors varied among studies, studies frequently considered SES indicators and/or 

19 residential allergen measurement. Low SES has been associated with higher blood Pb 

20 levels, poorer housing conditions, higher exposures to mouse and cockroach allergen, and 

21 with conditions such as asthma and allergy. Jedrychowski et al. (2011) found similar 

22 magnitudes of association between cord blood Pb level and cord blood IgE in models that 

23 did and did not adjust for potential confounding variables, including maternal education. 

24 Studies examining cognitive effects in children also found that associations with blood Pb 

25 levels with and without adjustment for SES-related variables (Section 5.3). Pugh Smith et 

26 al. (2011) and Jedrychowski et al. (2011) also indicated lack of confounding by allergen 

27 exposures and other indicators of housing condition by their inclusion as model 

28 covariates or analysis of their independent associations with outcomes. The robust 

29 association between blood Pb level and allergic sensitization (which indicates elevated 

30 allergen-specific IgE) observed in Jedrychowski et al. (2011), provides support for the 

31 associations observed between blood Pb and IgE, in children, in which confounding by 

32 SES and allergen was not considered. Collectively, these findings do not indicate that 

33 confounding by SES or allergen exposure alone accounts for associations observed 

34 between blood Pb levels and immune effects in children. This evidence for associations 

35 of blood Pb level with clinical conditions such as asthma, allergic sensitization, and 

36 allergic diseases in children expanded by results from recent studies also supports the 

37 public health significance for Pb-associated immune effects.
February 2012 5-393 Draft – Do Not Cite or Quote

1 With respect to lifestages of Pb exposure, animal studies found that gestational Pb 

2 exposures, encompassing a wide range of concentrations, affected endpoints such as IgE, 

3 cytokine levels, and DTH but also found postnatal short-term (multiple days) long-term 

4 (multiple weeks) Pb exposures to affect cytokine levels in adult animals. The blood Pb 

5 levels and Pb exposure lifestage, magnitude, frequency, and duration associated with 

6 immune effects are not well characterized in humans. Several epidemiologic studies 

7 examined associations (of IgE, asthma, and allergy) with concurrent blood Pb levels of 

8 children > 10 μg/dL. A few studies found cord blood or newborn hair Pb levels to be 

9 associated with endpoints such as Ig levels (Annesi-Maesano et al., 2003; Belles-Isles et 

10 al., 2002) and allergic sensitization (Jedrychowski et al., 2011). The small body of 

11 epidemiologic studies of nonoccupationally-exposed adults examined different endpoints 

12 and found associations with concurrent blood Pb levels, which are influenced by current 

13 Pb exposures as well as cumulative Pb stores in bone.

14 In summary, recent toxicological and epidemiologic studies support the strong body of 

15 evidence presented in the 2006 Pb AQCD that Pb exposure is associated with a broad 

16 spectrum of changes in both cell-mediated and humoral immunity that cumulatively 

17 promote a Th2 phenotype and hyperinflammatory state. The principal findings are Pb18 induced increased production of Th2 cytokines, suppressed production of Th1 cytokines, 

19 increased inflammation, and elevated IgE, with the weight of evidence provided by 

20 toxicological studies. Collectively, these findings are coherent with the observed effects 

21 of Pb exposure on decreasing responses to antigens (e.g., DTH, bacterial resistance) in 

22 animals. Both toxicological and epidemiologic studies in children provide evidence for 

23 Pb-associated increases in IgE. The toxicological and epidemiologic findings for Th2 

24 cytokines, IgE, and inflammation provide biological plausibility for associations 

25 observed for blood Pb levels with asthma and allergic conditions in children. 

26 Associations with asthma and allergy were observed after considering potential 

27 confounding by several factors, including, SES and allergen exposure. Animal studies 

28 found a range of immune effects with prenatal exposure in juvenile animals and long29 term postnatal (> 4 weeks) Pb exposures in adult animals. The blood Pb levels and Pb 

30 exposure lifestage, magnitude, frequency, and duration associated with immune effects 

31 are not well characterized in children or adults. Epidemiologic studies of children and 

32 adults primarily examined concurrent blood Pb levels. Little information was provided on 

33 concentration-response functions. In epidemiologic studies, higher IgE and higher asthma 

34 prevalence were examined and found in children with blood Pb levels > 10 μg/dL. In the 

35 large body of studies of adults (mostly males) with occupational Pb exposures, the most 

36 consistent findings were decreased neutrophil functionality in workers with mean blood 

37 Pb levels 21-71 μg/dL. Recent epidemiologic studies provided new evidence in adults 

38 without occupational Pb exposures; however, each examined a different immune 

39 endpoint, for example, IgE, eNO, IL-6. These endpoints were associated with concurrent 
February 2012 5-394 Draft – Do Not Cite or Quote

1 blood Pb levels in populations of adults with mean blood Pb levels of 1.9-7 μg/dL; 

2 however, there is uncertainty regarding the contributions of current Pb exposures and 

3 cumulative Pb stores in bone. The consistency and coherence of findings across the 

4 continuum of related immune parameters that demonstrate a stimulation of Th2 responses 

5 in toxicological studies combined with the supporting epidemiologic evidence in children 

6 are sufficient to conclude that there is a causal relationship between Pb exposures and 

7 immune system effects.

5.7 Effects on Heme Synthesis and Red Blood Cell Function

5.7.1 Summary of Findings from 2006 Pb AQCD

8 The 2006 Pb AQCD reported that Pb affects developing red blood cells (RBCs) in 

9 children and occupationally exposed adults as noted by anemia observed with blood Pb > 

10 40 μg/dL. Pb-induced anemia is thought to occur due to decreased RBC life span and 

11 effects on hemoglobin (Hb) synthesis. The exact mechanism for these effects was not 

12 known, although Pb-induced changes on iron uptake or inhibition of enzymes in the heme 

13 synthetic pathway may be responsible. 

14 The 2006 Pb AQCD indicated that Pb crosses RBC membranes through passive 

15 (i.e., energy-independent) carrier-mediated mechanisms including a vanadate-sensitive 

Ca2+ 16 pump. Once Pb enters the cells, it is predominantly found in protein-bound form, 

17 with Hb and aminolevulinic acid dehydratase (ALAD) both identified as targets. Pb 

18 poisoning (blood Pb levels > 100 μg/dL) was found to decrease RBC survival in 

19 laboratory animals, as well as alter RBC mobility and morphology, although the precise 

20 mechanisms by which it does so are not known. Pb exposure has been found to 

21 significantly decrease several hematological parameters including Hb, hematocrit (Hct), 

22 mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), and mean 

23 corpuscular hemoglobin concentration (MCHC). Pb has also been observed to exert 

24 multiple effects on RBC membranes, including altered microviscosity and fluidity, 

25 decreased sialic acid content, decreased lamellar organization, decreased lipid resistance 

26 to oxidation (possibly mediated by perturbations in RBC membrane lipid profiles), and 

27 increased permeability. These alterations to RBC membranes potentially lead to RBC 

28 fragility, abnormal cellular function, RBC destruction, and ultimately anemic conditions. 

29 Pb exposure also has been shown to result in increased activation of RBC scramblase, an 

30 enzyme responsible for the expression of phosphotidylserine (PS) on RBC membranes. 

31 This expression of PS decreases the life span of RBCs via phagocytosis by macrophages. 

32 Pb exposure has been observed to alter the phosphorylation profiles of membrane 
February 2012 5-395 Draft – Do Not Cite or Quote

1 proteins, which may influence the activity of membrane enzymes and the functioning of 

2 receptors and channels located on the membrane. 

3 The 2006 Pb AQCD reported that Pb affects heme synthesis through the inhibition of 

4 multiple key enzymes, most notably ALAD, the enzyme that catalyzes the second, rate5 limiting step in heme biosynthesis (Figure 5-45 presents a schematic representation of the 

6 heme biosynthetic pathway). The 2006 Pb AQCD further reported that decreased RBC 

7 ALAD activity is the most sensitive measure of human Pb exposure, in that measurement 

8 of ALAD activity is correlated with blood Pb levels. Concentration-response changes in 

9 the ratio of activated/nonactivated ALAD activity in avian RBCs was observed to be not 

10 dependent on the method of Pb administration. The inhibition of the ALAD enzyme was 

11 observed in RBCs from multiple species, including birds, Cynomolgous monkeys, and 

12 humans. Pb was also observed to inhibit other enzymes responsible for heme 

13 biosynthesis, including ferrochelatase, porphobilinogen (PBG) deaminase, and 

14 coproporphyrinogen oxidase. Pb also potentially alters heme biosynthesis through 

15 inhibition of transferrin (TF) endocytosis and iron transport. 

16 Pb has been found to alter RBC energy metabolism through inhibition of enzymes 

17 involved in anaerobic glycolysis and the pentose phosphate pathway. Pb was also found 

18 to inhibit pyrimidine 5'-nucleotidase (P5N) activity and the 2006 Pb AQCD indicated that 

19 this might be another possible biomarker of Pb exposure. Inhibition of P5N results in an 

20 intracellular increase in pyrimidine nucleotides leading to hemolysis and potentially 

21 ultimately resulting in anemic conditions. The 2006 Pb AQCD indicated that 

22 perturbations in RBC energy metabolism may be related to significant decreases in levels 

23 of nucleotide pools, including nicotinamide adenine nucleotide (NAD), possibly due to 

24 decreased NAD synthase activity, and nicotinamide adenine nucleotide phosphate 

25 (NADP) accompanying significant increases in purine degradation products. 

Pb was found to alter the activity of membrane-bound ion pumps. Potassium (K+ 26 ) 

27 permeability was found to be increased by Pb due to altered sensitivity of the membrane 

calcium (Ca2+)-binding site that caused selective efflux of K+ 28 ions from the RBC 

membrane. Inhibition of RBC sodium (Na+)-K+ 29 adenosine triphosphate synthase 

30 (ATPase), acetylcholinesterase (ACh), and NADH dehydrogenase was also observed. In 

human RBCs, Na+-K+ ATPase activity was more sensitive to Pb exposure than were Ca2+ 31

or magnesium (Mg2+ 32 ) ATPases.

33 The 2006 Pb AQCD identified oxidative stress as an important potential mechanism of 

34 action by which Pb exposure induced effects on RBCs. Increased lipid peroxidation and 

35 inhibition of antioxidant enzymes (e.g., superoxide dismutase [SOD], catalase [CAT]) 

36 were observed following exposure to Pb. 
February 2012 5-396 Draft – Do Not Cite or Quote

5.7.2 Red Blood Cell Functions

1 As stated in the 2006 Pb AQCD, Pb poisoning has been associated with anemia resulting 

2 from shortened RBC life span and Pb effects on Hb synthesis. As of 2006, the 

3 mechanism for this was not clear, but it was determined not to be due to iron deficiency, 

4 which can be found to occur independently of Pb exposure. However, Zimmerman et al.

5 (2006) found that blood Pb levels were statistically significantly lower in non- or mildly 

6 anemic, iron-deficient 5- to 9-year old children in India fed an iron-fortified diet for 30 

7 weeks compared to 14 weeks (mean [range]: 8.1 [3.1-219] μg/dL versus 12.1 [3.7-

8 26.8] μg/dL; p < 0.02); blood Pb levels were not lower in children receiving the no iron 

9 diet for 30 weeks compared to 14 weeks (mean [range]: 10.2 [4.4-25.3] μg/dL versus 

10 12.0 [3.8-25.5] μg/dL). Although a number of epidemiologic studies find decreases in 

11 RBCs and/or Hct levels associated with blood Pb, it is not known whether this is due to 

12 reduced cell survival or a decrease in RBC cell production. However, decreased RBC 

13 survival and hematopoiesis can be expected to occur simultaneously, and any effect on 

14 RBC numbers is likely a combination of the two modes of action.

5.7.2.1 Pb Uptake, Binding, and Transport into Red Blood Cells

15 The 2006 Pb AQCD reported that Pb uptake into human RBCs occurs via passive anion 

16 transport mechanisms. Although Pb can passively cross the membrane in both directions, 

17 little of the Pb is found to leave the cell after entry. Simons (1993b) found that in vitro 

uptake of 203 18 Pb (1-10 μM) occurred via an anion exchanger while the efflux occurred via 

19 a vanadate-sensitive pathway. After entry into the RBC, radioactive Pb was found to 

20 partition with Hb at a ratio estimated to be about 6000:1 bound to unbound (Simons, 

21 1986). However, Bergdahl et al. (1997a) suggested that ALAD was the primary Pb 

22 binding protein and not Hb. The 2006 Pb AQCD also reported that the majority 

23 (approximately 98%) of Pb accumulates in RBC cytoplasm bound to protein and only 

24 about 2% is found in the membrane. This is related to the high ratio of Pb in RBCs 

25 compared to plasma Pb. Further information on Pb binding and transport in blood can be 

26 found in the kinetics section of 0 (Section 4.2).

27 Although no studies were indentified that examined transport of Pb into RBCs, Lind et al.

28 (2009) recently observed that several zinc (Zn) ionophores (8-hydroxyquinoline 

29 derivatives and Zn and Na pyrithione) were able to effectively transport Pb out of RBCs 

30 into the extracellular space. 
February 2012 5-397 Draft – Do Not Cite or Quote

5.7.2.2 Red Blood Cell Survival, Mobility, and Membrane Integrity

1 Although Pb exposure has been consistently shown to shorten the RBC life span and alter 

2 RBC mobility, as of the 2006 Pb AQCD the mechanism of this was not well understood. 

3 While the mechanism is still not fully understood, there has been some indication for a 

role of free Ca2+ 4 . There are also newer studies that examine the relationship between Pb 

5 and RBC survival, mobility, and membrane integrity. Occupational studies investigating 

6 severely Pb-intoxicated worker populations (mean blood Pb > 28 μg/dL) observed 

increased intracellular RBC free calcium levels ([Ca2+ 7 ]i) and decreased RBC membrane 

Ca2+-Mg2+ 8 -ATPase activity in workers compared with unexposed controls (Abam et al., 

2008; Quintanar-Escorza et al., 2007). [Ca2+ 9 ]i levels were highly correlated with blood Pb 

10 levels even among unexposed control populations with blood Pb levels < 10 μg/dL (mean 

9.9 ± 2 μg/dL) (Quintanar-Escorza et al., 2007). Changes in [Ca2+ 11 ]i were associated with 

12 increased fragility of the RBCs and dramatic morphological alterations, including the 

13 increased presence of echinocytes (cells without normal biconcave shape) and crenocytes 

14 (speculated cells) in Pb-exposed workers. 

15 Similar to the associations observed in Quintanar-Escorza et al. (2007), concentration16 dependent effects were observed when RBCs from healthy human volunteers were 

incubated in vitro with Pb nitrate at physiologically-relevant concentrations. [Ca2+ 17 ]i 

18 increased in a concentration-dependent manner when RBCs were exposed to 0.2 or 

19 0.4 μM Pb nitrate for 24 or 120 hours (0.4 μM Pb nitrate roughly approximates 10 μg/dL 

20 Pb, although concentrations in exposure media are not directly comparable to blood Pb 

levels) (Quintanar-Escorza et al., 2010). [Ca

2+ 21 ]i levels were further increased at higher, 

22 less physiologically-relevant concentrations (i.e., 2-6 μM Pb nitrate). The increase in 

[Ca2+]i levels was observed to be related to increased Ca2+ 23 influx: RBCs exposed to 

0.4 μM Pb nitrate for 24 hours incorporated twice as much Ca2+ 24 as controls did, whereas 

25 exposure for 120 hours resulted in a fourfold increase. Concomitant to increased influx of 

Ca2+, efflux of Ca2+ was observed due to reduced Ca2+-Mg2+ 26 ATPase activity; exposure to 

27 0.4 μM Pb nitrate for 24 or 120 hours reduced activity by 33% and 83%, respectively, 

28 compared to controls. As was observed among severely Pb-intoxicated workers, changes 

in [Ca2+ 29 ]i were associated with increased fragility of the RBCs (measured as increased 

30 hemolysis) and dramatic morphological alterations, including the increased presence of 

31 echinocytes (cells without normal biconcave shape) and crenocytes (speculated cells) 

32 following exposure to 0.4 μM Pb nitrate. Similarly, Ciubar et al. (2007) found that RBC 

33 morphology was disrupted, with ≥ 50% RBCs having lost the typical discocytic 

34 morphology and displaying moderate to severe echinocytosis following exposure to Pb 

35 nitrate concentrations of 0.5 μM or higher for 24 hours at 37 °C. Exposure of RBCs to 

36 higher concentrations (concentrations not stated) of Pb nitrate resulted in cell shrinkage. 

37 Ademuyiwa et al. (2009) observed that the cholesterol content of RBC plasma 
February 2012 5-398 Draft – Do Not Cite or Quote

1 membranes, but not the phospholipid content, was statistically significantly higher in rats 

2 exposed to 200 ppm Pb-acetate (resulting in mean [SD] blood Pb: 40.63 [9.21] μg/dL) for 

3 three months through drinking water compared to controls. Further, the 

4 cholesterol/phospholipid ratio was increased in the Pb-exposed rats with increased 

5 cholesterol, indicating that RBC membrane fluidity was decreased. 

6 A number of studies have investigated the effect of occupational exposure to Pb on 

7 various inter-connected and related hematological parameters. Most occupationally8 exposed cohorts represent populations highly exposed to Pb, with mean blood Pb levels 

9 ranging from 26-74 μg/dL. Although effects observed within these cohorts may not be 

10 generalizable to the population as a whole, they are useful in demonstrating consistent 

11 effects on a number of hematological parameters, including Hb, MCV, MCH, MCHC, 

12 total RBCs, and packed cell volume (PCV) (Khan et al., 2008; Patil et al., 2006a; Patil et 

13 al., 2006b; Karita et al., 2005). Additionally, these studies were cross-sectional in design; 

14 thus, there is uncertainty regarding the magnitude, timing, frequency, and duration of Pb 

15 exposure that contributed to the observed observations. Workers occupationally exposed 

16 to Pb may also have been co-exposed to other contaminants than can affect the 

17 hematological system although the potential for co-exposure was not assessed in most 

18 studies. A few occupational studies did investigate the effect of moderate occupational Pb 

19 exposure on hematological parameters. Ukaejiofo et al. (2009) studied the hematological 

20 effects of Pb in 81 male subjects moderately exposed to Pb at three different 

21 manufacturing companies in Nigeria for durations between six months and 20 years. Two 

22 control groups were used for comparison (30 individuals from the same industries not 

23 involved in handling Pb and 20 individuals from the same locality but not involved in Pb 

24 handling). The exposed individuals had a mean blood Pb level of 7.00 μg/dL compared to 

25 3 μg/dL in controls drawn from industries not involved in Pb handling (control group I) 

26 and 2 μg/dL in controls drawn from the general population (control group II) (p < 0.05). 

27 Pb-exposed workers had significantly reduced Hb and PCV levels and increased 

28 percentage of reticulocytes. Although the differences were statistically significant 

29 between the exposed and control subjects, the study authors state that the levels in the 

30 exposed subjects were at the lower range of normal for Nigerians. The percent cell lysis 

31 did not differ between controls and exposed workers; however, when workers and 

32 controls were stratified by age, there was a significant increase in cell lysis in workers 

33 under age 30 compared to similarly aged controls in group II (p < 0.01). Conterato et al.

34 (In Press) investigated hematological parameters in automotive painters exposed to Pb in 

35 Brazil. Exposed painters had a mean [SEM] blood Pb concentration of 5.4 [0.4] μg/dL 

36 compared to 1.5 [0.1] μg/dL in controls. The mean [SEM] duration of exposure to Pb in 

37 painters was 133.9 [14.5] months, whereas the controls were not occupationally exposed 

38 to Pb. Although hematocrit, hemoglobin concentration, and the number of RBCs were 

39 statistically significantly decreased in painters compared to controls, they were not 
February 2012 5-399 Draft – Do Not Cite or Quote

1 correlated with blood Pb levels. These parameters were correlated with blood Cd levels 

2 which were also statistically significantly elevated in painters compared to controls 

3 (mean [SEM]: 1.606 [0.074] versus 0.003 [0.001] μg/dL). MCHC and RBC distribution 

4 width (a measure of the variability of RBC widths) were correlated (r = 0.23 and 0.44, 

5 respectively; p < 0.05) with blood Pb although these measures were not statistically 

6 different in painters compared to controls. In an investigation of petrol workers in 

7 Sarajevo, Yugoslavia, examinations on hematological parameters were performed on the 

8 same population five years apart, in 2003 and 2008. Workers (mean [SD] duration of 

9 exposure: 12.1 [9.1] years) had increased blood Pb (mean: 5.96 μg/dL) in 2008 compared 

10 to 2003 (mean: 4.07 μg/dL; mean [SD] duration of exposure: 10.4 [5.5]) (Cabaravdic et 

11 al., 2010). In 2008, levels of MCH and MCHC were significantly decreased from levels 

12 in 2003, whereas RBC numbers, Hb, Hct, and MCV were elevated. Positive correlations 

13 were observed in all subjects between blood Pb and RBC count, Hb, and MCH (r = 

14 0.241, 0.201, and 0.213, respectively; p < 0.05). Taken together, the above occupational 

15 studies provide consistent evidence that very high (blood Pb > 26 μg/dL) and moderate 

16 (blood Pb ~ 5.5-7.0 μg/dL) occupational exposure to Pb reduces the number of RBCs in 

17 circulation. Although this decrease in RBCs observed in highly exposed worker cohorts

18 may be explained by both decreased cell survival and/or disruption of hematopoiesis, the 

19 observation of increased reticulocytes in Ukaejiofo et al. (2009) seems to represent 

20 compensation for decreased RBC survival due to Pb exposure. In a non-occupational 

21 study, the associations between blood Pb levels, calcium, iron, and hemoglobin were 

22 investigated in 55 pregnant Brazilian women (21.9% 14-19 years old, 74.5% 20-34 years 

23 olds, and 3.6% ≥ 35 years old) (Zentner et al., 2008). The majority of women (across all 

24 age groups) had concurrent blood Pb levels below 5 μg/dL (58.2%), although the mean 

25 blood Pb level was not reported; only 5.4% of women had blood Pb levels above 

26 10 μg/dL. The vast majority of women (78.2%) were also observed to have adequate 

27 levels of hemoglobin (≥ 11 g/dL). In a multiple linear regression model, blood Pb was 

28 observed to be negatively correlated with hemoglobin (β = -0.359 g/dL). 

29 Studies in children were generally supportive of effects on hematological parameters 

30 (e.g., Hb, MCV, MCH) observed in occupational adult populations. In two cross31 sectional studies of children measuring blood hemoglobin as the independent variable, 

32 blood Pb levels were observed to decrease with increasing blood Hb. Riddell et al. (2007)

33 found that 21% of children 6 months to 5 years of age living in rural Philippines had 

34 concurrent blood Pb levels greater than 10 μg/dL (total population mean: 6.9 μg/dL). Hb 

35 levels were inversely related to blood Pb, with a decrease of 3% blood Pb associated with 

36 every 1 g/dL increase in Hb. Similarly, in children aged 6-36 months (n = 222) living in 

37 Montevideo, Uruguay, 32.9% of children had blood Pb greater than 10 μg/dL (population 

38 mean [SD]: 9.0 [6.0] μg/dL) (Queirolo et al., 2010). The mean [SD] Hb concentration 

39 was 10.5 [1.5] g/dL, and 44.1% of children were diagnosed as anemic (Hb < 10.5 g/dL). 
February 2012 5-400 Draft – Do Not Cite or Quote

1 Blood Pb levels were higher in anemic children compared to non-anemic (mean [SD]: 

2 10.4 [6.8] versus 7.9 [5.1] μg/dL), and in bivariate regression models, blood Pb decreased 

3 0.65 μg/dL for every 1 g/dL increase in Hb. In children younger than 18 months of age, 

4 blood Pb was, on average, 3.5 ± 1.1 μg/dL higher in anemic children. In children 

5 aged 5-9 years (n = 189) living in Cartegena, Columbia, a smaller percentage (4.7%) of 

6 children had blood Pb > 10 μg/dL (mean [SD]: 5.49 [0.23] μg/dL). The only 

7 hematological parameters that fell outside of their reference values were MCV and MCH, 

8 which were negatively correlated with blood Pb (r = -0.159 [p = 0.029] and -0.171 [p = 

9 0.019], respectively) (Olivero-Verbel et al., 2007). RBC count, which was not observed 

10 to differ from reference values, was positively correlated with blood Pb level (r = 0.208, p 

11 = 0.004). Ahamed et al. (2006) studied 39 male urban adolescents in India who were 

12 separated into groups according to their blood Pb level (group 1: <10 μg/dL [mean 

13 7.4 μg/dL], group 2: >10 μg/dL [mean 13.27 μg/dL]). Although the groups were similar 

14 in age (mean [SD]: 16.59 [0.91] versus 16.76 [0.90] years, respectively), height, weight, 

15 and body mass index, group 2 had a significantly lower PCV compared to group 1. In a 

16 related study, Ahamed et al. (2007) investigated the relationship between blood Pb, 

17 anemia, and other hematological parameters in urban children in India (n = 75). Children 

18 were split into two groups as above: group 1 had blood Pb <10 μg/dL (mean [SD]: 6.89 

19 [2.44] μg/dL, n = 19), whereas group 2 had blood Pb >10 μg/dL (mean [SD]: 21.86 

20 [7.58] μg/dL, n = 56). As with the earlier study, ages were similar between the two 

21 groups: mean [SD]: 4.68 [1.49] and 4.11 [1.77] years, respectively. Hb and Hct were 

22 significantly decreased in group 2, compared to group 1, and children in group 2 had an 

23 increased odds of anemia (OR: 2.87 [95% CI: 1.60, 2.87]) compared to group 1 after 

24 adjustment for age, sex, and area of residence. Similarly, in a study of 340 children (aged 

25 1-5 years) from Karachi, Pakistan, mildly and severely anemic children (mean [SD] Hb 

26 levels: 8.9 [0.9] and 7.4 [0.5] g/dL, respectively) had higher blood Pb levels compared to 

27 non-anemic children (mean [SD] Hb: 12.1 [1.3] g/dL). Mean [SD] blood Pb levels in the 

28 mildly anemic, severely anemic, and nonanemic children were 14.9 [0.81], 21.4 [2.7], 

29 and 7.9 [1.7] μg/dL, respectively (p < 0.01) (Shah et al., 2010). Additionally, Hct, RBC 

30 count, and MCV were all decreased in anemic children versus non-anemic children. 

31 Although statistical analyses were not reported, the levels of Hb, Hct, RBC count, and 

32 MCV in anemic children all fell outside of the reported normal range for these 

33 parameters, whereas the reported values in non-anemic children did not. Blood Pb was 

34 negatively correlated with Hb level in all groups, with the magnitude of negative 

35 correlation increasing with increasing severity of anemia: r = -0.315 (non-anemic 

36 children), -0.514 (mild anemia), and -0.685 (severe anemia). In iron-deficient anemic 

37 children (n = 23) from Denizli, Turkey, mean (SD) serum Pb levels were statistically 

38 increased compared to healthy children (n = 179): 0.013 (0.004) versus 0.008 

39 (0.001) μg/dL, respectively (Turgut et al., 2007). The iron-deficient children were 
February 2012 5-401 Draft – Do Not Cite or Quote

1 observed to have decreased Hb, MCV, and ferritin compared to controls, but increased 

2 RBC and RDW. In 140 children from southern Brazil aged 2-11 years, living within 25 

3 km of a Pb smelter, blood Pb levels were not observed to differ between anemic and 

4 non-anemic children (mean [SD]: 10.36 [6.8] versus 9.73 [5.8] μg/dL, p = 0.98) (Rondo 

5 et al., 2006). However, blood Pb levels were significantly negatively correlated with Hb 

6 in anemic children (r = -0.41, p = 0.01); this relationship was not observed in non-anemic 

7 children (r = 0.018, p = 0.84).

8 Contrary to the observations in the above studies, Huo et al. (2007) found that children 

9 (less than 6 years of age) living near an area where electronic waste was recycled in 

10 China had significantly higher mean blood Pb levels than did children in the neighboring 

11 town with no waste recycling (15.3 versus 9.94 μg/dL). No difference was detected in the 

12 mean Hb levels of the children in the two towns (127.55 g/L in children from the waste 

13 recycling town versus 123.46 g/L in children from the town with no recycling). 

14 Equivocal findings in studies investigating associations of blood Pb levels with 

15 hematological effects in children may be due to the comparatively shorter time period 

16 and magnitude of exposure versus those seen in occupational studies.

17 A number of animal toxicology studies support associations of blood Pb levels with 

18 hematological parameters observed in epidemiologic studies. Baranowska-Bosiacka et al.

19 (2009) examined the effects of Pb on RBC hemolysis both in vitro measuring lysate in 

20 human RBCs incubated with Pb at concentrations ranging from 0.1–100 μM for 5–

21 30 minutes, and in vivo using a rat RBC lysate from rats exposed to Pb-acetate (0.1 %) in 

22 drinking water for 9 months. Rats exposed to Pb in the in vivo portion of the study 

23 achieved a blood Pb concentration of 7.1 μg/dL. The concentration of Hb in the plasma 

24 of chronically-exposed rats, a marker of RBC hemolysis, was statistically significantly (p 

25 = 0.01) increased compared to that in control rats. The in vitro studies demonstrated a 

26 similar concentration-dependent increase in the amount of hemolysis, with a significant 

27 (fourfold) increase even at the lowest concentration tested (i.e., 0.1 μM). Pb-induced 

28 hemolysis in these experiments may be due to inhibition of RBC 

29 phosphoribosyltransferases (Section 5.7.5.1). Lee et al. (2005) observed that rats orally 

30 administered Pb (25 mg/kg) via gavage once a week for 4 weeks had an average plasma 

31 Pb level of 6.5 μg/dL (9.6-fold higher than that in controls, p < 0.05), and had significant 

32 decreases in Hct, Hb, and RBCs (p < 0.05). Male mice orally administered 50 mg/kg Pb 

33 nitrate in distilled water via gavage for 40 days had mean [SD] final blood Pb levels of 

34 1.72 [0.02] μg/dL versus 0.09 [0.011] μg/dL in control mice. Pb-exposed mice had 

35 significantly reduced total RBC counts, total leukocyte counts, Hb, lymphocytes, and 

36 monocytes compared to controls (p < 0.001) (Sharma et al., 2010b). Rats exposed to 2 

37 g/L Pb-acetate in drinking water for 30 days (blood Pb not reported) had significantly 

38 decreased RBCs, Hb, PCV, MCH, and MCHC compared to controls (p < 0.05) (Simsek 
February 2012 5-402 Draft – Do Not Cite or Quote

1 et al., 2009), and not a disruption of hematopoiesis. Mice exposed to 1 g/L Pb-acetate in 

2 drinking water (blood Pb not reported) for 90 days, but not those exposed for 15 or 45 

3 days`, had significantly decreased RBC counts and Hct compared to controls (p< 0.05)

4 (Marques et al., 2006). Spleen weights were observed to be increased relative to body 

5 weight in animals exposed to Pb for 45 days. Mice injected daily with 50 mg/kg 

6 Pb-acetate subcutaneously (blood Pb not reported) had significantly reduced Hb, MCV, 

7 MCH, and MCHC compared to controls injected with 5% dextrose (Wang et al.). In 

8 weanling rats (age = 25 days, n = 10) whose dams were exposed to 2.84 mg/mL 

9 Pb-acetate (approximating mean [SD] daily exposures of 342.57 [28.11] and 744.47 

10 [29.27] mg/kg during gestation and lactation, respectively), blood Pb was significantly 

11 elevated compared to controls (mean [SE]: 698.1 [78.2] versus 5.4 [0.8] ng/g). The only 

12 hematological parameter affected by Pb exposure was Hct, which was decreased in 

13 exposed rats (mean [SE]: 27.3 [0.5] versus 33.4 [0.3] percent) (Molina et al., 2011).

14 Some toxicological studies found no evidence of hematological effects in animals 

15 following exposure to Pb. Male rats administered Pb-acetate in the drinking water for 4 

16 weeks at concentrations ranging from 100–1,000 ppm had a concentration-dependent 

17 increase in blood Pb (range: 6.57–22.39 μg/dL) compared to controls (0.36 μg/dL), but 

18 there were no significant changes in any of the hematological parameters (complete blood 

19 cell count performed) measured at the end of treatment (Lee et al., 2006b). Slight, 

20 statistically nonsignificant increases in PS expression on RBC membranes were also 

21 observed. Similarly, exposure of male rats to 0.5% Pb nitrate in drinking water (blood Pb 

22 not reported) for three weeks had no affect on any measured hematological parameter

23 (Gautam and Flora, 2010). In vitro experiments with rat and human blood did not 

24 demonstrate a significant increase in hemolysis after 4 hours of treatment with Pb-acetate

25 at concentrations up to 10 μM. 

26 Khaĭrullina et al. (2008) observed that the surface profiles of RBC membrane shadows 

27 incubated with 0.5-10 μM Pb-acetate for three hours were much smoother than were 

28 untreated RBC membranes when examined by atomic force microscopy. The authors 

29 postulate that the observed smoothing in Pb-treated RBC membranes may be due to 

30 clusterization of band 3 protein. Band 3 (anion exchanger 1 [AE1]), is a 

chloride/bicarbonate (Cl -/HCO3

- 31 ) exchanger and is the most abundant protein in RBC 

32 membranes. AE1 is integral in carbon dioxide (CO2) transport and linkage of the cellular 

33 membrane to the underlying cytoskeleton (Akel et al., 2007; Su et al., 2007). The 

34 observed smoothing of the RBC membrane may due to Pb interfering with how the 

35 membrane attaches to the cytoskeletal structure of the RBC through perturbation of the 

36 normal activity of AE1.
February 2012 5-403 Draft – Do Not Cite or Quote

Eryptosis 

1 Eryptosis is the suicidal death of RBCs. It is characterized by cell shrinkage, membrane 

2 blebbing, and cell membrane phospholipid scrambling associated with PS exposure on 

3 the cell membrane that leads to cell destruction via macrophages (Föller et al., 2008; 

4 Lang et al., 2008). As previously reported in the 2006 Pb AQCD, Kempe et al. (2005)

5 found that exposing human RBCs to Pb at concentrations ranging from 0.3–3 μM caused 

increased activation of K+ 6 channels that led to cell shrinkage and scramblase activation. 

7 The activation of scramblase increased the exposure to PS on the cell membrane, which 

8 causes an increase in the destruction of the RBCs by macrophages. 

9 Shin et al. (2007) found that in vitro exposure of human RBCs to 1–5 μM Pb-acetate

10 increased PS expression in a time- and concentration-dependent manner. The maximum 

11 mean [SE] increase in expression of PS was 26.8 ± 3.15% (compared to deionized water), 

12 observed after incubation with 5 μM Pb for four hours. The expression of PS in RBCs is 

considered to be regulated through a Ca2+ 13 dependent mechanism and, correspondingly, 

[Ca2+ 14 ]i was observed to increase with exposure to Pb (mean [SE]: 0.24 [0.21] μM in 

15 controls to 6.88 [1.13] μM in RBCs treated with 5 μM Pb for one hour). Consistent with 

16 this finding, Shin et al. (2007) also observed that scramblase activity, which is important 

for induction of PS exposure and is activated by [Ca2+]i 17 , was increased in Pb-exposed 

18 RBCs. Flippase, which translates PS exposure to inner membranes, is inhibited by high 

levels of [Ca2+ 19 ]i and was observed to exhibit reduced activity following Pb exposure. The 

20 inhibition of flippase is additionally influenced by the depletion of cellular adenosine 

21 triphosphate (ATP). ATP levels were decreased in a concentration-dependent manner 

22 following exposure to Pb. To corroborate these findings in vivo, Shin et al. (2007) treated 

23 male rats i.p. to 25, 50, or 100 mg/kg Pb-acetate (blood Pb not reported). Expression of 

24 PS was observed to increase in a concentration-dependent manner at concentrations ≥ 

25 50 mg/kg, confirming the in vitro results. No hemolysis or microvesicle formation was 

26 observed in the in vitro and in vivo experiments. In a follow-up study, the same lab 

27 observed that exposure of human RBCs to low concentrations of Pb-acetate (0.1-0.5 μM) 

28 induced PS expression. Most notably, exposure to 0.1 μM Pb for 24 hours increased PS 

29 expression on RBC membranes by approximately 20% (Jang et al., 2011).

30 Accompanying the expression of PS were abnormal, echinocytic RBCs following 

31 incubation with Pb. Unlike the above study, incubation with low concentrations of Pb 

32 (0.1 μM) induced the generation of microvesicles, which also expressed PS on their 

33 membranes. Flippase was inhibited by 0.1 μM Pb following incubation for one hour, but 

34 scramblase activity was not changed at any Pb exposure concentration. The intracellular 

35 concentration of ATP was decreased at Pb concentrations 0.25 μM and greater, but 

[Ca2+ 36 ]i did not increase following exposure. This decrease in ATP levels, but lack of 

affect on [Ca2+ 37 ]i may explain why flippase activity, but not scramblase, was altered 
February 2012 5-404 Draft – Do Not Cite or Quote

1 following Pb incubation. At 0.5 μM, Pb-exposed RBCs with externalized PS were 

2 observed to be targeted and engulfed by differentiated macrophages. Similar in vitro 

3 effects were observed in rat erythrocytes, although higher concentrations were generally 

4 required. PS expression on rat erythrocytes was observed ex vivo in rats exposed to 10 or 

5 50 mg/kg Pb. To corroborate these in vitro and ex vivo findings, rats were exposed to 0, 

6 50, 250, or 1,000 ppm Pb-acetate in drinking water for 4 weeks. At 100 ppm, Hb and Hct 

7 were significantly decreased relative to control, and liver and spleen weights were 

8 increased. At doses greater than 50 ppm, iron accumulation was observed in the spleen, a 

9 clear sign of increased RBC clearance via phagocytosis.

10 Ciubar et al. (2007) also found that exposure to Pb nitrate (0.5–2 μM) resulted in an 

11 increase in PS exposure to RBCs and cell shrinkage, which authors stated were indicators 

12 of cell apoptosis. As reported above, Khaĭrullina et al. (2008) observed Pb-induced RBC 

13 membrane smoothing that may be due to alterations in AE1 activity. Disruptions in AE1 

14 activity may also result in enhanced PS exposure and premature cell death. Akel et al.

(2007) observed that in AE1-/- 15 knockout mice, Pb-induced PS exposure was much greater 

16 than that in wild type mice. Decreased RBCs and increased reticulocytes were also 

17 observed, an indication of high cell turnover.

5.7.2.3 Red Blood Cell Hematopoiesis

18 Erythropoietin is a glycoprotein hormone excreted by the kidney to promote the 

19 development of RBCs in the bone marrow. Sakata et al. (2007) examined the relationship 

20 between blood Pb level and serum erythropoietin levels in Pb-exposed nonanemic 

21 tricycle taxi drivers (n=27) working in Kathmandu, Nepal (mean [SD] age: 5.6 [2.6] 

22 years). The average blood Pb level in the taxi drivers was 6.4 μg/dL compared to 

23 2.4 μg/dL in nondrivers. Drivers had a significantly lower mean level of serum 

24 erythropoietin (12.7 versus 18.8 mU/mL) compared to the nondrivers and there was an 

25 inverse relationship between the level of serum erythropoietin and blood Pb (r = -0.68, p 

26 < 0.001). Blood Pb level was not associated with any other hematological effects. The 

27 Sakata et al. (2007) study demonstrated that serum erythropoietin levels are affected by 

28 Pb even at levels low enough not to cause anemia. While decreased erythropoietin is 

29 generally considered a measure of kidney toxicity, it can also indicate that Pb could 

30 possibly affect the level of RBCs through decreasing levels of serum erythropoietin.

31 Celik et al. (2005) observed that exposure of female rats to 140, 250, or 500 mg/kg 

32 Pb-acetate via gavage once per week for 10 weeks (blood Pb not reported) resulted in 

33 decreased numbers of polychromatic RBCs (PCE) and increased numbers of 

34 micronucleated PCEs, compared to controls (p < 0.001). Alghazal et al. (2008b) exposed 
February 2012 5-405 Draft – Do Not Cite or Quote

1 male and female rats to 100 mg/L Pb-acetate daily in drinking water for 125 days (blood 

2 Pb not reported) and observed increases in micronucleated PCEs in female rats (p = 0.02) 

3 but no significant reduction in the ratio of PCEs to normochromic RBCs (NCE). In male 

4 rats, an increase in micronucleated PCEs was observed (p < 0.001) along with a decrease

5 in the PCE/NCE ratio (p = 0.02). While the results from Alghazal et al. (2008b) indicate 

6 that Pb is cytotoxic in male rats only, but is genotoxic in both sexes, results from Celik et 

7 al. (2005) indicate that Pb is cytotoxic in female rats as well. Mice exposed to 1 g/L 

8 Pb-acetate in drinking water for 90 days (blood Pb not reported) had statistically 

9 significant increases in micronucleated PCEs; a small, but statistically nonsignificant 

10 decrease in the PCE/NCE ratio was also observed (Marques et al., 2006). Cyto- and 

11 geno-toxicity in RBC precursor cells are strong indications of altered hematopoiesis in 

12 bone marrow.

5.7.2.4 Membrane Proteins

13 There have been few studies examining the effects of Pb on membrane proteins since the 

14 2006 Pb AQCD. According to the 2006 Pb AQCD, Pb has been found to affect RBC 

15 membrane polypeptides in exposed workers (Apostoli et al., 1988; Fukumoto et al., 

16 1983). In Pb-exposed workers, Fukumoto et al. (1983) found decreased levels of 

17 polypeptides in band 3, which Apostoli et al. (1988) suggested may represent an anion 

18 channel protein, and increases in the level of polypeptides in bands 2, 4, 6, and 7. 

19 Fukumoto et al. (1983) suggested that the changes in the RBC membrane polypeptides 

20 may cause changes in membrane permeability. Apostoli et al. (1988) found that the 

21 changes in membrane polypeptides in association with blood Pb levels greater than 

22 50 μg/dL. Exposure to Pb-acetate at concentrations above 0.1 μM for 60 minutes has also 

23 been found to increase the phosphorylation of proteins in human RBC membranes in 

24 vitro (Belloni-Olivi et al., 1996). Phosphorylation did not occur in cells depleted of 

25 protein kinase C (PKC), indicating a PKC-dependent mechanism. 

26 Huel et al. (2008) found that newborn hair and cord blood Pb levels (mean [SD]: 1.22 

27 [1.41] μg/g and 3.54 [1.72] μg/dL) were negatively associated with Ca-ATPase activity 

28 in plasma membranes of RBCs isolated from cord blood; newborn hair Pb levels were 

29 more strongly associated with cord Ca pump activity than were cord blood Pb (p < 

30 0.0001 versus p < 0.05). Maternal blood Pb levels were not correlated with Ca pump 

31 activity in maternal or cord blood. Pb-induced disruptions in Ca homeostasis in RBCs can 

32 lead to cytotoxicity and necrosis, and these effects may be representative of cellular 

33 dysfunction in other organ systems. 
February 2012 5-406 Draft – Do Not Cite or Quote

5.7.3 Red Blood Cell Heme Metabolism

1 Pb has been found to inhibit several enzymes involved in heme synthesis, namely ALAD 

2 (cytoplasmic enzyme catalyzing the second, rate-limiting, step of the heme biosynthesis 

3 pathway), coporphyrinogen oxidase (catalyses the sixth step in heme biosynthesis 

4 converting coporphyrinogen III into protoporphyrinogen IX), and ferrochelatase 

5 (catalyses the terminal step in heme synthesis converting protoporphyrin IX into heme) 

6 (Figure 5-45). The observations of decreased Hb (measured as total Hb, MCH, or 

7 MCHC) in occupationally-exposed adults (Ukaejiofo et al., 2009; Khan et al., 2008; Patil 

8 et al., 2006b; Karita et al., 2005) and Pb-exposed experimental animal models (Sharma et 

9 al., 2010b; Simsek et al., 2009; Marques et al., 2006; Lee et al., 2005) and associations 

10 with blood Pb levels in children (Riddell et al., 2007), is a direct indicator of decreased 

11 heme synthesis due to Pb exposure.

Note: Steps in the pathway potentially affected by Pb are indicated with curved arrows pointing to the affected enzyme, and effects 

are represented by ↑ and ↓ arrows.

Figure 5-45 Schematic representation of the enzymatic steps involved in the 

heme synthetic pathway.
February 2012 5-407 Draft – Do Not Cite or Quote

5.7.3.1 Red Blood Cell 5-Aminolevulinic Acid Dehydratase 

1 Decreases in RBC 5-aminolevulinic acid dehydratase (ALAD) levels are strongly 

2 associated with Pb exposure in humans to such an extent that RBC ALAD activity has 

3 been used to assess Pb toxicity. Several epidemiologic studies published since the 2006 

4 Pb AQCD evaluated the relationship between Pb exposure, blood Pb levels and ALAD 

5 activity in adults and children (see below). These studies were cross-sectional in nature. 

6 This limits their utility in assessing the magnitude, timing, frequency, and duration of Pb 

7 exposure necessary to contribute to the observed effects.

8 As was seen with epidemiologic studies investigating Pb-induced deficits in 

9 hematological parameters, most occupationally-exposed cohorts investigated ALAD 

10 levels in association with high blood Pb levels, i.e., > 27 μg/dL (range: 27.0-74.4 μg/dL). 

11 Although effects observed within these cohorts regarding ALAD levels may not be 

12 generalizable to the population as a whole, they are useful in demonstrating consistent 

13 and negative effects of Pb on the activity of this enzyme (Quintanar-Escorza et al., 2007; 

14 Patil et al., 2006a; Patil et al., 2006b; Ademuyiwa et al., 2005b). Occupationally-exposed 

15 adults had levels of inhibition of ALAD that were as great as 90% relative to control

16 (Quintanar-Escorza et al., 2007). There were few studies that investigated Pb-associated 

17 decrements in ALAD levels among moderately-exposed workers. Painters in India with a 

18 mean blood Pb level of 21.92 μg/dL (mean [SD] duration of exposure: 126.08 [49.53] 

19 months) had lower ALAD levels (p < 0.01) compared to controls whose mean blood Pb 

20 level was 3.06 μg/dL (Mohammad et al., 2008). Stoleski et al. (2008) observed that 

21 workers in a Pb smelter in Macedonia (mean [SD]: 16.4 [8.5] μg/dL blood Pb; 18.8 [7.5] 

22 years employment) had lower ALAD activity (p < 0.001) and higher ALA levels (p < 

23 0.0005) compared to workers with no exposure to Pb (mean [SD] blood Pb: 7.0 

24 [5.4] μg/dL). In automotive painters exposed to Pb in Brazil (mean [SD]: 5.4 [0.4] μg/dL 

25 blood Pb; 133.9 [14.5] months duration of exposure), the ALAD reactivation index was 

26 increased over that in controls; ALAD activity did not differ between groups (Conterato 

27 et al., In Press). However, ALAD activity was negatively correlated with blood Pb (r = -

28 0.59, p < 0.05) but not blood Cd, whereas ALAD reactivation index was positively 

29 correlated with blood levels of both metals (Pb: r = 0.84, p < 0.05; Cd: r = 0.27, p < 

30 0.05). In a benchmark dose (BMD)-based analysis, Murata et al. (2009) calculated the 

31 BMD and 95% lower confidence limit of the BMD (BMDL) for decreased ALAD 

32 activity in RBCs of exposed Pb workers. The calculated BMD and BMDL values of 2.7 

33 and 2.3 μg/dL, respectively, were substantially lower than the BMDs (28.7-44.2 μg/dL) 

34 and BMDLs (19.4-29.6 μg/dL) for decreased Hb, Hct, and RBC count in similarly 

35 exposed workers, indicating decreases in ALAD activity can occur at blood Pb levels that 

36 do not decrease RBC survival.
February 2012 5-408 Draft – Do Not Cite or Quote

1 Wang et al. (2010f) found that there was also a concentration-dependent decrease in 

2 ALAD activity in both children (4-13 years old) and adults (16-77 years old) (mean blood 

3 Pb levels: 7.1 and 6.4 μg/dL, respectively) in rural southwest China. Further, Wang et al.

4 (2010f) observed that the relationship between blood Pb and ALAD activity was 

5 nonlinear and exponential, with larger decreases in ALAD activity occurring with blood 

6 Pb levels greater than 10 μg/dL. No correlation was observed between urinary ALA 

7 levels and blood Pb. Ahamed et al. (2006) studied male urban adolescents in India. The 

8 39 adolescents were separated into groups according to their blood Pb levels (group 1: 

9 <10 μg/dL [mean 7.4 μg/dL], group 2: >10 μg/dL [mean 13.27 μg/dL]). Although group 

10 1 and 2 were similar in age (mean [SD]: 16.59 [0.91] versus 16.76 [0.90] years, 

11 respectively), height, weight, and body mass index, group 2 had lower ALAD activity 

12 than did group 1 (p < 0.001). When all 39 adolescents were examined together, an inverse 

13 relationship was found between blood Pb and ALAD activity. Similar decreases in 

14 ALAD activity were observed in other populations of Indian children (aged 4-12 and 1-7 

15 years) with elevated blood Pb (mean [SD]: 11.39 [1.39] and 21.86 [7.58] μg/dL) 

16 compared to children with lower blood Pb levels (mean [SD]: 3.93 [0.61] and 6.89 

17 [2.44] μg/dL) (Ahamed et al., 2007; Ahamed et al., 2005). Decreases were also observed 

18 in children 3–6 years of age with > 10 μg/dL, compared to children < 10 μg/dL (mean 

19 blood Pb concentration for groups not reported) in northeastern China (Jin et al., 2006).

20 Decreased ALAD activity in response to Pb exposure is also observed in toxicological 

21 studies. Rats administered 500 ppm Pb-acetate in drinking water for 15 or 30 days had 

22 decreased blood ALAD activity that was related to duration of exposure and blood Pb

23 (Rendón-Ramirez et al., 2007). Administration of Pb (25 mg/kg) to rats once a week for 4 

24 weeks achieved a blood Pb level of 6.5 μg/dL, which was associated with statistically 

25 significant decreases (approximately 50% lower than control levels) in RBC ALAD 

26 activity (Lee et al., 2005). Exposure of male Wistar rats to 0.5% Pb-acetate via drinking 

27 water for three weeks statistically significantly decreased ALAD activity 72% compared 

28 to controls (mean [SD]: 7.35 [0.35] versus 26.14 [2.19] nM/min/mL RBCs) (Gautam and 

29 Flora, 2010).

5.7.4 Other Heme Metabolism Enzymes

30 The 2006 Pb AQCD indicated that Pb affects RBC PBG synthase (Simons, 1995; Farant 

31 and Wigfield, 1990, 1987), PBG deaminase (Tomokuni and Ichiba, 1990), and TF 

32 endocytosis and iron transport across membranes (Qian and Morgan, 1990), all of which 

33 are directly or indirectly involved in heme synthesis. Although there are no new studies 

34 that examine the effect Pb has on the activities of other heme metabolism enzymes, a 
February 2012 5-409 Draft – Do Not Cite or Quote

1 number of studies investigated associations of blood Pb with concentrations of various 

2 intermediate products in the heme biosynthetic pathway. 

3 Pb intoxication is known to inhibit the function of ferrochelatase, the enzyme that 

4 catalyzes the last step in the heme biosynthetic pathway. Under normal conditions, 

ferrochelatase incorporates ferrous iron (Fe2+ 5 ) into protoporphyrin IX, converting it into a 

6 heme molecule (Figure 5-45). However, Pb has been shown to inhibit this insertion of 

Fe2+ 7 into the protoporphyrin ring and instead, Zn is inserted into the ring creating ZPP. A 

8 number of recent studies have shown that blood Pb is statistically significantly associated 

9 with increased RBC ZPP levels in adults occupationally exposed to high levels of Pb 

10 (blood Pb level > 27-54 μg/dL) (Patil et al., 2006b; Ademuyiwa et al., 2005b), workers 

11 exposed to moderate levels of Pb (blood Pb level = 21.92 μg/dL) (Mohammad et al., 

12 2008), children aged 1-21 years (blood Pb 18-23 μg/dL) (Counter et al., 2009, 2008; 

13 Counter et al., 2007), and animals exposed to 500 ppm Pb via drinking water for 15 or 30 

14 days (Rendón-Ramirez et al., 2007). Interestingly, Wang et al. (2010f) found that in 

15 children and adults living in a rural area of Southwest China, ZPP levels were negatively 

16 correlated with blood Pb at blood Pb levels < 10 μg/dL and were only positively 

17 correlated with blood Pb at higher blood Pb concentrations (i.e., > 10 μg/dL). The authors 

18 suggest that this may be representative of ALAD activities at low blood Pb levels, which 

19 contributes to lower ZPP levels. Scinicariello et al. (2007) performed a meta-analysis and 

20 observed that Pb-exposed individuals who carried the ALAD2 allele had slightly lower 

21 concentrations of blood ZPP levels compared to carriers of the ALAD1 allele (overall 

22 pooled standardized mean estimate: -0.09 [units not specified]; 95% CI: -0.22, 0.03, p = 

23 0.13).

5.7.5 Other Hematological Parameters

5.7.5.1 Energy Metabolism

24 RBCs use high energy purine nucleotides (i.e., ATP and guanine triphosphate [GTP]) to 

25 support basic metabolic functions. In mature RBCs, these nucleotides are synthesized via 

26 salvage reactions via either an adenine pathway, which requires adenine 

27 phosphoribosyltransferase (APRT), or an adenosine pathway, which requires adenosine 

28 kinase. The 2006 Pb AQCD reported that Pb significantly reduces the nucleotide pool 

29 including NAD and NADP, as well as increases purine degradation products resulting in 

30 altered RBC energetics. Since the 2006 Pb AQCD, there have been few studies 

31 examining Pb effects on energy metabolism. Baranowska-Bosiacka et al. (2009)

32 examined the effects of Pb on RBC APRT and hypoxanthine-guanine 
February 2012 5-410 Draft – Do Not Cite or Quote

1 phosphoribosyltransferase (HPRT) due to in vitro and in vivo exposures. For the in vitro 

2 exposure, APRT and HPRT were measured in lysate of human RBCs after exposure to 

3 Pb at a concentration range from 0.1 to 100 μM for 5–30 minutes. In vivo tests measured 

4 APRT and HPRT in rat RBC lysate from rats exposed to Pb-acetate (0.1 %) in drinking 

5 water for 9 months. Both the in vivo and vitro studies found a significant decrease in both 

6 HPRT and APRT levels. The levels were significantly decreased in vitro after only 

7 5 minutes of exposure to the 0.1 μM concentration, but the decrease was also 

8 concentration-dependent. However, the study authors considered the inhibition moderate 

9 (30–35%) even with the highest levels used in vitro. Shin et al. (2007) found a 

10 concentration-dependent decrease in intracellular ATP in human RBCs in vitro with 

11 significant decreases found even with the lowest concentration (i.e., 1 μM).

5.7.5.2 Other Enzymes

The 2006 Pb AQCD reported that K+ 12 permeability was increased by Pb due to altered 

sensitivity of the membrane Ca2+-binding site that caused selective efflux of K+ 13 ions from 

the RBC membrane. However, inhibition of the RBC Na+-K+ 14 ATPase is more sensitive to 

Pb exposure than is the inhibition of Ca2+-Mg2+ 15 ATPase. Few new studies were found

16 that examined the effects of Pb exposure on other enzymes. Ekinci et al. (2007) tested the 

17 effects of Pb on two carbonic anhydrase isozymes (I and II) isolated from human RBCs. 

18 Carbonic anhydrases are metalloproteins that use Zn to catalyze the equilibrium between 

19 carbon dioxide and bicarbonate in the cells of higher invertebrates. Although 

20 investigators found that Pb nitrate inhibited both carbonic anhydrase isozymes in a 

concentration-dependent manner, the concentrations used (i.e., 200-1,000 μ 21 M) were above 

22 those that would be physiologically relevant. Inhibition of isozyme I was noncompetitive, 

23 while the inhibition for isozyme II was uncompetitive. Bitto et al. (2006) examined the 

24 mechanisms of action of Pb-induced inhibition of P5N, an enzyme important in the 

25 pyrimidine salvage pathway that requires manganese for normal activity. Pb was 

26 observed to bind directly to the active site of the enzyme in a different position than the 

27 manganese, thus possibly resulting in improper protein folding and inhibition of activity. 

5.7.6 Red Blood Cell Oxidative Stress

28 It has been suggested that the Pb-associated decreases in ALAD activity result in 

29 increased oxidative stress, owing to the buildup of ALA. ALA can act as an electron 

30 donor in the formation of reactive oxygen species (ROS) (Nemsadze et al., 2009; 

31 Ahamed and Siddiqui, 2007). Many epidemiologic and toxicological studies have found 
February 2012 5-411 Draft – Do Not Cite or Quote

1 an association between the level of blood Pb and lipid peroxidation, antioxidant levels, or 

2 indicators of ROS production.

5.7.6.1 Oxidative Stress, Lipid Peroxidation, and Antioxidant 

Enzymes

3 Malondialdehyde (MDA) is an end product of lipid peroxidation and is commonly used 

4 as an indicator of lipid peroxidation. Numerous occupational studies demonstrated 

5 increased lipid peroxidation in highly-exposed worker populations (blood Pb levels 

6 ranging from 29 to 74.4 μg/dL) (Kasperczyk et al., 2009; Khan et al., 2008; Quintanar7 Escorza et al., 2007; Patil et al., 2006a; Patil et al., 2006b). There was a correlation 

8 between MDA levels and blood Pb even in the unexposed workers who had lower 

9 (i.e., <12 μg/dL) blood Pb levels, although the magnitude of correlation in exposed 

10 workers was greater. (Quintanar-Escorza et al., 2007). Increases in C-reactive protein and 

11 decreases in RBC SOD, catalase, and plasma ceruloplasm were also observed in these 

12 workers, further indicating increased RBC oxidative stress due to higher Pb exposure.

13 Evidence of lipid peroxidation was also observed in occupational cohorts moderately 

14 exposed to Pb. These studies were cross-sectional in design; thus, there is uncertainty 

15 regarding the magnitude, timing, frequency, and duration of Pb exposure that contributed 

16 to the observed associations. In auto repair apprentices in Turkey (mean [SD]: 16.8 [1.2] 

17 years of age, 3.8 [1.8] years duration of exposure) with blood Pb levels as low as 

18 7.9 μg/dL (Ergurhan-Ilhan et al., 2008), increases in glutathione peroxidase (GPx) and 

19 MDA, as well as decreases in α-tocopherol and β-carotene were observed compared with 

20 controls (compared to 2.6 μg/dL in controls; mean [SD] age: 16.3 [1] years). Decreases 

21 were observed in SOD and CAT, but the results did not attain statistical significance. In 

22 painters in India (mean [SD] duration of exposure: 126.08 [49.53 months]) with a mean 

23 blood Pb level of 21.92 μg/dL (compared to 3.06 μg/dL in controls), there was a 

24 significant decrease in SOD, glutathione (GSH), and CAT accompanied by a significant 

25 increase in oxidized GSH (i.e., GSSG) and thiobarbituric acid reactive species (TBARS, 

26 expressed in terms of MDA) measured in plasma and RBC lysate (Mohammad et al., 

27 2008). In automotive painters in Brazil (mean [SE] blood Pb: 5.4 [0.4] μg/dL), 

28 glutathione-S-transferase, GPx, and SOD were positively correlated with blood Pb (r = 

29 0.34, 0.38, and 0.32, respectively; p < 0.05) (Conterato et al., In Press). Similar effects on 

30 indices of oxidative stress were observed in in vitro studies: increased MDA and 

31 decreased SOD and catalase in RBCs exposed to 2 μM Pb (Ciubar et al., 2007),

32 decreased glutathione reductase (GR) activity in human RBCs incubated with 5-18 μM 

33 Pb (Coban et al., 2007), and decreased GSH and increased GSSG and lipid peroxidation 
February 2012 5-412 Draft – Do Not Cite or Quote

1 in RBCs from healthy volunteers with no Pb exposure incubated with 0.4 μM Pb for 24-

2 120 hours (Quintanar-Escorza et al., 2010).

3 Evidence of lipid peroxidation was also observed in children moderately exposed to Pb; 

4 as with the occupational studies, their cross-sectional design limits their utility in 

5 assessing the magnitude, timing, frequency, and duration of Pb exposure necessary to 

6 elicit the observed effects. Ahamed et al. (2005) investigated the relationship between 

7 blood Pb levels and antioxidant enzyme levels and lipid peroxidation in children aged 4-

8 12 years in Lucknow, India. A total of 62 children, with a mean [SD] blood Pb level of 

9 7.47 [3.06] μg/dL, were included in the study. Children were separated into three groups 

10 based on their blood Pb levels. The means (SDs) were: 3.93 (0.61) μg/dL group I; 7.11 

11 (1.25) μg/dL group II; and 11.39 (1.39) μg/dL group III. Lipid peroxidation, measured as 

12 blood MDA, was statistically significantly greater in group III, compared to group II and 

13 I, whereas GSH was lower in group III relative to groups II and I. Catalase activity was 

14 the only measure of oxidative stress that was statistically significantly elevated in group 

15 II compared to group I. Additionally, blood Pb levels were found to be statistically 

16 significantly positively correlated with MDA and CAT and negatively correlated with 

17 GSH. Ahamed et al. (2006) additionally studied male urban adolescents in India. The 39 

18 adolescents were separated into groups according to their blood Pb level (group 1: 

19 <10 μg/dL [mean 7.4 μg/dL], group 2: >10 μg/dL [mean 13.27 μg/dL]). Although the 

20 groups were similar in age (mean [SD]: 16.59 [0.91] versus 16.76 [0.90] years, 

21 respectively), height, weight, and body mass index, group 2 had significantly higher 

22 levels of CAT and MDA compared to group 1. There were no significant differences in 

23 blood GSH levels. Examining all the study subjects together, investigators found a 

24 correlation between blood Pb level and blood MDA and RBC CAT levels, as well as an 

25 inverse relationship between ALAD activity and MDA and CAT levels. In a similar 

26 study, Ahamed et al. (2008) examined oxidative stress in Indian children (aged 3-12 

27 years) with neurological disorders. There was a significantly higher mean blood Pb level 

28 in the study population compared to the control healthy population (18.60 versus 

29 10.37 μg/dL). In addition, the following indicators of oxidative stress were observed in 

30 the study population: increased blood MDA, RBC SOD, and CAT levels and decreased 

31 blood GSH levels. GPx levels were similar between the two groups. Typical indicators of 

32 Pb exposure (active/nonactive ALAD ratio) were found to be correlated with lipid 

33 peroxidation and oxidative stress. Children aged 3–6 years old living near a steel refinery 

34 in China with blood Pb levels ≥ 10 μg/dL also had a significant increase in plasma MDA 

35 compared to the children with blood Pb levels < 10 μg/dL. However, levels of RBC 

36 SOD, GSH, and GPx were not different from those in controls (Jin et al., 2006).

37 Administration of Pb (25 mg/kg) to rats once a week for 4 weeks, which was related to a 

38 blood Pb level of about 6.5 μg/dL, caused a significant increase in RBC MDA levels (Lee 
February 2012 5-413 Draft – Do Not Cite or Quote

1 et al., 2005). Other indications of Pb-induced oxidative stress included significant 

2 increases in RBC SOD and CAT levels accompanied by significant decreases in GSH 

3 and GPx. Exposure of rats to 750 mg/kg Pb-acetate in drinking water for 11 weeks 

4 resulted in decreased concentrations of plasma vitamin C, vitamin E, nonprotein thiol, 

5 and RBC-reduced glutathione, with simultaneous increased activity of SOD and GPx

6 (Kharoubi et al., 2008b). CAT activity was also slightly elevated in Pb-exposed rats, but 

7 the increase failed to reach statistical significance. Exposure of male rats to 0.5% Pb 

8 nitrate in drinking water (blood Pb not reported) for three weeks decreased GSH levels 

9 compared to that in controls (mean [SE]: 1.91 [0.02] versus 2.44 [0.09] mg/mL, 

10 respectively) (Gautam and Flora, 2010). SOD activity was significantly decreased in rats 

11 injected with 15 mg/kg Pb i.p. for seven days, but not rats treated with 5 mg/kg Pb

12 (Berrahal et al., 2007). GPx activity and MDA concentrations were slightly elevated in 

13 the exposed groups, but differences with the control group failed to reach statistical 

14 significance. 

5.7.6.2 Antioxidant Defense

15 In addition to the studies listed above that examine lipid peroxidation and oxidative 

16 stress, there have been studies that indicate that the use of antioxidants and free radical 

17 reactions is protective against Pb-induced RBC oxidative stress. Rats treated with 

18 500 ppm Pb-acetate in drinking water for 15 or 30 days had an increase in free RBC 

19 protoporphyrin and TBARS that was related to length of exposure and blood Pb

20 (Rendón-Ramirez et al., 2007). Vitamin E administration after exposure to Pb 

21 significantly reduced the TBARS levels and increased ALAD activity, compared to 

22 exposure to Pb alone. Co-exposure to vitamin E and Pb simultaneously and exposure to 

23 vitamin E before Pb exposure also prevented Pb-induced oxidative stress. In vitro studies 

24 by Casado et al. (2007), found that Pb-induced hemolysis and RBC membrane damage 

25 was mediated via oxidative stress. The in vitro studies demonstrated a concentration- and 

26 time-dependent formation in lipid peroxide that was inhibited with a number of 

27 antioxidants, including desferrioxamine (iron chelator), trolox (chain breaking 

antioxidant), and mannitol and Na formate (• 28 OH scavengers). Results suggested the role 

29 of singlet oxygen in Pb-mediated membrane damage and hemolysis of exposed RBCs. In 

30 rats exposed to 2,000 ppm Pb in drinking water for 5 weeks, MDA levels were 

31 significantly increased, whereas vitamin E concentrations were significantly decreased

32 (Caylak et al., 2008). In the case of MDA, co-exposure to Pb and a number of sulfur33 containing antioxidants (e.g., L-methionine, N-acetylcysteine, and L-homocysteine) 

34 reduced concentrations to a level not statistically significantly different from that in 
February 2012 5-414 Draft – Do Not Cite or Quote

1 controls, but statistically smaller than concentrations observed with Pb alone. Exposure to 

2 L-methionine and N-acetylcysteine also reduced Pb-induced depletion of vitamin E. 

5.7.7 Summary and Causal Determination

3 There is consistent toxicological and epidemiologic evidence indicating that exposure to 

4 Pb affects hematological endpoints, including decreased RBC survival and function, 

5 altered heme synthesis, and increased RBC oxidative stress. Pb has been shown to 

6 preferentially partition into RBCs following exposure, with RBC concentrations 

7 approximately 100-fold greater than those observed in the plasma (Jin et al., 2008; 

8 Timchalk et al., 2006).

9 Elaborating on the body of evidence presented in the 2006 Pb AQCD, recent 

10 epidemiologic and toxicological studies continue to demonstrate that Pb exposure is 

11 associated with decreased RBC survival and function, with the largest body of evidence 

12 consisting of populations of adults and children in which Pb is associated with effects on 

13 several inter-connected and related hematological parameters such as Hb, PCV, MCV, 

14 MCH, and MCHC. For adult populations, the largest body of evidence consists of 

15 occupationally-exposed workers in which measures of RBC survival are altered when 

16 compared with unexposed control populations (Cabaravdic et al., 2010; Ukaejiofo et al., 

17 2009; Khan et al., 2008; Patil et al., 2006a; Patil et al., 2006b; Karita et al., 2005; 

18 Conterato et al., In Press). Although the mean blood Pb level in most occupationally 

19 exposed populations was in excess of 20 μg/dL, decreases in Hb and PCV were observed 

20 in adults with blood Pb levels of 7 μg/dL (compared to controls with blood Pb levels of 

21 3 μg/dL), and significant correlations were observed between RBC distribution width and 

22 MCHC and blood Pb levels in adult populations with mean blood Pb levels of 5.4 μg/dL. 

23 Only one non-occupational study was found investigating the association of Pb with 

24 hematological parameters; in pregnant women, blood Pb levels were found to be 

25 negatively correlated with Hb concentrations. Studies in children measuring concurrent 

26 blood Pb levels are generally in agreement with those investigating occupationally27 exposed adults regarding effects on hematological parameters (i.e., Hb, MCV, (Queirolo 

28 et al., 2010; Shah et al., 2010; Ahamed et al., 2007; Huo et al., 2007; Olivero-Verbel et 

29 al., 2007; Riddell et al., 2007; Turgut et al., 2007; Ahamed et al., 2006; Rondo et al., 

30 2006). Any differences in the effects on specific hematological parameters observed 

31 between study findings for adults and those for children may be due to the comparatively 

32 shorter duration and lower magnitude of Pb exposure experienced by children compared 

33 to adults, although there is uncertainty regarding the timing and duration of exposure 

needed to induce effects in adults. In addition, Pb was shown to reduce Ca2+- and Ca2+ 34 -

Mg2+-ATPase activity in RBC membranes, which leads to an increase in RBC [Ca2+]i 35 , 
February 2012 5-415 Draft – Do Not Cite or Quote

1 increased membrane fragility, and abnormal morphological changes in studies of 

2 occupationally exposed adults (Quintanar-Escorza et al., 2007) and in in vitro studies

3 (Quintanar-Escorza et al., 2010; Ciubar et al., 2007). Heul et al. (2008) observed a 

reduction of RBC Ca2+-Mg2+ 4 -ATPase activity in children in association with a concurrent 

5 group mean cord blood Pb level of 3.54 μg/dL. Toxicological studies have also observed 

6 decreases in Hct and Hb and increases in hemolysis and reticulocyte density in rats and 

7 mice with blood Pb levels as low as 6.6-7.1 μg/dL (Sharma et al., 2010b; Baranowska8 Bosiacka et al., 2009; Simsek et al., 2009; Lee et al., 2005). Pb exposure has also been 

9 observed to increase PS expression on RBC membranes, leading to cell shrinkage, 

10 erythropoiesis, and destruction of the RBCs by macrophages (Jang et al., 2011; Ciubar et 

11 al., 2007; Shin et al., 2007). Suggestive evidence of disrupted hematopoiesis evidenced 

12 by decreased serum erythropoietin was observed in occupationally-exposed adults with a 

13 mean blood Pb level of 6.4 μg/dL (Sakata et al., 2007); toxicological studies in rats also 

14 indicate that Pb is cytotoxic to RBC progenitor cells (Alghazal et al., 2008b; Celik et al., 

15 2005). Taken together, these studies provide consistent evidence that exposure to Pb 

16 affects RBC function and survival, and leads to the reduction of RBCs in circulation. 

17 Although this decrease in RBCs may be explained by both decreased cell survival and/or 

18 disruption of hematopoiesis, the observation of increased reticulocytes seems to represent 

19 compensation for decreased RBC survival due to Pb exposure. 

20 Confirming effects observed in the 2006 Pb AQCD, a large body of evidence consisting 

21 of cross-sectional epidemiologic studies measuring concurrent blood Pb in adults and 

22 children have found that decreases in RBC ALAD levels and activity are strongly 

23 associated with higher blood Pb levels (Wang et al., 2010f; Mohammad et al., 2008; 

24 Ahamed et al., 2007; Quintanar-Escorza et al., 2007; Ahamed et al., 2006; Patil et al., 

25 2006a; Patil et al., 2006b; Ademuyiwa et al., 2005b; Ahamed et al., 2005; Conterato et 

26 al., In Press). Although the body of evidence is smaller than for humans, decreases in 

27 blood ALAD activity were also seen in rats with increased blood Pb levels, compared to 

28 controls (Lee et al., 2005). In addition to ALAD, recent studies have shown that Pb 

29 exposure inhibits the activity of ferrochelatase, leading to increased RBC ZPP in children 

30 and occupationally-exposed adults (Counter et al., 2009, 2008; Mohammad et al., 2008; 

31 Counter et al., 2007; Patil et al., 2006b; Ademuyiwa et al., 2005b) and animals (Rendón32 Ramirez et al., 2007). Pb has also been shown to inhibit the in vitro activities of other 

33 enzymes in RBCs, including those involved in nucleotide scavenging, energy 

34 metabolism, and acid-base homeostasis (Baranowska-Bosiacka et al., 2009; Ekinci et al., 

35 2007).

36 Lastly, Pb exposure induces lipid peroxidation and oxidative stress in RBCs. 

37 Epidemiologic studies have observed increases in MDA in occupationally-exposed adult 

38 populations (Ergurhan-Ilhan et al., 2008; Khan et al., 2008; Mohammad et al., 2008; 
February 2012 5-416 Draft – Do Not Cite or Quote

1 Quintanar-Escorza et al., 2007; Patil et al., 2006a; Patil et al., 2006b). Other changes in 

2 oxidative stress parameters observed included lowered activities of SOD, GR, and CAT, 

3 and increased CRP. Altered indices of RBC oxidative stress were also seen in adolescents 

4 and children in association with blood Pb levels (Ahamed et al., 2008; Ahamed et al., 

5 2006; Jin et al., 2006). In vitro and vivo studies have also demonstrated that prior, 

6 concurrent, or subsequent treatment with various antioxidants has been shown to 

7 ameliorate at least partially Pb-induced oxidative stress in RBCs (Caylak et al., 2008; 

8 Casadoa et al., 2007; Rendón-Ramirez et al., 2007).

9 Similar to the epidemiologic and toxicological studies that demonstrate an association 

10 between Pb exposure and hematological effects in humans and laboratory animals, the 

11 ecological literature has consistently reported on hematological responses in aquatic and 

12 terrestrial invertebrates and vertebrates (Sections 7.4.1.2 and 7.4.2.2). The most 

13 consistently observed effect in metal impacted environments is decreased RBC ALAD 

14 activity. This effect has been observed across a wide range of taxa, including bivalves, 

15 fish, amphibians, birds, and mammals. More limited evidence exists regarding deleterious 

16 effects of Pb on serum enzyme levels and white blood cell counts in birds and mammals.

17 In summary, new epidemiologic and toxicological studies included in the current review 

18 provide strong evidence that exposure to Pb is associated with numerous deleterious 

19 effects on the hematological system, including effects on RBC survival and function, 

20 altered heme synthesis, and increased oxidative stress, and continue to confirm previous 

21 conclusions from the 2006 Pb AQCD. The principal finding regarding RBC survival and 

22 function are consistent Pb-induced alterations in several inter-connected and related 

23 hematological parameters such as Hb, Hct, and MCV across multiple studies, with the 

24 weight of evidence provided by epidemiological studies in occupationally-exposed adult 

25 populations and children. In occupationally-exposed adults, these findings are most 

26 substantiated in populations with current blood Pb levels > 20 μg/dL, although effects on 

27 hematological parameters were observed in some occupationally-exposed populations at 

28 concurrent blood Pb levels in the range of 5-7 μg/dL. In Pb-exposed children, effects on 

29 hematological parameters were most substantiated in populations with blood Pb levels 

30 less than 15 μg/dL. The weight of evidence in adult rodents exposed long-term to Pb (i.e, 

31 ≥ 4 weeks), although less than the weight of evidence in humans, is coherent with 

32 epidemiologic studies regarding decrements in hematological parameters at blood Pb 

33 levels as low as 6.6-7.1 μg/dL. Regarding alterations in heme synthesis, the largest body 

34 of evidence again is provided by decreased ALAD activity observed in epidemiologic 

35 studies in occupationally-exposed adult populations and children. In the occupationally36 exposed adult populations, the observation of decreased ALAD activity was most often 

37 observed in populations with concurrent blood Pb levels >15 μg/dL. In children, 

38 decreases in ALAD activity were observed in populations with concurrent blood Pb 
February 2012 5-417 Draft – Do Not Cite or Quote

1 levels in the range of 7-22 μg/dL. Animal toxicological studies also provide to the weight 

2 of evidence regarding altered ALAD activity, with effects seen in adult animals exposed 

3 for 3-4 weeks with blood Pb levels as low as 6.5 μg/dL. As for the effects listed above, 

4 the weight of evidence for oxidative stress (i.e., increased lipid peroxidation or alterations 

5 in antioxidant enzyme levels) primarily comes from epidemiological studies in 

6 occupationally-exposed adults and children. The majority of evidence for increased 

7 oxidative stress in Pb-exposed adults comes from occupational cohorts with concurrent 

8 blood Pb levels >15 μg/dL. In children, concurrent blood Pb levels of 7-22 μg/dL were 

9 associated with measures of oxidative stress. Due to the cross-sectional nature of the 

10 above epidemiologic studies in adults and children, and the measurement of concurrent 

11 blood Pb, the timing and duration of exposure necessary to alter RBC survival and 

12 function, heme synthesis, or the state of oxidative stress in RBCs is unclear. This 

13 uncertainty is greatest in adults as concurrent blood Pb levels reflecting recent exposures 

14 is likely to be less than blood Pb levels resulting from past exposures. The consistency of 

15 findings in epidemiologic studies investigating effects in adults and children, and the 

16 coherence of findings in the toxicological literature and coherence across the disciplines 

17 is sufficient to conclude that a causal relationship exists between Pb exposures and 

18 effects on heme synthesis and red blood cell function. 

5.8 Reproductive and Developmental Effects 

19 The effect of Pb on reproductive outcomes has been of interest for years, starting in 

20 cohorts of occupationally-exposed individuals. More recently, researchers have begun to 

21 focus on reproductive effects in populations without occupational exposures, with 

22 environmentally-relevant levels of Pb exposure. In the toxicological and epidemiologic 

23 literature, research on reproductive effects of Pb include female and male reproductive 

24 function (hormone levels, fertility, puberty, and effects on reproductive organs and 

25 estrus), birth defects, spontaneous abortions, infant mortality, preterm birth, low birth 

26 weight/fetal growth, and other developmental effects. In epidemiologic studies, various 

27 biological measures of Pb are used including Pb measured in blood and bone; 

28 toxicological studies only report exposure using blood Pb. Bone Pb is indicative of 

29 cumulative Pb exposure. Blood Pb can represent more recent exposure, although it can 

30 also represent remobilized Pb occurring during times of bone remodeling and pregnancy 

31 or lactation. More detailed discussion of these measures and Pb transfer via umbilical 

32 cord blood Pb across the placenta, and via lactation is given in Section 4.3.5.2 on Pb 

33 Toxicokinetics. A few studies of pregnancy-induced hypertension and eclampsia have 

34 been conducted and are reported on in the section on hypertension (Section 5.4.2.1). 
February 2012 5-418 Draft – Do Not Cite or Quote

1 Briefly, the relatively small number of studies found consistently positive associations 

2 between blood Pb levels and pregnancy-induced hypertension. 

3 Overall, the recent reproductive literature continues to support associations reported in 

4 earlier Pb AQCDs between Pb exposure and effects on various parameters of sperm 

5 (function, motility, count, integrity, histology). The toxicological and epidemiologic 

6 literature also indicates that Pb exposure is associated with delayed onset of puberty in 

7 both males and females. The new information from epidemiologic and toxicological 

8 studies is integrated with conclusions from previous Pb AQCDs below. 

5.8.1 Effects on Female Reproductive Function

9 The epidemiologic studies on Pb and female reproductive function presented in the 2006 

10 Pb AQCD (U.S. EPA, 2006b) provided little evidence for an association between Pb 

11 biomarkers and effects on female reproduction and fertility. However, the 1986 and 2006 

12 Pb AQCDs (U.S. EPA, 2006b, 1986a) reported toxicological findings that Pb exposure 

13 was associated with effects on female reproductive function that can be classified as 

14 alterations in female sexual maturation, effects on fertility and menstrual cycle, endocrine 

15 disruption, and changes in morphology or histology of female reproductive organs 

16 including the placenta. Since the 2006 Pb AQCD, many epidemiologic studies have been 

17 published regarding Pb biomarker levels in women and reproductive effects. For some 

18 effects, there are inconsistent findings, but for others, such as delayed puberty, there are 

19 clear associations with blood Pb levels. In addition, recent toxicological studies add 

20 further knowledge of Pb-related effects on the female reproductive system. 

5.8.1.1 Effects on Female Sex Endocrine System and Estrus 

Cycle

21 Multiple epidemiologic studies have examined the association between blood Pb levels 

22 and hormone levels and the estrus cycle. Epidemiologic studies (characterized in Table 

23 5-30; all studies included in the table used measures of Pb and hormones that were either 

24 concurrent or close in time) support the toxicological findings, which are the major body 

25 of evidence on endocrine effects of Pb.
February 2012 5-419 Draft – Do Not Cite or Quote

Table 5-30 Summary of recent epidemiologic studies of associations between 

Pb levels and hormones for females

Referencea

Study, 

Location, 

and Years Outcome Study population

Pb 

Biomarker

Mean Pb (SD) 

in μg/dL Adjusted Effect Estimates

Krieg et al. (2007) U.S.

1988-1994

FSH, LH Women aged 35-60 

from the NHANES III 

study

Blood Pb 2.8 Linear regression slope (95% CI) for log- transformed Pb

FSH:

Post-menopausal 22.2 (13.5, 30.8)

Pregnant 0.1 (-0.1, 0.3)

Menstruating at time of exam 2.1 (-2.1, 

6.3)

Both ovaries removed 32.6 (10.1, 55.1)

Birth control pills being used -6.3 (-10.0, -

2.5)

Pre-menopausal 8.3 (3.8, 12.7)

LH:

Post-menopausal 6.2 (3.0, 9.5)

Pregnant -0.8 (-1.9, 0.4)

Menstruating at time of exam -0.3 (-1.8, 

1.3)

Both ovaries removed 10.0 (1.1, 18.9)

Birth control pills being used: -0.6 (-2.9, 

1.6)

Pre-menopausal 1.7 (-0.6, 4.1)

Chang et al. (2006) Kaohsiung City, 

Taiwan

1999, 

2000-2001

Estradiol Women receiving care 

at a infertility clinic in 

2000-2001 or delivering 

a normal infant at a 

nearby medical center in 

1999

Blood Pb 3.12 (0.19) Linear regression β(SE) for Pb

1.18 (0.60)

p-value: 0.049

Pollack et al. (2011) Buffalo, NY

2005-2007

FSH, estradiol, 

LH, 

progesterone, 

and cycle 

length

Healthy, premenopausal 

women aged 18-44 with 

menstrual cycle length 

of 21-35 days, BMI of 

18-35 kg/m2

, not 

recently using birth 

control, not planning to 

become pregnant, and 

not breast feeding

Blood Pb 0.93

IQR: 0.68, 1.20

Mean % Estradiol

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: 8.2 (-1.2, 18.6) 

1.11-6.20 μg/dL: 4.7 (-4.7, 15.2) 

Amplitude Estradiol

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: -0.01 (-0.06, 0.04)

1.11-6.20 μg/dL: -0.02 (-0.7, 0.03)

Phase Shift Estradiol

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: -0.09 (-0.24, 0.05) 

1.11-6.20 μg/dL: 0.14 (-0.01, 0.29)

Mean % FSH

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: 8.0 (-0.9, 17.7) 

1.11-6.20 μg/dL: 3.6 (-5.3, 13.3) 

Amplitude FSH

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: -0.01 (-0.03, 0.02)

1.11-6.20 μg/dL: -0.02 (-0.04, 0.01)

Phase Shift FSH
February 2012 5-420 Draft – Do Not Cite or Quote

Referencea

Study, 

Location, 

and Years Outcome Study population

Pb 

Biomarker

Mean Pb (SD) 

in μg/dL Adjusted Effect Estimates

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: -0.06 (-0.25, 0.12) 

1.11-6.20 μg/dL: -0.02 (-0.21, 0.18)

Mean % LH

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: 5.1 (-5.1, 16.4) 

1.11-6.20 μg/dL: -0.5 (-10.5, 10.7) 

Amplitude LH

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: -0.01 (-0.03, 0.02)

1.11-6.20 μg/dL: -0.02 (-0.04, 0.01)

Phase Shift LH

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: -0.16 (-0.36, 0.03) 

1.11-6.20 μg/dL: -0.11 (-0.32, 0.10)

Mean % Progesterone

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: 7.5 (0.1, 15.4) 

1.11-6.20 μg/dL: 6.8 (-0.8, 14.9) 

Amplitude Progesterone

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: 0.07 (0.01, 0.15)

1.11-6.20 μg/dL: -0.06 (-0.13, 0.01)

Phase Shift Progesterone

0.30-0.72 μg/dL: Ref 

0.73-1.10 μg/dL: 0.04 (-0.06, 0.15) 

1.11-6.20 μg/dL: 0.15 (0.05, 0.26)

Linear models β (95% CI)

Estradiol

0.03 (-0.05, 0.11)

FSH

-0.01 (-0.07, 0.06)

LH

0.02 (-0.06, 0.10)

Progesterone

0.06 (-0.04, 0.17)

OR (95% CI) for anovulation per I μg/dL

1.20 (0.62, 2.34)
February 2012 5-421 Draft – Do Not Cite or Quote

Referencea

Study, 

Location, 

and Years Outcome Study population

Pb 

Biomarker

Mean Pb (SD) 

in μg/dL Adjusted Effect Estimates

Jackson et al. (2011) Buffalo, NY

2005-2007

FSH, estradiol, 

LH, 

progesterone, 

and cycle 

length

Healthy, premenopausal women 

aged 18-44 with 

menstrual cycle length 

of 21-35 days, BMI of 

18-35 kg/m2

, not 

recently using birth 

control, not planning to 

become pregnant, and 

not breast feeding

Blood Pb Median: 0.87

IQR: 0.68, 1.20

Adjusted percent change (95% CI) in 

serum hormone level for change in blood 

Pb

FSH: -2.5 (-11.2, 7.0)

Estradiol: 4.9 (-5.0, 15.9)

LH: 2.5 (-12.3, 19.9)

Progesterone: 4.6 (-12.2, 24.6)

Cycle length: 0.2 (-2.8, 3.3)

OR (95% CI) per unit Pb

<25 d vs. 25-35 d cycle length:

0.9 (0.4, 2.3)

>35 d vs. 25-35 d cycle length:

0.5 (0.1, 1.9)

a

Studies are presented in order of first appearance in the text of this section.

1 An epidemiologic study using the NHANES III data and including women aged 

2 35-60 years old examined the relationship between blood Pb levels (mean 2.8 μg/dL) and 

3 serum follicle stimulating hormone (FSH) and luteinizing hormone (LH) (Krieg, 2007).

4 Deviation from normal FSH and LH levels may indicate endocrine disruption related to 

5 ovary functioning. Researchers found that higher blood Pb levels were associated with 

6 higher levels of serum FSH and LH among both postmenopausal women and women 

7 with both ovaries removed. There was also a trend of increasing serum FSH with blood 

8 Pb levels for pre-menopausal women who were not menstruating at the time of the exam 

9 or pregnant, although the association was not statistically significant for LH. A limitation 

10 of this portion of the study is that FSH and LH were measured without attention to day of 

11 a woman’s menstrual cycle and LH and FSH are known to vary throughout the cycle of 

12 non-menopausal, cycling women who are not taking birth control pills. Higher blood Pb 

13 levels were associated with lower levels of serum FSH among women taking birth 

14 control pills. The inverse association was also present for LH, but it was not statistically 

15 significant. No associations between blood Pb and FSH or LH were apparent for women 

16 who were menstruating at the time of the exam or were pregnant. Further analysis 

17 indicated that the lowest level of blood Pb for which a statistically significant association 

18 between blood Pb and FSH could be observed was 1.7 μg/dL among women with their 

19 ovaries removed. For LH, the lowest level of blood Pb for which a statistically significant 

20 association between blood Pb and LH could be observed was 2.8 μg/dL among 

21 postmenopausal women. Another epidemiologic study was performed in Kaohsiung City, 

22 Taiwan among two groups of women aged 23-44 years: those who were seeking help at a 

23 fertility clinic after one year of trying to conceive, and those who had previously 

24 delivered an infant and were identified from medical records of a postpartum care unit

25 (Chang et al., 2006). The mean (SD) blood Pb in this study was 3.12 (0.19) μg/dL. The 

26 study reported a positive association between blood Pb levels and serum estradiol 
February 2012 5-422 Draft – Do Not Cite or Quote

1 concentrations during the early follicular phase, which reflects ovary activity. 

2 Associations between hormones and blood Pb level were also investigated using the 

3 BioCycle study cohort (Jackson et al., 2011; Pollack et al., 2011). These women were 

4 premenopausal with normal cycles and not on birth control. Neither study detected an 

5 association between unit change in blood Pb and hormone levels. However, when 

6 examining tertiles of Pb, women in the highest tertile blood Pb (1.11-6.20 μg/dL) had 

7 higher mean progesterone and longer length of a phase shift compared to women in the 

8 lowest tertile (0.30-0.72 μg/dL) (Pollack et al., 2011). Other associations were observed 

9 but were not statistically significant (Pollack et al., 2011). No associations were detected 

10 for anovulation (Pollack et al., 2011) or for cycle length (Jackson et al., 2011).

11 The effect of Pb exposure on the female endocrine system was demonstrated in 

12 toxicological studies reviewed in the 1986 and 2006 Pb AQCD (U.S. EPA, 2006b, 

13 1986a). However, the mechanism by which Pb affects the endocrine system has not been 

14 fully elucidated. Several recent articles continue to demonstrate that Pb alters the 

15 concentration of circulating hormones in female experimental animals. As mentioned in 

16 the previous AQCD, Pine et al. (2006) observed that maternal Pb exposure (during 

17 gestation and lactation) caused a decrease in basal LH levels in pre-pubertal female 

18 Fisher 344 rat pups as compared to control, non-Pb exposed pups. Dumitrescu et al.

19 (2008a) observed alteration of hormone levels in female Wistar rats after ingesting 

20 Pb-acetate (50, 100, 150 ppb) in drinking water for 6 months; measurements were made 

21 during the pro-estrous stage of the estrous cycle to allow for consistent timing for 

22 comparison of cyclic hormonal variation. The authors reported decreases in FSH, 

23 estradiol, and progesterone levels with increases in LH and testosterone levels. 

24 Nampoothiri and Gupta (2008) administered Pb-acetate at a concentration that did not 

25 affect reproductive performance, implantation or pregnancy outcome (0.05 mg/kg body 

26 weight) to Charles Foster female rats 5 days before mating and during the gestational 

27 period. They observed a decrease in steroidogenic enzymes, 3β- hydroxysteroid 

28 dehydrogenase (HSD) and 17β-HSD, activity in reproductive organs, as well as a 

29 decrease in steroid hormones (progesterone and estradiol), suggesting that chronic 

30 exposure to low levels of Pb may affect reproductive function of mothers and their 

31 offspring. Similarly, Pillai et al. (2010) reported impaired ovarian steroidogenesis in 

32 Charles Foster adult female rats (PND56) from dams exposed gestationally and 

33 lactationally to Pb-acetate (subcutaneous daily injections of 0.05 μg/kg BW). Pillai 

34 observed a decrease in steroidogenic enzymes, 3β-HSD and 17β-HSD, but saw no 

35 changes in ovarian steroidogenic acute regulatory protein (StAR) or CYP11 mRNA 

36 levels indicating Pb-induced inhibition of ovarian steroidogenesis.

37 Kolesarova et al. (2010) conducted an in vitro study to examine the secretory activity of 

38 porcine ovarian granulose cells after Pb administration. The results of the study showed 
February 2012 5-423 Draft – Do Not Cite or Quote

1 that Pb-acetate concentrations of 0.046 mg/mL and 0.063 mg/mL statistically 

2 significantly inhibited insulin-like growth factor-1 (IGF-1) release, but concentrations of 

3 0.25 mg/mL and 0.5 mg/mL did not influence IGF-1 release. Progesterone release was 

4 not affected by Pb treatment; however, Pb caused a reduction in LH and FSH binding in 

5 granulose cells and increased apoptosis as evidenced by increased expression of 

6 caspase-3 and cyclin B1, suggesting a Pb-induced alteration in the pathways of 

7 proliferation and apoptosis of porcine ovarian granulose cells. Decreased gonadotropin 

8 binding was also observed in rats after Pb exposure (Nampoothiri and Gupta, 2006).

9 No recent toxicological studies were found that examined Pb-induced effects on the 

10 estrus cycle.

11 Overall, toxicological studies report alterations in hormone levels related to blood Pb 

12 concentration. Similarly, epidemiologic studies reported associations between blood Pb 

13 levels and hormone levels in female adults. Although Pb-associated changes in hormone 

14 levels are observed, there are discrepancies about the direction of the hormone changes 

15 related to Pb. One explanation is that the direction of change could vary based on current 

16 hormonal and reproductive status.

5.8.1.2 Effects on Fertility

17 Previous studies indicated that Pb exposure does not produce total sterility, but it can 

18 disrupt female fertility (U.S. EPA, 2006b). Recent epidemiologic studies and studies in 

19 experimental animals support this finding. The epidemiologic studies are summarized in 

20 Table 5-31. Most of these studies examined biological measures of Pb collected at or 

21 during the period of possible fertilization, although Bloom et al. (2011a) measured blood 

22 Pb at baseline and followed women for at least 12 menstrual cycles (or until pregnancy). 
February 2012 5-424 Draft – Do Not Cite or Quote

Table 5-31 Summary of recent epidemiologic studies of associations between 

Pb levels and fertility for females

Referencea

Study, 

Location, 

and Years Outcome Study population Pb Biomarker

Mean Pb (SD) 

in μg/dL Adjusted Effect Estimates

Chang et al. (2006) Kaohsiung City, 

Taiwan

1999, 

2000-2001

Infertility Women receiving care 

at a infertility clinic in 

2000-2001 or delivering 

a normal infant at a 

nearby medical center in 

1999

Blood Pb 3.12 (0.19) OR (95% CI)

Infertility

≤ 2.5 μg/dL: 1.00 (Ref)

>2.5 μg/dL: 2.94 (1.18, 7.34)

Al-Saleh et al. (2008a) Riyadh, Saudi 

Arabia

2002-2003

Achieving 

pregnancy 

and/or 

fertilization

Women aged 19-50 

undergoing IVF

Blood Pb 

Follicular fluid Pb

Blood Pb 

3.34 (2.24)

Blood Pb levels >10 

μg/dL: 1.7%

Follicular fluid

0.68 (1.82)

OR (95% CI) (unit not given, 

assume results are per 1 

μg/dL)

Pregnancy 

Blood Pb 

0.55 (0.23, 1.31)

Follicular fluid Pb

1.36 (0.91, 2.02)

Fertilization

Blood Pb 

0.30 (0.08, 1.03)

Follicular fluid Pb

1.45 (0.69, 3.02)

Note: In a reduced adjusted 

model for fertilization, the OR 

for blood Pb was 0.38 (0.14, 

0.99)

Silberstein et al.

(2006)

Providence, RI

NS

Achieving 

pregnancy 

Women undergoing IVF 

at the study hospital

Follicular fluid Pb Not given 

quantitatively 

From a figure in the 

paper:

Median Pb in follicular 

fluid of pregnant 

women: ~1.3

Median Pb in follicular 

fluid of non-pregnant 

women: ~2.2

P-value for difference in 

medians by Mann-Whitney U 

test: 0.0059

*note, study only included 9 

women

Bloom et al. (2010) California

2007-2008

Oocyte 

maturity, oocyte 

fertilization

Women who were part 

of the Study of Metals 

and Assisted 

Reproductive 

Technologies (SMART): 

women referred to the 

Center for Reproductive 

Health of UCSF for 

infertility treatment and 

their first IVF procedure

Blood Pb 0.82 (0.32) RR per 1 μg/dL

Oocyte maturity (determined 

by Metaphase II arrest):

0.54 (0.31, 0.93)

0.25 (0.03, 2.50)*

Oocyte fertilization:

0.97 (0.66, 1.43)

1.09 (0.72, 1.65)*

*Controlling for Cd
February 2012 5-425 Draft – Do Not Cite or Quote

Referencea

Study, 

Location, 

and Years Outcome Study population Pb Biomarker

Mean Pb (SD) 

in μg/dL Adjusted Effect Estimates

Bloom et al. (2011b) California

2007-2008

Embryo cell 

number, 

embryo 

fragmentation 

score

Women who were part 

of the Study of Metals 

and Assisted 

Reproductive 

Technologies (SMART) 

(women referred to the 

Center for Reproductive 

Health of UCSF for 

infertility treatment and 

their first IVF procedure) 

and who generated 

embryos

Blood Pb 0.83 (0.30) OR per 1 μg/dL*

Embryo cell number:

0.25 (0.07, 0.86)

Embryo fragmentation score:

1.71 (0.45, 6.56)

*Adjusted for Hg and Cd

Bloom et al. (2011a) New York

1996-1997

Achieving 

pregnancy

Women who were aged 

18-34 years, were 

previously part of a 

study five years prior 

about fish consumption, 

and were not currently 

pregnant

Blood Pb No positive pregnancy 

test: 1.55 (0.16)

Positive pregnancy 

test: 1.54 (0.12)

β (95% CI)

-0.031 (-1.066, 1.004) per 0.6 

μg/dL 

a

Studies are presented in order of first appearance in the text of this section.

1 Epidemiologic studies examined women having difficulty conceiving by performing 

2 studies among patients of fertility clinics or undergoing in vitro fertilization (IVF). 

3 Among women aged 23-44 years, a difference in blood Pb was reported between women 

4 who were seeking help at a fertility clinic after one year of trying to conceive and women 

5 who had previously delivered an infant and were identified from medical records of a 

6 postpartum care unit at a medical center (Chang et al., 2006). Higher odds of infertility 

7 were observed when comparing women with blood Pb levels >2.5 μg/dL to those with 

8 blood Pb levels ≤ 2.5 μg/dL. Another study examining fertility reported on women in 

9 Saudi Arabia aged 19-50 years who were undergoing IVF treatment (Al-Saleh et al., 

10 2008a). Women were categorized as having achieved a pregnancy versus not having 

11 achieved a pregnancy and achieved fertilization versus not achieving fertilization. The 

12 majority of women had follicular Pb levels that were below the limit of detection, 

13 whereas less than 2% of women had blood Pb levels below the limit of detection. In 

14 addition, less than 2% of women had blood Pb levels that were above 10 μg/dL. 

15 Follicular Pb levels were not correlated with the blood Pb. No association was observed

16 between blood or follicular Pb and pregnancy outcomes in either crude or adjusted 

17 models. An association was not detected between follicular Pb and fertilization, but 

18 higher blood Pb was associated with lower rates of fertilization. Finally, a study that 

19 included nine women undergoing IVF treatment in Rhode Island (Silberstein et al., 2006)

20 found that median follicular Pb levels in women who achieved pregnancy were lower 

21 than the follicular Pb levels among nonpregnant women. One limitation present in these 

22 studies is that the participants, especially in the later two studies, are women who are 

23 seeking help for fertility problems. The participants are not samples of the general 

24 population and therefore cannot be generalized to all women of childbearing age. The 

25 Study of Metals and Assisted Reproductive Technologies (SMART) enrolled women 
February 2012 5-426 Draft – Do Not Cite or Quote

1 undergoing their first round of IVF and investigated multiple steps before pregnancy as 

2 the outcomes (Bloom et al., 2011b; Bloom et al., 2010). Higher blood Pb levels were 

3 associated with lower oocyte maturity although the lack of power made interpretation of 

4 models controlling for Cd difficult. No association was observed between blood Pb and 

5 oocyte fertilization (Bloom et al., 2010). When examining markers of IVF success, 

6 inconsistent results were observed. Embryo cell number was lower in association with 

7 higher blood Pb levels but no association was observed for embryo fragmentation score

8 (Bloom et al., 2011b). The study was limited by the small number of participants. 

9 Overall, these epidemiologic studies examine a variety of fertility-related endpoints and 

10 although some studies demonstrate an association between higher Pb levels and 

11 fertility/pregnancy, as a whole the results are inconsistent across studies. 

12 A prospective cohort study enrolled women who previously participated in a study of fish 

13 consumption for a length of <1 to >6 menstrual cycles and investigated the relationship 

14 between blood Pb levels and having a positive pregnancy test (Bloom et al., 2011a). No 

15 association was observed between blood Pb and achieving pregnancy. 

16 Animal toxicology studies following female fertility looked at various outcomes. Several 

17 studies observed a decrease in litter size when females were exposed to Pb before mating 

18 or during pregnancy (Dumitrescu et al., 2008b; Iavicoli et al., 2006b; Teijon et al., 2006).

19 Pups in a study by Teijon et al. (2006) receiving 400 ppm Pb-acetate in drinking water 

20 had blood Pb of 97 μg Pb/dL blood at 1 week post-weaning and 18.2 μg Pb/dL blood at 

21 2 week post-weaning. Dumitrescu et al. observed a modification in sex ratio of pups born 

22 to dams exposed to Pb before mating and during pregnancy. As the dose of Pb increased, 

23 the number of females per litter also increased (i.e., 1 male to 0.8 female in non-Pb 

24 exposed group; 1 male to 0.66 female in 50 ppb Pb-acetate group; 1 male to 2.25 females 

25 in 100 ppb group; and 1 male to 2.5 females in 150 ppb group). These results are not 

26 consistent with earlier results of Ronis et al. (1998b), who did not observe differences in 

27 sex ratio dams and offspring were exposed only during pregnancy. Thus, Pb exposure in 

28 animal studies during or before pregnancy have shown effects on litter size and mixed 

29 effects on sex ratio.

30 Nandi et al. (2010) demonstrated a concentration-dependent decline in viability rate, 

31 maturation, fertilization, and cleavage rates of buffalo oocytes cultured in medium 

32 containing 1-10 μg/mL Pb-acetate. Karaca and Şimşek (2007) observed an increase in the 

33 number of mast cells in ovary tissue after Pb exposure (2,000 μg/mL in drinking water) 

34 suggesting that Pb may stimulate an inflammatory response in the ovaries which may 

35 contribute to Pb-induced female infertility.

36 In contrast, Nampoothiri and Gupta (2008) did not observe any statistically significant 

37 change in fertility rate or litter size in female rats subcutaneously administered Pb 
February 2012 5-427 Draft – Do Not Cite or Quote

1 (0.05 mg/kg body weight daily before mating and during pregnancy) with a resulting 

2 blood Pb of 2.49 μg/mL. Although reproductive performance was not affected in this 

3 study, the authors did report an alteration in implantation enzymes. Cathepsin-D activity 

4 decreased and alkaline phosphatase activity increased after Pb exposure.

5 Recent epidemiologic and toxicological studies on the effect of Pb on fertility outcomes 

6 have generated inconsistent results. However, the bulk of the evidence including the 

7 current and historical Pb literature (U.S. EPA, 2006b) indicate that increased Pb exposure 

8 may decrease fertility.

5.8.1.3 Effects on Puberty

9 Recent toxicological studies of rodents have examined the effects of Pb on pubertal and 

10 reproductive organ development and on biomarkers of pubertal development. There have 

11 also been recent epidemiologic studies examining associations between blood Pb levels 

12 and onset of puberty, which are summarized in Table 5-32 and in the text below. All of 

13 the epidemiologic studies examined concurrently measured blood Pb and puberty and are 

14 reported below. Additionally, Naicker et al. (2010) followed girls to determine their age 

15 of menarche; however, blood Pb levels were measured once at 13 years of age.
February 2012 5-428 Draft – Do Not Cite or Quote

Table 5-32 Summary of recent epidemiologic studies of associations between 

Pb levels and puberty for females

Referencea Study Location and Years Outcome Study Population Pb Biomarker

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Tomoum et al.

(2010)

Cairo, Egypt

2007

Hormones and 

pubertal 

development

Healthy children aged 

10-13 yr; seeking 

treatment for minor 

health problems and

living in one of two 

designated areas (one 

with high-risk for Pb 

contamination and one 

with no Pb source)

Blood Pb NS for girls only 

(combined with 

boys in the study 

the mean was 9.46 

[3.08])

Breast Development 

<10 μg/dL: 

Stage 2: 36.4%

Stage 3: 63.6%

≥ 10 μg/dL:

Stage 2: 100%

Stage 3: 0%

Chi-square p-value<0.01

Pubic Hair Development

<10 μg/dL: 

Stage 2: 36.4%

Stage 3: 63.6%

≥ 10 μg/dL:

Stage 2: 77.8%

Stage 3: 22.2%

Chi-square p-value>0.05

*Quantitative results for 

hormones not provided

Denham et al.

(2005)

Akwesasne Mohawk 

Nation (boundaries of 

New York, Ontario, 

and Quebec

NS

Age at 

menarche

10- to 16.9-yr-old girls 

in the Akwesasne 

community 

Blood Pb 0.49 (0.905)

Median: 1.2

Coefficients for binary logistic 

regression predicting menarche 

with Pb centered at the mean:

log blood Pb -1.29 (p-value 0.01)

log blood Pb -squared: -1.01 (p- value 0.08)

Non-linear relationship observed 

and Pb below the mean did not 

appear to affect the odds of 

menarche. Increasing blood Pb 

from 0.49 to 0.98 μg/dL 

decreased the odds of menarche 

attainment by 72%

Naicker et al.

(2010)

Johannesburg/Soweto, 

South Africa

Born in 1990

Self-reported 

Tanner staging 

at age 13 and 

age at 

menarche 

Girls of black or mixed 

ancestry who were 

enrolled in the Birth to 

Twenty (Bt20) cohort 

(born in 1990) that lived 

in 

Johannesburg/Soweto 

for at least 6 mo after 

birth

Blood Pb at 13 yr 

of age

4.9 (1.9)

blood Pb levels ≥ 10 

μg/dL: 1% 

OR (95% CI)

Delay in breast development at 

age 13

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 2.34 (1.45, 3.79)

Delay in pubic hair development 

at age 13

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 1.81 (1.15, 2.84)

Delay in attainment of menarche 

at age 13

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 2.01 (1.38, 2.94)
February 2012 5-429 Draft – Do Not Cite or Quote

Referencea Study Location and Years Outcome Study Population Pb Biomarker

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Den Hond et al.

(2011)

Flanders

2003-2004

Tanner staging, 

age at 

menarche, 

regular menses

Girls ages 14 and 15, in 

their 3rd year of 

secondary education 

and living in the same 

study areas for at least 

5 years

Blood Pb Median: 1.81

10th percentile: 0.88

90th percentile: 3.81 

OR (95% CI) for pubic hair 

development with doubling of 

exposure

0.65 (0.45, 0.93)

*Association was no longer 

statistically significant when PCB 

marker included in the model

No association between Pb and 

breast development (results not 

given)

Wolff et al.

(2008); Wolf et al.

(2007)

New York City, NY

1996-1997

Pubertal stages 

defined using 

standard 

drawings

9-yr old girls from the 

study hospital and 

nearby pediatric offices

Blood Pb Median: 2.4 PR (95% CI) (unit not given, 

assume results are per 1 μg/dL)

Breast stage: 1.01 (0.79, 1.30)

Pubic hair stage: 1.25 (0.83, 

1.88)

Wu et al. (2003b) U.S.A.

1988-1994

Tanner staging 

and age at 

menarche

Girls ages 8-16 from 

the NHANES III study 

Blood Pb 2.5 (2.2)

Weighted proportion 

of the sample with 

blood Pb 5.0-21.7: 

5.9% 

OR (95% CI)

Breast development

0.7-2.0 μg/dL: 1.00 (Ref)

2.1-4.9 μg/dL: 1.51 (0.90, 2.53)

5.0-21.7 μg/dL: 1.20 (0.51, 2.85)

Pubic hair development

0.7-2.0 μg/dL: 1.00 (Ref)

2.1-4.9 μg/dL: 0.48 (0.25, 0.92)

5.0-21.7 μg/dL: 0.27 (0.08, 0.93)

Menarche

0.7-2.0 μg/dL: 1.00 (Ref)

2.1-4.9 μg/dL: 0.42 (0.18, 0.97)

5.0-21.7 μg/dL: 0.19 (0.08, 0.43)
February 2012 5-430 Draft – Do Not Cite or Quote

Referencea Study Location and Years Outcome Study Population Pb Biomarker

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Selevan et al.

(2003)

U.S.A.

1988-1994

Tanner staging 

and age at 

menarche

Girls ages 8-18 from 

the NHANES III study 

Blood Pb Geometric mean 

NHWhites: 1.4

NHBlacks: 2.1

Mexican-Americans: 

1.7

Blood Pb 

levels>5μg/dL:

NHWhites: 2.7%

NHBlacks: 11.6%

Mexican-Americans: 

12.8%

Blood Pb levels >10 

μg/dL:

NHWhites: 0.3%

NHBlacks: 1.6%

Mexican-Americans: 

2.3%

OR (95% CI)

Breast development

NH Whites:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.82 (0.47, 1.42)

NH Blacks:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.64 (0.42, 0.97)

Mexican Americans:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.76 (0.63, 0.91)

Pubic hair development

NH Whites:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.75 (0.37, 1.51)

NH Blacks:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.62 (0.41, 0.96)

Mexican Americans:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.70 (0.54, 0.91)

HR (95% CI) *included only girls 

8-16

Age at menarche

NH Whites:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.74 (0.55, 1.002)

NH Blacks:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.78 (0.63, 0.98)

Mexican Americans:

1 μg/dL: 1.00 (Ref)

3 μg/dL: 0.90 (0.73, 1.11)

Gollenberg et al.

(2010)

U.S.A.

1988-1994

Luteinizing 

hormone (LH) 

and inhibin B

Girls ages 6-11 from 

the NHANES III study 

Blood Pb Median 2.5 (range 

0.07, 29.4)

blood Pb

>10 μg/dL: 5%

OR (95% CI) for exceeding 

pubertal inhibin B cutoff 

(>35pg/mL)

<1 μg/dL: 1.00 (Ref)

1-4.9 μg/dL: 0.38 (0.12, 1.15)

≥ 5 μg/dL: 0.26 (0.11, 0.60)

OR (95% CI) for exceeding 

pubertal LH cutoff (>0.4 mIU/mL)

<1 μg/dL: 1.00 (Ref)

1-4.9 μg/dL: 0.98 (0.48, 1.99)

≥ 5 μg/dL: 0.83 (0.37, 1.87)

*a sensitivity analysis including 

only those with blood Pb <10 

μg/dL had similar results but ORs 

were slightly attenuated

a

Studies are presented in order of first appearance in the text of this section.

1 Several epidemiologic studies investigated the association between blood Pb and 

2 indicators of puberty onset. A study among girls aged 10-13 years (median: 12 years) 
February 2012 5-431 Draft – Do Not Cite or Quote

1 reported lower levels of FSH and LH levels in the group with blood Pb of at least 

2 10 μg/dL compared to the group with blood Pb less than 10 μg/dL (Tomoum et al., 

3 2010). In addition, there were some indications of lower Tanner stages of breast 

4 development associated with Pb levels of at least 10 μg/dL, but this relationship was not 

5 present for stages of pubic hair development. A study of girls aged 10-16.9 years of age 

6 in the Akwesasne Mohawk Nation reported a nonlinear positive association between 

7 blood Pb and age at menarche (Denham et al., 2005). No association was observed below 

8 blood Pb of 0.49 μg/dL in a nonlinear model of the Pb-menarche relationship. A study 

9 conducted in South Africa reported a positive association between blood Pb levels and 

10 age at first menarche and pubertal development (Naicker et al., 2010). Another study 

11 reporting on girls with low blood Pb concentrations observed an association between 

12 blood Pb and pubic hair but not breast development (Den Hond et al., 2011). The 

13 association was no longer statistically significant when a marker for polychlorinated 

14 biphenyl exposure was included in the model. A study performed in NYC among 9 year 

15 old girls reported no association between Pb levels and pubertal development (Wolff et 

16 al., 2008), but this age group may be too young to study when investigating delayed 

17 puberty as the outcome. 

18 Multiple studies have been performed examining blood Pb levels and puberty using 

19 NHANES III data (Gollenberg et al., 2010; Selevan et al., 2003; Wu et al., 2003b). A 

20 study that included girls aged 8-16 years and reported an association for delayed 

21 attainment of menarche and pubic hair development, but not for breast development (Wu 

22 et al., 2003b). The associations were observed even at blood Pb levels of 2.1-4.9 μg/dL 

23 compared to girls with blood Pb levels <2.1 μg/dL. Another NHANES III study included 

24 girls 8-18 years of age and reported the results stratified by race (Selevan et al., 2003).

25 Higher blood Pb levels were associated with lower Tanner stage of breast and pubic hair 

26 development and later age at menarche among African Americans and with lower stage 

27 of breast and pubic hair development among Mexican Americans. For whites, the 

28 associations were in the same directions, but none reached statistical significance. In a 

29 study of girls ages 6-11 years old from NHANES III data, higher blood Pb levels were 

30 associated with lower inhibin B, a protein that inhibits FSH production, but no 

31 association was observed for LH. (Gollenberg et al., 2010). The inverse association 

32 between blood Pb and inhibin B was greater among girls with iron deficiency compared 

33 to those with high Pb but sufficient iron levels. Inhibin B and LH were chosen for this 

34 study because, as the authors indicated, these hormones are, “believed to be relevant for 

35 younger girls... near the onset of puberty and...serve as markers for hypothalamic36 pituitary-gonadal functioning.” 

37
February 2012 5-432 Draft – Do Not Cite or Quote

Figure 5-46 Toxicological exposure-response array for reproductive effects 

of Pb.
February 2012 5-433 Draft – Do Not Cite or Quote

Table 5-33 Toxicological concentration-response array summary for 

reproductive effects of Pb presented in Figure 5-46

Reference

Blood Pb level with Effect 

(μg/dL) Altered Outcome

Iavicoli I. et al. (2006b) 8 & 13 Delayed onset female puberty

Leasure et al. (2008) 10 & 42

10, 24 & 42

10 & 42

Neurotransmitter, Dopamine homeostasis

Physical Development, Adult obesity (males)

Aberrant response to amphetamine

Fox et al. (2008) 12 Retinal aberrations

Nava-Hernandez et al. (2009) 19.5 Sperm affected via redox imbalance

Moorman et al. (1998) 25-130 Semen quality affected

Teijon et al. (2006) 40 & 100

40 & 100

40 & 100

100

Hematology

Histology-Offspring renal & hepatic

Biomarker-Offspring renal function

Physical development: birth weight

Fox et al. (2008) 12 Retinal aberrations

Nava-Hernandez et al. (2009) 19.5 Sperm affected via redox imbalance

Moorman et al. (1998) 25-130 Semen quality affected

Teijon et al. (2006) 40 & 100

40 & 100

40 & 100

100

Hematology

Histology-Offspring renal & hepatic

Biomarker-Offspring renal function

Physical development: birth weight

Fox et al. (2008) 12 Retinal aberrations

1 Earlier studies showed that prenatal and lactational exposures to Pb can cause a delay in 

2 the onset of female puberty in rodents. Recent studies corroborate these findings and 

3 show that puberty onset is one of the more sensitive markers of effects of Pb exposure as 

4 is demonstrated in the exposure response array (Figure 5-46 and Table 5-33 Figure 5-45; 

5 including outcomes described in sections that follow). Dumitrescu et al. (2008b) exposed 

6 adult Wistar female rats to varying doses of Pb-acetate (50-150 ppb) in drinking water for 

7 3 months before mating and during pregnancy. Vaginal opening, an indicator of sexual 

8 maturation, was statistically significantly delayed in pups from all Pb treated groups 

9 when compared to pups from non-treated dams. The age at vaginal opening in female 

10 pups from the Pb treated groups increased, in a concentration-dependent manner, from 39 

11 days to 43-47 days. The authors also observed a correlation between body weight and age 

12 at vaginal opening meaning that as body weight decreased the age at vaginal opening 

13 increased. This effect also exhibited a concentration-dependent relationship. 

14 In another recent study, Iavicoli et al. (2006b) reported a statistically significant delay in 

15 several indicators of sexual maturity in offspring (Swiss mice, F1 generation) born to 

16 dams that ingested 3.5-40 ppm Pb in their daily diet; offspring had continuous dietary 

17 exposure until the termination of the experiment. Maternal ingestion of Pb at the various 
February 2012 5-434 Draft – Do Not Cite or Quote

1 doses resulted in female pup blood Pb levels of 3.5-13 μg/dL. For all diet groups in this 

2 range (3.5-13 μg/dL), there was a delay in age at vaginal opening, age of first estrus, age 

3 of vaginal plug formation, and age of first parturition when compared to the group at 

4 background Pb concentration (2 μg/dL). A novel finding in the Iavicoli study was that 

5 very low dose Pb (blood Pb of 0.7 μg/dL, food concentration of 0.02 ppm continuous 

6 through gestation, lactation and until the termination of the experiment) induced 

7 statistically significant acceleration of markers of sexual maturation in female offspring 

8 versus background Pb level animals (blood Pb of 2 μg/dL). There were statistically 

9 significant increases in time of vaginal opening (30% earlier), first estrous, first vaginal 

10 plug formation, and first parturition at the very low Pb exposure versus 2 μg/dL animals. 

11 Thus, the timing of puberty is delayed in a concentration-dependent fashion with very 

12 low dose Pb having a statistically significant earlier onset of puberty than the background 

13 Pb animals (2 μg/dL). Also, the animals exposed to the higher dose of Pb (blood Pb up to 

14 13 μg/dL) had statistically significant delays in onset of puberty when compared to the 

15 other dose groups. 

16 In addition, Pb-induced shifts in sexual maturity were observed in the subsequent 

17 generation (F2 generation) across that dose range. These F2 animals continued to be 

18 exposed to same concentrations of Pb over multiple generations through the diet. Results 

19 in the F2 generation closely resembled those of the F1 generation, as both generations 

20 received Pb exposure. The authors concluded that a modest elevation in blood Pb level 

21 (13 μg/dL) over background (2-3 μg/dL) can result in a profound delay in the onset of 

22 puberty (15-20%). In the F2 generation, reduction in blood Pb (0.7 μg/dL) below 

23 background (2-3 μg/dL) was associated with an earlier onset of sexual maturity (30% 

24 increase) above background. 

25 In the 2006 Pb AQCD (U.S. EPA, 2006b), it was reported that a statistically significant 

26 reduction in the circulating levels of insulin-like growth factor 1 (IGF-1), LH, and 

27 estradiol (E2) was associated with Pb-induced delayed puberty in Fisher 344 pups. 

28 Subsequently, Pine et al. (2006) evaluated whether IGF-1 replacement could reverse the 

29 effects of Pb on delayed female puberty onset. The authors reported that offspring from 

30 dams exposed to Pb during gestation and lactation (daily oral gavage of dam with 1.0 mL

31 solution of Pb-acetate 12 mg/mL; mean maternal blood Pb level 40 μg/dL) exhibited a 

32 marked increase in LH and luteinizing hormone releasing hormone (LHRH) secretion 

after IGF-1 administration (200 ng3 33 /μL i.p. injection twice daily from PND23 until the 

34 appearance of vaginal opening which appears in control animals at ~ PND 40) resulting 

35 in restored timing of vaginal to that of control animals. It should be noted that, IGF-1 

36 replacement in Pb-exposed animals did not cause advanced puberty over non-Pb-exposed 

37 controls. The results of this study provide support to the theory that Pb-induced delayed 

38 onset of puberty may be due to disruption of pulsatile release of sex hormones (U.S. 
February 2012 5-435 Draft – Do Not Cite or Quote

1 EPA, 2006b) and not necessarily due to a direct toxic effect on the hypothalamic2 pituitary-gonadal axis (Salawu et al., 2009), and IGF-1 may play a prominent role in the 

3 process.

4 In sum, epidemiologic studies consistently show an association between higher 

5 concurrent blood Pb and delayed pubertal development in girls. This association is 

6 apparent even at low blood Pb levels. New evidence from the toxicology literature 

7 continues to support Pb-induced delays in the onset of puberty. Further, the biological 

8 plausibility of delayed puberty is expanded with the toxicological literature that shows 

9 this pathway is mediated by IGF-1. 

5.8.1.4 Effects on Lactation

10 Experiments in laboratory animals have shown that dietary manipulation of maternal 

11 fatty acid (FA) levels in diet can worsen Pb-related behavioral effects of offspring after 

12 lactational Pb exposure (Lim et al., 2005). To determine if components of dam milk 

13 contributed to this change, dam dietary fatty acids were altered via diet. Diets deficient in 

14 n-3 fatty acids can lead to a deficiency of DHA, which is essential for proper nervous 

15 system development. Lim et al. (2005) found that dam Pb exposure (Long-Evans rats, 

16 0.2% Pb-acetate trihydrate/BW) during lactation (PND 0-21) led to a decrement in 

17 non-essential fatty acids in the maternal organs at PND25 (mean [SD] blood Pb levels in 

18 dams: 308 [56] μg/dL). In animals with a diet deficient in n-3 FAs, there was a Pb-diet 

19 interaction on the 20-carbon n-6 PUFAs. In general, Pb exposure caused a decrement in 

20 shorter chain monounsaturated and saturated FAs in maternal organs. 

21 Dietary supplementation with calcium can be an especially important contributor to Pb 

22 mobilization during periods of high calcium demand including pregnancy/lactation. For 

23 example, mothers with elevated blood Pb levels given calcium phosphate and ascorbic 

24 acid supplementation during lactation had a 90% decrease in placental Pb content and a 

25 15% decrease in the concentration of Pb in breast milk (Altmann et al., 1981) versus the 

26 control group that did not receive dietary treatment. Another study (Gulson et al., 2004a)

27 has shown that calcium supplementation during the lactation is less beneficial in 

28 modulating maternal blood Pb levels (mean blood Pb at first sampling was 2.4 μg/dL); 

29 the Gulson cohort was limited by power (n=10 women). In a cohort of women from 

30 Mexico City, daily calcium supplementation during lactation reduced maternal blood lead 

31 by 15–20% and lead in breast milk by 5–10% (Ettinger et al., 2004a). Another study by 

32 the same investigators showed that using calcium supplements daily during pregnancy 

33 also reduced blood lead levels during pregnancy (Ettinger et al., 2009) with the effect 

34 strongest in women with higher biomarkers of Pb exposure (elevated baseline bone Pb or 
February 2012 5-436 Draft – Do Not Cite or Quote

1 >5 μg/dL blood Pb) or in women with higher Pb exposure (self-reported use of lead2 glazed ceramics). Thus, dietary modulation with calcium supplementation during 

3 pregnancy and lactation may decrease the amount of Pb to which the developing fetus of 

4 infant is exposed. The evidence for this seems especially strong for protection during 

5 pregnancy and more mixed for protective effects of calcium during lactation.

5.8.1.5 Summary of Effects on Female Reproductive Function

6 In summary, Pb exposure was found to affect female reproductive function as 

7 demonstrated by both epidemiologic and toxicological studies. At low concurrent blood 

8 Pb levels, associations are observed with delayed puberty. These associations are noted 

9 among girls with mean blood Pb levels of 10 μg/dL and lower. Some evidence is also 

10 available regarding blood Pb levels and altered hormone levels in adults, although the 

11 direction of the change varied among studies. Although studies reported inconsistent 

12 findings for the association between Pb and fertility, there is some evidence of a potential 

13 relationship. Most of the epidemiologic studies are cross-sectional, thus there is 

14 uncertainty regarding the lifestages of Pb exposure associated with the greatest risk. 

15 Toxicological studies are often dealing with prenatal or early postnatal exposures, except 

16 for puberty studies which use concurrent exposure. Although epidemiologic and 

17 toxicological studies provide information on different exposure periods, both types of 

18 studies support the conclusion that Pb affects at least some aspects of female reproductive 

19 function. 

5.8.2 Effects on Male Reproductive Function

20 The 2006 Pb AQCD (U.S. EPA, 2006b) reported on male Pb exposure or biomarker 

21 levels and reproductive functions as measured by sperm count/motility/morphology, time 

22 to pregnancy, reproductive history, and chromosomal aberrations. Despite limitations, 

23 most of the studies found slight associations between high blood Pb levels (i.e., ≥ 

24 45 μg/dL) and reduced male fecundity or fertility (U.S. EPA, 2006b). Evidence reviewed 

25 in the 1986 Pb AQCD (U.S. EPA, 1986a) also demonstrated that Pb exposure affects 

26 male reproductive function in humans and experimental animals. Recently published 

27 research has continued to support an association between Pb and reproductive function in 

28 males. These studies are described in the sections below.
February 2012 5-437 Draft – Do Not Cite or Quote

5.8.2.1 Effects on Sperm/Semen Production, Quality, and 

Function 

1 Multiple epidemiologic and toxicological studies have examined the relationship between 

2 Pb and sperm and semen production, quality, and function. These studies are summarized 

3 in the text below. In addition, recent epidemiologic studies are included in Table 5-34. 

4 The biological samples in these studies are from concurrent measures. The biological 

5 samples measuring Pb levels and outcomes were collected concurrently in the 

6 epidemiologic studies.

Table 5-34 Summary of recent epidemiologic studies of associations between 

Pb levels and effects on sperm and semen

Referencea

Study 

Location 

and Years

Study 

Population

Pb Biomarker or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Naha and Manna

(2007)

Bangalore, 

India

NS

Non-occupationally 

exposed controls 

and occupationally 

exposed workers 

Categorized by work history as 

controls, low exposure (7-10 yr 

of exposure for 8 h/day) and 

high exposure (>10 yr of 

exposure for 8 h/day)

Blood Pb 

measurement

Controls 10.25 

(2.26)

Low exposure 

50.29 (3.45)

High exposure 

68.26 (2.49)

Semen Pb 

measurement

Controls 2.99 

(0.76)

Low exposure 

15.85 (1.95)

High exposure 

25.30 (2.28)

p-values for difference across the three groups 

for mean values of semen profiles were <0.01 

for: liquefaction time, seminal volume, sperm 

count, sperm DNA hyploidy, sperm 

morphological abnormality, sperm motility, 

sperm ATPase activity, seminal plasma 

fructose, seminal plasma total protein, seminal 

plasma free amino acid, seminal plasma 

cholesterol 

Naha and 

Chowdhury (2006)

Kolkata, India

NS

Men aged 31-45 

that were nonoccupationally 

exposed controls 

and occupationally 

exposed workers)

Categorized by work history as 

controls, low exposure (7-10 yr 

of exposure for 8 h/day) and 

high exposure (>10 yr of 

exposure for 8 h/day) 

Blood Pb 

measurement

Controls 13.62 

(2.45)

Low exposure 

48.29 (4.91)

High exposure 

77.22 (1.25)

Semen Pb 

measurement

Controls 3.99 

(1.36)

Low exposure 

10.85 (0.75)

High exposure 

18.30 (2.08)

p-values for difference across the three groups 

for mean values of semen profiles were <0.01 

for: sperm count, sperm protein, sperm DNA 

hyploidy, sperm DNA, sperm RNA, sperm 

viability, sperm membrane lipid peroxidation, 

seminal plasma total ascorbate, seminal plasma 

DHAA, sperm ATPase activity, sperm motility, 

sperm velocity, seminal plasma fructose
February 2012 5-438 Draft – Do Not Cite or Quote

Referencea

Study 

Location 

and Years

Study 

Population

Pb Biomarker or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Hsu et al. (2009b) Taiwan

NS

Men working at a 

battery plant

Blood Pb

Categorized into 3 groups: 

<25 μg/dL, 

25-45 μg/dL, 

>45 μg/dL

40.2 p-values for difference across the three groups 

were <0.05 for: sperm head abnormalities, 

sperm neck abnormalities, sperm chromatin 

structure assay (αT, COMPαT)

p-values for difference across the three groups 

were >0.05 for: semen volume, sperm count, 

motility, sperm tail abnormalities, sperm 

immaturity, computer-assisted semen analysis, 

% sperm with ROS production

Coefficients for regression analysis with blood 

Pb:

Morphologic abnormality 0.271 (p-value 

<0.0001)

Head abnormality 0.237 (p-value 0.0002)

αT 1.468 (p-value 0.011)

COMPαT 0.233 (p-value 0.21)

Kasperczyk et al.

(2008)

Poland

NS

Healthy, nonsmoking, fertile men 

that worked at the 

Zn and Pb 

Metalworks

Blood Pb; seminal fluid Pb

Categorized as high exposure

workers (blood Pb 40-81 μg/dL), 

low exposed workers (blood Pb 

25-40 μg/dL), and controls 

(office workers with no history of 

occupational Pb exposure)

Blood Pb

High exposure 

workers: 53.1 

(2.05)

Low exposure 

workers: 34.7 

(0.83)

Controls: 8.47 

(0.54)

Seminal plasma 

Pb

High exposure 

workers: 2.02 

(0.23)

Low exposure 

workers: 2.06 

(0.40)

Controls: 1.73 

(0.16)

Mean (SE)

Sperm volume (mL) 

Controls: 2.94 (0.32)

Low exposure: 2.89 (0.22) 

High exposure: 2.98 (0.22) 

(p-value for ANOVA: 0.993)

Sperm cell count (mln/mL) 

Controls: 43.1 (7.0)

Low exposure: 44.6 (10.1)

High exposure: 42.2 (5.86)

(p-value for ANOVA: 0.400)

Normal morphology (%) 

Controls: 63.3 (2.7)

Low exposure: 57.3 (2.5)

High exposure: 58.4 (2.1)

(p-value for ANOVA: 0.266)

Progressively motile sperm after 1 h (%) 

Controls: 16.4 (3.2)

Low exposure: 14.8 (2.6)

High exposure: 10.5 (1.9)

(p-value for ANOVA: 0.217)

Motile sperm after 24 h (%) 

Controls: 4.4 (1.8)

Low exposure: 7.3 (1.7)

High exposure: 3.1 (0.8)

(p-value for ANOVA: 0.188)

p-value for correlation between blood Pb and 

sperm cell motility after 1 h: 0.011
February 2012 5-439 Draft – Do Not Cite or Quote

Referencea

Study 

Location 

and Years

Study 

Population

Pb Biomarker or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Telisman et al.

(2007)

Croatia

2002-2005

Men aged 19-55, 

never 

occupationally 

exposed to metals 

and going to a clinic 

for infertility 

examination or for 

semen donation to 

be used for artificial 

insemination

Blood Pb Median: 4.92 

(range 1.13- 14.91)

Standardized regression coefficients for log 

blood Pb (units not given)

Immature sperm: 0.13 (p-value <0.07)

Pathologic sperm: 0.31 (p-value <0.0002)

Wide sperm: 0.32 (p-value <0.0001)

Round sperm: 0.16 (p-value <0.03)

Coefficients and p-values not given if not 

statistically significant: semen volume, sperm 

concentration, slow sperm, short sperm, thin 

sperm, amorph sperm

Meeker et al. (2008) Michigan

NS

Men aged 18-55 

going to infertility 

clinics (distinction 

not made between 

clinic visits for male 

or female fertility 

issues)

Blood Pb Median: 1.50 (IQR 

1.10, 2.00)

OR (95% CI) for having below reference-level 

semen parameters

Concentration

1st quartile: 1.00 (ref)

2nd quartile: 0.88 (0.32, 2.44)

3rd quartile: 2.58 (0.86, 7.73)

4th quartile: 1.16 (0.37, 3.60)

Motility

1st quartile: 1.00 (ref)

2nd quartile: 1.04 (0.43, 2.53)

3rd quartile: 1.95 (0.70, 5.46)

4th quartile: 1.66 (0.64, 4.29)

Morphology

1st quartile: 1.00 (ref)

2nd quartile: 0.83 (0.37, 1.87)

3rd quartile: 1.41 (0.54, 3.67)

4th quartile: 1.18 (0.50, 2.79)

Models with adjustment for multiple metals

Concentration

1st quartile: 1.00 (ref)

2nd quartile: 0.89 (1.57, 2.89)

3rd quartile: 3.94 (1.15, 13.6)

4th quartile: 2.48 (0.59, 10.4)

Slivkova et al.

(2009)

NS Men aged 22-48 

undergoing semen 

analysis at an 

infertility clinic

Semen Pb 1.49 mg/kg (0.40 

mg/kg)

Correlation between Pb and flagellum ball : -

0.39 (p-value not given)

*correlations not given for any other sperm 

pathological changes (therefore assume not 

statistically significant): broken flagellum, 

separated flagellum, separated flagellum, small 

heads, retention of cytoplasmic drop, other 

pathological spermatozoa, large heads, 

acrosomal changes, and knob twisted flagellum
February 2012 5-440 Draft – Do Not Cite or Quote

Referencea

Study 

Location 

and Years

Study 

Population

Pb Biomarker or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Mendiola et al.

(2011)

Spain

2005-2007

Men attending 

infertility clinics and 

classified as either 

normal sperm 

(controls) or oligo- asthenoteratozoospermia 

(cases) based on 

WHO semen quality 

criteria

Seminal plasma Pb

Blood Pb

Seminal plasma: 

2.90 (IQR 2.70, 

3.20)

Whole blood: 9.50 

(IQR 7.50, 11.90)

Blood plasma: 

2.90 (IQR 2.70, 

3.10)

Cases:

Seminal plasma: 

3.0 (0.3)

Whole blood: 9.8 

(2.3)

Blood plasma: 2.9 

(0.2)

Controls:

Seminal plasma: 

2.9 (0.3)

Whole blood: 9.7 

(2.3)

Blood plasma: 2.9 

(0.3)

β (95% CI) 

Sperm concentration

Seminal plasma: -1.0 (-3.1, 2.3)

Whole blood: -0.2 (-1.7, 1.6)

Blood plasma: 0.08 (-4.1, 5.2)

% Immotile sperm 

Seminal plasma: 1.5 (0.37, 1.9)

Whole blood: 0.05 (-0.32, 0.43)

Blood plasma: -0.49 (-1.8, 6.2)

% Morphologically normal sperm 

Seminal plasma: -0.54 (-3.1, 2.0)

Whole blood: -0.31 (-1.5, 0.89)

Blood plasma: -0.08 (-3.5, 3.4)

*Units not given (assume 1 μg/dL)

Note: No correlation in Pb levels among bloods 

or seminal plasma. There was correlation 

between Pb and other metals (Cd and Hg) 

within each body fluid. Other metals were not 

controlled for in models

a

Studies are presented in order of first appearance in the text of this section.

1 International epidemiologic studies of men occupationally exposed to Pb have reported 

2 on associations between Pb exposure or biomarker levels and sperm count and quality 

3 and semen quality. In most of these occupational studies, blood Pb levels over 40 μg/dL 

4 have been reported for individuals occupationally exposed to Pb. For example, studies 

5 performed in India (Naha and Manna, 2007; Naha and Chowdhury, 2006) reported that 

6 men in the highest exposure group (men working in battery or paint manufacturing plants 

7 for 10-15 years for 8 hours/day) had mean blood Pb levels of 77.22 μg/dL (Naha and 

8 Chowdhury, 2006) and 68.26 μg/dL (Naha and Manna, 2007). Control groups in these 

9 studies (those without occupational Pb exposure) had mean blood Pb levels below 

10 15 μg/dL. Increases in levels of Pb in semen were also noted across exposure groups. 

11 Both studies report decreases in sperm count and in sperm velocity and motility with 

12 increasing Pb exposure. Higher Pb exposure was also associated with greater hyploidy of 

13 sperm DNA and morphologic abnormalities (Naha and Manna, 2007; Naha and 

14 Chowdhury, 2006). Decreased viability and increased lipid peroxidation were detected

15 (Naha and Chowdhury, 2006). A study performed in Taiwan among men with high levels 

16 of blood Pb reported that men with higher blood Pb levels had increased sperm head 

17 abnormalities, increased sperm DNA denaturation, and increased sensitivity to 

18 denaturation compared to men with lower blood Pb levels (Hsu et al., 2009b). No 

19 difference was detected between three Pb exposure groups and semen volume, sperm 

20 count, motility, velocity, and reactive oxygen species production. A similar study in 
February 2012 5-441 Draft – Do Not Cite or Quote

1 Poland included employees exposed to Pb and compared them with a group of male 

2 office workers (Kasperczyk et al., 2008). Pb levels measured in seminal fluid were 

3 slightly higher among those in the exposed groups although they were not statistically 

4 different from the levels in the control group. No difference was observed for semen 

5 volume, sperm count, or sperm morphology among the groups. Sperm motility was lower 

6 in the highest exposure group compared to both the control and moderate exposure 

7 groups. Lipid peroxidation, which can induce tissue damage in sperm via reactive oxygen 

8 species, was greater in the highest exposure group compared to the controls. 

9 A study performed in Croatia recruited men who had never been occupationally exposed 

10 to metals and therefore had lower blood Pb levels than the occupational studies (Telisman 

11 et al., 2007). Increased blood Pb was associated with increased percentages of pathologic 

12 sperm, wide sperm, and round sperm. There was also a slight increase in immature sperm 

13 although it was not statistically significant. Similar results were seen when other 

14 biomarkers for Pb (erythrocyte protoporphyrin and δ-aminolevulinic acid dehydratase 

15 [ALAD]) were used instead.

16 A few studies examined blood or seminal plasma Pb levels and semen quality of men at 

17 infertility clinics (Mendiola et al., 2011; Slivkova et al., 2009; Meeker et al., 2008). In 

18 general, these men had lower levels of Pb biomarkers than men that were occupationally 

19 exposed. Meeker et al. (2008) detected no associations between higher blood Pb and 

20 semen concentration, morphology, or motility (although a slight positive trend was 

21 observed between higher Pb levels and motility in unadjusted models). In models that 

22 include multiple metals, blood Pb was associated with being below the WHO limit of 

23 sperm concentration levels (less than 20 million sperm/mL), although the 95% CI was 

24 wide for the 4th quartile of Pb levels and included the null. Slivkova et al. (2009)

25 reported a negative correlation between semen Pb and pathological changes in sperm 

26 (specifically, flagellum ball), but no correlations were observed for other alterations in 

27 the sperm. Another study reported a positive association between seminal plasma Pb 

28 concentration and percentage of immotile sperm, but this analysis did not adjust for 

29 exposure to other metals reported to be correlated with Pb concentration in the seminal 

30 plasma (Mendiola et al., 2011). No association was observed for seminal plasma Pb 

31 concentration and sperm concentration or percentage of morphologically normal sperm. 

32 Additionally, neither Pb levels in whole blood or plasma were associated with sperm 

33 concentration, percentage of immotile sperm, or percentage of morphologically normal 

34 sperm. 

35 An abundance of evidence in the toxicological literature demonstrates that Pb exposure is 

36 detrimental to the quality and overall health of testicular germ cells. Earlier studies 

37 showed that chronic Pb exposure (15 weeks) in adult male rabbits, resulting in blood Pb 
February 2012 5-442 Draft – Do Not Cite or Quote

1 of 16-24 μg/dL, induced statistically significant decrements in semen quality and greater 

2 testicular pathology (Moorman et al., 1998). Recent studies corroborate earlier findings 

3 that Pb alters sperm parameters such as sperm count, viability, motility, and morphology. 

4 Anjum et al. (2010) exposed 50 day old male albino Wistar/NIN rats to Pb-acetate (273 

5 or 819 mg/L in drinking water, 0.05% or 0.15%, respectively) for 45 days. Affected 

6 endpoints included reduced epididymal sperm count, motile sperm, and viable sperm, 

7 indicating decreased sperm production and quality. Anjum did not report blood Pb 

8 values. Wistar/NIN rats (0.15% Pb-acetate in drinking water for 70 days) supplemented 

9 with the herb Centella asiatica had significant attenuation of the Pb-induced changes 

10 observed by Anjum et al. (2010). Pillai et al. (In Press) found gestational and lactational 

11 treatment with Pb-acetate in Charles Foster rats (subcutaneous injection of 0.05 mg/kg 

12 BW/day) induced effects on sperm in adults (PND65) including significant decreases in 

13 testicular sperm count, epididymal sperm count, and sperm motility. Oliveira et al. (2009)

14 observed a negative correlation between Pb dose and intact acrosomes. Rubio et al.

15 (2006), Biswas and Ghosh (2006), and Salawu et al. (2009) observed a decrease in 

16 absolute testicular weight after Pb exposure. Rubio et al. (2006) and Biswas and Ghosh

17 (2006) also observed a Pb-induced decrease in seminal vesicle and ventral prostate 

18 weights and Rubio et al. (2006) reported that Pb-acetate, in a exposure concentration19 dependent manner (8-24 mg/kg body weight), reduced the length of certain stages of the 

20 spermatogenic cycle of rat seminiferous tubules and thus affected spermatogenesis. Oral 

21 Pb-acetate exposure (25 mg/kg bw in drinking water for 3 months, resulting in blood Pb 

22 level of 5.3 μg/dL) to adult male albino rats produced significant histological 

23 seminiferous tubule damage (epithelium, spermatocytes, acrosomes) that was attenuated 

24 with ascorbic acid treatment (Pb exposure + 100 mg/kg bw/day ascorbic acid, resulting in 

25 blood Pb level of 4.7 μg/dL) (El Shafai et al., 2011). Reshma Anjum et al. (2010)

26 reported decreased testicular and epididymal weights, sperm count, and viable sperm of 

27 male rats exposed to Pb-acetate (273 mg/L or 819 mg/L in drinking water) which were 

28 significantly attenuated with Pb co-exposure to the herb Centella asiatica (Sainath et al., 

29 2011). Pb induced morphological abnormalities in sperm in a concentration-dependent 

30 manner (Allouche et al., 2009; Oliveira et al., 2009; Salawu et al., 2009; Shan et al., 

31 2009; Tapisso et al., 2009; Massanyi et al., 2007; Wang et al., 2006a). Sperm 

32 abnormalities reported after Pb exposures were amorphous sperm head, abnormal tail, 

33 and abnormal neck. Dong et al. (2009) reported decreased epididymis and body weights 

34 in mice after exposure to 0.6% Pb-acetate in drinking water. However, the majority of 

35 studies did not observe a statistically significant difference in body weight or 

36 reproductive organ weights after Pb exposure at the doses used in the studies. Not all of 

37 the aforementioned studies observed changes in every parameter. This may be due to the 

38 use of different strains or species, chemical form of the Pb compound administered, 
February 2012 5-443 Draft – Do Not Cite or Quote

1 dosage schedule, duration of exposure, and age of animals at the time of the study

2 (Oliveira et al., 2009).

3 Data from recent studies suggest that a component of Pb-induced toxicity is the 

4 generation of reactive oxygen species (ROS) which can then affect antioxidant defense 

5 systems of cells (Pandya et al., 2010). Salawu et al. (2009) observed a statistically 

6 significant increase in malondialdehyde (MDA, oxidative stress marker) and a significant 

7 decrease in the activity of antioxidant enzymes superoxide dismutase (SOD) and catalase 

8 (CAT) in plasma and testes of adult male Sprague Dawley rats after administration of 1% 

9 Pb-acetate in drinking water for 8 weeks. Supplementation with tomato paste (used as a 

10 source of antioxidants) reduced Pb-induced ROS production and prevented the Pb11 induced increase in MDA formation and decrease in SOD and CAT activity. 

12 Furthermore, co-treatment of Pb with substances that are known to have antioxidant 

13 properties [i.e., tomato paste, Maca (Lepidium meyenii), and ascorbic acid] prevented the 

14 Pb-induced reduction in sperm count, sperm motility, and sperm viability (Salawu et al., 

15 2009; Shan et al., 2009; Madhavi et al., 2007; Rubio et al., 2006; Wang et al., 2006a).

16 Recent studies also demonstrate that Pb may be directly toxic to mature spermatozoa

17 (Tapisso et al., 2009; Hernandez-Ochoa et al., 2006) as well as primary spermatocytes

18 (Nava-Hernandez et al., 2009; Rafique et al., 2009). Nava-Hernandez et al. (2009)

19 exposed two groups of rodent to Pb via drinking water (L1 and L2). In their study, all 

20 Pb-treated animals had blood Pb levels statistically significantly higher than controls 

21 (L1:19.54 μg/dL and L2:21.90 μg/dL); no statistically significant difference in blood Pb 

22 levels existed between the two Pb exposure groups likely because the L2 group drank less 

23 water than did the L1 group. Piao et al. (2007) reported that Pb exposure caused DNA 

24 damage to sperm; the Pb exposed group had a blood Pb of 67 μg/l. Piao et al. (2007) also 

25 examined the effect of Zn supplementation on Pb-induced sperm aberrations and found 

26 that the proportion of abnormal sperm was statistically significantly higher in the Pb 

27 group and the Pb+Zn group than in controls. However, the proportion of abnormal sperm 

28 in Pb+Zn group was statistically significantly lower than in Pb alone group. 

29 Hernandez-Ochoa et al. (2006) reported that Pb reaches the sperm nucleus in the 

30 epididymis of mice chronically exposed (16 weeks in adult animals) to Pb (resulting in 

31 mean blood Pb of 75.6 μg/dL) by binding to nuclear sulfhydryl groups from the 

32 DNA-protamine complex, increasing sperm chromatin condensation, and thereby 

33 interfering with the sperm maturation process without altering sperm quality parameters. 

34 Tapisso et al. (2009) observed a statistically significant increase in the number of 

35 micronuclei and frequency of sister chromatid exchange with increasing treatment 

36 duration in adult male mice administered 21.5 mg/kg body weight Pb-acetate by i.p. 

37 injection. Nava-Hernandez (2009) reported a concentration-dependent increase in DNA 

38 damage in rat primary spermatocytes after a 13-week exposure period to Pb-acetate in 
February 2012 5-444 Draft – Do Not Cite or Quote

1 drinking water (resulting in mean blood Pb levels between 19.5 and 21.9 μg/dL). Rafique 

2 et al. (2009) reported degenerative changes from pyknosis to apoptosis in primary 

3 spermatocytes. Hepatic expression of spermatogenic genes was transiently down4 regulated in 8 week old male Wistar-Kyoto (WKY) rats in response to Pb nitrate 

5 (100 μmol single i.v. injection) 3 hours after injection and recovered to baseline by 12 

6 hours (Nemoto et al., 2011); this effect was not seen in the stroke-prone spontaneously 

7 hypertensive rats, which are from a WKY background, or in Sprague-Dawley rats, 

8 demonstrating strain specificity.

9 Pb-induced apoptosis in germ cells within the seminiferous tubules is another suggested 

10 mechanism by which Pb exerts its toxic effects on sperm production and function (Wang 

11 et al., 2006a). Dong et al. (2009) reported a exposure concentration-related increase in 

12 apoptosis in spermatogonia and spermatocytes of Kunming mice after exposure to 

13 0.15-0.6% Pb-acetate in drinking water. Pb-induced testicular germ cell apoptosis was 

14 associated with up-regulation of genes involved in the signal pathway of MAPK and 

15 death receptor signaling pathway of FAS. For instance, up-regulation of K-ras and Fas 

16 expressions was concomitant with activation of c-fos and active caspase-3 proteins. 

17 Wang et al. (2006a) observed a exposure concentration-dependent increase in the 

18 expression of apoptotic markers TGFβ1 and caspase-3 in spermatogenic cells, Sertoli 

19 cells, and Leydig cells. Shan et al. (2009) also reported a statistically significant increase 

20 in mRNA expression and protein levels of Fas, Fas-L and caspase-3 after Pb exposure. 

21 Supplementation with ascorbic acid inhibited or reduced the Pb-induced apoptosis in 

22 germ cells and protected testicular structure and function (El Shafai et al., 2011; Shan et 

23 al., 2009; Wang et al., 2006a) suggesting ROS generation is a major contributing factor in 

24 decreased male fertility observed after chronic Pb exposure.

25 Similar to the results summarized in previous Pb AQCDs, recent epidemiologic and 

26 toxicological studies indicate that high levels of Pb exposure have effects on sperm and 

27 semen. In studies of men exposed to Pb in occupational settings, associations were 

28 observed between blood Pb levels as low as 20-45 μg/dL and sperm count and quality. 

29 Multiple epidemiologic studies of occupational cohorts included control populations with 

30 high blood Pb levels (close to or greater than 10 μg/dL), which makes identification of 

31 effects at lower levels difficult. Future studies are warranted to determine whether this 

32 association is observed at lower Pb levels.

5.8.2.2 Effects on Hormone Levels

33 The 2006 Pb AQCD (U.S. EPA, 2006b) provided evidence that Pb acts as an endocrine 

34 disruptor in males at various points along the hypothalamic-pituitary-gonadal axis. The 
February 2012 5-445 Draft – Do Not Cite or Quote

1 2006 Pb AQCD also reported inconsistencies in the effects of Pb exposure on circulating 

2 testosterone levels. Recent epidemiologic and toxicological studies are reported below. 

3 Epidemiologic studies are summarized in Table 5-35 (biological samples used for the 

4 measurement of Pb in these studies were measured concurrently with hormone levels).

Table 5-35 Summary of recent epidemiologic studies of associations between 

Pb levels and hormones for males

Referencea

Study 

Location and 

Years Outcome Study Population

Pb Biomarker 

or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Telisman et al.

(2007)

Croatia

2002-2005

FSH, LH, 

testosterone, 

estradiol, prolactin

Men aged 19-55, never 

occupationally exposed to metals 

and going to a clinic for infertility 

examination or for semen donation 

to be used for artificial 

insemination

Blood Pb Median: 4.92 

(range 1.13-14.91)

Standardized regression 

coefficients for log blood Pb 

(units not given)

Testosterone: 0.21 (p-value 

<0.003)

Estradiol: 0.22 (p-value 

<0.0008)

Prolactin:- 0.18 (p-value 

<0.007)

Coefficients and p-values not 

given if not statistically 

significant (LH, FSH)

Naha and 

Manna (2007)

Bangalore, 

India

NS

FSH, LH, 

testosterone 

Non-occupationally exposed 

controls and occupationally 

exposed workers 

Categorized by 

work history as 

controls, low 

exposure (7-10 

yr of exposure 

for 8 h/day) and 

high exposure 

(>10 yr of 

exposure for 8 

h/day)

Blood Pb 

measurement

Controls 10.25 

(2.26)

Low exposure 

50.29 (3.45)

High exposure 

68.26 (2.49)

Semen Pb 

measurement

Controls 2.99 

(0.76)

Low exposure 

15.85 (1.95)

High exposure 

25.30 (2.28)

Mean FSH (SD)

Control: 2.69 (1.22)

Low exposure: 2.58 (1.94)

High exposure: 2.16 (0.99)

p-values for difference >0.05 

Mean LH (SD)

Control: 5.14 (2.35)

Low exposure: 4.27 (2.52)

High exposure: 3.9 (1.69)

p-values for difference >0.05

Mean testosterone (SD)

Control: 5.24 (2.40)

Low exposure: 4.83 (1.21)

High exposure: 4.59 (1.27)

p-values for difference >0.05
February 2012 5-446 Draft – Do Not Cite or Quote

Referencea

Study 

Location and 

Years Outcome Study Population

Pb Biomarker 

or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Hsieh et al.

(2009a)

Taiwan

1991-NS

FSH, LH, 

testosterone, 

inhibin B

Workers at a Pb-acid battery 

factory with annual blood Pb 

measures

Current blood 

Pb, cumulative 

blood Pb, timeweighted 

cumulative blood 

Pb

Current blood Pb:

<10 μg/dL: 11.6%

>40 μg/dL: 17.1%

β from linear regression

Inhibin B

Current blood Pb: 0.40 (pvalue 0.40)

Cumulative blood Pb: 0.05 (pvalue 0.02)

Time-weighted cumulative 

blood Pb: 1.33 (p-value 0.007)

Pearson's correlations 

detected no correlations 

between current blood Pb 

levels and FSH, LH, or 

testosterone. Cumulative 

blood Pb levels were 

correlated with FSH and LH, 

but not testosterone. Timeweighted cumulative blood Pb 

levels were correlated with 

LH, but not FSH or 

testosterone.
February 2012 5-447 Draft – Do Not Cite or Quote

Referencea

Study 

Location and 

Years Outcome Study Population

Pb Biomarker 

or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Meeker et al.

(2010)

Michigan

NS

FSH, LH, inhibin 

B, testosterone, 

SHBG, FAI, 

testosterone/LH

Men aged 18-55 going to infertility 

clinics (distinction not made 

between clinic visits for male or 

female fertility issues)

Blood Pb Median: 1.50 (IQR 

1.10, 2.00)

Regression coefficients (95% 

CI) 

FSH

1st quartile: 1.00 (ref)

2nd quartile: 0.13 (-0.10, 0.37)

3rd quartile: 0.10 (-0.15, 0.35)

4th quartile: 0.07 (-0.18, 0.31)

LH

1st quartile: 1.00 (ref)

2nd quartile:0.004 (-0.20, 

0.21)

3rd quartile: 0.13 (-0.09, 0.35)

4th quartile: 0.88 (-0.14, 0.29)

Inhibin B

1st quartile: 1.00 (ref)

2nd quartile: -6.45 (-27.2, 

14.3)

3rd quartile: -4.62 (-26.6, 17.4)

4th quartile: -7.79 (-29.0, 13.4)

Testosterone

1st quartile: 1.00 (ref)

2nd quartile: 28.6 (-6.82, 64.1)

3rd quartile: 15.8 (-21.8, 53.3)

4th quartile: 39.9 (3.32, 76.4)

SHBG

1st quartile: 1.00 (ref)

2nd quartile:-0.01 (-0.16, 0.15)

3rd quartile: 0.04 (-0.12, 0.21)

4th quartile: 0.07 (-0.10, 0.23)

FAI

1st quartile: 1.00 (ref)

2nd quartile: 0.8 (-0.04, 0.20)

3rd quartile: 0.03 (-0.10, 0.17)

4th quartile: 0.08 (-0.05, 0.21)

Testosterone/LH

1st quartile: 1.00 (ref)

2nd quartile: 0.07 (-0.16, 0.30)

3rd quartile: -0.05 (-0.29, 0.19)

4th quartile: 0.07 (-0.17, 0.31)
February 2012 5-448 Draft – Do Not Cite or Quote

Referencea

Study 

Location and 

Years Outcome Study Population

Pb Biomarker 

or Exposure 

Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Mendiola et al.

(2011)

Spain

2005-2007

FSH, LH, 

testosterone

Men attending infertility clinics and 

classified as either normal sperm 

(controls) or oligo-asthenoteratozoospermia (cases) based 

on WHO semen quality criteria

Seminal plasma 

Pb

Blood Pb

Seminal plasma: 

2.90 (IQR 2.70, 

3.20)

Whole blood: 9.50 

(IQR 7.50, 11.90)

Blood plasma: 2.90 

(IQR 2.70, 3.10)

Cases:

Seminal plasma: 

3.0 (0.3)

Whole blood: 9.8 

(2.3)

Blood plasma: 2.9 

(0.2)

Controls:

Seminal plasma: 

2.9 (0.3)

Whole blood: 9.7 

(2.3)

Blood plasma: 2.9 

(0.3)

Linear regression β (95% CI) 

FSH

Seminal plasma: 0.05 (-0.24, 

0.39)

Whole blood: 0.04 (-0.03, 

0.04)

Blood plasma: -0.20 (-0.64, 

0.25)

LH

Seminal plasma: 0.14 (-0.13, 

0.41)

Whole blood: 0.05 (-0.05, 

0.07)

Blood plasma: -0.07 (-0.49, 

0.31)

Testosterone

Seminal plasma: 0.11 (-0.10, 

0.31)

Whole blood: 0.01 (-0.05, 

0.02)

Blood plasma: -0.12 (-0.40, 

0.14)

*Units not given (assume 1 

μg/dL)

a

Studies are presented in order of first appearance in the text of this section.

1 Hormone levels were measured in a few recent epidemiologic studies. In a study of men 

2 non-occupationally exposed to Pb in Croatia, increased blood Pb level was associated 

3 with increasing serum testosterone and estradiol but decreasing serum prolactin

4 (Telisman et al., 2007). In addition, the analysis of an interaction term for blood Pb and 

5 blood cadmium levels demonstrated a synergistic effect on increasing serum testosterone 

6 levels. No association was observed between blood Pb and FSH or LH. Another study of 

7 men with high blood Pb levels reported no difference in serum FSH, LH, and testosterone 

8 among the three groups (controls: mean blood Pb 10.25 μg/dL, low exposure: mean 

9 blood Pb 50.29 μg/dL, high exposure: mean blood Pb 68.26 μg/dL) (Naha and Manna, 

10 2007). A study of occupationally-exposed men in Taiwan reported an association 

11 between measures of cumulative blood Pb levels and inhibin B levels, but no association 

12 was detected when using current blood Pb levels (Hsieh et al., 2009a). A correlation 

13 between cumulative blood Pb measures and LH levels was detected but correlations were 

14 not present when examining FSH or testosterone levels. No correlations were apparent 

15 between FSH, LH, or testosterone and current blood Pb levels. 

16 Among men recruited from infertility clinics in Michigan, median blood Pb levels were 

17 much lower than those observed in the other studies of Pb and hormone levels among 
February 2012 5-449 Draft – Do Not Cite or Quote

1 men (Meeker et al., 2010). No association was detected between blood Pb and levels of 

2 FSH, LH, inhibin B, sex hormone-binding globulin (SHBG), free androgen index (FAI) 

3 or a measure of Leydig cell function (T/LH). A positive association between the highest 

4 quartile of blood Pb and testosterone was present, but this association did not persist 

5 when other metals were included in the model. Similarly, another study of men recruited 

6 from infertility clinics observed no association between Pb concentrations from seminal 

7 plasma, whole blood, or blood plasma and FSH, LH, or testosterone (Mendiola et al., 

8 2011).

9 In a recent toxicological study, Rubio et al. (2006) observed a decrease in testosterone 

10 levels in Pb-acetate-treated rats in a exposure concentration-related fashion (8-24 mg/kg 

11 body weight), and this decrease correlated with reduced lengths of spermatogenic cycle 

12 stages VII-VIII (spermiation) and IX-XI (onset of spermatogenesis). Anjum et al. (2010),

13 who dosed 50 day old male rats with 273 or 819 mg/L Pb-acetate in drinking water 

14 (0.05% or 0.15%, respectively; blood Pb not reported), found significant decreases in 

15 serum testosterone and testicular 3β-HSD and 17β-HSD levels in Pb-exposed animals 

16 versus controls. Pandya et al. (2010) reported altered hepatic steroidogenic enzyme 

17 activity. Pillai et al. (In Press) found gestational and lactational exposure to Pb-acetate in 

18 Charles Foster rats (subcutaneous injection of 0.05 mg/kg BW/day, blood Pb not 

19 reported) induced significant decreases in testicular 17β-HSD and serum testosterone. 

20 Biswas and Ghosh (2006) reported a Pb-induced decrease in serum testosterone and 

21 gonadotropins (FSH, LH) with inhibition of spermatogenesis, however, there was a 

22 statistically significant increase in adrenal steroidogenic enzyme, Δ5-3β-HSD activity 

23 and serum corticosterone levels indicating disruption of the adrenocortical process. 

24 Exposure concentration-dependent decreases in serum testosterone were reported in Pb25 exposed male rats (Reshma Anjum et al., 2010). In contrast, Salawu et al. (2009) did not 

26 observe a decrease in serum testosterone between control animals and animals 

27 administered 1% Pb-acetate in drinking water for 8 weeks. Allouche et al. (2009) not 

28 only did not observe any statistically significant changes in serum FSH or LH, but 

29 reported an increase in serum testosterone levels after 0.05-0.3% Pb-acetate treatment in 

30 drinking water (only statistically significant in animals administered 0.05% Pb-acetate). 

31 The results of these recent studies further support the theory that compensatory 

32 mechanisms in the hypothalamic-pituitary-gonadal axis may allow for the adaptation of 

33 exposed animals to the toxic endocrine effects of Pb (Rubio et al., 2006; U.S. EPA, 

34 2006b).

35 Overall, recent epidemiologic and toxicological studies report mixed findings regarding 

36 hormone aberrations in males associated with Pb exposure or Pb biomarker levels. These 

37 results are similar to those from the 2006 Pb AQCD on the effects of Pb exposure on 

38 circulating testosterone levels. 
February 2012 5-450 Draft – Do Not Cite or Quote

5.8.2.3 Fertility

1 Epidemiologic studies have been performed comparing Pb and infertility in men. A study 

2 conducted in Turkey reported that blood and seminal plasma Pb levels were different in 

3 fertile (mean [SD] blood Pb: 23.16 [5.59] μg/dL) and infertile men (mean [SD] blood Pb: 

4 36.82 [12.30] μg/dL)(p <0.001, ANOVA) (Kiziler et al., 2007). The SMART study 

5 examined the success of IVF treatment for women and their partners starting their first 

6 round of treatment (Bloom et al., 2011b; Bloom et al., 2010). A small number of the male 

7 partners participated. Their mean (SD) blood Pb level was 1.50 (0.80) μg/dL. Higher 

8 blood Pb concentration was associated with greater oocyte fertilization (OR 1.08 [95% 

9 CI: 0.97, 1.21] per 1 μg/dL increase in blood Pb when adjusted for Cd), which is not the 

10 expected direction (Bloom et al., 2010). However, higher blood Pb was associated with 

11 lower embryo cell number (a predictor of IVF success) and associated with higher 

12 embryo fragmentation score (an inverse predictor of IVF success) (OR for embryo cell 

13 number: 0.58 [95% CI: 0.37, 0.91]; OR for embryo fragmentation score: 1.47 [95% CI 

14 1.11, 1.94] per 1 μg/dL, controlled for Cd and Hg) (Bloom et al., 2011b). Another study 

15 examined occupational Pb exposure (determined by self-report of occupational exposure) 

16 and detected no difference in reported exposure for infertile versus fertile men (OR 0.95 

17 [95% CI: 0.6, 1.6]) (Gracia et al., 2005). Blood Pb was not measured but approximately 

18 5.0% of infertile men and 5.3% fertile men reported occupational exposure to Pb. As with 

19 the fertility studies among women, a limitation present in these studies is that the cases 

20 included are men who are seeking help at fertility clinics; the study populations are not a 

21 sample of the general population regarding fertility.

22 A couple of recent animal toxicology studies assessed paternal-mediated reproductive 

23 fitness by examining the reproductive success of Pb-exposed males with non-exposed 

24 control females. Anjum et al. (2010) found that adult male rats who were exposed to 273 

25 or 819 mg/L Pb-acetate in drinking water (0.05%or 0.15%, respectively; blood Pb not 

26 reported) spent a significantly longer time copulating than did their control littermates. 

27 The Pb-exposed males were less successful copulators with only 73% of the 0.05% 

28 Pb-acetate exposed males, and 53% of the 0.15% exposed males generating copulatory 

29 plugs in the unexposed female mates. While the number of pregnant females did not 

30 significantly differ from controls, Pb exposed males contributed to the formation of 

31 significantly fewer implantations/dam, and significantly fewer fetuses/dam. Pb-exposed 

32 males were able to sire offspring, but produced fewer offspring per litter. In a group of 

33 males rats with co-exposure to Pb and the herb Centella asiatica, these reproductive 

34 decrements were attenuated relative to rats exposed to Pb alone (Sainath et al., 2011).
February 2012 5-451 Draft – Do Not Cite or Quote

5.8.2.4 Puberty 

1 Research has also been published examining the association between blood Pb and onset 

2 of puberty in males. These epidemiological studies are summarized in Table 5-36. The 

3 majority of studies used concurrent measures of blood Pb and puberty (Den Hond et al., 

4 2011; Tomoum et al., 2010; Hauser et al., 2008), but Williams et al. (2010) performed a 

5 longitudinal analysis of blood Pb levels measured at ages 8-9 years and pubertal onset.
February 2012 5-452 Draft – Do Not Cite or Quote

Table 5-36 Summary of recent epidemiologic studies of associations between 

Pb levels and puberty for males

Referencea Study Location Outcome Study Population Pb Biomarker

Mean Pb (SD) 

in μg/dL Adjusted Effect Estimates

Hauser et al.

(2008)

Chapaevsk, 

Russia

2003-2005

Pubertal stages 

defined using 

standard 

drawings

Healthy boys 

aged 8-9 

Blood Pb Median: 3 (IQR 2-5)

blood Pb >10 μg/dL: 3%

OR (95% CI)

Pubertal onset based on testicular 

volume

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 0.83 (0.43, 1.59)

*after adjustment for macronutrients, 

the OR (95% CI) became 0.66 (0.44, 

1.00)

Genital development

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 0.57 (0.34, 0.95)

*after adjustment for macronutrients, 

the OR (95% CI) became 0.52 (0.31, 

0.88)

Pubic hair development

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 0.74 (0.34, 1.60)

Williams et al.

(2010)

Chapaevsk, 

Russia

2003-2008

Pubertal stages 

defined using 

standard 

drawings

Healthy boys 

aged 8-9 at 

enrollment who 

had annual followup evaluations

Blood Pb at ages 

8-9

Median: 3 (IQR 2-5)

Blood Pb level 

>10 μg/dL: 3%

HR (95% CI)

Pubertal onset based on testicular 

volume

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 0.73 (0.55, 0.97)

Genital development

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 0.76 (0.59, 0.98)

Pubic hair development

<5 μg/dL: 1.00 (Ref)

≥ 5 μg/dL: 0.69 (0.44, 1.07)
February 2012 5-453 Draft – Do Not Cite or Quote

Referencea Study Location Outcome Study Population Pb Biomarker

Mean Pb (SD) 

in μg/dL Adjusted Effect Estimates

Tomoum et al.

(2010)

Cairo, Egypt

2007

Hormones and 

pubertal 

development

Healthy children 

aged 10-13 

seeking treatment 

for minor health 

problems and 

living in one of two 

designated areas 

(one with high-risk 

for Pb 

contamination and 

one with no Pb 

source)

Blood Pb NS for boys only 

(combined with girls in 

the study the mean 

was 9.46 [3.08])

Testicular size

<10 μg/dL: 

Stage 1: 0%

Stage 2: 44.4%

Stage 3: 55.6%

≥ 10 μg/dL:

Stage 1: 33.3%

Stage 2: 66.7%

Stage 3: 0%

Chi-square p-value<0.01

Pubic Hair Development

<10 μg/dL: 

Stage 1: 0%

Stage 2: 55.6%

Stage 3: 44.4%

≥ 10 μg/dL:

Stage 1: 33.3%

Stage 2: 66.7%

Stage 3: 0%

Chi-square p-value<0.05

Penile staging

<10 μg/dL: 

Stage 1: 11.1%

Stage 2: 44.4%

Stage 3: 44.4%

≥ 10 μg/dL:

Stage 1: 58.3%

Stage 2: 41.7%

Stage 3: 0%

Chi-square p-value<0.05

Mean testosterone level

<10 μg/dL: 

4.72 (SD 1.52)

≥ 10 μg/dL:

1.84 (SD 1.04)

*Quantitative results for LH and FSH 

not provided

Den Hond et al.

(2011)

Flanders 

2003-2004

Tanner staging 

and 

gynecomastia

Boys ages 14 and 

15, in their 3rd 

year of secondary 

education and 

living in the same 

study areas for at 

least 5 years

Blood Pb Median: 2.50

10th percentile: 1.20

90th percentile: 5.12 

OR (95% CI) for gynecomastia with 

doubling of exposure

1.84 (1.11, 3.05)

No association between Pb and pubic 

hair or genital development (results not 

given)

a

Studies are presented in order of first appearance in the text of this section.

1 Studies were performed among a cohort of Russian boys enrolled between ages 8-9 years

2 (Williams et al., 2010; Hauser et al., 2008). The area where these studies were performed 

3 had various environmental contaminants such as dioxin, polychlorinated biphenyls, and 

4 other metals, present. Both the cross-sectional study (Hauser et al., 2008) and the 
February 2012 5-454 Draft – Do Not Cite or Quote

1 prospective study with annual follow-ups (Williams et al., 2010) demonstrated an 

2 association; higher blood Pb levels at 8-9 years of age was associated with later onset of 

3 puberty. In a study of boys in Egypt, boys with higher blood Pb had delayed pubertal 

4 development compared to those with lower levels (median age in the high blood Pb group 

5 was 12.5 years compared to 13.0 years in the low blood Pb group) (Tomoum et al., 

6 2010). In addition, compared to the low blood Pb group, those boys with higher blood Pb 

7 had lower testosterone, FSH, and LH levels. A study in Flanders reported no associations 

8 between blood Pb concentration and pubertal development among 14 and 15 year old 

9 boys (Den Hond et al., 2011). However, higher blood Pb levels were associated with an 

10 increased odds of gynecomastia.

11 No recent toxicological studies address Pb-induced male sexual maturation and 

12 development, but older studies do provide support to findings in epidemiologic cohorts. 

13 Pb exposure resulted in delayed sexual maturity as measured by prostate weight in male 

14 Sprague-Dawley pups at PND 35. These pups were exposed chronically to 0.15 or 0.45% 

15 Pb-acetate in dam or their own drinking water from GD5 until PND85 and had blood Pb 

16 ranges from low to high of 88-196 and 120-379 μg/dL, respectively (Ronis et al., 1998b).

17 Cynomolgus monkeys exposed to Pb over a lifetime (10 years, blood Pb levels ranging 

18 from 30–60 μg/dL) had altered pituitary and Sertoli cell function along with decreases in 

19 inhibin/FSH ratio and reduced gonadotropin-releasing hormone (GnRH) stimulation of 

20 LH release in adulthood (Foster et al., 1993; Foster, 1992), all indicators that are 

21 important in proper sexual maturation. Further mechanistic understanding of the effect of 

22 Pb can be gleaned from studies in adult male Wistar rats exposed to Pb for 1 month 

23 (starting at PND56, 0.1 or 0.3% Pb-acetate in drinking water, respective blood Pb levels 

24 of 34 or 60 μg/dL) that showed significant decreases in FSH, ventral prostate weight and 

25 serum testosterone but no change in serum LH (Sokol et al., 1985). These Pb-exposed 

26 adult male rats (0.3% Pb-acetate in drinking water starting at PND56 for 30 days) 

27 demonstrated an impaired pituitary release of LH in response to challenge of the 

28 hypothalamic–pituitary–adrenal (HPA) axis with the opiate antagonist naloxone, an 

29 enhanced release of LH from the pituitary in response to direct stimulation of the 

30 pituitary with luteinizing hormone-releasing hormone (LHRH), an enhanced response to 

31 human chorionic gonadotropin (hCG) by the testes, increased pituitary LH stores, and 

32 increased GnRH mRNA levels in the hypothalamus (Klein et al., 1994; Sokol, 1987).

33 Thus, Pb likely interferes with the male HPA axis, contributing to its reproductive 

34 toxicity.

35 In summary, recent epidemiologic studies have demonstrated an inverse effect of Pb on 

36 pubertal development among boys at low concurrent blood Pb levels. No recent 

37 toxicological studies were found that addressed the effect of Pb on male sexual 

38 development and maturation; however, the 2006 Pb AQCD (U.S. EPA, 2006b) supported 
February 2012 5-455 Draft – Do Not Cite or Quote

1 earlier findings that Pb exposure may result in delayed onset of male puberty and altered 

2 reproductive function later in life in experimental animals.

5.8.2.5 Effects on Morphology and Histology of Male Sex Organs

3 Recent toxicological studies further support historical findings that showed an association 

4 between Pb exposure and changes in the sex organs as well as germ cells. Histological 

5 changes of testes in Pb nitrate-treated animals (a single i.p. dose of 12.5, 25, or 50 mg/kg 

6 of BW and were sacrificed 48 hours later) included seminiferous tubule atrophy, Sertoli 

7 cell and Leydig cell shrinkage with pyknotic nuclei (Shan et al., 2009; Wang et al., 

8 2006a), dilatation of blood capillaries in the interstitium, undulation of basal membrane, 

9 and occurrence of empty spaces in seminiferous epithelium (Massanyi et al., 2007). Pillai 

10 et al. (In Press) found gestational and lactational exposure to Pb-acetate in Charles Foster 

11 rats (subcutaneous injection of 0.05 mg/kg BW/day) induced significant decreases in 

12 absolute organ weight (testes and epididymis) and significant decreases in relative 

13 epididymal weight. Anjum et al. (2010), who exposed 50 day old male albino Wistar/NIN 

14 rats to Pb-acetate (273 or 819 mg/L in drinking water, 0.05% or 0.15%, respectively, 

15 blood Pb levels not reported) for 45 days, reported significant decreases in relative 

16 reproductive organ weight (epididymis, testis, vas deferens, and seminal vesicle) in Pb17 exposed animals. .

5.8.2.6 Summary of Effects on Male Reproductive Function

18 Evidence of associations between Pb exposure and male reproductive function vary by 

19 outcome. The strongest evidence of an association is the relationship observed between 

20 Pb and negative effects on sperm and semen in both recent epidemiologic and 

21 toxicological studies and studies reviewed in previous Pb AQCDs. Many of the 

22 epidemiologic studies included occupational cohorts, which had high blood Pb levels. 

23 Recent toxicological studies also reported an association between Pb exposure and 

24 decreases in reproductive organ weight, organ histological changes in the testes and germ 

25 cells. Male rats exposed to Pb also showed subfecundity in that they produced smaller 

26 litters when mated with unexposed females. In addition, recent epidemiologic studies 

27 found blood Pb levels to be associated with delayed pubertal development in boys with 

28 low concurrent blood Pb levels (mean/median blood Pb levels <10 μg/dL). This is 

29 supported by earlier toxicological studies. Similar to the 2006 Pb AQCD (U.S. EPA, 

30 2006b), recent epidemiologic and toxicological studies reported inconsistent results 

31 regarding hormone aberrations associated with Pb exposure. Mixed findings were also 

32 apparent among epidemiologic studies of fertility among men.
February 2012 5-456 Draft – Do Not Cite or Quote

5.8.3 Effects on Ovaries, Embryo Development, Placental function, and 

Spontaneous Abortions

1 The 2006 Pb AQCD (U.S. EPA, 2006b) included studies of Pb exposure among men and 

2 women and their associations with spontaneous abortions. The 2006 Pb AQCD

3 concluded that overall there was little evidence to support an association between Pb 

4 exposure among women and spontaneous abortion (U.S. EPA, 2006b). Most of the 

5 studies examined in the 2006 Pb AQCD assigned exposure based on living near a smelter 

6 or working in occupations that often result in Pb exposure and the results of these studies 

7 were inconsistent. Little evidence was available in the 2006 Pb AQCD to suggest an 

8 association with paternal Pb levels (U.S. EPA, 2006b), and no recent studies have been 

9 performed to examine paternal Pb levels and spontaneous abortion. Since the 2006 Pb 

10 AQCD, multiple epidemiologic studies have been published that examine Pb levels in 

11 women and their possible association with spontaneous abortion. Additionally, 

12 toxicological studies have studied the effects of Pb on fetal loss and the contribution of 

13 the ovaries and placenta to fetal loss.
February 2012 5-457 Draft – Do Not Cite or Quote

Table 5-37 Summary of recent epidemiologic studies of associations between 

Pb levels and spontaneous abortions

Referencea Study Location Outcome Study population Pb Biomarker

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Yin et al. (2008) Shanxi 

Province, 

China

2004-2006

Anembryonic 

pregnancy

Women age 25-35 yr 

old and at 8-12 

weeks of gestation at 

study entry; cases 

were anembryonic 

pregnancies and 

controls were normal 

pregnancies that 

ended in a live birth 

between 37-42 

weeks 

Maternal blood Pb 

after miscarriage for 

cases and at study 

enrollment for 

controls

Cases: 5.3 (95% CI: 

5.2, 5.9)

Controls: 4.5 (95% 

CI: 3.7, 5.0)

Comparisons between logtransformed blood Pb levels of 

cases and controls performed via 

Student’s t-test had a p-value of 

0.03

Vigeh et al. (2010) Tehran, Iran

2006-2008

Pregnancy 

ended before 20 

weeks of 

gestation

Women who were 

non-smokers, nonobese, had no 

chronic health 

conditions, had their 

last menstrual period 

less than 12 weeks 

prior, and were 

pregnant with a 

singleton infant 

Maternal blood Pb 

during weeks 8-12 of 

pregnancy

3.8 (2.0)

Spontaneous 

abortion: 3.51 (1.42)

Non-spontaneous 

abortion: 3.83 (1.99)

T-test for difference in mean 

values: 0.41

OR: 0.331 (95% CI: 0.011, 

10.096) for an increase in logtransformed blood Pb (units not 

given, assume 1 μg/dL)

Lamadrid-Figueroa et 

al. (2007)

Mexico City, 

Mexico

1997-1999, 

2001-2004

Previous 

miscarriage

Women who had a 

previous pregnancy 

and were currently 

pregnant with 

gestational age of 

≤ 14 wks

Maternal and 

umbilical cord blood 

Pb, maternal bone Pb

Overall: 

Blood Pb: 6.2 (4.5)

Plasma Pb: 0.014 

(0.013) 

Cases: 

Blood Pb: 5.8 (3.4)

Plasma Pb: 0.014 

(0.013)

Controls: 

Blood Pb: 6.5 (4.9)

Plasma Pb: 0.013 

(0.013)

Categorized Plasma Blood Pb 

ratio:

1st tertile: 1.00 (Ref)

2nd tertile: 1.16 (p-value 0.61)

3rd tertile: 1.90 (p-value 0.015)

IRR (95%CI) Per 1 SD increase: 

Plasma Pb 1.12 (p-value 0.22)

Blood Pb 0.93 (p-value 0.56)

Plasma/Blood Pb ratio 1.18 (pvalue 0.02)

Patella Pb 1.15 (p-value 0.39)

Tibia Pb 1.07 (p-value 0.56)

Gundacker et al.

(2010)

Vienna, Austria

2005

Previous 

miscarriage

Women recruited 

during the second 

trimester of 

pregnancy

Whole placentas 

shortly after birth

Median (IQR):

25.8 (21.0, 36.8) 

Median Placenta Pb:

Women who had not previously 

miscarried: 27 μg/kg

Women who had previously 

miscarried: 39 μg/kg

(p-value for difference: 0.039)

a

Studies are presented in order of first appearance in the text of this section.

1 Table 5-37, above, provides a summary of the recent epidemiologic studies examining 

2 the association between Pb biomarker levels and past and current spontaneous abortion. 

3 Yin et al. (2008) performed a study in the Shanxi Province of China to examine if plasma 

4 Pb levels were associated with anembryonic pregnancies (spontaneous abortions during 

5 the first trimester, which account for 15% of all spontaneous abortions). Women were 

6 enrolled at 8-12 weeks of gestation. Women who delivered a term pregnancy had mean 

7 plasma Pb levels that were lower than those of women who had an anembryonic 

8 pregnancy. Of note, among cases plasma Pb level was inversely correlated with folate 

9 and vitamin B12, but this correlation was not observed among those who delivered at 

10 term; no models examining plasma Pb levels adjusted for nutrient status. Another study 
February 2012 5-458 Draft – Do Not Cite or Quote

1 examining spontaneous abortions occurring early in the pregnancy was conducted in Iran

2 (Vigeh et al., 2010). Mean blood Pb concentrations were similar in women who did and 

3 did not have spontaneous abortions. Higher blood Pb levels were not associated with 

4 greater odds of spontaneous abortions before 20 weeks of pregnancy. A study in Turkey 

5 reported on groups of women who either had a spontaneous abortion before the 

6 20th week of gestation or who had a viable pregnancy (Faikoglu et al., 2006). No 

7 difference was detected between the blood Pb levels of the two groups (Pb levels not 

8 reported here due to calculation errors discovered in the paper; errors do not appear to 

9 affect conclusions). A study in Mexico City examined a group of pregnant women 

10 (maximum gestational period at enrollment was 14 weeks) who had previously been 

11 pregnant and either given birth or had a spontaneous abortion (Lamadrid-Figueroa et al., 

12 2007). Women in the highest tertile of plasma/blood Pb ratio had higher rates of previous 

13 spontaneous abortions than did women in the lowest tertile. The authors state that the 

14 plasma/whole blood ratio represents the bioavailability of Pb, which is capable of 

15 crossing the placental barrier for a given blood concentration. No association was 

16 observed when examining the relationship between Pb and spontaneous abortions using 

17 whole blood, plasma, or bone Pb alone. Similarly, a study of placental Pb levels among 

18 pregnant women in Austria observed higher placenta Pb levels among women who had 

19 miscarried a previous pregnancy compared to women who had not miscarried a previous 

20 pregnancy (Gundacker et al., 2010). It is important to note that the number of women 

21 included in the study was small (only 8 women reported previously having a miscarriage) 

22 In toxicological studies, isolated embryo cultures are often used to understand the 

23 mechanisms responsible for aberrant embryo development as it may contribute to 

24 teratogenesis, fetal loss or negative postnatal pup outcomes. Nandi et al. (2010)

25 demonstrated an exposure concentration-dependent decline in embryo development of 

26 fertilized buffalo oocytes cultured in medium containing 0.05-10 μg/mL Pb-acetate as 

27 evidenced by reduced morula/blastocyst yield and increased four-to eight-cell arrest, 

28 embryo degeneration, and asynchronous division. This study provides evidence of the 

29 negative effect of Pb on embryo development and contributes mechanistic understand to 

30 Pb-dependent pregnancy loss. 

31 A possible explanation for reduced fertility and impaired female reproductive success as 

32 a result of Pb exposure is changes in morphology or histology in female sex organs and 

33 the placenta (Dumitrescu et al., 2007; U.S. EPA, 2006b). Wang et al. (2009e) observed 

34 that elevated maternal blood Pb (0.6-1.74 μM, ~12.4-36.0 μg/dL) compared to control 

35 (0.04 μM, ~0.83 μg/dL) were associated with decreased fetal body weight, pup body 

36 length, and placental weight in Wistar rats. The authors reported that placentae from Pb37 exposed groups showed concentration-dependent increasing pathology of 

38 cytoarchitecture and cytoplasmic organelles. The authors also reported a positive 
February 2012 5-459 Draft – Do Not Cite or Quote

1 expression of NF-κB, a transcription factor that controls the expression of genes involved 

2 in immune responses, apoptosis, and cell cycle, in the cytotrophoblasts, decidual cells, 

3 and small vascular endothelial cells in rat placenta under a low-level Pb exposure 

4 condition which correlated with low blood Pb levels. 

5 Pb-exposed (273 mg/L or 819 mg/L in drinking water, 0.05 or 0.15% Pb Acetate, 

6 respectively) male rats from Reshma Anjum et al. (2010) that had an exposure 

7 concentration-dependent decreases in serum testosterone, decreased male reproductive 

8 organ weight and decreased sperm were mated to untreated females. These untreated 

9 dams had male related exposure concentration-dependent decreased implantation rate and 

10 higher pre- and post-implantation loss, indicating paternally mediated fetal loss. The 

11 magnitude of these effects in dams was dependent on the concentration of Pb exposure in 

12 their male mating partners.

13 As observed in sperm cells, Pb stimulates changes in antioxidant enzyme activity in rat 

14 ovaries indicating that oxidative stress may be a contributing factor in Pb-induced 

15 ovarian. Nampoothiri et al. (2007) observed a reduction in SOD activity and an increase 

16 in CAT activity along with a decrease in glutathione content and an increase in lipid 

17 peroxidation in rat granulosa cells after 15 days of Pb treatment (0.05 mg/kg body 

18 weight).

19 Previous studies demonstrated that Pb accumulates in the ovaries and causes histological 

20 changes, thus contributing to Pb-induced effects on female fertility (U.S. EPA, 2006b). In 

21 support of historical studies, recent studies demonstrate Pb-induced histological changes 

22 in ovarian cells of pigs (Kolesarova et al., 2010) and rats (Nampoothiri et al., 2007; 

23 Nampoothiri and Gupta, 2006). Kolesarova et al. (2010) observed a reduction of the 

24 monolayer of granulosa cells after Pb addition (0.5 mg/mL). Nampoothiri and Gupta

25 (2006) reported that Pb exposure caused a decrease in cholesterol and total phospholipid 

26 content in the membranes of granulosa cells which resulted in increased membrane 

27 fluidity. A possible explanation for reduced fertility and impaired female reproductive 

28 success as a result of Pb exposure is changes in morphology or histology in female sex 

29 organs and the placenta (Dumitrescu et al., 2007; U.S. EPA, 2006b).

30 Overall, the recent studies support the conclusions of the 2006 Pb AQCD that there is 

31 insufficient evidence among epidemiologic studies to suggest an association between Pb 

32 and spontaneous abortions. It is important to note that studies of spontaneous abortions 

33 are difficult to conduct. The majority of spontaneous abortions are during the first 

34 trimester, which makes them difficult to capture. Women may miscarry before being 

35 enrolled in a study and many women may not have known they were pregnant when they 

36 miscarried. This limits the ability to detect subtle effects, especially if higher Pb levels do 

37 lead to increased risk of early spontaneous abortions. Toxicological data provide 
February 2012 5-460 Draft – Do Not Cite or Quote

1 mechanistic understanding of the contribution of Pb exposure to spontaneous abortions. 

2 These laboratory data show that Pb exposure impaired placental function, induced 

3 oxidative stress and histological changes in the ovaries, and affected embryo 

4 development. The toxicological and epidemiologic data provide mixed evidence on the 

5 role of Pb in spontaneous abortions. 

5.8.4 Infant Mortality and Embryogenesis

6 The 2006 Pb AQCD (U.S. EPA, 2006b) concluded that Pb exposure can increase fetal 

7 mortality and produce sublethal effects (disrupt growth and development) in offspring of 

8 Pb exposed dams at concentrations that do not result in clinical toxicity to the dams by 

9 disrupting implantation and pregnancy, particularly at the blastocyst stage of 

10 development. In rodent studies gestational exposure to Pb (blood Pb 32 to >70 μg/dL) 

11 resulted in smaller litters and fewer implantation sites and in non-human primates pre12 and perinatal mortality was reported in squirrel monkeys exposed to Pb (mean dam blood 

13 Pb of 54 μg/dL) in the last two-thirds of gestation (U.S. EPA, 2006b). There is substantial 

14 evidence to show that there is no apparent maternal-fetal barrier to Pb and it can easily 

15 cross the placenta and accumulate in fetal tissue during gestation (Pillai et al., 2009; 

16 Wang et al., 2009e; Uzbekov et al., 2007). No recent studies have reported on the 

17 relationship between Pb levels and infant mortality. 

5.8.5 Birth Defects

18 The 2006 Pb AQCD (U.S. EPA, 2006b) reported the possibility of small associations 

19 between high Pb exposure and birth defects, but many of the epidemiologic studies used 

20 occupational histories instead of actual measures of blood Pb levels. Among the studies 

21 included in the 2006 Pb AQCD, a couple studies reported possible associations between 

22 parental exposure to Pb and neural tube defects (Irgens et al., 1998; Bound et al., 1997).

23 Recent studies also examined indicators of Pb exposure and neural tube defects (Table 

24 5-38). No other recent epidemiologic studies of Pb exposure and birth defects were 

25 identified in the literature. No recent toxicological studies were found that investigated 

26 Pb-induced changes in morphology, teratology effects, or skeletal malformations of 

27 developing fetuses as a result of maternal Pb exposure; however, in the 2006 Pb AQCD 

28 toxicological studies demonstrated associations between exposure to high doses of Pb 

29 and increased incidences of teratogenic effect in experimental animals. 
February 2012 5-461 Draft – Do Not Cite or Quote

Table 5-38 Summary of recent epidemiologic studies of associations between 

Pb levels and neural tube defects

Referencea Study Location Study Population

Pb Biomarkers and 

Exposure Measurement Mean Pb (SD) in μg/dL Adjusted Effect Estimates

Zeyrek et al. (2009) Turkey

NS

Infants with 

gestational age of at 

least 20 wks

Maternal and umbilical cord 

blood Pb taken 0.5h after 

birth

Cases: Maternal: 15.5 (15.0)

Umbilical cord: 18.2 (17.8)

Controls: Maternal: 12.5 

(12.7)

Umbilical cord: 16.5 (16.1)

P-values for differences of 

Student’s t-test or Mann-Whitney U 

test (dependent on distribution) 

were 0.35 for maternal blood Pb 

and 0.63 for umbilical cord blood 

Pb

Brender et al. (2006) Texas

1995-2000

Infants of MexicanAmerican women Maternal blood Pb 

taken 5-6 wks post-partum

Cases: 2.4 (1.9) 

Controls: 2.5 (1.6)

Blood Pb<6.0 μg/dL: 1.0 (Ref)

Blood Pb≥ 6.0 μg/dL: 1.5 (95% CI: 

0.6, 4.3)

Huang et al. (2011b) China

2002-2004

Live and still births of 

women living in the 

study area (villages in 

the Lvliang region of 

Shanxi province)

2 soil samples from each 

village

56.14 μg/g (11.43 μg/g) N/A

a

Studies are presented in order of first appearance in the text of this section.

1 Among the recent epidemiologic studies (described in Table 5-38), a study of women in 

2 Turkey detected no difference between the blood Pb of mothers or the umbilical cord 

3 blood Pb of the newborns for healthy infants compared with infants with neural tube 

4 defects (cases of spina bifida occulta were excluded, but other forms of spina bifida were 

5 included) (Zeyrek et al., 2009). Brender et al. (2006) performed a study of Mexican6 American women living in Texas. Measurements were taken 5-6 weeks postpartum, 

7 which is a limitation of this study because the blood Pb levels may be different from 

8 those during the developmental period of gestation. The OR comparing women with at 

9 least 6 μg/dL blood Pb to those with less than 6 μg/dL blood Pb was 1.5 (95% CI: 0.6, 

10 4.3). This increased after adjusting for breast feeding, although this variable was not a 

11 confounder because it cannot be associated with neural tube defects. For these women, 

12 neither occupational exposure to Pb nor proximity of residence to a facility with Pb air 

13 emissions at the time of conception was associated with increased odds of neural tube 

14 defects. A study with an ecologic design was performed in China and did not use 

15 individual-level biomarkers to determine Pb levels (Huang et al., 2011b). A positive 

16 association between Pb levels in soil samples and neural tube defects was reported 

17 Exposure to multiple other trace elements also demonstrated a positive association but no 

18 control for co-exposures was included in the models for Pb.

19 Previous studies included in the 2006 Pb AQCD observed associations between Pb and 

20 neural tube defects but were limited due to the lack of biologically measured Pb [Pb was 

21 measured in drinking water (Bound et al., 1997) and estimated from occupational reports

22 (Irgens et al., 1998)]. A recent ecologic study reported an association between Pb in the 

23 soil and neural tube defects but was also limited by its lack of biological samples, as well 

24 as a lack of individual-level data and the prevalence of several other metals (Huang et al., 
February 2012 5-462 Draft – Do Not Cite or Quote

1 2011b). Other recent epidemiologic studies of maternal blood Pb levels and neural tube 

2 defects observed no associations (Zeyrek et al., 2009; Brender et al., 2006).

5.8.6 Preterm Birth

3 Epidemiologic studies on preterm birth included in the 2006 Pb AQCD (U.S. EPA, 

4 2006b) reported inconsistent findings regarding the relationship between Pb and 

5 gestational age. Recent studies have examined this potential association and again mixed 

6 results were reported (Table 5-39). Of these studies, the ones that categorized births as 

7 preterm or term all defined preterm birth as less than 37 weeks of gestation. One 

8 limitation to note for these studies is that if Pb affects spontaneous abortion and length of 

9 gestation via a similar pathway, then the studies that only collect data at delivery and not 

10 at earlier stages of pregnancy would be biased toward the null. 
February 2012 5-463 Draft – Do Not Cite or Quote

Table 5-39 Summary of recent epidemiologic studies of associations between 

Pb levels and preterm birth

Referencea Study Location Outcome Study Population

Pb Biomarkers or 

Exposure Measurement

Mean Pb (SD) 

in μg/dL Adjusted Effect estimates

Berkowitz et al.

(2006)

Idaho

1970-1981

Preterm birth 

(<37wk)

Singleton births 

with 28-45 wk 

gestation

Three time periods of two 

locations (unexposed and 

exposed/near smelter): pre- fire, “high-exposure period” 

(when a fire happened at 

the smelter and resulted in 

damages leading to high air 

Pb concentrations for 6 

mo), and “post-fire”

NS OR (90% CI) (unexposed location is 

referent group):

Pre-fire 0.93 (0.67, 1.28) 

High exposure 0.68 (0.34, 1.35)

Post-fire 1.17 (0.95, 1.45)

Patel and Prabhu

(2009)

Nagpur, 

India

NS

Gestational age Consecutive births at 

the study hospital

Umbilical cord blood Pb Umbilical cord 

blood Pb: 4.7 

(12.1)

>5 μg/dL: mean gestational age 38 

wks

≤ 5 μg/dL: mean gestational age 39 

wks

Linear regression: gestational age 

decreased 1 wk with every 1 μg/dL 

increase in umbilical cord blood Pb 

(exact values and 95% CI not given)

Jones et al. (2010) Tennessee

2006

Gestational Age: 

preterm (<37wk) 

, term (37-40 

wk), post-term 

(>40 wk)

Singleton births ≥ 27 

wk gestation from 

mothers aged 16-45 

living in the Shelby 

County area for at 

least 5 mo during 

pregnancy 

Umbilical cord blood Pb 2.4 (4.3)

Geometric 

mean: 1.3

Geometric Mean:

Preterm birth: 1.4

Term birth: 1.2 

Post-term birth: 1.3

p-value for difference: >0.10

Wells et al. (2011a) Baltimore, 

MD

2004-2005

Gestational age Singleton births from 

the Baltimore 

Tracking Health 

Related to 

Environmental 

Exposures (THREE) 

study

Umbilical cord Pb 0.84 (95%: CI 

0.72, 0.96)

≥ 5 μg/dL: 0.7%

Ratio for Pb concentration per 10 

days of gestation: 0.99 (0.93, 1.06)

Jelliffe-Pawlowski 

et al. (2006)

California

1995-2002

Preterm birth 

(<37 completed 

wk)

Singleton births to 

non-smoking 

mothers with blood 

Pb measures during 

pregnancy from 

either the California 

Childhood Lead 

Poisoning 

Prevention Branch 

or the California 

Occupational Lead 

Poisoning 

Prevention Program 

Maximum maternal blood 

Pb during pregnancy

≥ 10 μg/dL: 

30.9%

Odd Ratios:

≤ 5 μg/dL: 1.00 (Ref)

6-9 μg/dL: 0.8 (0.1, 6.4)

10-19 μg/dL: 1.1 (0.2, 5.2)

20-39 μg/dL: 4.5 (1.8, 10.9)

≥ 40 μg/dL: 4.7 (1.1, 19.9)

<10 μg/dL: 1.00 (Ref)

≥ 10 μg/dL: 3.2 (1.2, 7.4)

Vigeh et al. (2011) Tehran, Iran

2006

Preterm birth 

(20-37 wk)

Singleton births from 

non-smoking, nonobese mothers aged 

16-35 and referred 

for prenatal care 

during the 8th-12th 

week of gestation 

Maternal blood Pb 3.8 (2.0) Mean blood Pb (SD):

Preterm birth: 4.52 (1.63)

Term birth: 3.72 (2.03) 

p-value for difference: <0.05

OR (95% CI)

1.41 (1.08, 1.84)

(unit not given, assume per 1 μg/dL)
February 2012 5-464 Draft – Do Not Cite or Quote

Referencea Study Location Outcome Study Population

Pb Biomarkers or 

Exposure Measurement

Mean Pb (SD) 

in μg/dL Adjusted Effect estimates

Cantonwine et al.

(2010a)

Mexico City 

1997-1999

Preterm birth 

(<37wk), 

Gestational age

Births to mothers 

with at least 1 blood 

Pb measurement 

during pregnancy 

and no chronic 

diseases requiring 

medication

Maternal blood Pb during 

pregnancy

Blood Pb

Visit at <20wks 

pregnant 7.2 

(5.2)

Visit at 20-28 

weeks pregnant 

6.3 (4.3)

Visit at >28 

weeks pregnant 

6.8 (4.5)

Plasma Pb

Visit at <20wks 

pregnant 0.17 

(0.16)

Visit at 20-28 

weeks pregnant 

0.13 (0.10)

Visit at >28 

weeks pregnant 

0.16 (0.26)

Linear regression β (95% CI)

Blood Pb

Visit at <20wks -2.76 (-5.21, -0.31)

Visit at 20-28 weeks -1.77 (-3.39, -

0.15)

Visit at >28 weeks -0.47 (-1.78, 

0.84)

Average -1.49 (-3.63, 0.64)

Plasma Pb

Visit at <20wks -2.38 (-4.97, 0.21)

Visit at 20-28 weeks -1.34 (-2.98, 

0.29)

Visit at >28 weeks -1.28 (-2.63, 

0.06)

Average -0.28 (-2.81, 2.25)

Plasma-to-blood Pb ratio

Visit at <20wks -3.23 (-6.01, -0.44)

Visit at 20-28 weeks -1.41 (-3.10, 

0.29)

Visit at >28 weeks -1.30 (-2.67, 

0.07)

Average -1.27 (-3.89, 1.35)

Cord blood Pb 

-0.68 (-2.37, 1.00) 

Zhu et al. (2010) New York

2003-2005

Preterm birth 

(<37 completed 

wk)

Singleton births to 

mothers aged 15-49 

with blood Pb 

measures before or 

on the date of 

delivery and blood 

Pb measuring <10 

μg/dL

Maternal blood Pb 2.1 Odd Ratios:

≤ 1.0 μg/dL: 1.00 (Ref)

1.1-2.0 μg/dL: 1.03 (0.93, 1.13)

2.1-3.0 μg/dL: 1.01 (0.92, 1.10)

3.1-9.9 μg/dL: 1.04 (0.89, 1.22)

Chen et al. (2006a) Taiwan

1993-1997

Preterm birth 

(<37 wk)

Infants born to at 

least one parent who 

was part of the 

Program to Reduce 

Exposure by 

Surveillance System 

– Blood Lead Levels 

cohort that 

monitored workers 

occupationally 

exposed to Pb

Maternal blood Pb during 

pregnancy (or if that wasn’t 

available, the 1 year prior to 

fertilization) and/or paternal 

blood Pb during 

spermatogenesis (the 64 

days before fertilization, or if 

that wasn’t available, the 1 

year prior to 

spermatogenesis)

Maternal blood 

Pb 10.1 (10.4)

Paternal blood 

Pb 12.9 (13.8)

Risk Ratios

Maternal blood Pb

<10 μg/dL: 1.00 

10- 19 μg/dL: 1.97 (0.92, 3.86)

≥ 20 μg/dL: 1.86 (0.68, 4.28)

Paternal blood Pb

<10 μg/dL: 1.00 

10- 19 μg/dL: 1.17 (0.53, 2.32)

≥ 20 μg/dL: 0.55 (0.19, 1.28)

Orun et al. (2011) Turkey

NS

Preterm birth 

(<37 wk)

Births to mothers not 

occupationally 

exposed to toxic 

metals and living in a 

suburban but non- industrial area

Breast milk 2 months postpartumMedian: 20.6 

μg/L

>WHO limit (5 

μg/L): 87%

Median Pb (IQR)

>37 wk:20.6 (11.2, 29.2) μg/L

≤ 37 wk: 20.4 (14.4, 27.9) μg/L

p-value for Mann-Whitney U test: 

≥ 0.05

a

Studies are presented in order of first appearance in the text of this section.

1 A study of preterm birth included women living in two different residential areas over 

2 three different time periods (Berkowitz et al., 2006). One residential area had consistently 

3 lower exposures but the other had a period of high Pb emissions due to damage at a local 
February 2012 5-465 Draft – Do Not Cite or Quote

1 factory. Preterm birth rates were examined during three time periods: before, during, and 

2 after the time of higher Pb exposure. No association was observed between women living 

3 in the high exposure area compared to those in the low exposure area during any of the 

4 exposure time periods, but the number of preterm infants born during the period of higher 

5 exposure was small. 

6 In another study, measurements of umbilical cord blood were taken after birth at a 

7 hospital in Nagpur, India (Patel and Prabhu, 2009). A sample of women had their blood 

8 Pb measured and among this sample, maternal blood Pb was correlated with the umbilical 

9 cord Pb levels. Mean gestational age differed between infants with >5 μg/dL cord blood 

10 Pb and infants with ≤ 5 μg/dL cord blood Pb. In a linear regression model, gestational 

11 age was found to decrease with increasing umbilical cord Pb levels. A study of women in 

12 Tennessee consisted primarily of African American women living in an urban setting

13 (Jones et al., 2010). The mean level of umbilical cord blood Pb was slightly higher 

14 among infants born preterm but the difference was not statistically significant. Using 

15 umbilical cord blood Pb measures, a study reported no association between cord blood Pb 

16 levels and gestational age. The concentrations of cord blood Pb among study participants 

17 were overall low (99.3% had umbilical cord blood Pb ≤ 5 μg/dL) (Wells et al., 2011a).

18 In a study taking place in California, women with information on blood Pb levels during 

19 pregnancy based on their participation in a surveillance program (reason for participation 

20 in the surveillance program was unknown but the authors speculate it was likely because 

21 of potential Pb exposure) were matched with the birth certificates of their infants

22 (Jelliffe-Pawlowski et al., 2006). Almost 70% of women had maximum blood Pb 

23 measurements <10 μg/dL with the majority being <5 μg/dL. Preterm birth was associated 

24 with higher blood Pb when comparing women with maximum blood Pb levels 

25 ≥ 10 μg/dL to women with blood Pb levels <10 μg/dL in adjusted analyses. In analyses 

26 of maximum Pb levels further refined into additional categories, the odds of preterm birth 

27 were elevated among women with maximum blood Pb measurement ≥ 20 μg/dL 

28 compared with women with maximum blood Pb levels ≤ 5 μg/dL. A study in Iran also 

29 reported higher maternal blood Pb for preterm births than for term births (Vigeh et al., 

30 2011). The women in this study had lower blood Pb levels than did those observed in the 

31 Jelliffe-Pawlowski et al. study (Jelliffe-Pawlowski et al., 2006). Higher maternal blood 

32 Pb level was associated with higher odds of preterm birth. Another study examining 

33 blood Pb and gestational age among women with lower blood Pb levels reported an 

34 inverse association between maternal blood Pb concentration and gestational age, 

35 especially for blood Pb levels early in pregnancy (Cantonwine et al., 2010a). However, a 

36 study conducted in New York among women with lower blood Pb levels (inclusion 

37 criteria mandated that blood Pb concentration be less than 10 μg/dL), no association was 

38 observed between blood Pb levels and preterm birth (Zhu et al., 2010). Similarly, a study 
February 2012 5-466 Draft – Do Not Cite or Quote

1 of maternal and paternal blood Pb concentrations reported no association between 

2 maternal or paternal blood Pb levels and preterm birth (Chen et al., 2006a).

3 A study of breast milk in the second month postpartum reported no difference in breast 

4 milk Pb levels for those infants born preterm or term; however, a limitation of this study 

5 is that Pb levels were not measured until two months after the birth (Örün et al., 2011).

6 In sum, as in the 2006 Pb AQCD, recent epidemiologic studies report inconsistent 

7 findings for a relationship between indicators of Pb exposure and preterm birth. No 

8 patterns were apparent within type of exposure measurement or Pb level. 

5.8.7 Low Birth Weight/Fetal Growth

9 The 2006 Pb AQCD reported inconsistent epidemiologic study results for the 

10 associations between Pb and birth weight/fetal growth and concluded that there could be 

11 a small effect of Pb exposure on birth weight and fetal growth (U.S. EPA, 2006b). Since 

12 then, multiple epidemiologic studies on the relationship between Pb exposure and birth 

13 weight and fetal growth have been published using various measures of exposure, such as 

14 air levels, umbilical cord blood, and maternal blood and bone. These studies are 

15 summarized in Table 5-40 below. Additionally, there have been a few recent 

16 toxicological studies evaluating the effect of Pb exposure during gestation on birth 

17 weight.
February 2012 5-467 Draft – Do Not Cite or Quote

Table 5-40 Summary of recent epidemiologic studies of associations between 

Pb levels and low birth weight and fetal growth

Referencea Study Location Outcome Study population

Pb Biomarkers and 

Exposure Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Lamb et al. (2008) Mitrovica 

and 

Pristina, 

Yugoslavia

1985-1986

Height and BMI at birth Participants of the 

Yugoslavia Study 

of Environmental 

Lead Exposure, 

Pregnancy 

Outcomes, and 

Childhood 

Development

Mid-pregnancy blood Pb Mitrovica: 20.56 

(7.38) 

Pristina: 5.60 

(1.99)

Regression Coefficients (95% 

CI) for 1 μg/dL increase in Pb:

BMI

Mitrovica: -0.18 (-0.69, 0.33)

Pristina: -0.14 (-0.69, 0.42)

Height

Mitrovica: 0.43 (-0.83, 1.69)

Pristina: 0.35 (-0.64, 1.34)

Jelliffe-Pawlowski et 

al. (2006)

California

1995-2002

Low birth weight 

(<2,500g)

Small for gestational 

age (birth weight for 

gestational age <10th 

percentile of race- and 

gender- specific norms

Singleton births to 

non-smoking 

mothers with blood 

Pb measures 

during pregnancy 

from either the 

California 

Childhood Lead 

Poisoning 

Prevention Branch 

or the California 

Occupational Lead 

Poisoning 

Prevention 

Program and 

matched to birth 

records

Maximum maternal blood 

Pb during pregnancy

≥ 10 μg/dL: 

30.9%

Odd Ratios:

Low birth weight ≤ 5 μg/dL: 

1.00 (Ref)

6-9 μg/dL: --

10-19 μg/dL: 2.7 (0.5, 14.8)

20-39 μg/dL: 1.5 (0.3, 7.7)

≥ 40 μg/dL: --

<10 μg/dL: 1.00 (Ref)

≥ 10 μg/dL: 3.6 (0.3, 40.0)

Small for gestational age

≤ 5 μg/dL: 1.00 (Ref)

6-9 μg/dL: --

10-19 μg/dL: 2.3 (0.6, 9.2)

20-39 μg/dL: 2.1 (0.7, 6.7)

≥ 40 μg/dL: --

<10 μg/dL: 1.00 (Ref)

≥ 10 μg/dL: 4.2 (1.3, 13.9)

Iranpour et al.

(2007)

Isfahan, 

Iran

2005

Low birth weight 

(≤ 2,500g, >37wk)

Full-term infants 

born at a hospital 

affiliated with 

Isfahan University

Umbilical cord and 

maternal blood Pb within 

12 h of delivery

Maternal blood 

Pb:

Cases: 12.5 (2.0)

Controls: 13.5 

(2.7)

Umbilical cord 

blood Pb:

Cases: 10.7 (1.7)

Controls: 11.3 

(1.9)

P-values for t-tests:

Maternal blood Pb 0.07

Umbilical cord blood Pb:

0.20

P-values for correlations: 

Maternal blood Pb and Birth 

weight:

Low birth weight: 0.17

Normal birth weight: 0.3

P-values for correlations: 

Umbilical cord blood Pb and 

birth weight:

Low birth weight: 0.84

normal birth weight: 0.26
February 2012 5-468 Draft – Do Not Cite or Quote

Referencea Study 

Location Outcome Study population

Pb Biomarkers and 

Exposure Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Gundacker et al.

(2010)

Vienna, 

Austria

2005

Birth length, birth 

weight, head 

circumference

Infants of women recruited during 

their second 

trimester

Maternal blood Pb between 

wk 34-38 of gestation, 

whole placentas and 

umbilical cord Pb shortly 

after birth, meconium 

samples in first five days 

after birth

Median (IQR):

Maternal blood 

Pb: 2.5 (1.8, 3.5)

Umbilical cord 

blood Pb: 1.3 

(0.8, 2.4)

Placenta Pb: 25.8 

μg/kg (21.0, 36.8 

μg/kg) 

Meconium Pb: 

15.5 μg/kg (9.8, 

27.9 μg/kg) 

Regression coefficients (units 

not given, assume results are 

per 10 μg/dL or 1 μg/kg)

Birth length: 

Placenta Pb: 0.599 (SE 

0.154, p-value <0.001)

Meconium Pb: -0.385 (SE 

0.157, p-value 0.012)

Birth weight: 

Placenta Pb: 0.658 (SE 

0.136, p-value <0.001)

Maternal blood Pb: -0.262 

(SE 0.131, p-value 0.058)

Zhu et al. (2010) New York

2003-2005

Birth weight, small for 

gestational age (birth 

weight for gestational 

age <10th percentile 

based on national birth 

weight by gestational 

week from weeks 25-42

Singleton births to 

mothers aged 15- 49 with blood Pb 

measures before 

or on the date of 

delivery and blood 

Pb measuring <10 

μg/dL

Maternal blood Pb 2.1 Difference in birthweight in 

grams:

0 μg/dL: Ref

1 μg/dL: -27.4 (-37.8, -17.1)

2 μg/dL: -38.8 (-53.4, -24.1)

3 μg/dL: -47.5 (-65.4, -29.6)

4 μg/dL: -54.8 (-75.5, -34.2)

5 μg/dL: -61.3 (-84.4, -38.2)

6 μg/dL: -67.2 (-92.5, -41.8)

7 μg/dL: -72.5 (-99.9, -45.2)

8 μg/dL: -77.6 (-106.8, -48.3)

9 μg/dL: -82.3 (-113.3, -51.2)

10 μg/dL: -86.7 (-119.4, -54.0)

After exclusion of blood Pb <1 

μg/dL, a 1 μg/dL increase in 

blood Pb was associated with 

a 7.0 g decrease in 

birthweight

Odd Ratios for small for 

gestational age:

≤ 1.0 μg/dL: 1.00 (Ref)

1.1-2.0 μg/dL: 1.07 (0.98, 

1.17)

2.1-3.0 μg/dL: 1.06 (0.98, 

1.16)

3.1-9.9 μg/dL: 1.07 (0.93, 

1.23)
February 2012 5-469 Draft – Do Not Cite or Quote

Referencea Study 

Location Outcome Study population

Pb Biomarkers and 

Exposure Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Chen et al. (2006a) Taiwan

1993-1997

Low birth weight 

(<2,500 g), small for 

gestational age (birth 

weight ≤ 10th percentile 

of sex- and gestational 

wk weights for 

singletons in 1993- 1996)

Infants born to at 

least one parent 

who was part of 

the Program to 

Reduce Exposure 

by Surveillance 

System – Blood 

Lead Levels cohort 

that monitored 

workers 

occupationally 

exposed to Pb

Maternal blood Pb during 

pregnancy (or if that wasn’t 

available, the 1 year prior 

to fertilization) and/or 

paternal blood Pb during 

spermatogenesis (the 64 

days before fertilization, or 

if that wasn’t available, the 

1 year prior to 

spermatogenesis)

Maternal blood 

Pb 10.1 (10.4)

Paternal blood Pb 

12.9 (13.8)

Risk Ratios

Low birth weight

Maternal blood Pb

<10 μg/dL: 1.00 (Ref)

10- 19 μg/dL: 2.22 (1.06, 

4.26)

≥ 20 μg/dL: 1.83 (0.67, 4.20)

Paternal blood Pb

<10 μg/dL: 1.00 (Ref)

10- 19 μg/dL: 0.83 (0.34, 

1.75)

≥ 20μg/dL: 0.42 (0.12, 1.06)

SGA

Maternal blood Pb

<10 μg/dL: 1.00 (Ref)

10- 19 μg/dL: 1.62 (0.91, 

2.75)

≥ 20μg/dL: 2.15 (1.15, 3.83)

Paternal blood Pb

<10 μg/dL: 1.00 (Ref)

10- 19 μg/dL: 0.94 (0.49, 

1.66)

≥ 20 μg/dL: 0.94 (0.51, 1.62)

Kordas et al. (2009) Mexico City, 

Mexico

1994-1995

Head circumference, 

birth weight, birth length

Infants of mothers 

receiving antenatal 

care at hospitals 

serving low-tomiddle income 

populations (cross- sectional study of 

baseline info from 

Ca 

supplementation 

trial)

Umbilical cord and 

maternal blood Pb within 

12 h of delivery; maternal 

tibia Pb

Maternal tibia Pb: 

9.9 μg/g (9.8 

μg/g)

Maternal blood 

Pb ≥ 10μg/dL: 

27%

Umbilical cord 

blood Pb ≥ 

10μg/dL: 13.7%

Regression coefficients (SE) 

(adjusted for maternal BMI, 

maternal height, infant 

gestational age, and other 

variables) for each 1 μg/g 

increase in tibia Pb:

Birth weight: -4.9 (1.8)

Birth length: -0.02 (0.01)

Head circumference: -0.01 

(0.01; p-value<0.05)

Women with 4th quartile tibia 

Pb (15.6-76.5 μg/g) delivered 

infants 140 g less than 

women with tibia Pb in the 

lowest quartile

Afeiche et al. (2011) Mexico City

1994-2005

Birth weight Term, singleton 

births, at least 

2,500 grams 

enrolled in one of 

three birth cohorts 

recruited for other 

longitudinal studies

Maternal patella and tibia 

Pb measured at 1 month 

postpartum

Patella Pb 10.4 

(11.8) μg/g

Tibia Pb 8.7 (9.7) 

μg/g

β (95% CI) for 1 SD increase 

in maternal patella Pb

Girls

-45.7 (-131.7, 40.2)

Boys

72.3 (-9.8, 154.4)

No association for birth 

weight and tibia Pb among 

girls. A positive association 

was observed for tibia Pb and 

birth weight among boys. 

(results not given) 
February 2012 5-470 Draft – Do Not Cite or Quote

Referencea Study 

Location Outcome Study population

Pb Biomarkers and 

Exposure Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Cantonwine et al.

(2010b)

Mexico City

1994-1995

Birthweight Infants who were 

part of a clinical 

trial to assess 

maternal calcium 

supplementation 

on bone lead 

mobilization during 

lactation

Umbilical cord blood Pb

Maternal tibia and patella 

Pb one month after delivery

Umbilical cord 

blood Pb varied 

by genotype from 

6.3 to 6.9

Umbilical cord 

blood Pb ≥ 10 

μg/dL: 12.6%

Regression models

β (95% CI)

Umbilical cord blood Pb -31.1 

(-105.4, 43.3)

Maternal tibia Pb 

Overall -4.4 (-7.9, -0.9)

<1-4.1 μg/g: Ref

4.1-9.2 μg/g: 17.2 (-75.6, 

110.1)

9.2-15.4 μg/g: -19.1 (-112.1, 

73.9)

15.4-43.2 μg/g: -95.4 (-189.9, 

-0.8)

Llanos and Ronco

(2009)

Santiago, 

Chile

NS

Fetal growth restriction 

(1,000-2,500g) 

*note normal birth 

weights were >3,000g

Term births (37-40 

wks) from nonsmoking mothers

Placenta Pb Fetal growth 

restricted: 0.21 

μg/g (0.04 μg/g) 

Controls: 0.04 

μg/g (0.009 μg/g) 

P-value for Mann-Whitney 

U-test <0.01

Zentner et al. (2006) Santo 

Amaro, 

Brazil 

2002

Birth weight and length Singleton births 

with maternal 

residence within 

5 km of Pb smelter

Umbilical cord blood Pb 

from delivery

Umbilical cord 

blood Pb: 3.9 

(3.6)

Linear regression coefficient 

with umbilical cord blood Pb 

as the dependent variable in 

model with only length and 

weight (unit not given, 

assume per 1 μg/dL): Length 

-0.46 (p-value 0.003) and 

Weight -0.275 (0.048) (i.e., in 

this study, Pb is assessed as 

the outcome)

Atabek et al. (2007) Turkey

NS

Birth weight, birth 

length, head 

circumference, mid-arm 

circumference

Term, singleton 

infants born to 

healthy mothers 

living in urban 

areas and 

assumed to have 

high Pb 

concentrations

Umbilical cord blood Pb 14.4 (8.9)

Umbilical cord 

blood Pb ≥ 10 

μg/dL: 53.7%

Umbilical cord 

blood Pb ≥ 25 

μg/dL: 9.2%

Regression models

β (p-value)

Birth weight 

-0.81 (0.01)

Birth length

0.41 (0.05)

Mid-arm circumference

0.30 (0.05)

Al-Saleh et al.

(2008b)

Saudi 

Arabia

2004

Head circumference Infants with a 

gestational age of 

at least 34 weeks 

born to healthy 

mothers aged 17-

46 years and non- occupationally 

exposed to Pb 

Umbilical cord blood Pb 2.210 (1.691)

Umbilical cord 

blood Pb >10 

μg/dL: 1.23%

Regression models for those 

above the 75th percentile of 

cord blood Pb levels

β (SE) per unit of logtransformed Pb

-0.158 (0.718), p-value: 0.036

Janjua et al. (2009) Karachi, 

Pakistan

2005

Low birth weight 

(≤ 2,500g)

Infants of randomly 

selected women 

who planned to 

deliver between 

37-42 wk

Umbilical cord blood Pb Umbilical cord 

blood Pb: 10.8 

(0.2)

Prevalence ratio: 

<10 μg/dL: 1.00 (Ref)

≥ 10 μg/dL: 0.82 (0.57, 1.17)

Jones et al. (2010) Tennessee

2006

Low birth weight 

(<2.500g)

Singleton births 

≥ 27 wks gestation 

from mothers aged 

16-45 living in the 

Shelby County 

area for at least 5 

mo during 

pregnancy 

Umbilical cord blood Pb 2.4 (4.3)

Geometric mean: 

1.3

Geometric Mean:

Low birth weight: 1.2

Normal birth weight: 1.3

p-value for difference: >0.10
February 2012 5-471 Draft – Do Not Cite or Quote

Referencea Study 

Location Outcome Study population

Pb Biomarkers and 

Exposure Measurement

Mean Pb (SD) in 

μg/dL Adjusted Effect Estimates

Wells et al. (2011a) Baltimore, 

MD

2004-2005

Birth weight Singleton births 

from the Baltimore 

Tracking Health 

Related to 

Environmental 

Exposures 

(THREE) study

Umbilical cord Pb 0.84 (95%: CI 

0.72, 0.96)

≥ 5 μg/dL: 0.7%

Ratio for Pb concentration per 

100g birth weight: 1.01 (0.99, 

1.02)

Örün et al. (2011) Turkey

NS

Birth weight and head 

circumference Births to mothers not occupationally 

exposed to toxic 

metals and living in 

a suburban but 

non-industrial area

Breast milk 2 months post- partumMedian: 20.6 μg/L

>WHO limit (5 

μg/L): 87%

Median (IQR)

<2500g: 20.4 

(8.5, 27.1) μg/L

≥ 2500g: 20.6 

(11.8, 29.5) μg/L

Correlations for breast milk 

Pb and z-scores of head 

circumference

Girls 0.087

Boys 0.029

Correlations for breast milk 

Pb and z-scores of birth 

weight 

Girls 0.097

Boys 0.045

*All p-values for 

correlations>0.05

Williams et al.

(2007)

Tennessee

2002

Birth weight Infants from 

singleton births or 

the firstborn infant 

in a set of multiples

Air Pb levels during first 

trimester of pregnancy

0.12 μg/m3 (0.04 

μg/m3)

p-value for multilevel 

regression of Pb with birth 

weight: 0.002

Increase of Pb from 0 to 0.04 

relates to a 38g decrease in 

birth weight

Increase of Pb from 0 to 0.13 

(maximum) relates to a 124g 

decrease in birth weight

Berkowitz et al.

(2006)

Idaho

1970-1981

Low birth weight 

(<2,500 g and ≥ 37 wk)

Small for gestational 

age (birth weight ≤ 5th 

percentile of sex- and 

gestational wk weights 

for singletons in Idaho)

Singleton infants 

with 28-45 wk 

gestation

Three time periods of two 

locations (unexposed and 

exposed/near smelter): prefire, “high-exposure period” 

(when a fire happened at 

the smelter and resulted in 

damages leading to high air 

Pb concentrations for 6 

mo), and “post-fire”

Not specified Term Low birth weight:

OR (90% CI) (unexposed 

location is referent group):

Pre-fire 0.81 (0.55, 1.20) 

High exposure 2.39 (1.57, 

3.64)

Post-fire 1.28 (0.95, 1.74)

Small for gestational age:

OR (90% CI) (unexposed 

location is referent group):

Pre-fire 0.98 (0.73, 1.32) 

High exposure 1.92 (1.33, 

2.76)

Post-fire 1.32 (1.05, 1.67)

a

Studies are presented in order of first appearance in the text of this section.

1 Women residing in two different towns in Yugoslavia (one with a Pb smelter and one 

2 without a Pb smelter) were recruited during their first prenatal visit (Lamb et al., 2008)

3 (study based on previous work by Factor-Litvak et al. (1991). The mid-pregnancy blood 

4 Pb levels were greater in women from the town with a Pb smelter. No association was 

5 reported between maternal blood Pb and height or BMI at birth for the infants of these 

6 women despite the differences in maternal blood Pb between the two towns. Multiple 

7 studies were conducted that examined the association between maternal blood Pb and 

8 birth weight/fetal growth. In California, blood Pb measurements of women during 

9 pregnancy were matched with the corresponding birth certificates (Jelliffe-Pawlowski et 
February 2012 5-472 Draft – Do Not Cite or Quote

1 al., 2006). The adjusted OR for low birth weight that compared women with blood Pb 

2 levels ≥ 10 μg/dL to women with levels <10 μg/dL was elevated. However, it was 

3 difficult to draw conclusions about the relationship between blood Pb and birth weight 

4 due to small numbers (n = 9 for low birth weight) and the subsequently wide 95% CI. An 

5 association was detected for high blood Pb and having an infant who was small of his/her

6 gestational age (SGA). A study of term births in Iran reported no difference in blood Pb 

7 of women giving birth to a normal weight infant and women giving birth to an infant with 

8 low birth weight (Iranpour et al., 2007). A study in Vienna, Austria reported an inverse 

9 association between maternal blood Pb levels and birth weight but no associations for 

10 birth length or head circumference (Gundacker et al., 2010). Similarly, increased 

11 maternal blood Pb was associated with decreased birth weight, with the strongest 

12 associations observed at the lowest blood Pb level (Zhu et al., 2010). No association was 

13 observed between maternal blood Pb and SGA. A study in Taiwan examined both 

14 maternal and paternal blood Pb levels among those occupationally exposed to Pb and 

15 their associations with birth weight and SGA (Chen et al., 2006a). Paternal blood Pb 

16 levels were not associated with increased risk of low birth weight or SGA. Higher 

17 maternal blood Pb concentration was associated with higher risk of low birth weight and 

18 SGA, although not all of the associations were statistically significant. There were small 

19 numbers of infants with low birth weight or SGA, especially at the highest blood Pb 

20 levels (≥ 20 μg/dL). 

21 A study examining the association between Pb biomarker levels and birth weight used 

22 tibia bone measurements from mothers living in Mexico City (Kordas et al., 2009). Tibia 

23 Pb levels were inversely associated with birth weight but not with birth length. This 

24 association between Pb and birth weight was not modified by maternal folate 

25 consumption or maternal or infant MTHFR genotype, although the association between 

26 tibia Pb levels and birth weight was greater in magnitude among women with certain 

27 genotypes (statistical tests not reported). Another study in Mexico City reported no 

28 association between maternal tibia Pb levels and birth weight among girls but reported a 

29 positive association for boys (Afeiche et al., 2011). No associations were observed with 

30 maternal patella Pb concentration, although among boys, the relationship was positive but 

31 not statistically significant. One of the cohorts used by Afeiche et al. (2011) was also 

32 evaluated in another study (Cantonwine et al., 2010b). An inverse association was 

33 observed between tibia Pb and birth weight, especially at higher levels. This association 

34 was stronger among those mothers with variants of the hemochromatosis iron gene 

35 (HFE).

36 Multiple studies examined the relationship between Pb level and birth weight using Pb 

37 measured from the placenta or umbilical cord. Researchers in Chile collected the 

38 placentas from term births and compared the Pb levels for those born with normal birth 
February 2012 5-473 Draft – Do Not Cite or Quote

1 weights to those with low birth weights (Llanos and Ronco, 2009). Pb levels were greater 

2 in the placentas of infants with low birth weights. In addition, the authors note that 3 low 

3 birth weight infants had extremely high Pb levels in the placentas (>1.5 μg/g) and were 

4 excluded from these analyses. A study in Brazil examined Pb levels in umbilical cord 

5 blood from term births of women residing within 5 km of a Pb smelter (Zentner et al., 

6 2006). The cord blood Pb level was found to be inversely correlated with length and 

7 weight of the infants. A study with high Pb concentrations in umbilical cord blood 

8 reported an inverse association between Pb levels and birth weight (Atabek et al., 2007).

9 However, no correlation was detected in an analysis restricted to umbilical cord Pb less 

10 than 10 μg/dL. No association with other measures of growth, such as birth length and 

11 mid-arm circumference, were detected. In Saudi Arabia, a study was conducted among 

12 non-occupationally exposed women (Al-Saleh et al., 2008b). Umbilical cord blood Pb 

13 concentrations were low and an association was observed between umbilical cord Pb and 

14 head circumference. Another study recruited women in Pakistan (Janjua et al., 2009).

15 Umbilical cord blood Pb levels were not associated with low birth weight. The study by 

16 Iranpour et al. (2007) discussed above investigated the association with umbilical cord 

17 blood Pb levels in addition to their examination of maternal whole blood Pb. They again 

18 report no difference in levels between term infants of normal and low birth weight. A 

19 study comparing geometric mean umbilical cord blood Pb levels reported no difference in 

20 the levels for normal and low birth weight infants born to women living primarily in 

21 urban areas of Memphis, TN (Jones et al., 2010). A study performed in Baltimore, MD 

22 also reported no association between umbilical cord blood Pb concentration and birth 

23 weight (Wells et al., 2011a). This study had low blood Pb levels, with only 0.7% of 

24 participants having umbilical cord blood Pb measuring > 5 μg/dL. A study previously 

25 mentioned that observed an inverse association between maternal tibia Pb and birth 

26 weight in Mexico City reported no association between umbilical cord blood Pb 

27 concentration and birth weight (Cantonwine et al., 2010b). Finally, a study in Vienna 

28 measured Pb in the placenta (Gundacker et al., 2010). A positive correlation was 

29 observed between placenta Pb and birth length and weight; however, in the same study, 

30 maternal blood Pb was inversely related to birth weight. 

31 A study performed in Turkey examined the relationship between Pb levels in breast milk 

32 two months postpartum and size at birth (Örün et al., 2011). No association was observed 

33 between breast milk Pb concentration and birth weight or head circumference.

34 A few studies examined air exposures and reported inverse associations between air Pb 

35 concentrations and birth weight. Williams et al. (2007) examined Pb concentrations in the 

36 air during the first trimester. The purpose of their study was to demonstrate the use of 

37 hierarchical linear models and they used the example of air pollution and birth weight in 

38 Tennessee. The model results showed an association between ambient Pb concentration 
February 2012 5-474 Draft – Do Not Cite or Quote

1 and birth weight, with an estimated decrease in birth weight of 38 grams for every 

0.04 μg/m3 2 (i.e., one standard deviation) increase in Pb concentration. Another study of 

3 air Pb levels was conducted in Idaho and included two areas over three time periods. One 

4 study area was affected by damage to a local factory that lead to high Pb emissions 

5 during one of the time periods under study (Berkowitz et al., 2006). No levels of Pb are 

6 provided. Mean birth weight for term births was decreased among infants born to women 

7 living in the high exposure area during the period of high exposure compared to those 

8 living in the lower exposure area. The difference in birth weight of term births remained, 

9 but was reduced, between the two areas during the time period after the exposure ended. 

10 During the period of higher exposure, the odds of low birth weight among term births 

11 was increased among those living in the higher exposed area compared to those in the 

12 lower exposed area, but the odds were not different between the two study areas during 

13 the time periods before or after the high level of exposure. An increase in SGA infants 

14 (defined as infants with weights less than or equal to the lowest 5th percentile of birth 

15 weight for their sex and age) was also associated with living in the higher exposed area 

16 during the time period of higher exposure. The odds of SGA infants decreased during the 

17 time period after the exposure but the odds were still elevated compared to those residing 

18 in the lower exposed area. 

19 Evidence from previous toxicological studies has shown an association between 

20 gestational Pb exposure and reduced birth weight and impaired postnatal growth (U.S. 

21 EPA, 2006b). More recent studies have reported conflicting results. Wang et al. (2009e)

22 demonstrated a statistically significant decrease in fetal body weight and body length of 

23 Wistar rats after maternal exposure to 0.025% Pb-acetate during gestation days 1-10, 11-

24 20, or 1-20. The greatest decrease in fetal body weight and length was observed in the 

25 group exposed to Pb during gestation days 1-20 followed by the group exposed to Pb 

26 during gestation days 11-20. Teijón et al. (2006) observed that when pregnant dams were 

27 administered 200 ppm or 400 ppm Pb-acetate in drinking water, litter weight was 

28 significantly decreased (400 ppm Pb only) versus controls due to significant decrements 

29 in female pup birth weight; male birth weight was unaffected. This effect did not persist 

30 in the postnatal growth of the rats. The results of these studies indicate that as Pb 

31 exposure increases, the body weight of exposed offspring decreases. Massó-González and 

32 Antonia-García (2009) also observed an 8-20% decrease in body weight of pups from rat 

33 dams given 300 mg/L Pb-acetate in drinking water (exposure during gestation and 

34 lactation resulting in mean blood Pb level of 22.8 μg/dL), but no changes in body length 

35 were reported. In contrast, Leasure et al. (2008) reported a statistically significant inverse 

36 relationship between Pb exposure and body weight for male mice exposed to low 

37 (27 ppm)-and high (55 ppm)-levels of Pb during gestation. Male mice exposed to the low 

38 and high Pb concentrations during gestation were 26% and 13% heavier than controls at 

39 1 year of age, respectively. In this study, dams were administered 27 ppm (low), 55 ppm
February 2012 5-475 Draft – Do Not Cite or Quote

1 (moderate), and 109 ppm (high) Pb in drinking water beginning 2 weeks before mating 

2 and continuing until PND10. Resulting blood Pb levels ranged from 10 μg/dL or less in 

3 the low-exposure offspring to 42 μg/dL in the high-exposure offspring at PND10. The 

4 authors also reported that when dams received low or moderate levels of Pb in drinking 

5 water from birth to weaning neither male nor female offspring exposed to Pb postnatally 

6 exhibited a difference in body weight when compared to control offspring.

7 In summary, associations were observed between Pb and low birth weight in a study of 

8 maternal bone Pb and studies of Pb air exposures and birth weight. However, the 

9 associations were less consistent when using maternal blood Pb or umbilical cord and 

10 placenta Pb as the exposure measurement. Previous toxicological studies observed an 

11 association between gestational Pb exposure and reduced birth weight with moderate to 

12 high dose Pb. More recent findings using low dose Pb exposure reported increased 

13 offspring body weight after developmental Pb exposure. 

5.8.8 Effects on Postnatal Stature and Body Weight

14 Findings from previous toxicological studies of rodents and primates have demonstrated 

15 Pb induced impairment of postnatal growth (U.S. EPA, 2006b). Several recent 

16 epidemiologic studies examining the association of various biomarkers of Pb exposure 

17 with stature and body weight have been conducted and the evidence reported is mixed.
February 2012 5-476 Draft – Do Not Cite or Quote

Table 5-41 Summary of recent epidemiologic studies of associations between 

Pb levels and postnatal growth

Referencea Study Location and Years Study Population Pb Biomarker Mean Pb (SD) Effect Estimates

Afieche et al. (2011) Mexico City, Mexico

Children born between 

1994 and 2005

n=522 boys 

n=477 girls

Maternal bone Pb Patella: 10.4 (11.8) μg/g Change in weight at 5 years of age 

(g) per 1 SD increase in maternal 

bone Pb

Girls: 130.9 (95% CI: -227.4 to - 34.4) 

Boys: 13.0 (95%CI -73.7, 99.9)

Adjusted for cohort, maternal age, 

calf circumference, height, 

education, number of pregnancies, 

breast feeding for 6 months, 

calcium treatment, child’s 

gestational age at birth, height.

Sanna et al. (2011) Sardinia, Italy

Data collected in 1998, 

2002 and 2007

n=825 children 11-14 yrs 

old

Pb in hair 1998: 5.84 (6.56) μg/g

2002: 1.49 (1.72) μg/g

2007: 0.78 (0.93) μg/g

Height

1998: β log Pb= -0.121 (p=0.0021)

2002: β log Pb= -0.115 (p=0.0349)

2007: β log Pb= -0.011 (p=0.8665)

Sitting Height

1998: β log Pb=-0.117 (p=0.0017)

2002: β log Pb=-0.036 (p=0.5149)

2007: β log Pb=0.028 (p=0.6633)

ELL

1998: β log Pb=-0.103 (p=0.0209)

2002: β log Pb=-0.164 (p=0.0057)

2007: β log Pb=-0.008 (p=0.9058) 

Adjusted for age and sex

Ignasiak et al.

(2006)

South-western Poland

1995

(Industrial area with copper 

smelters and refineries)

school children 7-15 years

n=463 boys

n= 436 girls 

Concurrent blood Pb 7.7 (3.5) μg/dL Estimated decrement per 10 μg/dL 

increase in blood Pb

Weight:

Boys: 2.8 kg

Girls: 3.5 kg

Height:

Boys: 3.2 cm

Girls: 4.0 cm

Trunk length:

Boys: 1.2 cm

Girls: 1.1 cm

Leg length:

Boys: 2.1 cm

Girls: 2.9 cm

Arm length:

Boys: 1.8 cm

Girls: 1.9 cm

Adjusted for age, age2, and 

education level of mother
February 2012 5-477 Draft – Do Not Cite or Quote

Referencea Study Location and Years Study Population Pb Biomarker Mean Pb (SD) Effect Estimates

Hauser et al. (2008) Chapaevsk, Russia

May 2003 – May 2005

n=489 boys 

8-9 yrs old

Concurrent blood Pb 3 (2-5) μg/dL

Median (25-75 percentile)

Regression coefficient (95% CI)

Height (cm): -1.439 (-2,25, -0.63)

Weight (kg): -0.761 (-1.54, 0.02)

BMI: -0.107 (-0.44, 0.23)

Adjusted for birth weight, 

gestational age, and age at exam

Little et al. (2009) Dallas, Texas

1980-1989 and 2002

n=191 (1980s)

n=169 (2002)

2-12 yrs old

Concurrent blood Pb 1980s: 23.6 (1.3 SE) 

μg/dL

2002: 1.6 (0.2 SE) μg/dL

Blood Pb effect per 10μg/dL Pb 

increase – Mean(95%CI)

Height (cm): -2.1 (-1.9, -2.3)

Weight (kg): -1.9 (-1.7, -2.1)

BMI (kg/m2): -0.5 (-0.4, -0.7)

Adjusted for age, age2, sex and 

cohort effect

Min et al. (2008b) Seoul, South Korea

Date(s) not specified

n=62 boys

n= 46 girls

5-13 yrs

Concurrent blood Pb 2.4 (0.7) μg/dL Linear model estimate (SE; p)

Height: -1.449 (0.639; p=0.026)

Total arm length: -1.804 (0.702; 

p=0.012)

Body weight: -0.646 (0.718; 

p=0.370)

BMI: -0.006 (0.272; p=0.982)

Adjusted for age, sex, and father's 

education

Schell et al. (2009) Albany, New York

1986-1992, 1992-1998

n=244 Maternal blood Pb 

during second 

trimester, third 

trimester, and 

delivery; Infant blood 

Pb at delivery, 6 

months, and 12 

months

Maternal blood Pb during 

second trimester 2.8 (2.6) 

μg/dL, maternal blood Pb 

during third trimester: 2.6 

(2.2) μg/dL, maternal 

blood Pb at delivery: 2.8 

(2.4) μg/dL

Infant blood Pb at 

delivery: 2.3 (2.7) μg/dL, 

infant blood Pb at 6 

months: 3.2 (3.3) μg/dL, 

and infant blood Pb at 12 

months: 6.3 (4.8) μg/dL 

Linear model for maternal second 

trimester Pb: -0.242 (p-value 0.01) 

for 6-month head circumference. 

When examining second trimester 

maternal Pb ≥ 3 μg/dL, 

associations were observed for 

6 mo weight for age, 6 mo weight 

for length, 6 and 12 mo head 

circumference, and 12 mo upper 

arm circumference for age

Adjusted for infant sex, infant birth 

weight, infant nutrition, maternal 

age, marital status, employment, 

race, height, parity, second 

trimester smoking, and education. 
February 2012 5-478 Draft – Do Not Cite or Quote

Referencea Study Location and Years Study Population Pb Biomarker Mean Pb (SD) Effect Estimates

Lamb et al. (2008) Kosovo, Yugoslavia

1985-1986

n=309 mother child pairs Maternal blood Pb Pristina: 5.60 (1.99) μg/dL

Mitrovica: 20.56 (7.38) 

μg/dL

Regression coefficients relating 

maternal blood Pb to height (95% 

CI):

Pristina

Birth: 0.35 (-0.64, 1.34)

1 yr: -0.61 (-2.24, 1.03)

4 yr: 0.79 (-1.71, 3.29)

6.5 yr: 0.15 (-2.43, 2.74)

10 yr: -0.09 (-3.69, 3.52)

Mitrovica

Birth: 0.43 (-0.83, 1.69)

1 yr: -0.30 (-2.55, 1.96)

4 yr: -0.72 (-3.26, 1.82)

6.5 yr: -1.87 (-4.38, 0.64)

10 yr: -2.87 (-6.21, 0.47)

To BMI (95% CI):

Pristina

Birth: -0.14 (-0.69, 0.42)

1 yr: 0.61 (-0.28, 1.50)

4 yr: 0.17 (-0.67, 1.00)

6.5 yr: 0.61 (-0.09, 1.30)

10 yr: -0.49 (-1.45, 0.46)

Mitrovica

Birth: -0.18 (-0.69, 0.33)

1 yr: 0.23 (-0.84, 1.30)

4 yr: 0.16 (-0.66, 0.98)

6.5 yr: -0.12 (-0.90, 0.66)

10 yr: 1.31 (-0.95, 3.57)

Adjusted for sex, ethnicity, parity, 

maternal height or maternal BMI, 

maternal education, gestational age 

at delivery, gestational age at blood 

sample, and HOMES score

Zalina et al. (2008) Kuala Lumpur, Malaysia n=269 children 6.5-8.5 yrs 

old

n=169 urban

n=100 industrial

Concurrent blood Pb Industrial: 3.75 μg/dL

Urban: 3.56 μg/dL

Correlation with blood lead:

Height for age:

Urban: -0.095 (p=0.219)

Industrial: -0.037 (p=0.716)

Weight for age:

Urban: 0.019 (p=0.806)

Industrial: -0.063 (p=0.535)

Weight for height:

Urban: 0.136 (p=0.079)

Industrial: -0.069 (p=0.493)

Left arm circumference:

Urban: 0.041 (p=0.595)

Industrial: -0.055 (p=0.587)
February 2012 5-479 Draft – Do Not Cite or Quote

Referencea Study Location and Years Study Population Pb Biomarker Mean Pb (SD) Effect Estimates

Tomoum et al.

(2010)

Cairo, Egypt

Jan-Jun 2007

n=45 boys and girls 

10-13 yrs old

Concurrent blood Pb 9.46 (3.08) μg/dL Percentage of the median (SD):

Pb<10 μg/dL

Weight:

Boys: 127.56 (16.26)%

Girls: 114.8 (10.8)%

Height:

Boys: 98.06 (3.19)%

Girls: 96.75 (2.91)%

Pb≥ 10 μg/dL

Weight:

Boys: 122.0 (16.71)%

Girls: 123.11 (12.52)%

Height:

Boys: 99.5 (5.04)%

Girls: 100.33 (4.53)%

Adjusted for age and sex

Olivero-Verbel et al.

(2007)

Cartegena, Columbia

Jun-Aug 2004

n=189 children 5-9 yrs old Concurrent blood Pb 5.49 (0.23) μg/dL Spearman correlation coefficient (pvalue) between blood Pb and body 

size: -0.224 (0.002)

*no significance in partial 

correlation between blood Pb and 

size when controlled for age:

-0.096 (0.189)

Liu et al. (2011a) Guiyu, China

Chendian, China

Jan-Feb 2008

n=303

3-7 yrs old

Concurrent blood Pb Guiyu: 13.2 (4.0-48.5) 

μg/dL

Chendian: 8.2 (0-21.3) 

μg/dL

Median (range)

Mean chest circumference in girls 

was lower among those with higher 

blood Pb levels (>10μg/dL)

Mean chest and head 

circumference in children > 6 years 

old greater among those with 

higher blood Pb levels (>10 μg/dL)

No multivariate adjustment, 

stratification by age/sex

Mahram et al.

(2007)

Zanjan province,, Iran

Date(s) not specified

n=42 boys

n= 39 girls

7-11 yrs

Concurrent blood Pb Area with lead smelters: 

37.0 (24.7) μg/dL

Area without lead 

smelters: 15.6 (13.4) 

μg/dL

Comparison of control and study 

groups Height, standardized for 

age: p-value 0.52

Weight, standardized for age: pvalue 0.8

*Estimated Lower Limb Length

a

Studies are presented in order of first appearance in the text of this section.

1 Results from recent epidemiologic studies of postnatal growth are summarized in Table 

2 5-41. Afeiche et al. (2011) conducted a longitudinal study of children in Mexico City, 

3 born between 1994 and 2005. Maternal bone Pb during pregnancy was associated with a 

4 significant decrease in BW at age 5 years in girls but not in boys. The findings were 

5 robust to additional adjustment for child’s blood Pb level. In a study of children in 

6 Sardinia Italy, Sanna et al. (2011) measured Pb in hair at three points in time (1997, 2002, 

7 and 2007) and reported cross-sectional results from regression analyses for each of these 

8 time periods. Pb in hair decreased over time and significant associations of Pb in hair 
February 2012 5-480 Draft – Do Not Cite or Quote

1 with height were observed only in earlier time periods when hair Pb levels were relatively 

2 high. 

3 Ignasiak et al. (2006) studied school children aged 7-15 years living close to copper 

4 smelters and refineries in Poland to assess the impact of Pb exposure on their growth 

5 status. There was a statistically significant linear relationship between concurrent blood 

6 Pb and reduced weight, height, trunk, leg and arm lengths. This decrease in height was 

7 more influenced by decreases in leg length than trunk length. These results also indicated 

8 that there was attenuation in osteoblast activity associated with higher blood Pb levels, 

9 consistent with animal toxicological studies (Long et al., 1990). Hauser et al. (2008)

10 investigated the relationship between blood Pb and height in boys living in Chapaevsk, 

11 Russia. In a multivariate adjusted regression analysis, height significantly decreased with 

12 increasing blood Pb. Statistically nonsignificant decreases in weight and BMI were also 

13 observed. The association of blood Pb with height, weight and BMI was examined 

14 among two cohorts of children living near Pb smelters in Texas (Little et al., 2009). The 

15 first cohort included children 2-12 years old in 1980 and the second cohort included 

16 children of the same age in 2002 when blood Pb levels were substantially lower. 

17 Decreases in height, weight and BMI with increasing blood Pb levels were observed

18 among children in both cohorts and increases in height and weight were observed 

19 comparing children from the 2002 cohort to those from the 1980 cohort. In a study with 

20 Korean children, Min et al. (2008b) observed that height and total arm length decreased 

21 significantly with increasing blood Pb in multivariate adjusted regression models. A 

22 statistically nonsignificant decrease in body weight was observed with increasing blood 

23 Pb while no effect on BMI reported. A study in New York reported an association 

24 between maternal blood Pb during the second trimester of pregnancy and various 

25 measures of growth, especially among those mothers with blood Pb levels of at least 

26 3 μg/dL (Schell et al., 2009). These associations did not persist for those with maternal 

27 blood Pb levels less than 3 μg/dL. Among infants, 6 month blood Pb levels were not 

28 associated with 12 month measures of growth. In comparisons of changes in blood Pb 

29 levels over time, high maternal blood Pb combined with low 12 month blood Pb among 

30 infants (indicating a decrease in blood Pb over time) resulted in the greatest growth, even 

31 compared to those with both low maternal and infant blood Pb measures.

32 Contrary to the results summarized above, several studies do not observe associations 

33 between blood Pb levels and impaired growth. In a prospective study of 309 mother-child 

34 pairs from Yugoslavia, the relationship between maternal blood Pb and attained height in 

35 children was investigated in those living in an highly exposed town with a smelter and 

36 battery plant and those living in a relatively lower exposed town (Lamb et al., 2008). In 

37 multivariate adjusted regression models, neither attained height (at birth, 1, 4, 6.6, or 

38 10 years age) nor rate of height change per month (at birth-1 year, 1-4 years, 4-6.5 years, 
February 2012 5-481 Draft – Do Not Cite or Quote

1 6.5-10 years age) was associated with maternal pregnancy blood Pb levels in either the 

2 industrial or less exposed town. Weight was also not associated with maternal blood Pb 

3 in this study. In a study with a similar design, Zailina et al. (2008) studied the relationship 

4 of blood Pb and height in 7 year-old Malaysian school children comparing those 

5 attending two schools in an urban setting to those attending a school near an industrial 

6 area. After adjustment for age no statistically significant associations between concurrent 

7 blood Pb and physical development were observed. Tomoum et al. (2010) investigated 

8 the association between blood Pb and height in pubertal children in Cairo, Egypt. Neither 

9 boys nor girls with concurrent blood Pb levels > 10 μg/dL differed significantly in height 

10 or weight when compared to those with blood Pb < 10 μg/dL. In a simple correlation 

11 analysis of children aged 5-9 years in Colombia, Olivero-Verbel et al. (2007) reported 

12 that concurrent blood Pb levels were negatively associated with body size (r = -0.224, p < 

13 0.002). However, when a partial correlation analysis was performed controlling for age, 

14 the association between blood Pb and body size was no longer statistically significant. In 

15 a study of school children in China chest and head circumference were found to differ 

16 between high (>10μg/dL) and low concurrent blood Pb level groups; however, the 

17 direction of the difference was not consistent (Liu et al., 2011a). Among girls, comparing 

18 those with high and low blood Pb levels, a reduction in head circumference was 

19 observed. Among children greater than 6 years of age, those with higher blood Pb levels 

20 were reported to have greater head and chest circumferences. In a study of children aged 

21 7-11 years and living in an area of Iran with or without Pb smelters, age-standardized 

22 weight and height did not vary by study area (Mahram et al., 2007).

23 Recent toxicological studies report significant changes in postnatal or adult body weight 

24 after Pb exposure during different developmental windows. Masso-Gonzalez and 

25 Antonio-Garcia (2009) found Pb-induced decreased body weights at weaning (PND21) in 

26 rat pups from dams exposed to Pb during pregnancy and lactation. Blood Pb level in the 

27 control group was 1.43 μg/dL, in the Pb group it was 22.8 μg/dL. Dong et al. (2009)

28 reported decreased body weight in adult Kunming mice after exposure to 0.6% Pb-acetate

29 in drinking water for 8 weeks. In contrast, Leasure et al. (2008) reported a statistically 

30 significant inverse relationship between Pb exposure and body weight for male mice 

31 exposed to lower (27 ppm), moderate (55 ppm) and higher levels (109 ppm) levels of Pb 

32 during gestation and lactation (2 weeks before mating, through gestation and to PND10) 

33 with those exposed to the lowest dose having the highest adult body weight among the 

34 overweight Pb-exposed animals. Male mice exposed to the lower and higher Pb 

35 concentrations during gestation were 26% and 13% heavier than were controls at 1 year 

36 of age, respectively. In this study, dams were administered 27 ppm (low), 55 ppm

37 (moderate), and 109 ppm (high) Pb in drinking water beginning which resulted in 

38 respective blood Pb levels from 10 μg/dL or less in the low-exposure offspring to 42 

39 μg/dL in the high-exposure offspring at PND10. Leasure et al. (2008) also exposed a 
February 2012 5-482 Draft – Do Not Cite or Quote

1 separate group of mice to Pb only during the postnatal period (PND0-PND21, lactation 

2 only exposure) and mice exposed to the same aforementioned low or high dose of Pb did 

3 not exhibit a difference in body weight when compared to control offspring. Wang et al.

4 (2009e) observed a statistically significant decrease in fetal body weight and body length 

5 of Wistar rats at GD20 after maternal exposure to 0.025% Pb-acetate during gestation 

6 days 1-10, 11-20, or 1-20. Also, associations were reported between elevated maternal 

7 blood Pb levels (0.6, 1.3, or 1.74 μM, respectively or ~12.4, 26.9, or 36.0 μg/dL, 

8 respectively) compared to control (0.04 μM or ~ 0.83 μg/dL) and decreased pup body 

9 length, and placental weight in Wistar rats at GD20. The greatest decrease in fetal body 

10 weight and length was observed in the group exposed to Pb during gestation days 1-20 

11 followed by the group exposed to Pb during gestation days 11-20. Teijón et al. (2006)

12 observed reductions in birthweight of litters administered 200 ppm or 400 ppm Pb-acetate

13 in drinking water, but found that this effect did not persist in the postnatal growth of the

14 rats. 

15 Notably, previous toxicological studies observed reductions in postnatal weight as well as 

16 birth weight after exposure to Pb, albeit often at higher concentrations of Pb exposure. 

17 Ronis et al. (2001; 1998a; 1998b; 1996) have published a series of papers exposing rats to 

18 Pb over different developmental windows, showing associations between Pb exposure 

19 and deficits in growth. Sprague-Dawley rats with lifetime Pb exposure to 0.6% Pb-acetate

20 in drinking water (gestational-termination of experiment Pb exposure, maximum blood 

21 Pb of 316 μg/dL in males and 264 μg/dL in females) had sex-independent pre-pubertal 

22 growth suppression, male-specific suppression of pubertal growth and loss of growth 

23 effects postnatally but still maintained an overall decreased body size out to PND60 due 

24 to earlier deficits. In a follow up study using the same exposure duration with a dose of 

25 0.45% Pb-acetate (resulting in blood Pb of 263 μg/dL at PND85) yielded the same results

26 (Ronis et al., 1996) with mechanistic insight showing decrements in insulin-like growth 

27 factor 1 (IGF1) accompanying the decreases in growth rates. 

28 The body of toxicological literature on postnatal growth with Pb exposure indicates that 

29 Pb exposure can induce decrements in both height/body length and BW that may be 

30 persistent and differ by sex. However, findings from epidemiologic studies of postnatal 

31 growth are not consistent. Animal toxicology studies give insight to mechanistic changes 

32 that may contribute to this Pb-induced decrement and to the windows of exposure that 

33 may contribute greatest to these decrements. 
February 2012 5-483 Draft – Do Not Cite or Quote

5.8.9 Toxicological Studies of Developmental Effects

5.8.9.1 Developmental Effects on Blood and Liver

1 The 1986 and 2006 Pb AQCDs reported studies that suggest Pb may alter hematopoietic 

2 and hepatic function during development. Some recent studies provide evidence that 

3 support these findings; however recent results are not consistent among the studies.

4 Massó et al. (2007) reported a decrease in liver weights of pups born to dams that 

5 consumed 300 mg/L Pb in drinking water during gestation and lactation. They also 

6 reported an increase in the number of erythrocytes; however the erythrocyte size was 

7 diminished by 62%. Pb produced microcytic anemia as evidenced by decreased 

8 hemoglobin content and hematocrit values without changes in mean corpuscular 

9 hemoglobin (MCH) concentration. Alkaline phosphatase (ALP) activity, CAT activity, or 

10 thiobarbituric acid reactive substances (TBARS) production did not change in pups at 

11 postnatal 0, but increased statistically significantly by PND21 indicating reactive oxygen 

12 generation. No change in acid phosphatase (ACP) activity was observed in the livers of 

13 pups at PND0 or 21.

14 Massó-González and Antonia-García (2009) reported normochromic and microcytic 

15 anemia and a significant decrease in hematocrit values and blood δ-aminolevulinic acid 

16 dehydratase (ALAD) activity (90% reduction) in pups from dams administered 300 mg/L 

17 Pb-acetate in drinking water during gestation. The authors also reported that erythrocyte 

18 osmotic fragility was four times greater in Pb-exposed pups than in control pups. Massó19 González and Antonia-García (2009) reported increases in TBARS and CAT activity in 

20 the liver after Pb exposure. Intoxication with Pb also resulted in decreased liver protein 

21 concentrations and manganese-dependent SOD activity. Abnormalities in liver function 

22 were further exemplified by increases in liver concentrations of ALP and ACP.

23 Teijón et al. (2006) observed that gestational exposure to Pb caused a decrease in 

24 erythrocytes, hemoglobin, and MCH at weaning; however, by 1 and 3 months 

25 postweaning, these parameters had returned to normal values. The authors observed a 

26 slight increase in serum ALP, alanine aminotransferase (ALT), and aspartate 

27 aminotransferase (AST) levels after Pb exposure in the absence of liver histological 

28 changes. 

29 Pb-induced effects on SOD activity in the liver of fetuses after Pb intoxication was 

30 supported by a study by Uzbekov et al. (2007). The authors reported an initial increase in 

31 SOD activity in livers of pups exposed to 0.3 mg/L and 3.0 mg/L Pb nitrate during 

32 gestation for 1 month (mean daily consumption 27 μg/kg). In contrast, long-term 
February 2012 5-484 Draft – Do Not Cite or Quote

1 exposure (5 months) to the same concentrations of Pb nitrate concentration during 

2 gestation resulted in decreased hepatic SOD activity.

3 Effects on hepatic Phase I and Phase II enzymes after early developmental exposure of 

4 offspring to Pb during gestation and lactation was evaluated by Pillai et al. (2009). In the 

5 study, pregnant Charles Foster rats were administered 0.05 mg/kg body weight Pb 

6 subcutaneously throughout gestation until PND21. Pups were evaluated on PND56. 

7 Results of the study show that Phase I xenobiotic-metabolizing enzymes (NADPH- and 

8 NADH cytochrome c reductase) and Phase II xenobiotic- and steroid-metabolizing 

9 enzymes (δ-glutamyl transpeptidase, UDPGT, glutathione-s-transferase, and 17β10 hydroxysteroid oxidoreductase) were reduced in both male and female pups by PND56. 

11 Only inhibition in glutathione-s-transferase and 17β-hydroxysteroid oxidoreductase 

12 activities demonstrated a sex-specific pattern (glutathione-s-transferase inhibition in 

13 males; 17β-hydroxysteroid oxidoreductase inhibition greater in females). Observed Pb14 induced histological changes included massive fatty degeneration in hepatocytes, large 

15 vacuoles in cytoplasm, appearance of pyknotic nuclei, and infiltration of lymphocytes in 

16 the liver. Antioxidant enzymes (SOD, CAT, glutathione peroxidase, and glutathione 

17 reductase) were also reduced after Pb intoxication. Alterations in biochemical parameters 

18 included decreased DNA, RNA, and cholesterol content.

5.8.9.2 Developmental Effects on Skin

19 The 2006 Pb AQCD (U.S. EPA, 2006b) reported a study that demonstrated Pb-induced 

20 abnormalities in skin development. No current studies were identified that addressed Pb21 induced skin alterations.

5.8.9.3 Developmental Effects on the Retina

22 The 2006 Pb AQCD concluded that Pb exposure during early postnatal development 

23 (resulting in blood Pb levels ~20 μg/dL) impaired retinal development in female Long24 Evans hooded rats. A more recent study (Fox et al., 2008) exposed female Long-Evans 

25 hooded rats to low (27 ppm), moderate (55 ppm), and high (109 ppm) levels of 

26 Pb-acetate in drinking water beginning 2 weeks before mating, throughout gestation, and 

27 until PND10. Blood Pb levels measured in these pups on postnatal days 0-10 were 

28 10-12 μg/dL (low), 21-24 μg/dL (moderate), and 40-46 μg/dL (high). Results of the study

29 demonstrated supernormal persistent rod photoreceptor-mediated (scotopic) 

30 electroretinograms (ERGs) (Figure 5-46, Table 5-33) in adult rats similar to ERG 

31 findings in male and female children after gestational exposure to low- and moderate-
February 2012 5-485 Draft – Do Not Cite or Quote

1 levels of Pb. Low- and moderate-levels of Pb increased neurogenesis of rod 

2 photoreceptors and rod bipolar cells without affecting Müller glial cells and statistically 

3 significantly increased the number of rods in central and peripheral retina (Figure 5-46, 

4 and Table 5-33). High-level Pb exposure (109 ppm) statistically significantly decreased 

5 the number of rods in central and peripheral retina. Pb-exposure induced concentration6 dependent decreases in adult rat retinal dopamine synthesis and utilization/release.

5.8.9.4 Developmental Effects on Teeth 

7 Pb has been associated with multiple health effects including dental caries, however, 

8 there is very limited information available on the temporal and spatial incorporation of Pb 

9 in dental tissue (Arora et al., 2005). Arora et al. (2005) demonstrated that Wistar rat pups 

10 exposed to Pb during gestation and lactation (40 mg/L of Pb nitrate in drinking water of 

11 pregnant dams) had higher concentrations of Pb on the surface of enamel and in the 

12 dentine immediately adjacent to the pulp. The authors concluded that additional research 

13 is needed on the intracellular uptake of Pb during tooth development to fully understand 

14 the spatial distribution of Pb in teeth. 

5.8.10 Summary and Causal Determination

15 Many epidemiologic and toxicological studies of the effects of Pb on reproductive 

16 outcomes have been conducted since the 2006 Pb AQCD. These studies covered 

17 outcomes such as female and male reproductive function, birth defects, spontaneous 

18 abortions, infant mortality, preterm birth, low birth weight, and developmental effects. 

19 There is an abundance of evidence in the literature demonstrating that Pb induces 

20 reproductive and developmental effects in laboratory animals exposed to Pb during 

21 gestation and/or lactation. Many of the Pb-induced effects occur in a concentration22 dependent manner and have been observed at maternal blood Pb levels that do not result 

23 in clinical toxicity in the dams. Additionally, epidemiologic studies have demonstrated 

24 strong evidence of an association between Pb and delayed puberty as well as decrements 

25 to sperm/semen quality and function.

26 Many of the animal toxicology studies included in the 2006 Pb AQCD examined the 

27 effect of Pb on reproduction and development at blood Pb levels greater than 40 μg/dL, a 

28 dose where maternal toxicity can develop during pregnancy. Data from the 2006 Pb 

29 AQCD on male fertility showed Pb exposure or biomarkers of Pb-exposure were 

30 associated with decrements in semen quality. Recent studies have shown the effects of Pb 

31 exposure during early development to include disruption of endocrine function; delay in 
February 2012 5-486 Draft – Do Not Cite or Quote

1 the onset of puberty and alteration in reproductive function later in life; and changes in 

2 morphology or histology in sex organs and placenta. Additionally, recent epidemiologic 

3 studies of reproductive factors among males and females investigated whether Pb 

4 biomarker levels were associated with hormone levels, fertility, and onset of puberty. 

5 Epidemiologic studies showed associations between blood Pb and hormone levels for 

6 females. Studies of Pb and fertility are limited and inconsistent for females and males. 

7 Strong and consistent associations were observed between Pb levels in males in 

8 occupational settings with blood Pb levels as low as 20-45 μg/dL and sperm count and 

9 quality. 

10 Delayed puberty has been linked to decreased peak bone mass and increased risk of 

11 osteoporotic fractures (Gilsanz et al., 2011; Naves et al., 2005). Multiple epidemiologic 

12 studies of Pb and puberty have shown associations between concurrent blood Pb levels 

13 and delayed pubertal development for girls and boys (evidence is stronger among girls). 

14 These associations are consistently observed in multiple epidemiologic studies in 

15 populations with blood Pb levels <10μg/dL. Confounders considered in the 

16 epidemiologic studies that performed regression analyses varied. Most studies controlled 

17 for age and BMI. Other variables, such as measures of diet and SES and race/ethnicity, 

18 were included in some of the studies. Many of the studies demonstrating positive 

19 associations adjusted for many of the potential confounders. No patterns were detected in 

20 the associations between Pb and puberty based on inclusion of specific confounders. 

21 Pb-mediated changes in levels or function of reproductive and growth hormones have 

22 been demonstrated in past and more recent toxicological studies; however the findings 

23 are inconsistent. More data are needed to determine whether Pb exerts its toxic effects on 

24 the reproductive system by affecting the responsiveness of the hypothalamic-pituitary25 gonad axis or by suppressing circulating hormone levels. More recent toxicological 

26 studies suggest that oxidative stress is a major contributor to the toxic effects of Pb on 

27 male and female reproductive systems. The effects of ROS may involve interference with 

28 cellular defense systems leading to increased lipid peroxidation and free radical attack on 

29 lipids, proteins, and DNA. Several recent studies showed an association between 

30 increased generation of ROS and germ cell injury as evidenced by destruction of germ 

31 cell structure and function. Co-administration of Pb with various antioxidant compounds 

32 either eliminated Pb-induced injury or greatly attenuated its effects. In addition, many 

33 studies that observed increased oxidative stress also observed increased apoptosis which 

34 is likely a critical underlying mechanism in Pb-induced germ cell DNA damage and 

35 dysfunction. 

36 Overall, results of pregnancy outcomes were similar to those of the 2006 Pb AQCD; 

37 inconsistent evidence of a relationship with Pb was available for preterm birth and little 
February 2012 5-487 Draft – Do Not Cite or Quote

1 evidence was available to study the associations with spontaneous abortions. The 2006 

2 Pb AQCD included a few studies that reported potential associations between Pb and 

3 neural tube defects, but the recent epidemiologic studies found no association. Some 

4 associations were observed between Pb and low birth weight when epidemiologic studies 

5 used measures of maternal bone Pb or air exposures, but the associations were less 

6 consistent when using maternal blood Pb or umbilical cord and placenta Pb (maternal 

7 blood Pb or umbilical cord and placenta Pb were the biomarkers most commonly used in 

8 studies of low birth weight). Effects of Pb exposure during early development on 

9 toxicological studies included reduction in litter size, implantation, birth weight and 

10 postnatal growth. Findings from epidemiologic studies of postnatal growth are 

11 inconsistent.

12 Toxicological studies demonstrated that the effects of Pb exposure during early 

13 development include impairment of retinal development and alterations in the developing 

14 hematopoietic and hepatic systems. Negative developmental outcomes were also noted 

15 including effects on the eyes and teeth.

16 Similar to toxicological and epidemiologic studies that observed Pb to be associated with 

17 delayed puberty, delays of dynamic changes in the HPT axis are seen in the ecological 

18 literature, i.e., delayed metamorphosis in Pb exposed frogs. Additionally, Pb exposure 

19 has been shown to have detrimental effects on sperm, albeit often at higher blood Pb 

20 levels in epidemiology studies but in lower doses in the toxicology literature. Again, 

21 these findings agree with the ecological literature where Pb-dependent sperm effects are 

22 seen in rotifers, earthworms, and trout (Sections 7.3.5.2, 7.2.4.2, and 7.3.5.3). 

23 In conclusion, the recent toxicological and epidemiologic literature provides strong 

24 evidence that Pb exposure is associated with effects on reproduction and development. 

25 The weight of the evidence supports the association of Pb exposure with delayed onset of 

26 puberty in both males and females and detrimental effects on sperm and semen quality in 

27 occupationally-exposed males and in laboratory animals. In cross-sectional 

28 epidemiologic studies of girls (ages 6-18 years) with mean and/or median concurrent 

29 blood Pb levels less than 5 μg/dL consistent associations with delayed pubertal 

30 development (measured by age at menarche, pubic hair development, and breast 

31 development) were observed. Toxicological studies indicate that prenatal and lactational 

32 exposures to Pb can cause a delay in the onset of female puberty at blood Pb levels as low 

33 as 8 μg/dL (Iavicoli et al., 2006b; Iavicoli et al., 2004). Recent studies show that pubertal 

34 onset is one of the more sensitive markers of Pb exposure with effects observed after 

35 maternal exposures leading to blood Pb levels in the pup of 3.5-13 μg/dL (Iavicoli and 

36 Carelli, 2007). In boys (ages 8-15 years), fewer studies were conducted but associations 

37 were observed in most. Male animal toxicology studies have reported delayed sexual 
February 2012 5-488 Draft – Do Not Cite or Quote

1 maturity as measured with prostate weight, among other outcomes, seeing significant 

2 decrements at blood Pb levels of 34 μg/dL (Sokol et al., 1985). Additionally, Pb exposure 

3 has been shown to have detrimental effects on sperm. These were observed in 

4 epidemiologic studies at population mean blood Pb levels of 30 μg/dL and greater among 

5 men occupationally exposed (mean blood Pb levels in study controls around 10 μg/dL) 

6 and in animal toxicological studies with rabbits exposed to subcutaneous Pb 3 times 

7 per week for 15 weeks with blood Pb levels of 20 μg/dL (Moorman et al., 1998). The 

8 data on preterm birth, low birth weight, spontaneous abortions, birth defects, hormonal 

9 influences, and fecundity are less consistent between the toxicological and epidemiologic 

10 literature. The collective body of evidence integrated across epidemiologic and 

11 toxicological studies with a focus on the strong relationship observed with detrimental 

12 effects on sperm and delayed pubertal onset is sufficient to conclude that there is a causal 

13 relationship between Pb exposures and reproductive and developmental effects.

5.9 Effects on Other Organ Systems

5.9.1 Effects on the Hepatic System

14 Hepatotoxic effects of Pb indicated in various animal models and human populations 

15 include alterations in hepatic metabolism, hepatic cell proliferation, changes in 

16 cholesterol metabolism, as well as oxidative stress-related injury. Animal studies have 

17 also shown that exposure to Pb causes a decrease in Phase I along with a simultaneous 

18 increase in Phase II enzymes following exposure to Pb. Induction of oxidative stress by 

19 Pb exposure is well supported by an increase in lipid peroxidation along with a decrease 

20 in glutathione (GSH) levels and catalase (CAT), superoxide dismutase (SOD) and 

21 glutathione peroxidase (GPx) activities. 

5.9.1.1 Summary of Key Findings of the Effects on the Hepatic 

System from the 2006 Lead AQCD

22 The 2006 Pb AQCD stated that the large experimental animal database indicated 

23 hepatotoxic effects, including liver hyperplasia, at very high dose Pb exposures. Other 

24 effects noted in the liver following exposure to Pb included altered cholesterol synthesis, 

25 DNA synthesis and glucose-6-phosphotase dehydrogenase (G6DP) activity . The 2006 Pb 

26 AQCD reported that cytochrome (CYP) P450 levels decreased following single doses of 

27 Pb nitrate. Inhibition of induced and constitutive expression of microsomal CYP 1A1 and 

28 1A2 was observed among various P450 isozymes. Inhibition of Phase I enzymes was 
February 2012 5-489 Draft – Do Not Cite or Quote

1 accompanied by an increase in Phase II enzymes following exposure to Pb nitrate and 

2 other Pb compounds, suggesting that Pb is capable of causing a biochemical phenotype 

3 similar to hepatic nodules. Studies relating to Pb-induced hepatic hyperplasia suggested 

4 alterations in the gluconeogenic mechanism, DNA hypomethylation along with changes 

5 in proto-oncogene expression as well as cholesterol synthesis. Cholesterol metabolism 

6 changes following exposure to Pb were reportedly mediated as a result of induction of 

7 several enzymes related to cholesterol metabolism as well as a decrease in the cholesterol 

8 catabolizing enzyme, 7 α-hydroxylase. Tumor necrosis factor αlpha (TNF-α) was 

9 reported to be one of the major mitogenic signals that mediated Pb nitrate-induced 

10 hepatic hyperplasia in studies using inhibitors to block TNF-α activity. Other Pb-related 

11 effects presented in the 2006 Pb AQCD include liver cell apoptosis mediated by Kupffer 

12 cell derived signals and Pb-induced oxidative stress in vitro cell cultures. The 2006 Pb 

13 AQCD further suggested that alterations in liver heme metabolism may involve changes 

14 in 5-aminolevulinic acid dehydrogenase (ALAD) activity, porphyrin metabolism, 

15 transferrin (TF) gene expression and changes in iron metabolism. 

16 With regard to human studies, the 2006 Pb AQCD stated that nonspecific liver injury 

17 generally observed as increases in liver enzymes in the serum was reported in adults with 

18 occupational Pb exposure, although associations specifically with Pb exposures have not 

19 been well established. In addition, similar to effects noted in animal studies, cytochrome 

20 P450 activity was also suppressed in children and adults (drawn from the general 

21 population) following exposure to Pb. The 2006 Pb AQCD reported that hepatic effects 

22 occurred only at high Pb exposure levels (blood Pb levels > 30 μg/dL).

5.9.1.2 New Epidemiologic Studies

23 A few occupational epidemiologic studies examined liver biochemical parameters effects 

24 on antioxidant status and oxidative stress resulting from exposure to Pb. However all of 

25 these occupationally-exposed cohorts represented populations highly exposed to Pb, with 

26 blood Pb levels ranging from 29 to 53 μg/dL. Although the hepatotoxicity observed 

27 within these cohorts may not be generalizable to the general population as a whole, they 

28 are useful in demonstrating consistent effects on a number of liver outcomes, including 

29 altered liver function (i.e., changes in the level of liver function enzymes), oxidative 

30 stress, and antioxidant status (Can et al., 2008; Khan et al., 2008; Patil et al., 2007).

31 Additionally, these studies were cross-sectional in design; thus, there is uncertainty 

32 regarding the magnitude, timing, frequency, and duration of Pb exposure that contributed 

33 to the observed associations.
February 2012 5-490 Draft – Do Not Cite or Quote

1 In one of the occupational cohorts (spray painters in western Maharashtra, India, exposed 

2 to Pb for > 6 hours/day for 2 to 20 years) examined by Patil et al. (2007), mean (SD) 

3 blood Pb levels in workers were 22.32 (8.87) μg/dL. The blood Pb levels in workers were 

4 statistically significantly higher compared to those in the concurrent control group (mean 

5 [SD]: 12.52 [4.08] μg/dL), who had no history of Pb exposure. Liver function enzymes, 

6 including serum glutamic oxaloacetic transaminase (SGOT)/AST, and serum glutamic 

7 pyruvic transaminase (SGPT)/ALT levels were statistically significantly increased in 

8 spray painters compared to those in controls, whereas total serum protein levels were 

9 decreased. In another occupational epidemiolgic study, Conterato et al. (In Press)

10 investigated liver function parameters in automotive painters exposed to Pb in Brazil. 

11 Exposed painters had a mean (SD) blood Pb concentration of 5.4 (0.4) μg/dL compared 

12 to 1.5 (0.1) μg/dL in controls. The mean (SD) duration of exposure to Pb in painters was 

13 133.9 (14.5) months, whereas the controls were not occupationally exposed to Pb. In 

14 exposed workers, the levels of aspartate aminotransferase (AST), but not γ15 glutamyltranseferase, were increased approximately 2-fold compared to levels in controls 

16 (p < 0.05). The activity of AST was positively correlated with blood Pb levels (r = 0.26, p 

17 < 0.05). The authors suggested that confounding exposures to toxic constituents of the 

18 paints regularly used by painters, and not Pb, may be the etiological cause of decrements 

19 in AST function as these effects were not also seen in battery workers with much higher 

20 blood Pb levels (49.8 μg/dL). Co-exposure to other environmental contaminants may also 

21 explain the effects seen in occupationally-exposed spray-painters in Patil et al. (2007).

5.9.1.3 New Toxicological Studies

Hepatic Metabolism

22 As stated in the 2006 Pb AQCD, acute exposures of rodents to Pb nitrate and other Pb 

23 compounds cause a decrease in Phase I enzymes accompanied by a simultaneous increase 

24 in Phase II enzymes. The conclusions presented in the 2006 Pb AQCD were also 

25 reviewed by Mudipalli (2007).

26 Changes in biochemical parameters, suggestive of liver damage, in undernourished male 

27 Wistar rats (fed low-protein diet without mineral supplements) treated with 500 ppm

28 Pb-acetate in drinking water over a 10 month period, included decreases in serum protein 

29 and albumin levels as well as increases in aspartate aminotransferase (AST), alanine 

30 aminotransferase (ALT), serum alkaline phosphatase (ALP), and gamma glutamyl 

31 transpeptidase (GGT) levels (DS et al., 2009). In Pb-treated animals, the blood Pb levels 

32 steadily increased throughout the initial portion of the study period, reaching a maximum 

33 of approximately 30 μg/dL after 2 months. After this time, blood Pb levels rapidly 
February 2012 5-491 Draft – Do Not Cite or Quote

1 increased to approximately 110 μg/dL (greatly in excess of blood levels in the general 

2 human population) by six months time and remained at this level until the termination of 

3 exposure at 10 months. The study authors reported that similar biochemical changes were 

4 not observed in animals treated with Pb-acetate maintained on protein-adequate, mineral 

5 rich diet and concluded that nutritional management is important in managing Pb-related 

6 poisoning. Mice gavaged with 50 mg/kg Pb nitrate for 40 days demonstrated similarly 

7 increased activities of AST, ALT, ALP, and acid phosphatase (ACP) compared to 

8 controls (Sharma et al., 2010a). Upadhyay et al. (2009) reported that treatment of 

9 Sprague-Dawley rats with 35 mg/kg Pb via i.p. injection for 3 days (blood Pb not 

10 reported) significantly increased the activities of ALT, AST, serum ALP, and acid 

11 phosphatase over those in controls, whereas alkaline phosphatase activity was decreased 

12 in Pb-treated animals. Concomitant treatment with zinc and varying levels of ascorbic 

13 acid were observed to ameliorate the toxic effects of Pb. The serum activities of glutamic 

14 pyruvic transaminase (GPT) and lactate dehydrogenase (LDH) were similarly 

15 significantly increased over those in controls in mice subcutaneously injected with 

16 50 mg/kg Pb-acetate daily for 15 days (blood Pb not reported) (Wang et al.). Swarup et 

17 al. (2007) investigated serum biochemical changes in cows living in Pb-contaminated 

18 environments. Serum levels of ALT, AST, alkaline phosphatase, total protein, albumin, 

19 globulin, and A/G ratio were statistically significantly altered in cows living near Pb-Zn 

20 smelters (mean [SD] blood Pb: 86 [6] μg/dL, greatly in excess of blood levels in general 

21 human populations) compared to control cows (mean [SD] blood Pb: 7 [1] μg/dL). 

22 Significant positive correlations were found between blood Pb and ALT and AST, 

23 whereas a negative correlation was observed between blood Pb and total lipids, protein, 

24 and albumin. 

25 Pillai et al. (2009) investigated effects on hepatic phase I and II enzymes in male and 

26 female rats born to dams that were treated with 50 μg/kg Pb-acetate via subcutaneous 

27 injection daily throughout gestation and continuing until PND21. Thus, the offspring of 

28 treated dams were exposed to Pb via placental and lactational transfer. The female and 

29 male pups were then allowed to reach sexual maturity (PND55-56) to assess continuing 

30 exposure to bioaccumulated Pb. The activities of hepatic phase I enzymes NADPH- and 

31 NADH-cytochrome c reductase were statistically significantly reduced in Pb-exposed 

32 male and female rats on PND56 (blood Pb not reported), compared to controls. In rats 

33 treated with 25 μg/kg Pb and Cd, the effect on phase I enzymes was increased. Pb 

34 treatment additionally decreased the activities of phase II enzymes uridine diphosphate35 glucoronyl transferase and GST in males and females, but no effect was observed on 

36 GGT or 17β-hydroxysteroid oxidoreductase. Additionally, no effect was observed in Pb37 treated rats on serum glutamate pyruvate dehydrogenase or ALP activities in males or 

38 females. Histological observations in both male and female rats demonstrated fatty 

39 degeneration, vacuolization, and pycnotic nuclei, indicating general hepatotoxicity 
February 2012 5-492 Draft – Do Not Cite or Quote

1 following Pb treatment. In a similar study, Teijon et al. (2006) exposed Wistar rats to 200 

2 or 400 ppm throughout gestation, lactation, and 3 months postweaning, or only 1 month 

3 postweaning. In the animals exposed continuously throughout gestation and lactation, the 

4 concentrations of Pb in the liver were elevated in the 200- and 400-ppm groups 1 and 3 

5 months postweaning. Liver concentrations of Pb were greater in the 200 ppm animals 

6 compared to the 400 ppm animals at one month postweaning (mean [SE]: 1.19 μg Pb/g 

7 tissue [0.30] versus 0.76 [0.06], respectively), but were similar between the 2 dosing 

8 regimens at 3 months postweaning (mean [SE]: 0.054 [0.06] versus 0.55 [0.07], 

9 respectively). ALP activity was increased at 2 weeks postweaning in animals 

10 continuously exposed to Pb throughout gestation and lactation, whereas ALT activity was 

11 decreased only at 2 and 3 months postweaning. In animals exposed only for 1 month 

12 postweaning, serum ALP activity was significantly increased, although not in a 

13 concentration-dependent manner. ALT and AST activities were not affected.

14 Cheng et al. (2006) studied the mechanism of Pb effects on bacterial lipopolysaccharide 

15 (LPS)-induced TNF-α expression. A/J mice were injected via i.p with 100 μmol/kg Pb, 

16 with or without 5 mg/kg LPS. Pb alone did not affect liver function (measured as AST or

17 ALT activity) or the level of TNF-α in the serum. In comparison, treatment of mice with 

18 low doses of Pb and LPS together caused a statistically significant increase in TNF-α 

19 induction as well as enhanced liver injury, suggesting that Pb potentiated LPS-induced 

20 inflammation. In an in vitro study, the authors reported that co-exposure of Pb and LPS 

21 stimulated the phosphorylation of p42/44 mitogen-activated protein kinase (MAPK) and 

22 increased TNF-α expression in mouse whole blood cells, peritoneal macrophages, and 

23 RAW264.7 cells (a macrophage cell line) and concluded that monocytes/macrophages 

24 (rather than hepatocytes) were primarily responsible for Pb increasing LPS- induced 

25 TNF-α levels via the protein kinase C (PKC)/MAPK pathway. Similarly, Pb chloride 

26 potentiated bovine serum albumin (BSA)-induced inflammation in the livers of mice 

27 subcutaneously injected with Pb (Sá et al., In Press).

Lipid Metabolism

28 In a lipid metabolism study, Ademuyiwa et al. (2009) reported that male albino Sprague 

29 Dawley rats exposed to 200, 300 and 400 ppm Pb in drinking water had mean (SD) blood 

30 Pb levels of 40.63 (9.21), 61.44 (4.63), and 39.00 (7.90) μg/dL, respectively. Animals 

31 exposed to 200 ppm Pb had mean (SD) liver Pb concentrations of 10.04 (1.14) μg/g, 

32 compared to 3.24 (1.19) and 2.41 (0.31) in animals exposed to 300 or 400 ppm Pb, 

33 respectively. Animals exposed to Pb exhibited increased hepatic cholesterogenesis at all 

34 doses tested compared to controls. Additionally, a decrease in triglyceride was observed 

35 at 300 and 400 ppm Pb; a decrease in phospholipid levels was observed at 400 ppm Pb. 

36 The authors also reported positive correlations between tissue cholesterol and 
February 2012 5-493 Draft – Do Not Cite or Quote

1 phospholipids and Pb accumulation in liver across all doses. In contrast, the association 

2 between tissue triglyceride levels and Pb accumulation was negative. In related studies, 

3 Khotimchenko and Kolenchenko (2007) reported that adult male albino rats treated with 

4 100 mg/kg Pb-acetate for as little as 14 days (blood Pb not reported) exhibited disorders 

5 in lipid metabolism that were supported by increased levels of total cholesterol and 

6 triglyceride levels in the liver tissue, whereas Sharma et al. (2010a) reported increased 

7 liver cholesterol in mice gavaged with 50 mg/kg Pb nitrate for 40 days. Pillai et al. (2009)

8 observed decreases in total liver cholesterol in PND56 male and female rats that had been 

9 exposed to 50 μg/kg Pb-acetate continuously throughout gestation and lactation. These 

10 results suggest that induction of cholesterogenesis and phospholipidosis in the liver by Pb 

11 may cause subtle effects at the cellular level that may lead to hepatotoxicity. Kojima and 

12 Degawa (2006) examined the sex-related differences in the hepatic sterol regulatory 

13 element binding protein-2 (SREBP-2) and 3-hydroxy-3-methylglutaryl-CoA reductase 

14 (HMGR) gene expressions in male and female Sprague Dawley rats injected with 

15 100 μmol/kg body weight of Pb nitrate intravenously (blood Pb not reported). The 

16 SREBP-2 expression, which is a transcription factor for the HMGR gene, was 

17 significantly increased in males and females with the increase occurring earlier in male 

18 rats (6-12 hours, compared to 24-36 hours in females). In contrast, expression of the 

19 HMGR gene, a rate limiting enzyme in cholesterol biosynthesis, was significantly 

20 increased in both Pb-exposed males and females at earlier time frames (3-48 hours in 

21 males; 12-48 hours in females) compared to the SREBP-2 gene expression. Significant 

22 increases in total liver cholesterol were also observed in Pb-exposed males and females at 

23 3-48 and 24-48 hours, respectively. These results suggest that the SREBP-2 and HMGR 

24 gene expressions and increase in total cholesterol levels in the liver in response to Pb 

25 occur earlier in males compared to females and also suggest that the HMGR gene 

26 expression and increase in total cholesterol levels in the liver occur before an increase in 

27 the SREBP-2 gene expression in either sex. 

Hepatic Oxidative Stress

28 A number of studies pertaining to hepatic oxidative stress as a result of exposure to 

29 various Pb compounds were identified. Adegbesan and Adenuga (2007) reported that 

30 protein undernourished male Wistar rats injected with 100 μmol/kg Pb nitrate (blood Pb 

31 not reported) exhibited increased lipid peroxidation, increased CAT activity, decreased 

32 SOD activity, and increased GSH levels, compared to undernourished rats not exposed to 

33 Pb. Increased lipid peroxidation and decreased CAT and SOD activity were also 

34 observed when comparing undernourished Pb-exposed rats to well-nourished control rats. 

35 Study authors concluded that malnutrition exacerbated Pb exposure effects on liver lipid 

36 peroxidation and the involvement of free radicals in Pb toxicity. Male Foster rats treated 
February 2012 5-494 Draft – Do Not Cite or Quote

1 with 0.025 mg/kg Pb via i.p. injection (blood Pb not reported) also exhibited statistically 

2 significant increases in lipid peroxidation levels and decreases in SOD, CAT, and 

3 glucose-6-phosphatase dehydrogenase (G6PD) levels in liver mitochondrial and 

4 postmitochondrial fractions (Pandya et al., 2010). Statistically nonsignificant decreases 

5 were also observed in GSH levels and GPx and GR activities in Pb-treated animals. In 

6 mice gavaged with 50 mg/kg Pb nitrate for 40 days, lipid peroxidation was increased and 

7 SOD, CAT, and GSH were decreased compared to controls (Sharma et al., 2010a).

8 Additionally, exposure to Pb nitrate resulted in histopathological changes in the structure 

9 of the liver: hepatocytes were damaged and were marked by cytoplasmic vacuolization 

10 and pycnotic nuclei. Yu et al. (2008) reported similar concentration-dependent increases 

11 in lipid peroxide levels and decreases in GSH levels and CAT, SOD and GPx activities in 

12 castrated boars that received a supplemental diet with 0, 5, 10, or 20 mg/kg Pb. The level 

13 of hepatic CuZnSOD mRNA was also reduced in Pb-treated animals. The study authors 

14 suggested that this decrease in SOD mRNA expression and activity of antioxidant 

15 enzymes may lead to a reduction free radical scavenging capability, along with increased 

16 lipid peroxidation, potentially causing serious damage to hepatic function and structure. 

17 Khotimchenko and Kolinchenko (2007) also reported an increase in lipid peroxidation 

18 and development of hepatitis in male albino rat liver parenchyma following treatment 

19 with 100 mg/kg Pb-acetate for as little as 14 days. Lipid peroxidation was demonstrated 

20 by increases in malondialdehyde (MDA) levels along with decreases in GSH and thiol 

21 groups indicating injury in the liver antioxidant system. In another experiment, Jurczuk et 

22 al. (2007) reported that male Wistar rats treated with 500 mg/L Pb in drinking water 

23 (blood Pb not reported) exhibited decreases in liver vitamin E and GSH levels along with 

24 an increase in lipid peroxidation. The study authors hypothesized that vitamin E is 

25 involved in the mechanism of peroxidative action of Pb in the liver, and concluded that 

26 the suggested protective role of vitamin E in the potential toxicity by Pb may be related to 

27 scavenging of free radicals that are generated either directly or indirectly by Pb. In a 

28 study examining the role of low molecular weight thiols on peroxidative mechanisms, 

29 Jurczuk et al. (2006) stated that male Wistar rats treated with 500 mg/L Pb-acetate in 

30 drinking water exhibited a decrease in blood ALAD as well as decreases in GSH and 

31 nonprotein sulfhydryl (NPSH) levels in the liver. Metallthionein levels were also reported 

32 to be higher in the liver following exposure to Pb. Levels of hepatic lipid peroxidation 

33 were observed to be significantly increased in rats treated with 35 mg/kg Pb via i.p. 

34 injection (blood Pb not reported), whereas hepatic GSH was significantly decreased

35 (Upadhyay et al., 2009). In vitro exposure of human embryonic hepatocytes (WRL-68) to 

36 5 μM Pb-acetate for 30 days resulted in increase production of reactive oxygen species 

37 (ROS) throughout the incubation period (Hernández-Franco et al., 2011). Concurrent 

38 with this increase in ROS generation, the activities of SOD and the levels of membrane 
February 2012 5-495 Draft – Do Not Cite or Quote

1 lipid peroxidative damage also increased throughout the first 24 days of exposure, but 

2 returned to normal levels by the end of the incubation period. 

3 In a study examining the effects of Pb exposure to fetuses, Massó et al. (2007) exposed 

4 pregnant Wistar rats with 300 mg/L Pb in drinking water starting at day 1 of pregnancy to 

5 parturition or until weaning to determine the effects of Pb exposure in the fetal liver. 

6 Blood Pb levels were higher at parturition (mean [SD]: 31.5 [0.80] μg/dL) than at 

7 weaning (mean [SD]: 22.8 [0.50] μg/dL). Pups exhibited liver damage that was 

8 accompanied by an increase in thiobarbituric acid-reactive species (TBARS) production 

9 and increased CAT activity compared to controls. In addition, increased ALP and acid 

10 phosphatase activity was also observed. Uzbekov et al. (2007) exposed female Wistar rats 

11 to 0.3 and 3.0 mg/L Pb nitrate for 1 and 5 months prior to, and continuing during 

12 pregnancy, and measured fetal hepatic SOD activity on GD20. Control rats had a mean 

13 (SD) blood Pb level of 16.1 (0.63) μg/dL, whereas rats exposed to 0.3 and 3.0 mg/L Pb 

14 had mean blood Pb levels that were 26.5% (20.4 μg/dL) and 51.8% (24.4 μg/dL) higher, 

15 respectively. In the fetuses from dams exposed for 1 month prior to pregnancy, a 

16 concentration-dependent increase in liver SOD activity was observed, whereas SOD 

17 activity was decreased in the fetuses from dams exposed for 5 months prior to pregnancy. 

18 The increase in SOD activity in the livers of fetuses from dams exposed to 0.3 or 

19 3.0 mg/L Pb nitrate for one month suggests that activation of SOD in response to 

20 increased free radical production, while the decrease in SOD production in fetal livers 

21 from dams exposed to the same concentrations for 5 months suggests that longer 

22 durations of Pb exposure impairs the antioxidant defense mechanism. No effects on GSH 

23 or MDA levels were observed in PND56 male and female rats following continuous 

24 exposure to 50 μg/kg Pb-acetate throughout gestation and lactation (blood Pb not 

25 reported) (Pillai et al., 2009).

26 The studies presented above all support the possible oxidative stress impacts following 

27 exposure to various doses of Pb administered in various forms and the potential for 

28 hepatotoxicity as a result of oxidative stress. 

Hepatic Apoptosis

29 Fan et al. (2009b) reported that a single i.v. injection (tail vein) of 200 μmol/kg Pb nitrate 

30 resulted in an increase in the expression of ferritin light-chain (FLT) in rats (mean [SD]: 

31 3.5 [1.0]-fold increase) over that in controls. Immunohistochemical analysis revealed that 

32 hepatocytes around the central vein were heavily stained by anti-FTL antibodies, as were 

33 nonparenchymal cells identified as Kupffer cells. The authors hypothesized that the 

34 expression of FTL in Kupffer cells may be the result of phagocytosis of apoptotic cells:

35 in Pb-treated rats, apoptotic hepatocytes represented a mean (SD) 2.5 (1.4)% of total cells 
February 2012 5-496 Draft – Do Not Cite or Quote

1 whereas only a mean (SD) 0.31 (0.31)% of hepatocytes were apoptotic in control 

2 animals. FTL expression in Kupffer cells was not increased in rats treated with clofibrate, 

3 which induced hepatocellular proliferation, but not apoptosis. 

5.9.2 Effects on the Gastrointestinal System

4 Gastrointestinal effects of Pb exposure resulting in blood Pb levels ranging from 30 up to 

5 80 μg/dL in humans primarily include abdominal pain, constipation, and internal 

6 paralysis. In animals, degeneration of the intestinal epithelial mucosa and a decrease in 

7 duodenal motility has been reported following Pb exposure.

5.9.2.1 Summary of Key Findings on the Effects on the 

Gastrointestinal System from the 2006 Lead AQCD

8 The 2006 Pb AQCD stated that a number of factors influence the gastrointestinal 

9 absorption of Pb including the chemical and physical form of Pb, the age at Pb intake, as 

10 well as various nutritional factors. Potential malabsorption of Pb as a result of 

11 degeneration of the intestinal epithelial mucosa has been observed in rats exposed to Pb. 

12 In suckling rat pups, casein micelles incidences were reported as a result of Pb present in 

13 bovine and rat milk and in infant milk formula. Pb ingestion through water was more 

14 toxic compared to Pb ingestion via milk. Pb ingested in milk was reported to be taken up 

15 by the ileal tissue, whereas Pb administered intragastrically as a soluble salt was 

16 primarily accumulated in the duodenum irrespective of vehicle used for administration. 

17 Decreases in duodenal motility and the amplitude of contractility in the intestinal tract 

18 were observed in rats following Pb exposure. Nutritional studies examining different 

19 dietary levels of Pb in rats, calcium, and vitamin D indicated competition in absorption 

20 between Pb and calcium. Dietary supplement with vitamin D led to an increase in 

21 intestinal absorption of Pb and calcium. In instances where severe calcium deficiency 

22 was noted, ingestion of Pb caused a clear decrease in 1,25-dihydroxy vitamin D (1,25-

23 (OH)2D3) levels. Overall, the 2006 Pb AQCD stated that studies in rat intestine have 

24 shown that the largest amount of Pb absorption occurs in the duodenum with the 

25 mechanisms of absorption involving active transport and diffusion via the intestinal 

26 epithelial cells. Absorption has been reported to occur, through both saturable and 

27 nonsaturable pathways based on results from various animal studies. The 2006 Pb AQCD 

28 reported evidence that symptoms associated with gastrointestinal colic (abdominal pain, 

29 constipation, intestinal paralysis) were prevalent in occupationally exposed adults with 

30 blood Pb levels ≥ 50 μg/dL.
February 2012 5-497 Draft – Do Not Cite or Quote

5.9.2.2 New Epidemiologic Studies

1 The 2006 Pb AQCD reported that in humans, gastrointestinal effects generally include 

2 abdominal pain, constipation, and internal paralysis. Kuruvilla et al. (2006) reported 

3 gastrointestinal effects including stomach pain and gastritis along with other Pb-related 

4 clinical manifestations in painters (mean [SD] blood Pb: 8.04 [5.04] μg/dL) 

5 occupationally exposed to Pb in India. Case reports involving children and individuals 

6 occupationally exposed to Pb with very high blood Pb levels (> 15 μg/dL) were 

7 consistent with these observations of GI pain and gastritis (Fonte et al., 2007).

5.9.2.3 New Toxicological Studies

8 A few new studies pertaining to gastrointestinal effects of Pb exposure were identified. 

9 Santos et al. (2006b) examined the impact of Pb exposure on nonadrenergic 

10 noncholinergic (NANC) relaxations in rat gastric fundus. Male Wistar rats treated with 

11 0.008% Pb-acetate (80 ppm) via drinking water for 15, 30, and 120 days (blood Pb not 

12 reported) exhibited a significant difference in NANC relaxations in the gastric fundus 

13 following electrical field stimulus (EFS). While frequency-dependent relaxations were 

14 observed in all groups, including the control group, the relaxations were significantly 

15 inhibited in rats treated with Pb-acetate for all three durations. When gastric fundus strips 

16 from rats were incubated with L-nitroarginine (L-NOARG), a nitric oxide synthase 

17 (NOS) inhibiter, no additional inhibition in relaxations was observed. In contrast, 

18 incubation with sodium nitroprusside and 8-Br-GMPc (a Cyclic guanosine 

19 monophosphate [cGMP] analog), resulted in a concentration-dependent relaxation in 

20 strips in the control group and group exposed to Pb-acetate for 120 days. Study authors 

21 concluded that chronic exposure to Pb causes inhibition in NANC relaxation probably

22 due to the modulated release of NO from the NANC nerves or due to interaction with the 

23 intracellular transducer mechanism in the rat gastric fundus. 

24 In another study examining Pb-induced oxidative stress in the gastric mucosa, Olaleye et 

25 al. (2007) treated Albino Wistar rats with 100 or 5,000 mg/L of Pb-acetate for 15 weeks 

26 (blood Pb not reported). Exposure to Pb-acetate caused a significant increase in gastric 

27 mucosal damage caused by pretreatment with acidified ethanol. Study authors reported 

28 that though the basal gastric acid secretory rate was not altered, stomach response to 

29 histamine was significantly higher in animals treated with Pb-acetate compared to that in 

30 the controls. Additionally, there was a significant increase in gastric lipid peroxidation at 

31 both the 100 and 5,000 mg/L dose levels. In contrast, CAT, and SOD activities and nitrite 

32 levels were significantly decreased in the gastric mucosa. Study authors concluded that 
February 2012 5-498 Draft – Do Not Cite or Quote

1 exposure to Pb may increase the formation of gastric ulcers as a result of changes in the 

2 oxidative metabolism in the stomach.

5.9.3 Effects on the Endocrine System

3 Endocrine processes that are most commonly found to be impacted by Pb exposure 

4 include changes in the thyroid, such as changes in the thyroid stimulating hormone 

5 (TSH), triiodothyronine (T3), and thyroxine (T4). 

5.9.3.1 Summary of Key Findings of the Effects on the Endocrine 

System from the 2006 Lead AQCD

6 The 2006 Pb AQCD reported that endocrine processes impacted by occupational Pb 

7 exposure include thyroid hormone levels, changes in male sex hormone levels, as well as 

8 changes in the production of 1,25-(OH)2D3 levels. However, these effects were reported 

9 to be observed only with blood Pb levels exceeding 30–40 μg/dL. In addition, alterations 

10 in calcitropic hormones were affected in children with blood Pb levels ranging from 10-

11 120 μg/dL. A summary of key findings pertaining to reproductive hormones in males and 

12 females in the current document is presented in the section on reproductive and 

13 developmental effects (Sections 5.8.1 and 5.8.2). 

5.9.3.2 New Epidemiologic Studies

14 Recent epidemiologic studies have reported associations between indicators of exposure 

15 to Pb and thyroid hormone levels. In workers highly exposed to Pb (blood Pb = 

16 71.1 μg/dL) thyroid stimulating hormone (TSH) and free T4, but not free T3, were 

17 increased over levels in controls (mean blood Pb level of 0.2 μg/dL), although these 

18 results are most likely not generalizable to the general public due to the high blood Pb 

19 levels of exposed workers (Pekcici et al., 2010). Abdelouahab et al. (2008) performed a 

20 cross-sectional study in a Canadian population characterized by high consumption of 

21 freshwater fish. The median concurrent blood Pb level was 3.1 μg/dL for men and 

22 1.7 μg/dL for women. It is important to note that the median blood Pb level for women 

23 was lower than the limit of detection for Pb in the blood (2.1 μg/dL), effectively meaning 

24 that greater than 50% of women in the study had nondetectable levels of Pb in their 

25 blood. The study authors conducted a stratified analysis and concluded that TSH levels 

26 were negatively correlated with blood Pb in women who consumed fish contaminated 

27 with Pb and other environmental pollutants. No associations T3 and T4 levels were 
February 2012 5-499 Draft – Do Not Cite or Quote

1 reported in women. TSH, T3 and T4 levels were not observed to be correlated with blood 

2 Pb in males. However, study authors stated that occupational exposure to Pb in men can 

3 affect pituitary thyroid axis homeostasis and the relation between low-level Pb exposure 

4 thyroid hormone homeostasis in men and women needs to be investigated further. The 

5 authors also concluded that environmental contaminants not investigated (e.g., As) may 

6 be influencing TSH levels. Dundar et al. (2006) examined associations of blood Pb with 

7 thyroid function in 42 male adolescent auto repair workers exposed long term to Pb. A 

8 control group comprising 55 healthy subjects was also used for comparison purposes. 

9 Mean blood Pb levels were reported to be higher in the auto repair workers compared to 

10 the control subjects (mean [SD]: 7.3 [2.92] versus 2.08 [1.24] μg/dL). Free T4 (FT4) 

11 levels were significantly lower in the study group compared to the control group, which 

12 had no abnormal FT4 levels reported. In contrast, free T3 (FT3) and TSH levels were 

13 comparable between the study and control group. Blood Pb level was reported to be 

14 negatively correlated with FT4 levels. Based on the study outcome, the study authors 

15 reported that long-term Pb exposures that result in the studied blood Pb levels may lead 

16 to lower FT4 levels without impact on T3 and TSH levels in adolescents. The study 

17 authors stated that this effect is likely secondary to the toxic effects of Pb on the pituitary18 thyroid axis and to the earlier findings of primary hypothyroidism as a result of impaired 

19 production of peripheral thyroid hormones. Similar findings were reported by Croes et al.

20 (2009) in a study conducted in Belgium. Croes et al. (2009) examined the hormone levels 

21 in 1,679 adolescents residing in nine study areas with varying exposures to multiple 

22 industrial pollutants including Pb. The median concurrent blood Pb of the participants 

23 from the nine different regions ranged from 1.6 to 2.8 μg/dL. The study authors reported 

24 that, after adjustment for potential confounding, significant interregional differences were 

25 observed FT3 hormone levels. When individual neighborhoods were analyzed within the 

26 larger study areas, altered levels of FT3 levels were also observed. Though varying levels 

27 of FT3 levels were observed, the study authors reported that these changes were not 

28 wholly due to exposure to various pollutants, including Pb that were measured in the 

29 study and stated that other pollutants and environmental factors may also have 

30 contributed to the effects noted. In a prospective study of 309 mother-child pairs from 

31 Yugoslavia, the relationship between maternal TSH and T4 and blood Pb was 

32 investigated in those living in a highly exposed town with a smelter and battery plant (n = 

33 156 mother-child pairs) and those living in a relatively unexposed town (n = 153 mother34 child pairs) (Lamb et al., 2008). The mid-pregnancy blood Pb levels were highly elevated 

35 in the industrial town compared to the unexposed town (mean [SD]: 20.56 [7.38] versus 

36 5.60 [1.99] μg/dL). Mid-pregnancy maternal free T4 levels were observed to be inversely 

37 related with maternal blood Pb levels, but this association was not observed in the 

38 unexposed town. In 24 newborns delivered in Tokyo, Japan, neither TSH nor free T4 
February 2012 5-500 Draft – Do Not Cite or Quote

1 (sampled 4-6 days postpartum) was associated with cord blood Pb sampled at delivery 

2 (mean: 0.67 μg/dL) (Iijima et al., 2007).

3 Gump et al. (2008) examined cortisol response to acute stress in children (aged 9.5 years) 

4 whose prenatal and postnatal blood Pb levels had been determined prior to the study at 

5 birth (from cord blood) and at a mean (SD) age of 2.62 (1.2) years, respectively. For 

6 prenatal blood Pb, the children were divided into the following quartiles: ≤ 1, 1.1-1.4, 

7 1.5-1.9, and 2.0-6.3 μg/dL. For postnatal blood Pb, the quartiles were 1.5-2.8, 2.9-4.1, 

8 4.2-5.4, and 5.5-13.1 μg/dL. The study authors reported that blood Pb was not associated 

9 with initial salivary cortisol levels. However following an acute stressor, which 

10 comprised submerging the dominant arm for a minute in a gallon of one part ice to one 

11 part water, increasing prenatal and postnatal blood Pb levels were statistically 

12 significantly associated with increases in salivary cortisol responses. Children in the 2nd, 

13 3rd and 4th prenatal blood Pb quartiles and in the 4th postnatal quartile had increased 

14 salivary cortisol responses compared to children in the 1st quartile. When blood Pb was 

15 treated as a continuous variable, regression analysis showed that both prenatal and 

16 postnatal blood Pb levels were significantly correlated to salivary cortisol reactivity. 

17 Based on these results, the study authors reported that relatively low prenatal and 

18 postnatal blood Pb levels, notably those well below 10 μg/dL, can alter adrenocortical 

19 responses of children following acute stress and the health impact and behavioral aspects 

20 of this Pb-induced HPA deregulation in children needs to be further examined. 

21 In another study on the impact of Pb in children, Kemp et al. (2007) examined the blood 

22 Pb levels in 142 young, U.S. urban African-American and Hispanic children in winter 

23 and summer to determine the seasonal increase in blood Pb and its association with 

24 vitamin D (1,25-(OH)2D3), age and race. There was a winter/summer (W/S) increase in 

25 blood Pb levels in children aged between 1 and 3 years (mean [SE]: 4.94 [0.45] μg/dL in 

26 winter, 6.54 [0.82] μg/dL in summer), with a smaller W/S increase observed in children 

27 aged between 4 and 8 years (mean [SE]: 3.68 [0.31] μg/dL in winter, 4.16 [0.36] μg/dL 

28 in summer). Additionally, the winter and summer blood Pb levels were highly correlated 

29 with one another. The percentage of African-American children with blood Pb levels ≥ 

30 10 μg/dL increased from 12.2% in winter to 22.5% in summer. In children aged 4-8 

31 years, the concentrations of 1,25-(OH)2D3 were greater in the summer compared to the 

32 winter (mean [SE]: 33.8 [1.1] μg/L in summer versus 25.3 [1.2] μg/L in winter). No 

33 difference in seasonal 1,25-(OH)2D3 was observed in children 1-3 years old.

34 Additionally, winter and summer concentrations of 1,25-(OH)2D3 were highly correlated 

35 (r = 0.635, p < 0.0001). There was a significant correlation between seasonal differences 

36 in blood Pb and serum 1,25-(OH)2D3 in all children and African-American children 

37 between 4 and 8 years. Based on these results, the study authors concluded that higher 

38 summertime increase in serum 1,25-(OH)2D3 levels in children between 4 and 8 years is 
February 2012 5-501 Draft – Do Not Cite or Quote

1 most likely due to increased sunlight-induced vitamin D synthesis and may be a 

2 contributing factor to seasonal changes in blood Pb levels. 

5.9.3.3 New Toxicological Studies

3 . In a study examining the effects of Pb and cadmium in adult cows reared in a polluted 

4 environment in India, Swarup et al. (2007) stated that the mean plasma T3 and T4 levels 

5 were significantly higher in cows near Pb and zinc smelters (mean [SD] blood Pb: 86 

6 [6] μg/dL) and near closed Pb and operational zinc smelters (mean [SD] blood Pb: 51 

7 [9] μg/dL) when compared to cows in unpolluted areas (mean [SD] blood Pb: 7 

8 [1] μg/dL). Regression analyses from 269 cows examined in the study showed a 

9 significant positive correlation between blood Pb and plasma T3 and T4 levels, whereas 

10 the correlation between blood Pb and plasma cortisol was nonsignificant. Mean plasma 

11 estradiol level was significantly higher in cows near closed Pb and operational zinc 

12 smelter industries compared to the control group. Based on these results, the study 

13 authors concluded that endocrine profile in animals can be impacted following exposure 

14 to Pb in polluted environments. 

15 Biswas and Ghosh (2006) investigated the effect of Pb treatment on adrenal and male 

16 gonadal functions in Wistar rats treated with 8.0 mg/kg Pb-acetate via i.p. injection for 21 

17 days (blood Pb not reported). Pb treatment was observed to significantly increase adrenal 

18 steroidogenic enzyme activity and serum corticosterone levels. Accessory sex organ 

19 (prostate and seminal vesicle) weights were decreased in Pb-treated animals, whereas 

20 adrenal weights were increased. Spermatogenesis was decreased and the percent of 

21 spermatid degeneration was increased in animals treated with Pb. Lastly, serum 

22 concentrations of testosterone, FSH, and LH, were decreased in Pb-treated animals. 

23 Supplementation with testosterone during the last 14 days of Pb treatment was observed 

24 to ameliorate these effects.

5.9.4 Effects on Bone and Teeth

25 Primary effects on bone as a result of Pb exposure have included an increase in 

26 osteoporosis, increased frequencies of falls and fractures, changes in bone cell function as 

27 a result of replacement of bone calcium with Pb and depression in early bone growth. 

28 Similar to bone, calcium in the teeth can be easily substituted by Pb following Pb 

29 exposure. Exposure of animals to high levels of Pb (30 mg/kg body weight) may result in 

30 the formation of “Pb line” and Pb can also cause a decrease in cell proliferation, 

31 procollagen type I production, intracellular protein, and osteocalcin in human dental pulp 
February 2012 5-502 Draft – Do Not Cite or Quote

1 cell cultures. Accumulation of Pb was also associated with tooth loss and higher 

2 incidence of periodontitis.

5.9.4.1 Summary of Key Findings of the Effects on Bone and 

Teeth from the 2006 Lead AQCD

3 The 2006 Pb AQCD reported many effects on bone and some in teeth in animals and 

4 following exposure to Pb. Pb easily substituted for calcium in bone and was taken up by 

5 the bone causing changes in bone cell function. Exposure of animals to Pb during 

6 gestation and immediate postnatal period was reported to significantly depress early bone 

7 growth with the effects showing a concentration-dependent trends,. In mature animals, 

8 long-term exposure (up to one year) to Pb, along with poor nutrition (low calcium) 

9 affected bone growth as well bone density. Systemic effects of Pb exposure included 

10 disruption in bone mineralization during growth, alteration in bone cell differentiation 

11 and function due to alterations in plasma levels of growth hormones and calcitropic 

12 hormones such as 1,2-[OH]2D3 and impact on calcium binding proteins and increases in 

13 calcium and phosphorus concentrations in the bloodstream. Bone cell cultures exposed to 

14 Pb had altered vitamin D-stimulated production of osteocalcin accompanied by inhibited 

15 secretion of bone-related proteins such as osteonectin and collagen. In addition, Pb 

16 exposure caused suppression in bone cell proliferation most likely due to interference 

17 from factors such as growth hormone (GH), epidermal growth factor (EGF), transforming 

18 growth factor-beta 1 (TGF-β1), and parathyroid hormone-related protein (PTHrP).

19 As in bone, Pb can easily substitute for calcium in the teeth and is taken and incorporated 

20 into developing teeth in experimental animals. Since teeth do not undergo remodeling 

21 like the bone does during growth, most of the Pb in the teeth remains in a state of 

22 permanent storage. High dose exposure of Pb to animals (30 mg/kg body weight) has 

23 lead to the formation of a “Pb line” that is visible in both the enamel and dentin and is 

24 localized in areas of recently formed tooth structure. Areas of mineralization are easily 

25 evident in the enamel and the dentin within these “Pb lines.” Pb has also been shown to 

26 decrease cell proliferation, procollagen type I production, intracellular protein, and 

27 osteocalcin in human dental pulp cell cultures. Adult rats exposed to Pb have exhibited an 

28 inhibition of the posteruptive enamel proteinases, delayed teeth eruption times, as well as 

29 decrease in microhardness of surface enamel. Pb was reported to be widely dispersed and 

30 incorporated into developing apatite crystal during enamel formation process; however, 

31 post formation, Pb was reported to be capable of entering and concentrating in enamel

32 areas that were calcium deficient. The 2006 Pb AQCD also reported that a number of 

33 epidemiologic and animal studies have both separately suggested that Pb is a caries34 promoting element. 
February 2012 5-503 Draft – Do Not Cite or Quote

5.9.4.2 New Toxicological and Epidemiologic Studies

1 As reported in the 2006 Pb AQCD, Pb appears to be capable of causing effects in bones 

2 of humans and animals following exposure. The association between blood Pb levels and 

3 osteoporosis was examined in several epidemiologic studies. Most studies were cross4 sectional in design; thus, there is uncertainty regarding the magnitude, timing, frequency, 

5 and duration of Pb exposure that contributed to the observed associations. Campbell and 

6 Auigner (2007) examined subjects ≥ 50 years of age using NHANES III for association 

7 between concurrent blood Pb level and osteoporosis. The study authors used the bone 

8 mineral density in the hip as the primary outcome in groups comprising non-Hispanic 

9 white men (mean blood Pb: 4.9, range: 0.7 to 48.1 μg/dL), non-Hispanic white women 

10 (mean blood Pb: 3.6, range: 0.7 to 28.7 μg/dL), African-American men (mean blood Pb: 

11 7.7, range: 0.7 to 52.9 μg/dL), and African-American women (mean blood Pb: 4.5, range: 

12 0.7 to 23.3 μg/dL). The results indicated that the adjusted mean total hip bone mineral 

13 density in the non-Hispanic white males who had the lowest blood Pb levels (actual 

14 concentration not reported) was statistically significantly higher than that in the males 

15 with higher blood Pb levels. Similar associations, although not statistically significant, 

16 were reported among white females. Likely due to the small sample size, similar results 

17 were not observed among African-American men and women. No association was 

18 observed between blood Pb and osteoporotic fractures in any sex/race group. Since the 

19 NHANES study comprised a cross-sectional design, no inferences could be made 

20 regarding the temporal sequence of the observed association. The study authors 

21 concluded that further inquiry was needed to study the possible causal association 

22 between Pb exposure and osteoporosis. In a similar study, Sun et al. (2008b) examined 

23 the association between concurrent blood Pb levels and osteoporosis in 155 males and 37 

24 females in China occupationally-exposed to Pb (mean blood Pb: 20.22 and 15.5 μg/dL, 

25 respectively). Bone mineral density was reported to be statistically significantly lower in 

26 exposed females compared to exposed males. When all participants (including 36 male 

27 and 21 female unexposed controls) were divided into groups according to blood Pb and 

28 urinary Pb levels, the study authors reported that there were significant decreases in bone 

29 mineral density in groups that had high urinary Pb levels (≥ 5 μg/g creatinine) compared 

30 to groups with low urinary Pb in both sexes. In contrast, a significant difference was 

31 observed between blood Pb and bone mineral density only in males with blood Pb 

32 >30 μg/dL. Prevalence of osteoporosis was reported to increase significantly with 

33 increasing blood Pb in a linear manner. Khalil et al. (2008) reported similar associations 

34 between blood Pb level and osteoporosis in older women. The study authors conducted a 

35 prospective study using 533 women aged 65-87 years with a mean (SD) blood Pb of 5.3 

36 (2.3) μg/dL to determine the association between blood Pb and recurring fractures. 

37 Analysis of bone mineral density was conducted in 1986-1990 and 1993-1994, while 
February 2012 5-504 Draft – Do Not Cite or Quote

1 blood Pb was measured from 1990-1991. The bone mineral density was 7% lower in the 

2 total hip (p <0.02) and 5% lower in the femoral neck (p <0.03) in the highest blood Pb 

3 group (≥ 8 μg/dL) compared to the lowest blood Pb group (≤ 3 μg/dL). The trend across 

4 all dose groups was also observed to be statistically significant for hip and femoral neck 

5 bone mineral density. In addition, hip, femoral neck, and calcaneus bone loss was 

6 observed to be greater in the medium (blood Pb: 4-7 μg/dL) and high Pb groups 

7 compared to the low Pb group, but the observed trend was only significant for calcaneus 

8 bone loss. Multivariate analysis indicated that women with high blood Pb levels had an 

9 increased risk of non-spine fracture and women with medium or high blood Pb levels had 

10 a higher risk of falls compared to the low blood Pb level group. Based on these results, 

11 the study authors concluded that blood Pb is associated with an increased risk of falls and 

12 fractures leading to osteoporosis-related fractures. 

13 To examine the association between biomarkers of joint tissue metabolism and blood Pb 

14 levels, Nelson et al. (In Press) performed a cross-sectional analysis of 329 male and 342 

15 female participants in the Johnson County Osteoarthritis Project Metals Exposure Sub16 study. In women (mean age = 62 years), the median concurrent blood Pb level was 

17 1.9 μg/dL (range: 0.5-25.4 μg/dL). Blood Pb levels were higher in African-American 

18 compared to Caucasian women. Unadjusted correlation analyses demonstrated significant 

19 positive correlations between blood Pb and uNTX-I (a marker of bone 

20 resorption/turnover), uCTX-II (a marker associated with the progression of radiographic 

21 knee and hip osteoarthritis), and COMP (a cartilage biomarker related to osteroarthritis); 

22 however, significant associations only remained for uNTX-I and uCTX-II after adjusting 

23 for age, BMI, race, and smoking status. In men (median age = 65), the median blood Pb 

24 level was 2.2 μg/dL (range: 0.5-25.1 μg/dL). As with women, African-American men 

25 had higher blood Pb levels than did Caucasian men. In unadjusted correlation analyses, 

26 blood Pb levels in men were positively associated with uCTX-II, COMP, and C2C:CPII 

27 ratio (an indication of the balance between cartilage collagen degradation and synthesis), 

28 and negatively associated with CPII (a marker of collagen synthesis). After controlling 

29 for age, BMI, race, and smoking, only the positive association between blood Pb and 

30 COMP remained borderline statistically significant in men. The authors concluded that 

31 blood Pb is associated with bone turnover and mineralized cartilage turnover in women 

32 and non-mineralized cartilage turnover in men; however, as this study was cross-sectional 

33 in nature, it is impossible to conclude whether increased cartilage turnover is a product of 

34 increased blood Pb, or whether cartilage turnover itself results in increased Pb. Similarly, 

35 Machida et al. (2009) investigated bone matrix turnover rates in Japanese women related 

36 to menopause status and blood Pb. Perimenopausal women (n = 319) were observed to 

37 have significantly higher geometric mean blood Pb (2.0 μg/dL), than did premenopausal 

38 women (n = 261, blood Pb = 1.6 μg/dL), younger postmenopausal women (n = 397, 

39 blood Pb = 1.8 μg/dL), or older postmenopausal women (n = 248, blood Pb = 1.7 μg/dL). 
February 2012 5-505 Draft – Do Not Cite or Quote

1 In all subjects and perimenopausal women, markers of bone matrix turnover (osteocalcin 

2 [OC], bone-specific alkaline phosphatase [BALP], and N-telopeptide cross-linked 

3 collagen type I [NTx]) were significantly positively associated with blood Pb in 

4 unadjusted correlation analyses. In multivariate regression models controlling for bone5 mineral density, NTx, and age, OC was additionally statistically positively associated 

6 with blood Pb in all subjects and perimenopausal women. Bone mineral density and NTx 

7 were also positively associated with blood Pb in these models for all subjects. As with 

8 Nelson et al. (In Press), the cross-sectional nature of this study precludes a determination 

9 whether higher blood Pb is cause of increased bone matrix turnover biomarkers, or a 

10 consequence of increased bone turn over.

11 To understand the importance of bone as a target tissue of Pb toxicity, Jang et al. (2008)

12 studied the effect of Pb on calcium release activated calcium influx (CRACI) using 

13 primary cultures of human osteoblast-like cells (OLC). When cells were incubated with 

14 1,000 or 3,000 μM Pb, a concentration-dependent impact on the CRACI was observed, as 

15 was a concentration-dependent increase in the influx of Pb into human OLC. These 

16 results suggest that Pb interferes with CRACI in human OLCs by initiating the CRACI 

17 (i.e., the measurable influx of calcium upon re-addition of calcium is partially inhibited 

18 by Pb) and the influx of Pb is enhanced after CRACI is induced. Since studies have found 

19 associations between higher blood Pb level and reduced skeletal growth in children, 

20 Zuscik et al. (2007) conducted a study using murine limb bud mesenchymal cells (MSCs) 

21 to test the hypothesis that Pb alters chondrogenic commitment of mesenchymal cells and 

22 also to assess the effects of Pb on various signaling pathways. Exposure to 1 μM Pb 

23 caused increased basal and TGF-β/BMP induction of chondrogenesis in MSCs which was 

24 supported by nodule formation and upregulation of Sox-9, type 2 collagen, and aggrecan 

25 which are all key markers of chondrogenesis. The study authors also observed enhanced 

26 chondrogenesis during ectopic bone formation in mice that had been pre-exposed to Pb in 

27 drinking water (55 or 233 ppm, corresponding to 14 or 40 μg/dL blood Pb). MSCs 

28 exposed to Pb exhibited an increase in TGF-β, but BMP-2 signaling was inhibited. Pb 

29 was also reported to induce NF-κB and inhibit AP-1 signaling. Based on these results, the 

30 study authors concluded that chondrogenesis following exposure to Pb most likely 

31 involved modulation and integration of multiple signaling pathways including TGF-β, 

32 BMP, AP-1, and NF-κB.

33 Effects of Pb exposure on teeth were examined in a few epidemiologic studies. Since 

34 individuals may be impacted by the release of Pb stored in their skeletal compartments, 

35 Arora et al. (2009) examined the association between bone Pb concentrations and loss of 

36 natural teeth in 333 male participants of the NAS. Tooth loss in men was categorized as 

37 0, 1-8 or ≥ 9. Individuals with ≥ 9 teeth missing had significantly higher tibia and patella 

38 Pb concentrations (measured within 3 years of dental assessment) compared to those with 
February 2012 5-506 Draft – Do Not Cite or Quote

1 no tooth loss; no significant difference in blood Pb levels (measured within 3 years of 

2 dental assessment [(Hu et al., 1996a)]) was observed between the categories of teeth loss. 

3 Following adjustment for age, education, smoking status, pack-years of smoking, and 

4 diabetes, men with the highest tibia Pb concentrations (>23 μg/g) had higher odds of 

5 tooth loss (OR: 3.03 [95% CI: 1.60, 5.75]) compared to men with tibia Pb ≤ 15 μg/g. 

6 Men with the highest patellar Pb (>36 μg/g) also had higher odds of tooth loss (OR: 2.41 

7 [95% CI: 1.30, 4.49]) compared to men with patellar Pb ≤ 22.0 μg/g. Tooth loss was not 

8 statistically associated with blood Pb levels. Based on these results, the study authors 

9 concluded that long-term cumulative exposure to Pb is associated with increased odds of 

10 tooth loss. In a study examining the effects of Pb exposure on periodontitis in the U.S, 

11 Saraiva et al. (2007) analyzed data for 2,500 men and 2,399 women aged between 30 and 

12 56 years from NHANES III. The analysis took into account various covariates including 

13 age, NHANESIII phase, cotinine levels, poverty ration, race/ethnicity, education, bone 

14 mineral density, diabetes, calcium intake, dental visits, and menopause in women. After 

15 adjusting for these covariates and comparing individuals with a concurrent blood Pb level 

16 of >7 μg/dL to those with a blood Pb level of <3 μg/dL, the prevalence ratios of 

17 periodontitis was 1.70 (95% CI: 1.02, 2.85) for men and 3.80 (95% CI: 1.66, 8.73) for 

18 women. Based on these results, the study authors concluded that there was a positive 

19 association between periodontitis and blood Pb levels for both men and women. In a 

20 similar study, Yetkin et al. (2007) recruited 60 male subjects (30 apprentices with Pb 

21 exposure, 30 controls), to examine the impact of occupational exposure to Pb on 

22 periodontal status and association between periodontitis and blood Pb or oxidative stress. 

23 The results of their analysis indicated that blood Pb was significantly higher in 

24 apprentices exposed to Pb compared to controls (mean [SD]: 7.38 [4.41] versus 2.27 

25 [1.49] μg/dL, respectively). No clinical periodontal or oxidative stress parameters were 

26 significantly different between apprentices and controls. While the correlation between 

27 blood Pb and periodontal parameters was not reported, significant correlations between 

28 plaque index and CAT, probing depth and SOD, clinical attachment level and SOD, and 

29 clinical attachment level and malondialdehyde in Pb-exposed apprentices were observed. 

30 These results demonstrate that there is significant association between clinical 

31 periodontal parameters and oxidative stress/damage indices in Pb-exposed apprentices. In 

32 a multiple regression analysis, a statistically significant association between gingival 

33 index and working status, family income and either probing depth or clinical attachment 

34 level was noted. 
February 2012 5-507 Draft – Do Not Cite or Quote

5.9.5 Effects on Ocular Health

1 Ocular effects most commonly indicated to be associated with exposure to Pb include 

2 formation of cataract, impaired vision, edema and retinal stippling.

5.9.5.1 Summary of Key Findings of the Effects on Ocular Health 

from the 2006 Lead AQCD

3 The 2006 Pb AQCD stated that various changes in the visual system were observed with 

4 Pb poisoning including retinal stippling and edema, cataract, ocular muscle paralysis and 

5 impaired vision. The 2006 Pb AQCD reported that retinal responses were observed in 

6 children of mothers with a blood Pb range of 10.5 to 32.5 μg/dL during pregnancy, while 

7 cataracts were noted in middle-aged male with tibia bone Pb levels of 31-126 μg/g.

5.9.5.2 New Toxicological and Epidemiologic Studies

8 A small number of human studies pertaining to ocular effects of Pb were identified. 

9 Mosad et al. (2010) studied the association between subcapsular cataract and Pb, 

10 cadmium, vitamin C, vitamin E, and beta carotene blood levels in middle-aged male 

11 smokers compared to nonsmokers. Blood Pb was statistically significantly elevated in 

12 light (mean [SD]: 14.5 [0.41] μg/dL), moderate (14.5 [0.41] μg/dL), and heavy smokers 

13 (18.7 [1.24] μg/dL) compared to nonsmokers (12.2 [0.21] μg/dL). Blood Pb 

14 concentrations were also observed to be statistically higher in the cataracts of smokers 

15 versus nonsmokers. Similar associations were also observed for cadmium blood and lens 

16 levels, while vitamins C, E, and beta carotene levels were significantly decreased in 

17 smokers. Based on these results, the study authors concluded that the Pb and cadmium 

18 present in high concentration in smokers were associated with cataracts due to oxidative 

19 stress which was indicated by reduced levels of antioxidants such as vitamins C, and E 

20 and beta carotene. Erie et al. (2009) investigated the association between age-related 

21 macular degeneration and Pb and Cd in retinal tissue of human eye donors. The authors 

22 observed that Pb, but not Cd, was significantly elevated in the neural retina tissue of 

23 donors with age-related macular degeneration (n = 36 donors, 72 eyes; median [IQR]: 

24 12.0 [8-18] ng/g Pb) versus normal control donors (n = 25 donors, 50 eyes; median 

25 [IQR]: 8.0 [0-11] ng/g Pb). Neither heavy metal was significantly elevated in the retinal 

26 pigment epithelium-choroid complex in donors with age-related macular degeneration 

27 and normal controls.
February 2012 5-508 Draft – Do Not Cite or Quote

1 New animal studies pertaining to the ocular effects of Pb have investigated endpoints 

2 such as retinal progenitor cell proliferation and neurogenesis,(Section 5.3.4.3). An in 

3 vitro study was found that investigated whether exposure of cultured lenses from rats (4-6 

4 weeks age) to 1 μM Pb nitrate increased opacity with or without secondary oxidative 

5 challenge (Neal et al., 2010b). Lenses incubated for 3 days in the presence of 1 μM Pb 

6 were transparent, with no difference in amino acid incorporation compared to control 

7 lenses. The authors concluded this indicated that short-term Pb exposure does not induce 

8 osmotic swelling or lens shrinkage. However, exposure to lenses to Pb for 5 days 

9 dramatically decreased the percentage of transparent lenses (30%) compared to controls 

10 (80%). In Pb-exposed lenses, 30% displayed "definite cataracts" compared to only 2.5% 

11 in control lenses. By culture day 8, all exposed lenses were described either as clearly 

12 opaque or definite cataracts, while only 7% of control lenses displayed these 

13 characteristics, indicating that prolonged exposure of lenses to Pb induced an accelerated 

14 formation of opacity/cataract compared to unexposed lenses. Pb-exposed lenses cleared 

15 the media of hydrogen peroxide more rapidly than did control lenses, potentially due to 

16 increased CAT activity. Exposure to hydrogen peroxide resulted in total opacity in Pb17 exposed lenses at culture day 7, compared to less than 20% in control cells. Exposure to 

18 Pb additionally altered epithelial nutrient transport and lens histology relative to that in 

19 controls.

5.9.6 Effects on the Respiratory System

20 The collective body of toxicological and epidemiologic studies demonstrates Pb21 associated effects on multiple immunological pathways, including a shift from a Th1 to a 

22 Th2 phenotype, increased IgE antibody production, and increased inflammatory 

23 responses (Sections 5.2.5.1. and 5.6). These are well recognized pathways that contribute 

24 to increased susceptibility to infections and also to the development of respiratory 

25 diseases such as asthma. Recent investigation of the respiratory effects of Pb exposure 

26 has been limited; however, cross-sectional studies have indicated an association of 

27 increasing blood Pb level with increased prevalence of asthma in children 

28 (Section 5.6.4.2). As described in Section 5.2.4, Pb has been shown to induce the 

29 generation of ROS. ROS are implicated in mediating increases in bronchial 

30 responsiveness and activating neural reflexes leading to decrements in lung function. 

31 Studies investigating these airway responses also are limited in number and collectively 

32 do not provide strong evidence of an association with blood Pb (Section 5.6.6). 

33 Collectively, panel and time-series epidemiologic studies demonstrate associations 

34 between Pb measured in PM2.5 or PM10 air samples and decreases in lung function and 

35 increases in respiratory symptoms, and asthma hospitalizations in children but not adults 
February 2012 5-509 Draft – Do Not Cite or Quote

1 (Section 5.6.4.3). Toxicological studies have found pulmonary inflammation induced by 

2 CAPs in which Pb was one of numerous components (Wei et al., 2011; Duvall et al., 

3 2008; Godleski et al., 2002; Saldiva et al., 2002). Despite this evidence for respiratory 

4 effects related to air-Pb concentrations, it is important to note the limitations of air-Pb 

5 studies, including the limited data on the size distribution of Pb-PM (Section 3.5.3), the 

6 uncertain relationships of Pb-PM10 and Pb-PM2.5 with blood Pb levels, and the lack of 

7 adjustment for other correlated PM chemical components.

5.9.7 Summary

8 There is evidence from epidemiologic and toxicological studies that substantial exposure 

9 to Pb can result in altered liver function and hepatic toxicity. Biochemical changes 

10 indicative of liver injury, including decreases in serum protein and albumin levels and 

11 increased AST, ALT, ALP, and GGT activities, have been observed in occupationally12 exposed humans (blood Pb > 22 μg/dL) (Can et al., 2008; Khan et al., 2008; Patil et al., 

13 2007) mature animals exposed to high levels of Pb during adulthood (Sharma et al., 

14 2010a; Wang et al.; DS et al., 2009; Cheng et al., 2006), and animals exposed during 

15 gestation and lactation (Pillai et al., 2009; Teijon et al., 2006). In humans with mean 

16 blood Pb levels 5.4 μg/dL, altered AST levels were observed, but increases may be 

17 related to exposure to contaminants other than Pb (Conterato et al., In Press). Increased 

18 hepatic cholesterogenesis, altered triglyceride and phospholipid levels, and disorders in 

19 lipid metabolism accompanied by increased levels of total cholesterol and triglycerides 

20 have been reported in the animal literature (Ademuyiwa et al., 2009; Khotimchenko and 

21 Kolenchenko, 2007). These results suggest that induction of cholesterogenesis and 

22 phospholipids in the liver may cause subtle effects at the cellular level, leading to hepatic 

23 injury. Multiple studies in humans and animals have observed hepatic oxidative stress, 

24 generally indicated by an increase in lipid peroxidation along with a decrease in GSH 

25 levels and CAT, SOD, and GPx activities following exposure to Pb (Pandya et al., 2010; 

26 Sharma et al., 2010a; Khan et al., 2008; Yu et al., 2008; Adegbesan and Adenuga, 2007; 

27 Jurczuk et al., 2007; Khotimchenko and Kolenchenko, 2007; Jurczuk et al., 2006).

28 Indices of increased oxidative stress were additionally observed in the livers of fetuses 

29 exposed to Pb throughout gestation (Massó et al., 2007).

30 Relatively few human studies have been conducted on the gastrointestinal toxicity of Pb 

31 since the completion of the 2006 Pb AQCD. A case study reporting on GI symptoms in a 

32 child reported that elevated blood Pb was associated with nonlocalized abdominal pain, 

33 vomiting, nausea, constipation, lack of appetite, fatigue, and headaches (Cabb et al., 

34 2008). Symptoms were reported to diminish following cessation of exposure. Similar GI 

35 symptoms (stomach pain and gastritis) were observed in battery works and painters 
February 2012 5-510 Draft – Do Not Cite or Quote

1 exposed to Pb in India with blood Pb levels ranging from 0.4-116.6 μg/dL (Kuruvilla et 

2 al., 2006). Toxicological evidence for Pb-induced GI effects in rats includes inhibition of 

3 NANC relaxations in the gastric fundus and the observation of oxidative stress (lipid 

4 peroxidation, decreased SOD and CAT) in the gastric mucosa (Olaleye et al., 2007; 

5 Santos et al., 2006b). The observation of oxidative stress was accompanied gastric 

6 mucosal damage.

7 The endocrine processes most impacted by exposure to Pb include changes in thyroid 

8 function. FT4, but not FT3, was decreased in adolescent male auto repair workers

9 (Dundar et al., 2006) and mid-pregnancy T4 levels were negatively associated in women 

10 living in a highly contaminated town in Yugoslavia, whereas this association was not 

11 observed in a non-contaminated town (Lamb et al., 2008). Toxicological evidence for 

12 similar effects was found in adult cows reared in an environment contaminated with Pb. 

13 A positive correlation was reported between blood Pb and plasma T3 and T4 levels

14 (Swarup et al., 2007). In children challenged with an acute stressor, increasing prenatal 

15 maternal and age 2-year blood Pb levels were associated with significant increases in 

16 salivary cortisol responses at age 9 years, even with blood Pb levels less than 10 μg/dL

17 (Gump et al., 2008). A summary of key findings pertaining to reproductive hormones in 

18 males and females in the current document is presented in the section on reproductive and 

19 developmental effects (Sections 5.8.1 and 5.8.2).

20 Numerous epidemiologic studies investigated the association between Pb biomarkers and 

21 osteoporosis in adults. Higher blood Pb was observed to be associated with decreased 

22 bone mineral density in non-Hispanic white males (Campbell and Auinger, 2007),

23 whereas urinary Pb, but not blood Pb, was associated with decreased bone mineral 

24 density in Chinese individuals occupationally exposed to Pb (Sun et al., 2008b). In 

25 elderly women, blood Pb levels were positively associated with risk of falls and 

26 osteoporosis-related fractures (Khalil et al., 2008). Linear skeletal growth was reduced in 

27 children living near copper smelters and refiners (concurrent mean blood Pb level = 

28 7.7 μg/dL) (Ignasiak et al., 2006). In vitro studies indicate that Pb interferes with CARCI 

29 in human OLCs and that Pb perturbs multiple signaling pathways during murine limb bud 

30 growth, potentially resulting in altered skeletal development (Jang et al., 2008; Zuscik et 

31 al., 2007). Blood Pb levels have also been shown to be related to biomarkers of joint 

32 tissue metabolism in elderly populations, but the cross-sectional nature of these analyses 

33 prevents conclusions being drawn on whether Pb increases bone and joint turnover 

34 biomarkers, or whether increased turnover releases Pb into the bloodstream (Machida et 

35 al., 2009; Nelson et al., In Press). Epidemiologic studies investigating Pb exposure and 

36 tooth loss indicate that long-term, cumulative exposure to Pb is associated with increased 

37 odds of tooth loss, periodontitis in men and women, and that periodontitis is associated 
February 2012 5-511 Draft – Do Not Cite or Quote

1 with oxidative stress/damage in individuals exposed in an occupational setting (Arora et 

2 al., 2009; Saraiva et al., 2007; Yetkin-Ay et al., 2007).

3 New toxicology studies have reported ocular effects (i.e., retinal progenitor cell 

4 proliferation) due to Pb exposure (Section 5.3.4.3). and studies in humans report 

5 associations between heavy smoking, increased blood Pb levels, and cataracts (Mosad et 

6 al., 2010) and retinal Pb concentrations and age-related macular degeneration (Erie et al., 

7 2009). Investigation of the respiratory effects of Pb exposure has been limited; however, 

8 cross-sectional studies have indicated an association of higher blood Pb levels with 

9 increased prevalence asthma in children (Section 5.6.4.2). 

10 In summary, recent toxicological and epidemiologic evidence regarding the effects of Pb 

11 exposure on the liver, GI tract, endocrine system, bone and teeth, eyes, and respiratory 

12 tract largely are supportive of those effects noted in the 2006 Pb AQCD. However, recent 

13 evidence of these effects is relatively limited, and therefore no causal determinations are 

14 made regarding Pb-induced effects in these organ systems.

5.10 Cancer

15 The previous epidemiologic studies included in the 2006 Pb AQCD (U.S. EPA, 2006b)

16 “provide[d] only very limited evidence suggestive of Pb exposure associations with 

17 carcinogenic or genotoxic effects in humans” and the studies were summarized as 

18 follows:

“The epidemiologic data ...suggest a relationship between Pb exposure and cancers of the 

lung and the stomach... Studies of genotoxicity consistently link Pb-exposed populations 

with DNA damage and micronuclei formation, although less consistently with 

chromosomal aberrations.”

19 The International Agency for Research on Cancer (IARC) recently classified inorganic 

20 Pb compounds as probable human carcinogens (Group 2A of IARC classifications) based 

21 on stronger evidence in animal studies than human studies, and organic Pb compounds as 

22 not classifiable (Group 3 of IARC classifications) (IARC, 2006a; Rousseau et al., 2005).

23 Additionally, the National Toxicology Program has listed Pb and Pb compounds as 

24 “reasonably anticipated to be human carcinogens” (NTP, 2004). The typical cancer 

25 bioassays used by IARC or NTP as evidence of Pb-induced carcinogenicity used rodents 

26 that were continuously exposed to Pb-acetate in chow or drinking water for 18 months to 

27 two years in duration. These two year cancer bioassays and the doses administered are 

28 typical of cancer bioassays used with other chemicals.

29 In the following sections, recent epidemiologic and toxicological studies published since 

30 the 2006 Pb AQCD regarding Pb and cancer mortality and incidence are examined. In 
February 2012 5-512 Draft – Do Not Cite or Quote

1 addition, recent studies of Pb and DNA and cellular damage, as well as epigenetics 

2 studies, are summarized. When the information is available, the form of the Pb compound 

3 under study (e.g., inorganic, organic) is indicated. In epidemiologic studies, various 

4 biological measures of Pb are used including Pb measured in blood and bone. Bone Pb is 

5 indicative of cumulative Pb exposure. Blood Pb can represent more recent exposure, 

6 although it can also represent remobilized Pb occurring during times of bone remodeling. 

7 Toxicological studies only report exposure by blood Pb or exposure dose. More detailed 

8 discussion of these measures is given in Section 4.3.5.

5.10.1 Cancer Incidence and Mortality

9 Recent studies have included epidemiologic evaluations of the associations between Pb 

10 and both specific cancers, such as lung cancer and brain cancer, and overall cancer. Table 

11 5-42 provides an overview of the study characteristics and results for the epidemiologic 

12 studies that reported effect estimates. This section also evaluates toxicological evidence 

13 on the potential carcinogenicity of Pb. 
February 2012 5-513 Draft – Do Not Cite or Quote

Table 5-42 Summary of recent epidemiologic studiesa of cancer incidence and 

mortality

Referencea Study Location

Cancer 

Outcome Study Population

Measure of Pb 

Exposure Mean Pb (SD) Adjusted Effect Estimates

Cancer Mortality:

Menke et al.

(2006)

Multiple U.S. 

locations

Overall 

cancer 

mortality

NHANES III cohort 

with Blood Pb 

measures in 1988-

1994 

At least 12 years of 

follow-up

Blood Pb <10 μg/dL

Blood Pb at

baseline

2.58 μg/dL 

(geometric mean)

Tertile 1: <1.93 μg/dL

Tertile 2: 

1.94-3.62 μg/dL

Tertile 3: ≥ 3.63 

μg/dL

Tertile 1: 1.00

Tertile 2: 0.72 (95% CI: 0.46, 1.12)

Tertile 3: 1.10 (95% CI: 0.82, 1.47)

Schober et al.

(2006)

Multiple U.S. 

locations

Overall 

cancer 

mortality

NHANES III cohort

At least 40 years of 

age

Blood Pb at 

baseline

Blood Pb<5 μg/dL: 

67.7%

Blood Pb 5-9 μg/dL: 

26.0%

Blood Pb>=10 μg/dL: 

6.3%

Blood Pb<5 μg/dL: 1.00

Blood Pb 5-9 μg/dL: 1.44 (95% CI: 1.12, 1.86)

Blood Pb≥ 10 μg/dL: 1.69 (95% CI: 1.14, 2.52)

Note: Modification by age assessed and 

associations varied slightly 

Weisskopf et al.

(2009)

Boston, MA Overall 

cancer 

mortality

NAS

Included men only

Mean follow-up 

period for this study: 

8.9 yr 

Blood Pb at 

baseline, 

Patella Pb at 

baseline

Blood Pb: 5.6 μg/dL

(3.4)

Tertile 1 of Blood Pb: 

<4 μg/dL

Tertile 2 of Blood Pb: 

4-6 μg/dL

Tertile 3 of Blood Pb: 

>6 μg/dL

Tertile 1 of patella 

Pb: <22μg/g

Tertile 2 of patella 

Pb: 22-35μg/g 

Tertile 3 of patella 

Pb: >35 μg/g

Blood Pb Tertile 1: 1.00

Blood Pb Tertile 2: 1.03 (95% CI: 0.42, 2.55)

Blood Pb Tertile 3: 0.53 (95% CI: 0.20, 1.39)

Patella Pb Tertile 1: 1.00

Patella Pb Tertile 2: 0.82 (95% CI: 0.26, 2.59)

Patella Pb Tertile 3: 0.32 (95% CI: 0.08, 1.35)

Khalil et al.

(2009a)

Baltimore, 

MD, and 

Monongahela 

Valley, PA

Overall 

cancer 

mortality

Subgroup of the 

Study of 

Osteoporotic 

Fractures cohort

Included white 

women aged 65-87;

12 yr (+/- 3 yr) 

follow-up

Blood Pb at 

baseline

Blood Pb Level 

5.3 (2.3)μg/dL

Blood Pb<8 μg/dL: 1.00

Blood Pb≥ 8 μg/dL: 1.64 (95% CI: 0.73, 3.71)

Lung Cancer:

Lundstrom et al.

(2006)

Sweden Lung cancer 

(incidence 

and mortality)

Male Pb smelter 

workers first 

employed for ≥ 3

months between 

1928 and 1979

Followed up for 

mortality from 1955 -

1987

Median peak 

blood Pb level

Median number 

of yr with at least 

one blood sample 

obtained

Median 

cumulative blood 

Pb index (sum of 

annual blood Pb 

Level)

Median peak blood 

Pb Level: cases 2.4 

μmol/L, controls 2.7 

μmol/L

Median number of yr 

with at least one 

blood sample 

obtained: cases 4.5 

yr, controls 6.0 yr

Median cumulative 

blood Pb index: 

cases 9.0 μmol/Pb, 

controls 11.9 

μmol/Pb

Median peak blood Pb Level: 1.00 (95% CI: 

0.71, 1.42)

Median number of yr with at least one blood 

sample obtained: 0.96 (95% CI: 0.91, 1.02) 

per μmol/L

Median cumulative Blood Pb index: 0.99 (95% 

CI: 0.96, 1.02) per μmol/L

Note: similar results were observed when 

restricted to smokers only 
February 2012 5-514 Draft – Do Not Cite or Quote

Referencea Study 

Location

Cancer 

Outcome Study Population

Measure of Pb 

Exposure Mean Pb (SD) Adjusted Effect Estimates

Jones et al.

(2007)

Humberside, 

UK

Lung cancer 

mortality

Male tin smelter 

employees

Personnel record 

cards and air 

sampling 

conducted from 

1972-1991

Three exposure 

scenarios 

determined for 

working lifetime 

cumulative 

exposure – all 

have similar 

medians of 

approximately 2 

mg yr/m3

NA RR for Pb exposure weighted age and time 

since exposure: 1.54 (90% CI: 1.14, 2.08)

Note: Similar results for other exposure 

determination scenarios.

Rousseau et al.

(2007)

Montreal, 

Canada

Lung cancer 

and other 

cancer 

incidence

Men aged 35-79 Interview of job 

history and 

exposure matrix

Ever exposed to:

Organic Pb 3.0%

Inorganic Pb 17.0%

Pb in gasoline 

emissions 38.6%

Organic Pb exposure compared to no 

exposure: 

Lung 1.3 (95% CI: 0.5, 3.1)

Inorganic Pb exposure compared to no 

exposure: 

Lung 1.1 (95% CI: 0.7, 1.7)

Pb in gasoline emissions exposure compared 

to no exposure: 

Lung 0.8 (95% CI: 0.6, 1.1)

Note: results are for comparisons using 

population-based controls; results for controls 

with other types of cancers were similar 

Brain Cancer:

van Wijngaarden 

and Dosemeci

(2006)

Multiple U.S

locations

Brain cancer 

mortality

National 

Longitudinal 

Mortality Study – included individuals 

with occupational 

information

-included follow-up 

from 1970-1989

Interview about 

current or most 

recent job within 

the past 5 years 

and a job 

exposure matrix

NA Any Pb exposure compared to no exposure 

1.56 (95% CI: 1.00, 2.43)

Note: HRs were greatest among those with 

high probabilities of exposure and 

medium/high exposure intensity

Rajaraman et al.

(2006)

Phoenix, AZ, 

Boston, MA, 

and 

Pittsburgh, 

PA

Brain cancer 

incidence

NCI Brain Tumor 

Study

– included 

individuals >=18 yr 

diagnosed with brain 

cancer less than 8 

wk before 

hospitalization; 

frequency-matched 

controls were 

individuals admitted 

to the same 

hospitals for nonneoplastic 

conditions

Interviews of 

lifetime work 

history and 

exposure 

databases

NA Meningioma:

Ever exposure to Pb 0.8 (95% CI: 0.5, 1.3)

Glioma: Ever exposure to Pb 0.8 (95% CI: 0.6, 

1.1)

Note: positive associations between Pb 

exposure and meningioma incidence was 

observed among individuals with ALAD2 

genotypes, but not individuals with ALAD1 

genotypes; these associations were not 

observed for glioma incidence
February 2012 5-515 Draft – Do Not Cite or Quote

Referencea Study 

Location

Cancer 

Outcome Study Population

Measure of Pb 

Exposure Mean Pb (SD) Adjusted Effect Estimates

Bhatti et al.

(2009)

Phoenix, AZ, 

Boston, MA, 

and 

Pittsburgh, 

PA

Brain cancer 

incidence

NCI Brain Tumor 

Study 

– included nonHispanic whites ≥

18 yr diagnosed 

with brain cancer 

less than 8 wk 

before 

hospitalization; 

frequency-matched 

controls were 

individuals admitted 

to the same 

hospitals for nonneoplastic 

conditions

Interviews of 

lifetime work 

history and 

exposure 

databases

Glioma: 70.5 μg/m3y 

(193.8 μg/m3

y)

Glioblastoma 

multiform: 97.5 

μg/m3y (233.9 

μg/m3

y)

Meningioma: 101.1 

μg/m3

y (408.7 

μg/m3

y)

Controls: 69.7 

μg/m3

y (248.8 

μg/m3

y) 

Per 100 μg/m3y increase in cumulative Pb 

exposure

Glioma: 1.0 (95% CI: 0.9, 1.1)

Glioblastoma multiform: 1.0 (95% CI 0.9, 1.1)

Meningioma: 1.1 (95% CI: 1.0, 1.2)

Note: modification by SNPs was conducted 

and associations varied by SNP 

Breast Cancer:

Pan et al.

(2011)

Canada Breast 

cancer 

incidence

National Enhanced 

Cancer Surveillance 

System (NECSS) –

population-based 

sample of cancer 

cases and controls 

with information 

collected from 1994-

1997

Self-reported 

previous 

addresses and 

their proximity to 

Pb smelters 

(determined 

using 

Environmental 

Quality Database 

[EQDB])

NA Residing >3.2 km from Pb smelter or no 

nearby smelter: 1.00

Residing 0.8-3.2 km from Pb smelter: 0.41 

(95% CI: 0.11, 1.51)

Residing <0.8 km from Pb smelter: 0.61 (95% 

CI: 0.11, 3.42)

Multiple and Other Cancers:

Absalon and 

Slesak (2010)

Silesia 

province, 

Poland

Overall 

cancer 

incidence

Children living in this 

province at least five 

years 

Pb-related air 

pollution 

measures

NA Reported correlations between changes in Pb 

and cancer incidence – no/low correlations 

observed (correlation coefficients between -0.3 

and 0.2)

Obhodas et al.

(2007)

Island of Krk, 

Croatia

Incidence 

rates for 

neoplasms

Individuals living in 

the Island of Krk 

from 1997-2001

Soil and 

vegetation 

samples, 

household 

potable water 

samples, 

children’s hair 

samples

NA No association observed between Pb in the 

samples and incidence of neoplasm 

(numerical results not provided)

Mendey et al. 

(In Press)

Multiple U.S. 

locations

Incidence of 

cancer or 

“malignancy 

of any kind”

2007-2008 

NHANES cohort – at 

least 20 years of 

age

Concurrently 

measured 

creatinineadjusted urinary 

Pb

Geometric mean for 

creatinine-adjusted 

urinary Pb marker: 

0.59 μg/g (95% CI: 

0.57, 0.61)

Greater than log-transformed mean creatinine- adjusted urinary Pb level compared to less 

than log-transformed mean creatinineadjusted urinary Pb level: 0.76 (95% CI: 0.44, 

1.33)
February 2012 5-516 Draft – Do Not Cite or Quote

Referencea Study 

Location

Cancer 

Outcome Study Population

Measure of Pb 

Exposure Mean Pb (SD) Adjusted Effect Estimates

Rousseau et al.

(2007)

Montreal, 

Canada

Lung cancer 

and other 

cancer 

incidence

Men aged 35-79 Interview of job 

history and 

exposure matrix

Ever exposed to:

Organic Pb 3.0%

Inorganic Pb 17.0%

Pb in gasoline 

emissions 38.6%

Never exposed is referent group

Organic Pb: 

Esophageal 1.7 (95% CI: 0.5, 6.4)

Stomach 3.0 (95% CI: 1.2, 7.3)

Colon 1.5 (95% CI: 0.7, 3.6)

Rectum 3.0 (95% CI: 1.2, 7.5)

Pancreas 0.9 (95% CI: 0.1, 5.2)

Prostate 1.9 (95% CI: 0.8, 4.6)

Bladder 1.7 (95% CI: 0.7, 4.2)

Kidney 2.3 (95% CI: 0.8, 6.7)

Non-Hodgkin’s lymphoma 0.4 (95% CI: 0.1, 

2.2)

Inorganic Pb: 

Esophageal 0.6 (95% CI: 0.3, 1.2)

Stomach 0.9 (95% CI: 0.6, 1.5)

Colon 0.8 (95% CI: 0.5, 1.1)

Rectum 0.8 (95% CI: 0.5, 1.3)

Pancreas 0.9 (95% CI: 0.4, 1.8)

Prostate 1.1 (95% CI: 0.7, 1.6)

Bladder 1.1 (95% CI: 0.7, 1.5)

Kidney 1.0 (95% CI: 0.6, 1.7)

Melanoma 0.4 (95% CI: 0.2, 1.0)

Non-Hodgkin’s lymphoma 0.7 (95% CI: 0.4, 

1.2)

Pb in gasoline emissions: 

Esophageal 0.6 (95% CI: 0.4, 1.1)

Stomach 1.0 (95% CI: 0.7, 1.4)

Colon 0.8 (95% CI: 0.6, 1.1)

Rectum 1.0 (95% CI: 0.7, 1.4)

Pancreas 0.9 (95% CI: 0.5, 1.4)

Prostate 0.9 (95% CI: 0.7, 1.2)

Bladder 0.8 (95% CI: 0.6, 1.1)

Kidney 1.0 (95% CI: 0.7, 1.5)

Melanoma 0.8 (95% CI: 0.5, 1.4)

Non-Hodgkin’s lymphoma 0.7 (95% CI: 0.5, 

1.0)

Note: results are for comparisons using 

population-based controls; results for controls 

with other types of cancers were similar 

except no association was present between 

organic Pb and rectal cancer
February 2012 5-517 Draft – Do Not Cite or Quote

Referencea Study 

Location

Cancer 

Outcome Study Population

Measure of Pb 

Exposure Mean Pb (SD) Adjusted Effect Estimates

Santibanez et al.

(2008)

Valencia and 

Alicante, 

Spain

Esophageal 

cancer 

incidence

PANESOES study 

included 30-80 yr 

old men hospitalized 

in any of the 

participating study 

hospitals

Interviews to 

determine 

occupational 

history and a job 

exposure matrix

NA All esophageal cancers:

Unexposed: 1.00

Low workplace Pb exposure (≤ 0.237 μmol/L): 

0.79 (95% CI: 0.43, 1.46)

High workplace Pb exposure (>0.237 μmol/L): 

1.69 (95% CI: 0.57, 5.03)

Esophageal squamous cell carcinoma:

Unexposed: 1.00

Low workplace Pb exposure (≤ 0.237 μmol/L): 

0.70 (95% CI: 0.34, 1.43)

High workplace Pb exposure (>0.237 μmol/L): 

0.91 (95% CI: 0.22, 3.75)

Adenocarcinoma:

Unexposed: 1.00

Low workplace Pb exposure (≤ 0.237 μmol/L): 

0.95 (95% CI: 0.32, 2.82)

High workplace Pb exposure (>0.237 μmol/L): 

5.30 (95% CI: 1.39, 20.22)

*associations not changed or slightly 

increased when restricted to occupational 

exposures ≥ 15yr

a

Studies listed in order of appearance in the text.

5.10.1.1 Overall Cancer Mortality

1 Several recent epidemiologic studies examined the association between Pb levels and 

2 cancer mortality, including multiple analyses of the NHANES III population. In one 

3 NHANES III analysis, the cohort was followed for 12 years and individuals with blood 

4 Pb levels greater than 10 μg/dL were excluded from the study (mean baseline blood Pb 

5 level was 2.58 μg/dL). No association was observed between blood Pb and cancer 

6 mortality (HR of highest tertile [≥ 3.63 μg/dL]compared to lowest tertile [<1.93 μg/dL]: 

7 1.10 [95% CI: 0.82, 1.47]) (Menke et al., 2006). Another analysis of the NHANES III 

8 population, which was restricted to individuals 40 years and older at the time of blood Pb 

9 collection and included individuals with all blood Pb levels (including those greater than 

10 10 μg/dL), reported associations between blood Pb and cancer mortality (Schober et al., 

11 2006). In this study, median follow-up time was 8.6 years. The RRs were 1.69 (95% CI: 

12 1.14, 2.52) for individuals with blood Pb levels of at least 10 μg/dL and 1.44 (95% CI: 

13 1.12, 1.86) for blood Pb levels of 5-9 μg/dL compared to individuals with blood Pb levels 

14 less than 5 μg/dL. When stratified by age, point estimates comparing blood Pb levels 

15 of 5-9 versus less than 5 μg/dL were similar across all age groups but only statistically 

16 significant among 75-84 year olds. The odds of mortality associated with blood Pb levels 

17 ≥ 10 μg/dL in the groups aged 40-74 years and 85 years and older were elevated. A study 

18 of men from the greater Boston area enrolled in the NAS found no association between 
February 2012 5-518 Draft – Do Not Cite or Quote

1 blood or bone Pb and cancer mortality in adjusted analyses. The mean (SD) blood Pb 

2 level for this population was 5.6 (3.4) μg/dL and blood Pb was poorly correlated with 

3 measured bone Pb (Weisskopf et al., 2009). As part of the Study of Osteoporotic 

4 Fractures , white women aged 65-87 were included in a sub-study of blood Pb level and 

5 cancer mortality and were followed for approximately 12 years (Khalil et al., 2009a). The 

6 mean (SD) blood Pb levels were 5.3 (2.3) μg/dL and no association was detected between 

7 blood Pb and cancer mortality in the study population. 

8 Overall, epidemiologic studies of blood Pb levels and cancer mortality reported 

9 inconsistent results. One epidemiologic study using NHANES III data demonstrated an 

10 association between blood Pb and increased cancer mortality; however, other studies 

11 reported weak or no associations. 

5.10.1.2 Lung Cancer

12 Most of the evidence regarding lung cancer incidence is provided by studies of 

13 occupationally-exposed adults. In a study of smelter workers, no association was 

14 observed between several metrics of Pb exposure (peak blood Pb values, number of years 

15 Pb samples were obtained, and cumulative blood Pb index) and lung cancer incidence 

16 and mortality combined (Lundstrom et al., 2006). The median follow-up in the study was 

17 about 30 years and the median peak blood Pb values during employment were 49.7 μg/dL 

18 for lung cancer cases and 55.9 μg/dL for controls. In a study of tin smelters workers, no 

19 association was observed between Pb exposure and lung cancer mortality in unweighted 

20 analyses, but when the analyses were weighted by age and time since exposure, positive 

21 associations were apparent (Jones et al., 2007). In this study, Pb exposure was calculated 

22 by combining historical air sampling data and personnel record cards, which specified 

23 work histories. The median Pb exposure was estimated to be approximately 2 mgyear/m3 24 . It is important to note that the smelter workers were exposed to other metals as 

25 well, such as arsenic and antimony. A population-based case-control study 

26 performed among men in Montreal, Canada assessed Pb exposure via interviews 

27 regarding job histories and calculated the likely Pb exposures associated with the job 

28 activities (Rousseau et al., 2007). No association was apparent between organic Pb, 

29 inorganic Pb, or Pb from gasoline emissions and lung cancer. 

30 Studies of Pb and lung cancer that compared the lung tissue of individuals with lung 

31 cancer to those without lung cancer were also conducted. The controls for these studies 

32 were individuals with metastases in the lung from other primary cancers (De Palma et al., 

33 2008) and individuals with non-cancerous lung diseases (De Palma et al., 2008; Kuo et 

34 al., 2006). Findings are mixed among the studies. De Palma et al. (De Palma et al., 2008)
February 2012 5-519 Draft – Do Not Cite or Quote

1 reported higher Pb concentrations in the cancerous and non-cancerous lung tissue of 

2 individuals with non-small cell lung cancer compared to control groups (although the 

3 authors report these results may be confounded by smoking). Kuo et al. (Kuo et al., 2006)

4 found no statistical difference in Pb levels for lung tissue of individuals with lung cancer 

5 compared to controls . 

6 Some studies in the 2006 Pb AQCD reported associations between Pb exposure and lung 

7 cancer; however, more recent epidemiologic studies of lung cancer reported no 

8 associations. Overall, these recent epidemiologic studies included only men, limiting the 

9 generalizability. The studies by Jones et al. (2007) and Rousseau et al. (2007) also have 

10 the disadvantage of not obtaining actual measures of Pb levels. In addition, studies 

11 limited to occupational exposures may be confounded by other workplace exposures.

5.10.1.3 Brain Cancer

12 A few studies of brain cancer examined the association between cancer and Pb using 

13 exposures determined via exposure databases and patient interviews about past jobs and 

14 known exposures. Interpretation of these results is limited due to the lack of biological Pb 

15 measures and the potential confounding by other occupational exposures. The National 

16 Longitudinal Mortality Study, a study that included a national sample of the U.S. 

17 population, estimated Pb exposure based on current/most recent employment among 

18 individuals (Van Wijngaarden and Dosemeci, 2006). Although not all estimates are 

19 statistically significant, a pattern of increased associations between Pb exposure and brain 

20 cancer mortality was observed in the study population. In a case-control study of brain 

21 tumors, glioma was reported to have no association with any Pb exposure metric; 

22 however, positive associations were observed between high cumulative Pb exposure and 

23 meningioma among individuals with ALAD2 genotypes (OR 2.4 [95% CI 0.7, 8.8] 

24 comparing individuals ever exposed to Pb with those not exposed to Pb; OR 12.8 [95% 

CI 1.4, 120.8] comparing individuals with cumulative Pb exposure ≥ 100 μg/m3 25 y to those 

26 not exposed to Pb) (Rajaraman et al., 2006). This association was not present among 

27 individuals with the ALAD1 genotypes (OR 0.5 [95% CI 0.3, 1.0] comparing individuals 

28 ever exposed to Pb with those not exposed to Pb; OR 0.7 [95% CI 0.2, 1.8] comparing 

individuals with cumulative Pb exposure ≥ 100 μg/m3 29 y to those not exposed to Pb). 

30 Another study of the association between Pb exposure (measured using self-reported 

31 occupational exposure history) and brain tumors reported none or slight overall 

32 associations with types of brain tumors; however, positive associations were observed 

33 among individuals with certain genetic single nucleotide polymorphisms (SNPs) (Bhatti 

34 et al., 2009). After control for multiple comparisons, individuals with GPX1 variants 

35 (rs1050450) had positive associations between cumulative Pb exposure and glioblastoma 
February 2012 5-520 Draft – Do Not Cite or Quote

1 multiforme and meningioma. Individuals without RAC2 variants (rs2239774) showed a 

2 positive association between Pb and glioblastoma multiforme. Also, individuals without 

3 XDH variants (rs7574920) displayed a positive association between Pb and meningioma.

4 Overall, associations between Pb exposure and brain cancer incidence and mortality were 

5 found to vary according to several genetic variants. These studies were limited in their 

6 methods because they do not have individual level biological measures and the potential 

7 for confounding by other workplace exposures exist. Future research will be important in 

8 confirming these associations and the modification by various genetic variants.

5.10.1.4 Breast Cancer

9 A population-based case-control study in Canada examined the proximity to a Pb smelter 

10 based on residential addresses (Pan et al., 2011). No association was reported between 

11 proximity of a Pb smelter and breast cancer incidence, but the study was limited by the 

12 small number of women who resided near a Pb smelter (n=13 lived ≤ 3.2 km from Pb 

13 smelter).

14 A few studies examined Pb levels and breast tumors among individuals with and without 

15 breast tumor and/or cancer present. A study of newly diagnosed breast cancer patients 

16 and controls examined Pb levels in blood and hair samples and reported higher levels of 

17 both for cancer cases, although the difference in the Pb content in hair samples was not 

18 statistically significant (Alatise and Schrauzer, 2010). Siddiqui et al. (2006) observed 

19 higher blood Pb levels in women with benign and malignant tumors compared to 

20 controls. Additionally, although blood Pb levels were higher among those with malignant 

21 breast tumors compared to those with benign tumors, both had similar levels of Pb 

22 detected in breast tissues. Another study of Pb levels present in breast tissue also reported 

23 no statistical difference in Pb levels (Pasha et al., 2008b). However, one study of breast 

24 tissue did observe a statistically significant difference between Pb levels in the breast 

25 tissue of cancer cases and controls (Ionescu et al., 2007). Finally, a study of Pb levels in 

26 urine reported a positive association between urine Pb and breast cancer, but this 

27 association became null when women taking nonsteroidal aromatase inhibitors but not 

28 taking bisphostphonates (a combination responsible for bone loss) were excluded from 

29 the analysis (McElroy et al., 2008).

30 Overall, these studies demonstrate the possibility that women with breast cancer may 

31 have higher Pb levels in blood measurement, whereas the results for actual breast tissue 

32 are mixed. However, these studies are limited by their study design. The samples are 

33 taken after cancer is already present in the cases, leading to issues of temporality for the 
February 2012 5-521 Draft – Do Not Cite or Quote

1 Pb levels. Additionally, the sample sizes are often small and the studies may be 

2 underpowered. 

5.10.1.5 Other Cancers

3 Studies of all cancers combined, multiple different cancers, or cancers not listed above 

4 have also been performed. An ecologic analysis compared levels of Pb in the air from 

5 1990 to 2005 with incidence rates of cancer (cancer sites not specified) among children 

6 during this time period (Absalon and Slesak, 2010). The highest Pb levels were measured 

in 1990 when over 50% of the study area exceeded the limit of 1 μg/m2 7 ∙year. No 

8 correlation was observed both overall and in sex-specific analyses. A similar study 

9 examined correlations between Pb concentrations in soil, water, vegetation, and hair 

10 samples with incidence of neoplasms (Obhodas, 2007). The Pb concentrations were not 

11 correlated with incidence of neoplasms. A recent study using the 2007-2008 NHANES 

12 cohort reported no association between higher creatinine-adjusted urine Pb levels and 

13 having ever had cancer or a malignancy (Mendy et al., In Press). The timing of cancer 

14 diagnosis in relation to the urine sample collection was not identified.

15 A study performed among men evaluated multiple cancer outcomes and determined 

16 exposures to organic Pb, inorganic Pb, and Pb from gasoline emissions via interviews 

17 regarding job histories and then subsequent exposure approximations by chemists and 

18 hygienists (Rousseau et al., 2007). Adults exposed to organic Pb exposure had greater 

19 odds of stomach cancer compared to adults never exposed to organic Pb. A positive 

20 association was also observed for rectal cancer when population-based controls were 

21 used but was null when the control population was limited to individuals with other types 

22 of cancers. No association was detected for cancers of the esophagus, colon, pancreas, 

23 prostate, bladder, kidney, melanoma, or non-Hodgkin’s lymphoma. None of the cancers 

24 were associated with exposure to inorganic Pb. When occupational exposure to Pb in 

25 gasoline was categorized as unexposed, nonsubstantial level, and substantial level, a 

26 positive association with stomach cancer was observed when cancer controls were used 

27 as the comparison group; however the association was not present when population 

28 controls were utilized as the control group). Another case-control study using participant 

29 interviews and a job exposure matrix, including only men, reported no association 

30 between Pb exposure and esophageal squamous cell carcinomas, but an association was 

31 present between high Pb exposure and adenocarcinoma of the esophagus (Santibanez et 

32 al., 2008). However, neither of these studies was able to quantify Pb levels using 

33 biological measurements.
February 2012 5-522 Draft – Do Not Cite or Quote

1 Several studies compared Pb levels in blood, tissue, and urine of individuals who have 

2 cancer with individuals who are cancer-free. Compared to control groups, higher Pb 

3 levels were observed in the blood and bladder tissue of individuals with bladder cancer

4 (Golabek et al., 2009), the kidney tissue of individuals with renal cell carcinoma (with 

5 highest levels among those with the highest stage tumors) (Calvo et al., 2009), the tissue 

6 (but not serum) of individuals with laryngeal cancer (Olszewski et al., 2006), the blood of 

7 individuals with gastric cancer (Khorasani et al., 2008), the plasma and hair of 

8 individuals with gastrointestinal cancer (Pasha et al., 2010), the blood and hair of 

9 individuals with non-specified types of cancer (Pasha et al., 2008c; Pasha et al., 2007),

10 and the hair of individuals with benign tumors (Pasha et al., 2008a). No statistical 

11 difference in Pb levels was reported for colon tissue of individuals with colorectal polyps

12 (Alimonti et al., 2008) or urine of individuals with bladder cancer (Lin et al., 2009)

13 compared to control groups. A study examining Pb levels in kidney tissue reported the 

14 highest levels of Pb in normal kidney tissue samples that were adjacent to neoplastic 

15 tumors. The Pb levels reported in the kidney tissue of neoplastic tumors were elevated 

16 compared to those detected in corpses without neoplastic tumors of the kidney (Cerulli et 

17 al., 2006). All of these comparison studies are limited by the inability to examine 

18 temporality as Pb biomarkers were measured after the cancer diagnosis; the level of Pb 

19 may be due to changes that result from having cancer, not changes that result in cancer. 

20 Many of these studies attempted to control for this by including only cases who have not 

21 undergone certain treatments. Additionally, studies are limited by their small sample size 

22 and the selection of the control populations. Control populations are supposed to 

23 represent the general population from which the cases are drawn; some of the control 

24 subjects in these studies are individuals with diseases/conditions warranting tissue 

25 resections, which are not prevalent in the general population. 

26 In sum, epidemiologic studies reported no associations between various measures of Pb 

27 exposure and overall cancer incidence. Studies examining specific cancers reported 

28 varying associations. Associations were null for Pb and most cancer sites examined; 

29 however a positive association was observed between Pb exposure and adenocarcinoma 

30 of the esophagus as well as exposure to organic Pb and stomach cancer. Associations 

31 between organic Pb exposure and rectal cancer and exposure to Pb in gasoline and 

32 stomach cancer were inconsistent. These conclusions are limited by the small number of 

33 studies and a lack of biological measurements of Pb.
February 2012 5-523 Draft – Do Not Cite or Quote

5.10.1.6 Toxicological Models of Carcinogenicity

Carcinogenicity in Animal Models

1 Previous AQCDs have established that Pb has been shown to act as a carcinogen in 

2 animal toxicology models, albeit at relatively high concentrations. The 2006 Pb AQCD 

3 pointed out that because Pb is a "well-established animal carcinogen...., focus has been 

4 more on the mechanism of neoplasia and possible immunomodulatory effects of Pb in the 

5 promotion of cancer." This focus continues to date. The kidneys are the most common 

6 target of Pb-dependent carcinogenicity (Kasprzak et al., 1985; Koller et al., 1985; Azar et 

7 al., 1973; Van Esch and Kroes, 1969) but the testes, brain, adrenals, prostate, pituitary, 

8 and mammary gland have also been affected (IARC, 2006a). The typical cancer 

9 bioassays used by IARC or NTP as evidence of Pb-induced carcinogenicity were 

10 designed using rodents, typically males but sometimes animals of both sexes, that were 

11 continuously exposed to Pb-acetate in chow (i.e., 0.1 or 1% Pb-acetate) or drinking water 

12 (i.e., 26 or 2,600 ppm Pb-acetate) for 18 months to two years in duration (Kasprzak et al., 

13 1985; Koller et al., 1985; Azar et al., 1973; Van Esch and Kroes, 1969). These two year 

14 cancer bioassays and the doses employed are typical of cancer bioassays employed by 

15 other chemicals. Recognition of the importance of windows of exposure in Pb-induced 

16 cancer bioassays is a focus of more recent studies. In one study, early life gestational and 

17 lactational exposure of laboratory rodents to inorganic Pb induced carcinogenicity in 

18 adult offspring (Waalkes et al., 1995). Another recent study considered Pb-dependent 

19 carcinogenesis in laboratory animals with early life Pb exposure. Tokar et al. (2010)

20 considered tumorigenesis in homozygous metallothionein I/II knockout mice and their 

21 corresponding wild type controls (groups of ten mice each) that were exposed by 

22 drinking water to 2,000 or 4,000 ppm Pb-acetate and compared to untreated controls. 

23 Study animals were exposed in utero, through birth and lactation, and then postnatally to 

24 drinking water until 8 weeks old. The Pb-exposed metallothionein I/II knockout mice had 

25 increased testicular teratomas and renal and urinary bladder preneoplasia. Pb exposed 

26 wild-type mice were not statistically significantly different than controls. The data 

27 suggest that metallothionein can protect against Pb-induced tumorigenesis. Concerns with 

28 the study are that the doses are at levels of Pb to which humans would not likely be 

29 exposed and there is no metallothionein null condition in humans, though there is 

30 variability in the expression of metallothionein. The data do not address whether this 

31 variability would have any impact on Pb-dependent carcinogenesis in humans. Thus, the 

32 animal toxicology data demonstrate that Pb is a well-established animal carcinogen in 

33 studies employing high dose Pb exposure over a continuous extended duration of 

34 exposure (i.e., 2 years), which is typical of cancer bioassays. Newer studies are showing 
February 2012 5-524 Draft – Do Not Cite or Quote

1 early life maternal Pb exposure can contribute to carcinogenicity in offspring and have 

2 shown that metallothionein is protective against cancer in this pathway.

Neoplastic Transformation Studies, Human Cell Cultures

3 Carcinogenesis can be measured in cell culture systems through neoplastic transformation 

4 models that monitor change by following morphological transformation of cells, 

5 i.e., formation of a focus (or foci) of cell growth.. Xie et al. (2007) treated BEP2D cells 

6 (human papilloma virus- immortalized human bronchial cells) with 0, 1, 5, or 10 μg/cm2 

7 PbCrO4 for 120 h. PbCrO4 induced foci formation in a concentration-dependent manner. 

8 Xie et al. (2008) treated BJhTERT cells (hTERT-immortalized human skin fibroblasts) 

9 and ATLD-2 cells (hTERT-immortalized human skin fibroblasts deficient in Mre11) with 

0, 0.1, 0.5, and 1 μg/cm2 10 PbCrO4 for 120 h. PbCrO4 induced foci formation in a 

11 concentration-dependent manner in the Mre11 deficient cells. Mre11 was required to 

12 prevent PbCrO4-induced neoplastic transformation.

Immune Modulation of Tumorigenesis by Pb

13 As described in the 2006 Pb AQCD (U.S. EPA, 2006b), Pb-induced immunotoxicity can 

14 contribute to increased risk of cancer, primarily due to the intersection of suppressed Th1 

15 responses and misregulated inflammation. First, Pb-induced misregulation of 

16 inflammation involving innate immune cells has been shown to result in chronic insult to 

17 tissues. These insults, excessive lipid and DNA oxidation production by overproduction 

18 of ROS and weakened anti-oxidant defenses, can increase the likelihood of mutagenesis, 

19 cellular instability, and tumor cell formation. For example, Xu et al. (2008) found 

20 toxicological evidence that supports the association with Pb exposure and DNA damage 

21 and concluded that it is a possible route to increased Pb-induced tumorigenesis. The 

22 second component of increased risk of cancer involves Pb-induced suppression of Th1-

23 dependent anti-tumor immunity as acquired immunity shifts statistically significantly 

24 toward Th2 responses. With cytotoxic T lymphocytes and other cell-mediated defenses 

25 dramatically lessened, the capacity to resist cancer may be compromised.

5.10.2 Cancer Biomarkers

26 A study of men aged 21-40 years without occupational history of metals exposure 

27 examined prostate specific antigen (PSA), a biomarker for prostate cancer. This study 

28 reported a positive association between Pb levels and PSA levels (measured in the same 

29 blood samples) in regression models adjusted for confounders, including other metals 
February 2012 5-525 Draft – Do Not Cite or Quote

1 (Cd, Zn, Se, and Cu) (Pizent et al., 2009). The median blood Pb level was 2.6 μg/dL 

2 (range 1.0-10.8 μg/dL). The authors note that the study population was young and at 

3 lower risk of prostate cancer than are older men.

5.10.3 DNA and Cellular Damage

4 Multiple studies have been performed examining the relationship between Pb and DNA 

5 and cellular damage. Details of the recent epidemiologic and toxicological studies follow.

5.10.3.1 Epidemiologic Evidence for DNA and Cellular Damage

6 Multiple studies examined the relationship between Pb and sister chromatid exchange 

7 (SCE). SCEs are exchanges of homologous DNA material between chromatids on a 

8 chromosome and is a test for mutagenicity or DNA damage. A study of male policemen 

9 reported mean blood Pb levels for the study population of 43.5 μg/dL (Wiwanitkit et al., 

10 2008). When dichotomized as having high or low blood Pb levels (cut-off at 49.7 μg/dL), 

11 the higher blood Pb group was observed to have higher mean SCE. Another study of 

12 adult males compared the SCE of storage battery manufacturing workers (mean blood Pb 

13 levels of 40.14 μg/dL) and office workers (mean blood Pb levels of 9.77 μg/dL) (Duydu 

14 et al., 2005). The exposed workers had higher SCE levels and also a greater number of 

15 cells in which the SCEs per cell were higher than the 95th percentile of the population. 

16 Finally, a study of children aged 5-14 years old (mean [SD] blood Pb levels of 7.69 

17 [4.29] μg/dL) reported no correlation between blood Pb levels and SCE (Mielzyńska et 

18 al., 2006). However, the study did report a positive association between blood Pb and 

19 micronuclei (MN) levels. 

20 Other studies of DNA damage have reported mixed results. A study of children ages 6-11 

21 years old and environmentally-exposed to Pb (children attending a school far from a Pb 

22 smelter: median blood Pb levels 4.6 μg/dL; children attending a school near a Pb smelter: 

23 median blood Pb levels 28.6 μg/dL for) reported no association between blood Pb and 

24 baseline DNA damage or repair ability after a peroxide challenge (Méndez-Gómez et al., 

25 2008). Another study included adult participants aged 50-65 years and reported an 

26 association between blood Pb and carcinoembryonic antigen (CEA) but not with DNA27 strand breaks, MN frequency, or oxidative DNA damage (median blood Pb level of the 

28 study population: 3.92 μg/dL) (De Coster et al., 2008). A study conducted among 

29 workers exposed to Pb (mean blood Pb level: 30.3 μg/dL) and unexposed controls (mean 

30 blood Pb level: 3.2 μg/dL) reported greater cytogenetic damage (measured by MN 

31 frequency), chromosomal aberrations, and DNA damage in the Pb-exposed group 
February 2012 5-526 Draft – Do Not Cite or Quote

1 (although this was not statistically significant in linear regression models controlling for 

2 age) (Grover et al., 2010). A study of painters in India, where Pb concentrations in paint 

3 are high, reported a mean (SD) blood Pb level of 21.56 (6.43) μg/dL among painters who 

4 reported painting houses for 8-9 hours/day for 5-10 years (Khan et al., 2010b); the mean 

5 (SD) blood Pb level was 2.84 (0.96) μg/dL for healthy workers who had not been 

6 occupationally exposed to Pb. Cytogenetic damage was higher among the painters 

7 compared to the healthy controls. Another study compared the blood Pb of metal workers 

8 and office workers and reported higher blood Pb levels (both current and 2 year average) 

9 among the metal workers (blood Pb level ≥ 20 μg/dL) compared to the office workers 

10 (blood Pb level < 10 μg/dL for) (Olewińska et al., 2010). Overall, the workers had 

11 increased DNA strand breaks versus the office workers (this held true at various blood Pb 

12 levels). Finally, a study of Pb battery workers with symptoms of Pb toxicity and a group 

13 of controls were examined (Shaik and Jamil, 2009). Higher chromosomal aberrations, 

14 MN frequency, and DNA damage were reported for the battery workers as compared to 

15 the controls.

5.10.3.2 Toxicological Evidence for DNA and Cellular Damage

Sister Chromatid Exchanges

16 Tapisso et al. (2009), considered sister chromatid exchanges (SCE) in Algerian mice 

17 (groups of six mice each) that were exposed by i.p. injection to 5 or 10 doses of 

18 0.46 mg/kg Pb-acetate. The SCE in bone marrow were elevated after Pb exposure alone, 

19 which increased with time. Co-exposure with cadmium or zinc further increased SCE 

20 levels. 

21 SCE was also followed in cultured human cells. Ustundag and Duydu (2007), considered 

22 the ability of N-acetylcysteine and melatonin to reduce Pb nitrate-induced SCE in a 

23 single human donor. Cells were treated with 0, 1, 5, 10, or 50 μM Pb nitrate. SCE 

24 statistically significantly increased at every Pb concentration in a concentration 

25 dependent manner. Both 1 and 2 mM N-acetylcysteine and melatonin were able to 

26 statistically significantly reduce SCE levels in Pb-exposed animals. In another study, 

27 Turkez et al. (In Press) considered the ability of boron compounds to prevent Pb chloride28 induced SCE in human lymphocytes. Cells were obtained from 4 non-smoking donors. 

29 Both 3 and 5 ppm Pb chloride induced a statistically significant increase in SCE levels 

30 over controls. Boron was able to statistically significantly diminish these levels. For both 

31 studies, exposure times were not provided and the full interpretation of these data is 

32 limited by the limited number of donors and the absence of an exposure time for the SCE 

33 assay.
February 2012 5-527 Draft – Do Not Cite or Quote

Micronuclei Formation

1 The 2006 Pb AQCD stated "studies of genotoxicity consistently find associations of Pb 

2 exposure with DNA damage and MN formation" and the current document continues to 

3 report these associations. Alghazal et al. (2008b), considered the ability of Pb-acetate

4 trihydrate to induce MN in bone marrow of Wistar rats. Animals were given a daily dose 

5 of 100 mg/l in their drinking water for 125 days. The mean number of MN in male and 

6 female rats was statistically significantly higher in Pb-exposed animals than in unexposed 

7 controls. Tapisso et al., (2009), considered Pb alone, Pb plus zinc and Pb plus cadmium8 induced MN in rodents. Algerian mice were exposed by i.p. injection to 5 or 10 doses of 

9 0.46 mg/kg Pb-acetate and compared to untreated controls. The MN in bone marrow 

10 were elevated after Pb exposure and increased with time. Co-exposure with cadmium or 

11 zinc did not further increase MN levels.

12 MN formation has also been followed in cultured human cells. Ustundag and Duydu

13 (2007) considered the ability of N-acetylcysteine and melatonin to reduce Pb nitrate14 induced MN in a single human donor. Cells were treated with 0, 1, 5, 10, or 50 μM Pb 

15 nitrate. MN formation statistically significantly increased at the two highest Pb 

16 concentrations in a concentration dependent manner. Both 1 and 2 mM N-acetylcysteine 

17 and melatonin were not able to statistically significantly reduce MN levels. In another 

18 study, Turkez et al. (In Press) considered the ability of boron compounds to prevent Pb 

19 chloride-induced MN in human lymphocytes. Cells were obtained from 4 non-smoking 

20 donors. Both 3 and 5 ppm Pb chloride induced a statistically significant increase in MN 

21 levels over controls. Boron induced a statistically significant attenuation of these Pb22 induced levels. For both studies, exposure times were not provided, and the full 

23 interpretation of these data is limited by the limited number of donors and the absence of 

24 an exposure time for the MN assay. Gastaldo et al. (2007) evaluated the ability of Pb to 

25 induce MN. Human endothelial HMEC cell line was treated with 1–1,000 μM Pb nitrate 

26 for 24 hours. MN increased in a statistically significant, concentration-dependent manner.

Hypoxanthine-guanine phosphoribosyltransferase Mutations

27 The potential mutagenicity of Pb in human or animal cells was evaluated by monitoring 

28 mutations at the hypoxanthine-guanine phosphoribosytransferase (HPRT) locus. Li et al.

29 (2008a) evaluated Pb-acetate-induced HPRT in the non-small-cell lung carcinoma tumor 

30 cell line, CL3 and in normal human diploid fibroblasts (specific tissue source not 

31 reported). All cells were exposed to 0, 100, 300 or 500 μM Pb-acetate for 24 hours in 

32 serum-free medium ± a 1-hour pretreatment with a MKK1/2 inhibitor or a PKC-alpha 

33 inhibitor. Pb alone did not induce HPRT mutations. Inhibiting the ERK pathway via 

34 either inhibitor statistically significantly increased Pb-induced mutagenesis. Wang et al.
February 2012 5-528 Draft – Do Not Cite or Quote

1 (2008c), investigated Pb-acetate -induced HPRT mutations in CL3 cells. All cells were 

2 exposed to 0, 100, 300 or 500 μM Pb-acetate for 24 hours in serum-free medium ± a 

3 1-hour pretreatment with a PKC-alpha inhibitor or siRNA for PKC-alpha. Pb alone did 

4 not induce HPRT mutations. Inhibiting PKC-alpha via either inhibitor statistically 

5 significantly increased Pb-induced mutagenesis. McNeill et al. (2007) considered 

6 Pb-acetate induced HPRT mutations in Chinese hamster ovary AA8 cells and AA8 cells 

7 overexpressing human Ape1. Cells were treated with 5 μM Pb-acetate for 6 hours. No 

8 increases in HPRT mutations were observed after Pb exposure in either cell line but with 

9 specific pathway perturbations (PKC-alpha or ERK) Pb was able to induce HPRT 

10 mutations.

Chromosomal Aberrations

11 Chromosomal aberrations, an indicator of cancer risk, were followed in Pb-exposed 

12 rodents (El-Ashmawy et al., 2006). Dietary exposure to Pb-acetate administered as a 

13 single dose of 0.5% w/w to male Swiss albino mice caused statistically significant 

14 increased levels of chromosomal aberrations in the Pb treatment alone group, particularly 

15 with respect to fragments, deletions, ring chromosomes, gaps, and end-to-end 

16 associations. In addition, the authors found turmeric and myrrh powders were protective. 

17 Concerns with the study include the use of only a single dose of Pb-acetate along with the 

18 high levels of unusual aberrations such as ring chromosomes and end-to-end associations. 

19 Typically, these aberrations are rare after metal exposure, but were the most commonly 

20 observed aberration in this study raising questions about the quality of the metaphase 

21 preparations. An additional concern was that only 50 metaphases per dose were analyzed 

22 instead of the more common 100 metaphases per dose. The authors did not explain why 

23 their spectrum of aberrations was so different, why they only used one dose, or analyzed 

24 fewer metaphases per dose. 

25 Multiple studies considered the ability of Pb to induce chromosomal aberrations in 

26 cultured human cells. The ability of Pb nitrate to induce chromosomal aberrations was 

27 examined in primary human peripheral blood lymphocytes obtained from healthy, 

28 nonsmoking donors (Pasha Shaik et al., 2006). Cells were treated with 0, 1.2 or 2 mM 

29 Pb-nitrate for 2 hours. No increase in chromosomal aberrations was reported. Some 

30 aneuploidy was observed. Concerns with the study are that only a 2 hour exposure was 

31 used, which may not be long enough for DNA damage to be expressed as a chromosomal 

32 aberration. It also appears from the data presentation that only three subjects were used; 

33 one for a control, one for the low dose and one for the high dose. Experiments were not 

34 repeated, thus given the small number of subjects, this study may not have had sufficient 

35 power to detect any effects. Holmes et al. (2006a), treated WHTBF-6 cells (hTERTimmortalized human lung cells) with 0, 0.1, 0.5, or 1 μg/cm2 36 Pb chromate for 24-120 
February 2012 5-529 Draft – Do Not Cite or Quote

hours or with 0, 0.1, 0.5, 1, 5 or 10 μg/cm2 1 Pb oxide for 24 or 120 hours. Pb chromate 

2 induced statistically significant, concentration-dependent increases in centrosome 

3 abnormalities and aneuploidy. Wise et al. (2006b) treated BEP2D cells with 0, 0.5, 1, 5, 

or 10 μg/cm2 4 Pb chromate for 24 hours. Pb chromate induced statistically significant 

5 concentration-dependent increases in chromosomal aberrations. Holmes et al. (2006b),

treated WHTBF-6 cells with 0, 0.1, 0.5, or 1 μg/cm2 6 Pb chromate for 24-72 hours. Pb 

7 chromate induced statistically significant, concentration-dependent increases in 

8 chromosomal aberrations. The effects of the chromate anion cannot be ruled out as 

9 causative in inducing these chromosomal aberrations. Wise et al. (2006a), treated 

WHTBF-6 cells with 0, 0.1, 0.5, or 1 μg/cm2 10 Pb chromate for 24-120 hours. Pb chromate 

11 induced statistically significant, concentration-dependent increases in spindle assembly 

12 checkpoint disruption, effects of mitosis and aneuploidy. By contrast, chromate-free Pb 

13 oxide did not induce centrosome amplification. The effects were likely attributable to the 

chromate anion. Xie et al. (2007) treated BEP2D cells with 0, 1, 5, or 10 μg/cm2 14 Pb 

15 chromate for 24 hours. Pb chromate induced statistically significant, concentration16 dependent increases in chromosomal aberrations and aneuploidy. Wise et al. (2010)

treated WHTBF-6 cells with 0, 0.1, 0.5, or 1 μg/cm2 17 Pb chromate for 24 hours in a study 

18 comparing 4 chromate compounds. Pb chromate induced statistically significant, 

19 concentration-dependent increases in chromosomal aberrations 

20 Multiple investigators considered the ability of Pb chromate to induce chromosome 

21 aberrations in rodent cell cultures. Grlickova Duzevik et al. (2006) treated Chinese 

hamster ovary (CHO) cells with 0, 0.1, 0.5, or 1 μg/cm2 22 Pb chromate for 24 h. Specific 

23 CHO lines used included AA8 (wildtype) EM9 (XRCC1-deficient), and H9T3 (EM9 

24 complemented with human XRCC1 gene). Pb chromate induced statistically significant, 

25 concentration-dependent increases in chromosomal aberrations that were statistically 

26 significantly increased by XRCC1 deficiency. Nestmann and Zhang (2007) treated 

Chinese hamster ovary cells (clone WB(L)) with 0, 0.1, 0.5, 1, 5, or 10 μg/cm2 27 Pb 

28 chromate (as pigment yellow) for 18 h. No increases in chromosomal aberrations were 

observed. Savery et al. (2007) treated CHO cells with 0, 0.1, 0.5, 1, or 5 μg/cm2 29 Pb 

30 chromate for 24 h. Specific CHO lines used included AA8 (wildtype), KO40 

31 (Fancg-deficient), and 40BP6 (Fancg-complemented). Pb chromate induced statistically 

32 significant, concentration-dependent increases in chromosomal aberrations that were 

33 increased by Fancg-deficiency. The Fancg gene plays an important role in cellular 

34 resistance to DNA interstrand crosslinks, protecting against genetic instability. Camrye et 

al., (2007) treated CHO cells with 0, 0.1, 0.5, 1, 5, or 10 μg/cm2 35 Pb chromate for 24 

36 hours. Specific CHO lines used included CHO-K1 (parental), xrs-6 (Ku80 deficient), and 

37 2E (xrs-6 complemented with Ku80 gene). Pb chromate induced statistically significant, 

38 concentration-dependent increases in chromosomal aberrations that were not affected by 

39 Ku80 deficiency. Ku80 is a gene a gene involved in nonhomologous end-joining repair 
February 2012 5-530 Draft – Do Not Cite or Quote

1 and its absence can contribute to genetic instability. Stackpole et al. (2007) treated CHO 

and Chinese hamster lung (CHL) cells with 0, 0.1, 0.5, or 1 μg/cm2 2 Pb chromate for 24 

3 hours. Specific CHO lines used included AA8 (wildtype), irs1SF (XRCC3-deficient), and 

4 1SFwt8 (XRCC3 complemented). XRCC3 is DNA repair enzyme involved in 

5 homologous recombination. CHL lines used included V79 (wildtype), irs3 (Rad51C 

6 deficient) and irs3#6 (Rad51C complemented). Rad51C is a gene which encodes strand7 transfer proteins that are thought to be involved in recombinational repair of damaged 

8 DNA and in meiotic recombination. Pb chromate induced statistically significant, 

9 concentration-dependent increases in chromosomal aberrations that were statistically 

10 significantly increased by both XRCC3 and Rad51C deficiency.

11 Multiple studies considered the ability of Pb chromate to induce chromosome aberrations 

12 in marine mammal cell cultures. Li Chen et al. (2009) treated primary North Atlantic 

right whale lung and skin fibroblasts with 0, 0.5, 1.0, 2.0, and 4.0 μg/cm2 13 Pb chromate for 

14 24 hours. Wise et al. (2009) treated primary Steller sea lion lung fibroblasts with 0, 0.1, 

0.5, 1 and 5 μg/cm2 15 Pb chromate for 24 hours. Wise et al. (2011) treated primary sperm 

whale skin fibroblasts with 0, 0.5, 1, 3, 5, and 10 μg/cm2 16 Pb chromate for 24 hours. In all 

17 three studies, Pb chromate induced statistically significant, concentration-dependent 

18 increases in chromosomal aberrations.

19 In summary, exposure of various cell models and an in vivo model to Pb (acetate, 

20 chromate, or nitrate) induced significant increases in chromosomal aberration that often 

21 responded in a concentration dependent manner. The use of various cell lines deficient in 

22 specific DNA repair enzymes helped to elucidate which pathways may be most sensitive 

23 to Pb-dependent chromosomal aberration.

COMET Assay

24 Multiple studies considered the ability of Pb to induce DNA single strand breaks in 

25 laboratory animals using the comet assays. The COMET assay measures DNA damage 

26 assessed by single cell electrophoresis of a lysed cell and measurement of the fragmented 

27 DNA or tail length. Xu et al. (2008) considered the ability of Pb-acetate to induce DNA 

28 damage measured by the comet assay in lymphocytes of male ICR mice. Animals (5 per 

29 group) were given Pb-acetate by gavage at doses of 0, 10, 50, or 100 mg/kg body weight 

30 every other day for 4 weeks. Pb exposure statistically significantly increased both tail 

31 length and tail moment in a dose-dependent manner. Nava-Hernandez et al. (2009)

32 considered the ability of Pb-acetate to induce DNA damage in primary spermatocyte 

33 DNA of male Wistar rats. Animals (3 per group) were treated for 13 weeks with 0, 250 or 

34 500 mg/L Pb in their drinking water. There was statistically significantly less DNA 

35 damage in the controls compared to the two treatment groups. Narayana and Al-Bader
February 2012 5-531 Draft – Do Not Cite or Quote

1 (2011) considered the ability of Pb nitrate to induce DNA damage in liver tissue of adult 

2 male Wistar rats. Animals (8 per group) were treated for 60 days with doses of 0, 0.5 or 

3 1% Pb(NO3)2 in their drinking water. There were no statistical differences between 

4 treated groups and controls. Drosophila melanogaster larvae (72 hours old) exposed to Pb 

5 nitrate (2,000, 4,000, and 8,000 μM in culture media for 24 hours) yielded haemocytes 

6 that tested positive in the comet assay; Pb chloride (8,000 μM) did not cause DNA 

7 damage with the comet assay (Carmona et al.).

8 Other studies used the COMET assay in cultured human cells. Pasha Shaik et al. (2006)

9 treated primary human peripheral blood lymphocytes obtained from healthy, nonsmoking 

10 donors with 0, 2.1, 2.4, 2.7, 3.0, 3.3 Pb nitrate for 2 hours and found dose-dependent 

11 increases in Comet tail length. Concerns with the study are that apparently no untreated 

12 control was used. It also appears from the data presentation that only five subjects were 

13 used; one for each dose. Experiments were not repeated. Thus, given the small number of 

14 subjects and the absence of a negative control, this study may only be detecting 

15 background levels. Xie et al. (2008) treated BJhTERT cells (hTERT-immortalized human 

16 skin fibroblasts) and ATLD-2 cells (hTERT-immortalized human skin fibroblasts 

deficient in Mre11) with 0, 0.1, 0.5, and 1 μg/cm2 17 Pb chromate for 24 hours. Mre11 is a 

18 component of the MRN complex and plays a role in telomere maintenance and double19 strand break repair. Pb chromate induced a concentration-dependent increase in DNA 

20 double strand breaks measured by the comet assay. In another study, Pb nitrate exposure 

21 (30 μg/mL) induced statistically significant increased DNA damage in human liver 

22 HepG2 cells that was statistically significantly attenuated with co-exposure with the 

23 antioxidant NAC (500 μM) (Yedjou et al., 2010).

24 Other studies used the comet assay to examine Pb-induced DNA single strand breaks in 

25 rodent cell cultures. Xu et al. (2006), treated PC12 cells with 0, 0.1, 1 or 10 μM 

26 Pb-acetate. Both tail length and tail moment statistically significantly increased in a 

27 concentration-dependent manner. Kermani et al. (2008) exposed mouse bone marrow28 mesenchymal stem cells to 60 μM Pb-acetate for 48 hours. There was an increase in 

29 several comet assay measurements including tail length.

Other Assays

30 Other studies considered the ability of Pb to induce DNA double strand breaks by 

31 measuring gamma-H2A.X foci formation in cultured human cells. Xie et al. (2008)

32 treated BJhTERT cells (hTERT-immortalized human skin fibroblasts) and ATLD-2 cells 

33 (hTERT-immortalized human skin fibroblasts deficient in Mre11) with 0, 0.1, 0.5, and 

1 μg/cm2 34 Pb chromate for 24 hours. Pb chromate induced a concentration-dependent 

35 increase in DNA double strand breaks measured by gamma-H2A.X foci formation. 
February 2012 5-532 Draft – Do Not Cite or Quote

1 Gastaldo et al. (2007) evaluated the ability of Pb to induce DNA double strand breaks 

2 with both gamma-H2A.X foci formation and pulse-field gel electrophoresis in cultured 

3 human cells. The human endothelial HMEC cell line was treated with 1 to 1,000 μM Pb 

4 nitrate for 24 hours. DNA double strand breaks increased in a concentration-dependent 

manner. Wise et al. (2010) treated WHTBF-6 cells with 0, 0.1, 0.5, or 1 μg/cm2 5 Pb 

6 chromate for 24 hours in a study comparing four chromate compounds. Pb chromate 

7 induced statistically significant, concentration-dependent increases in DNA double strand 

8 breaks measured by gamma-H2A.X foci formation, at a similar level to the other three 

9 compounds. Two studies demonstrated the ability of Pb to destabilize DNA by forming 

10 DNA-histone cross links, which can lead to histone aggregation. Extracts of rat liver co11 incubated with Pb nitrate (<300 μM) were shown to react with chromatin components 

12 and induce chromatin aggregation via histone-DNA cross links (Rabbani-Chadegani et 

13 al., 2011; Rabbani-Chadegani et al., 2009).

14 Genotoxicity testing of Drosophila melanogaster larvae (72 hours old) using the Wing 

15 Spot test showed that neither Pb chloride nor Pb nitrate (at concentrations of 2,000, 4,000 

16 and 8,000 μM in culture media with exposure until pupation) was able to induce 

17 significant increases in the frequency of wing spots (Carmona et al.). Further, wing spot 

18 assays employing Pb co-exposure with gamma radiation showed no effect of Pb on 

19 gamma radiation induced spotting frequency. The wing spot test can detect mitotic 

20 recombination and multiple mutational events such as point mutations, deletions, and 

21 certain types of chromosome aberrations (Graf and Würgler, 1986).

22 Multiple studies considered Pb and DNA repair. Most were conducted in cultured cells, 

23 and one was done in an animal model. El-Ghor et al. (In Press) followed microsatellite 

24 instability (MSI) in Pb-acetate trihydrate exposed adult male rats; MSI reflects impaired 

25 DNA mismatch repair, and contributes to an increased risk of cancer. DNA from 

26 leukocytes of male albino rats exposed to Pb-acetate (acute: single oral dose of 

27 467 mg/kg BW or sub-chronic: 47 mg/kg BW six d/wk for 4 wk) showed increased MSI 

28 at three microsatellite loci (D6mit3, D9mit2, and D15Mgh1). This study is limited by its 

29 small sample size (n=2 to 3 rodents per treatment group). Li et al., (2008a), evaluated 

30 Pb-acetate-induced effects on nucleotide excision repair efficiency in CL3 cells. All cells 

31 were exposed to 0, 100, 300 or 500 μM Pb-acetate for 24 hours in serum-free medium. 

32 Pb increased nucleotide excision repair efficiency. Gastaldo et al. (2007) evaluated the 

33 ability of Pb to affect DNA repair in cultured human cells. The human endothelial HMEC 

34 cell line was treated with 100 μM Pb nitrate for 24 hours. Pb inhibited non-homologous 

35 end joining (NHEJ) repair, over activated MRE11-dependent repair and increased Rad51-

36 related repair. Xie et al. (2008) treated BJhTERT cells (hTERT-immortalized human skin 

37 fibroblasts) and ATLD-2 cells (hTERT-immortalized human skin fibroblasts deficient in 

Mre11) with 0, 0.1, 0.5, and 1 μg/cm2 38 Pb chromate for 24 or 120 hours. Mre11 was 
February 2012 5-533 Draft – Do Not Cite or Quote

1 required to prevent Pb chromate-induced DNA double strand breaks. McNeill et al,

2 (2007) considered Pb-acetate effects on Ape1. Chinese hamster ovary cells (AA8) were 

3 treated with 0, 0.5, 5, 50, or 500 μM Pb-acetate and then whole cell extracts were used to 

4 determine AP site incision activity. The data show that Pb reduced AP endonuclease 

5 function. Finally, studies considered Pb-induced cellular proliferation in laboratory 

6 animals. An earlier study in rats showed Pb nitrate-induced increased proliferation of 

7 liver cells after a partial hepatectomy, effects that were more prominent in males than 

8 females (sexual dimorphism) (Tessitore et al., 1995). The newer studies showed similar 

9 trends in males. Fortoul et al. (2005) exposed adult male CD1 mice (24 animals per 

10 group) to 0.01 M Pb-acetate, 0.006 M cadmium chloride or a mixture of the two 

11 chemicals for 1 h twice a week for 4 weeks by inhalation. The lungs were then examined 

12 by electron microscopy for changes. Pb induced cellular proliferation in the lungs. 

13 Kermani et al. (2008) exposed mouse bone marrow-mesenchymal stem cells to 0-100 μM 

14 Pb-acetate for 48 hours. As measured by the MTT assay, Pb decreased cell proliferation 

15 at all concentrations tested.

5.10.4 Effects of Lead within Mixtures

16 Several studies considered the impact of mixtures with Pb. All considered genotoxicity. 

17 Mendez-Gomez et al., (2008), evaluated 65 children from Mexico exposed to both 

18 arsenic and Pb. DNA damage and decreased DNA repair were seen using the comet assay 

19 and other assays, but did not correlate with urinary arsenic or blood Pb levels. Tapisso et 

20 al., (2009), considered Pb alone, Pb plus zinc and Pb plus cadmium-induced MN in 

21 rodents. Algerian mice (groups of six mice each) were exposed (i.p.) to 5 or 10 doses of 

22 0.46 mg/kg Pb-acetate and compared to untreated controls. The MN in bone marrow 

23 were elevated after Pb treatment alone and increased with time. Co-exposure with 

24 cadmium or zinc did not further increase MN levels but did increase SCE levels. Glahn et 

25 al., (2008) performed a gene array study in primary normal human bronchial epithelial 

26 cells from four donors treated with 550 μg/L Pb chloride, 15 μg/L cadmium sulfate, 

27 25 μg/L cobalt chloride or all three combined for 72 hours. There was a clear interaction 

28 of all three metals impacting RNA expression.

29 No recent studies of the protective role of calcium in Pb-dependent carcinogenesis or 

30 genotoxicity were found. There were some data suggesting that boron, melatonin, 

31 N-acetylcysteine, turmeric and myrrh protect cells against Pb-induced genotoxicity 

32 (Sections 5.10.3.2 and 5.10.6).

33 A new study details Pb and selenium interactions in virus-dependent carcinogenesis in 

34 laboratory animals. Schrauzer (2008) considered the impact of selenium on 
February 2012 5-534 Draft – Do Not Cite or Quote

1 carcinogenesis by studying 4 groups of weanling virgin female C3H/St mice infected 

2 with murine mammary tumor virus (groups of 20-30 mice), which induces mammary 

3 tumor formation. One set of two groups were fed a diet containing 0.15 ppm selenium 

4 and then were exposed via drinking water to acetic acid (control group) or 0.5 ppm

5 Pb-acetate (treated group). The second set of two groups were fed a diet containing 

6 0.65 ppm selenium and then similarly exposed to acetic acid or 0.5 ppm Pb-acetate. The 

7 study was primarily focused on the general effects of a low selenium diet. The data 

8 suggest that selenium is anticarcinogenic as in the groups without Pb exposure, the 

9 animals exposed to the higher selenium levels had fewer mammary tumors and these 

10 tumors had a delayed onset of appearance. Pb exposure with low selenium caused the 

11 same delayed onset as did the higher dose of selenium and also caused some reduction in 

12 the tumor frequency. Pb exposure with higher selenium increased the tumor frequency 

13 and the onset of the tumors. Pb also induced weight loss at 14 months in both exposed 

14 groups. The data suggest that there may be interactions of Pb and selenium, but they 

15 suggest that Pb mimics or antagonizes selenium. They do not suggest that selenium is 

16 protective of Pb-induced toxicity or carcinogenesis.

5.10.5 Modes of Action

17 The carcinogenic mechanism of action of Pb is poorly understood. It is unclear whether 

18 the mechanism of action of Pb is best understood within the framework of multistage 

19 carcinogenesis, genomic instability or epigenetic modification. For example, multistage 

20 carcinogenesis involves a series of cellular and molecular changes that result from the 

21 progressive accumulation of mutations that induce alterations in cancer-related genes. Pb 

22 does not appear to follow this paradigm and the literature suggests it is weakly 

23 mutagenic. Pb does appear to have some ability to induce chromosomal mutation and 

24 DNA damage, i.e., clastogenicity. However, the ability of Pb to alter gene expression 

25 (epigenetic effects) and to interact with proteins may be a means by which Pb induces its 

26 carcinogenicity. It is known that Pb can replace zinc in zinc-binding (zinc-finger) 

27 proteins, which include hormone receptors, cell-cycle regulatory proteins, the Ah 

28 receptor, estrogen receptor, p53, DNA repair proteins, protamines, and histones. These 

29 zinc-finger proteins all bind to specific recognition elements in DNA. Thus, Pb may act at 

30 a post-translational stage to alter protein structure of Zn-finger proteins, which can in turn 

31 alter gene expression, DNA repair and other cellular functions. To recapitulate, cancer 

32 develops from one or a combination of multiple mechanisms including modification of 

33 DNA via epigenetics or enzyme dysfunction and genetic instability or mutation(s). These 

34 modifications then provide the cancer cells with a selective growth advantage. In this 

35 schematic, Pb appears to contribute to epigenetic changes, and chromosomal aberrations.
February 2012 5-535 Draft – Do Not Cite or Quote

1 The genomic instability paradigm requires a cascade of genome-wide changes caused by 

2 interfering with DNA repair, kinetochore assembly, cellular checkpoints, centrosome 

3 duplication, microtubule dynamics or a number of cell maintenance processes. These 

4 processes have been rarely studied for Pb, thus there are few data that suggest Pb may 

5 interfere with some of these processes. Furthermore, the bulk of the literature in this area 

6 involves Pb chromate and it is unclear if the effects are due to Pb or chromate. Epigenetic 

7 modifications can lead to cancer by altering cellular functions without altering the genetic 

8 material. The most commonly studied epigenetic change is methylation alterations. A 

9 small number of studies show that Pb can induce epigenetic changes, but studies are still 

10 missing to clearly tie these effects to Pb-induced carcinogenesis and genotoxicity. Thus, 

11 the mechanism is difficult to define but, if Pb is a human carcinogen, the mechanism 

12 likely involves either genomic instability or epigenetic modification paradigms or some 

13 combination of the two. However, Pb-dependent carcinogenicity is it not likely to occur 

14 by a multistage paradigm. More work is needed to determine the mechanism. 

15 Exposure to mixtures can also contribute to understanding of modes of action. No recent 

16 studies of the protective role of calcium or zinc in Pb-dependent carcinogenesis or 

17 genotoxicity were found. There were some data suggesting that metallothionein protects 

18 rodents from Pb-induced cancers. There were some data suggesting that boron, 

19 melatonin, N-acetylcysteine, turmeric and myrrh protected cells against Pb-induced 

20 genotoxicity. There were some data suggesting that Pb mimics or antagonizes selenium 

21 in rodents. These data are discussed in more detail elsewhere in the cancer section and 

22 point to the relevance of mixtures in assessing toxicity.

5.10.5.1 Epigenetics

23 Air pollution exposure is being linked increasingly with epigenetic changes (Pavanello et 

24 al., 2010; Baccarelli and Bollati, 2009; Tarantini et al., 2009; Bollati et al., 2007).

25 Epigenetic changes involve changes in DNA expression without actual changes in the 

26 DNA sequence and these changes may be heritable. Epigenetic changes are mediated by 

27 histone modification, DNA methylation, miRNA changes, or pathways that affect these 

28 three mediators. Differential epigenetic modification has the possibility to contribute to 

29 disease. Epigenetic studies have been conducted to examine the associations between Pb 

30 biomarker levels and global DNA methylation markers [Alu and long interspersed 

31 nuclear element-1 (LINE-1)] in humans (Wright et al., 2010; Pilsner et al., 2009). Wright 

32 et al. (2010) utilized a sample of participants from the NAS with mean (SD) Pb levels of 

33 20.5(14.8)g/g for tibia measures, 27.4 (19.7)g/g for patella measures, and 4.1 (2.4) μg/dL 

34 for blood measures. In both crude and adjusted analyses, patella Pb levels were inversely 

35 associated with LINE-1 methylation but not with Alu, both of which are indicators of 
February 2012 5-536 Draft – Do Not Cite or Quote

1 global methylation. When examining the relationship between patella Pb and LINE-1 

2 more closely, a non-linear trend was observed with a smaller magnitude of effect 

3 estimated for higher patella Pb (≥ 40 μg/g). No associations were observed for tibia or 

4 blood Pb and either LINE-1 or Alu. Another study included maternal-infant pairs from 

5 the Early Life Exposures in Mexico to Environmental Toxicants study and measured 

6 LINE-1 and Alu methylation in umbilical cord blood samples (Pilsner et al., 2009). In 

7 unadjusted models, maternal pregnancy tibia Pb levels [mean (SD) 10.5 (8.4) μg/g] were 

8 inversely associated with Alu methylation; maternal patella Pb levels [mean (SD) 12.9 

9 (14.3) μg/g] were inversely associated with LINE-1 methylation. The associations

10 persisted in adjusted models although the association between patella Pb and LINE-1 was 

11 only apparent when the adjusted models also included umbilical cord blood Pb levels. No 

12 association was detected between umbilical cord Pb levels and the DNA methylation 

13 markers. Overall, the studies consistently demonstrate an association between patella Pb 

14 levels and LINE-1 methylation. 

15 Toxicological studies have been performed examining Pb-dependent epigenetic changes 

16 and gene expression, DNA repair, and mitogenesis. Glahn et al., (2008) performed a gene 

17 array study in primary normal human bronchial epithelial cells from four donors after in 

18 vitro treatment of the cells with 550 μg/L Pb chloride, 15 μg/L cadmium sulfate, 25 μg/L 

19 cobalt chloride or all three combined for 72 hours. The authors describe a pattern of RNA 

20 expression changes indicating “ ...coordinated stress-response and cell-survival signaling, 

21 deregulation of cell proliferation, increased steroid metabolism, and increased expression 

22 of xenobiotic metabolizing enzymes.” These are all known targets of possible epigenetic 

23 changes, but full interpretations of the data as epigenetic changes are complicated by the 

24 absence of a measure to determine if these changes were a result of genotoxic effects. A 

25 recent publication using HepG2 cells in tissue culture showed that cells exposed to a high 

26 dose of Pb (100 μM Pb-acetate) experienced ALAD gene promoter hypermethylation and 

27 decreased ALAD transcription. This was in agreement with findings in battery plant 

28 workers who showed ALAD hypermethylation (versus non-occupationally exposed 

29 controls) and an association of this hypermethylation with elevated risk of Pb poisoning

30 (Li et al., 2011).

31

5.10.6 Summary and Causal Determination

32 In summary, the toxicological literature on the genotoxic, mutagenic, and carcinogenic 

33 potential of Pb provide strong evidence of effects in laboratory animals. In laboratory 

34 studies, high-dose Pb has been demonstrated to be an animal carcinogen. There are data 
February 2012 5-537 Draft – Do Not Cite or Quote

1 to suggest Pb is a human carcinogen among toxicological studies, but they are not 

2 definitive. Multiple toxicological studies showed neoplastic transformation in cultured 

3 cells, but all focused on Pb chromate and the positive response to the chromate ion and 

4 not the Pb likely contributed to these findings. Multiple epidemiologic studies have been 

5 performed examining the association between cancer incidence and mortality and Pb 

6 exposures, estimated with biological measures and exposure databases. Mixed results 

7 have been reported for cancer mortality studies; a large NHANES epidemiologic study 

8 demonstrated a positive association between blood Pb and cancer mortality, but the other 

9 studies reported null results. Although the 2006 Pb AQCD reported some studies that 

10 found an association between Pb exposure indicators and lung cancer, current studies 

11 mostly included occupationally-exposed adults and observed no associations. Most 

12 studies of Pb and brain cancer were null among the overall study population, but positive 

13 associations were observed among individuals with certain genetic variants. However the 

14 studies of Pb and brain cancer were all performed among occupational cohorts using 

15 interviews instead of biological measurements to represent Pb exposure. A limited 

16 amount of research has been performed on other types of cancer. The 2006 Pb AQCD

17 reported evidence that suggested an association between Pb exposure and stomach 

18 cancer, but recent studies of stomach cancer are lacking. One study examining Pb and 

19 stomach cancer has been performed since the last AQCD and the results of the study are 

20 mixed

21 Among epidemiologic studies, high Pb levels were associated with SCEs among adults 

22 but not children. Other epidemiologic studies of DNA damage reported inconsistent 

23 results. Consistent with previous toxicological findings, Pb does appear to have genotoxic 

24 activity in animal and in vitro models, inducing SCE, MN and DNA strand breaks, but 

25 continues to not produce chromosomal aberrations except for Pb chromate; this again is 

26 likely due to the chromate. Pb does not appear to be very mutagenic as the HPRT assays 

27 were typically negative unless a cell signaling pathway was disturbed. 

28 Mechanistic understanding of the carcinogenicity of Pb is expanding with work on the 

29 antioxidant selenium and metallothionein, a protein that binds Pb and reduces its 

30 bioavailability. Metallothionein has been shown to be protective against the effect of Pb 

31 on carcinogenicity. Pb is clastogenic and mutagenic in some but not all models. 

32 Clastogenicity and mutagenicity may be possible mechanisms contributing to cancer but 

33 are not absolutely associated with the induction of cancer. Because Pb has a higher 

34 atomic weight than does zinc, Pb replaces zinc at many zinc binding sites or zinc finger 

35 proteins. This substitution has the potential to induce indirect effects that can contribute 

36 to carcinogenicity via interactions at hormone receptors, at cell-cycle regulatory proteins, 

37 with tumor suppressor genes like p53, with DNA repair enzymes, with histones, etc. 
February 2012 5-538 Draft – Do Not Cite or Quote

1 These indirect effects may act at a post-translational level to negatively alter protein 

2 structure and DNA repair.

3 Epigenetic changes associated with Pb exposure, particularly with respect to methylation 

4 and effects on DNA repair, are beginning to appear in the literature. These modifications 

5 may further alter DNA repair or change the expression of a tumor suppressor gene or 

6 oncogene. A small number of epidemiologic studies examining Pb and global epigenetic 

7 changes including LINE-1 and Alu demonstrated an inverse association between patella 

8 Pb and LINE-1 methylation, an emerging area of research. Toxicological studies show 

9 that Pb can activate or interfere with a number of signaling and repair pathways, though it 

10 is unclear whether these are due to epigenetic responses or genotoxicity. Thus, an 

11 underlying mechanism is still uncertain, but likely involves either genomic instability, 

12 epigenetic modifications, or both.

13 In conclusion, the toxicological literature provides the strongest evidence for Pb exposure 

14 and cancer with supporting evidence provided by the epidemiologic literature. This is 

15 substantiated by the findings of other agencies including IARC, which has classified 

16 inorganic Pb compounds as a probable human carcinogen and the National Toxicology 

17 Program, which has listed Pb and Pb compounds as “reasonably anticipated to be human 

18 carcinogens.” Strong evidence from toxicological studies demonstrates an association 

19 between Pb and cancer, genotoxicity/clastogenicity or epigenetic modification.

20 Carcinogenicity in historical animal toxicology studies with Pb exposure has been 

21 reported in the kidneys, testes, brain, adrenals, prostate, pituitary, and mammary gland, 

22 albeit at high doses of Pb. Epidemiologic studies of cancer incidence and mortality 

23 reported inconsistent results; one strong epidemiologic study demonstrated an association 

24 between blood Pb and increased cancer mortality, but the other studies reported weak or 

25 no associations. In the 2006 Pb AQCD, Pb exposure was found to be associated with 

26 stomach cancer, but there was only one recent study on stomach cancer and Pb exposure, 

27 which reported mixed findings. Similarly, some studies in the 2006 Pb AQCD reported 

28 associations between Pb exposure and lung cancer. More recent epidemiologic studies of 

29 lung cancer focused on occupational exposures and reported no associations. The 

30 majority of epidemiologic studies of brain cancer had null results overall, but positive 

31 associations between Pb exposure and brain cancer were observed among individuals 

32 with certain genotypes. In toxicological studies, chromosomal aberrations after Pb 

33 exposure are most often reported with Pb chromate exposure, which is likely due to 

34 toxicity of the chromate moiety. Mechanistic understanding of Pb and its effect on cancer 

35 and genotoxicity is expanding through toxicological work focusing on antioxidants and 

36 other proteins that sequester Pb or reduce its bioavailability. The collective body of 

37 evidence integrated across toxicological and epidemiologic studies is sufficient to 

38 conclude that there is a likely causal relationship between Pb exposure and cancer.
February 2012 5-539 Draft – Do Not Cite or Quote

1

2

3

References

. (2011). Correspondence between Molini Patel and Todd Jusko. (EPA-HQ-ORD-2011-0051-0018). 

http://www.regulations.gov/#!documentDetail;D=EPA-HQ-ORD-2011-0051-0018.

Abam, E; Okediran, BS; Odukoya, OO; Adamson, I; Ademuyiwa, O. (2008). Reversal of ionoregulatory disruptions 

in occupational lead exposure by vitamin C. Environ Toxicol Pharmacol 26: 297-304. 

http://dx.doi.org/10.1016/j.etap.2008.05.008.

Abdallah, GM; El-Sayed, ES; Abo-Salem, OM. (2010). Effect of lead toxicity on coenzyme Q levels in rat tissues. 

Food Chem Toxicol 48: 1753-1756. http://dx.doi.org/10.1016/j.fct.2010.04.006.

Abdel Moneim, AE; Dkhil, MA; Al-Quraishy, S. (2011a). Effects of Flaxseed Oil on Lead Acetate-Induced 

Neurotoxicity in Rats. Biol Trace Elem Res 144: 904-913. http://dx.doi.org/10.1007/s12011-011-9055-4.

Abdel Moneim, AE; Dkhil, MA; Al-Quraishy, S. (2011b). The protective effect of flaxseed oil on lead acetateinduced renal toxicity in rats. J Hazard Mater 194: 250-255. 

http://dx.doi.org/10.1016/j.jhazmat.2011.07.097.

Abdelouahab, N; Mergler, D; Takser, L; Vanier, C; St-Jean, M; Baldwin, M; Spear, PA; Chan, HM. (2008). Gender 

differences in the effects of organochlorines, mercury, and lead on thyroid hormone levels in lakeside 

communities of Quebec (Canada). Environ Res 107: 380-392. 

http://dx.doi.org/10.1016/j.envres.2008.01.006.

Abe, A; Hiraoka, M; Shayman, JA. (2007). A role for lysosomal phospholipase A2 in drug induced 

phospholipidosis. Drug Metab Lett 1: 49-53. http://dx.doi.org/10.2174/187231207779814292.

Aboul-Ela, EI. (2002). The protective effect of calcium against genotoxicity of lead acetate administration on bone 

marrow and spermatocyte cells of mice in vivo. Mutat Res Genet Toxicol Environ Mutagen 516: 1-9. 

http://dx.doi.org/10.1016/S1383-5718(01)00332-1.

Absalon, D; Slesak, B. (2010). The effects of changes in cadmium and lead air pollution on cancer incidence in 

children. Sci Total Environ 408: 4420-4428. http://dx.doi.org/10.1016/j.scitotenv.2010.06.030.

Adegbesan, BO; Adenuga, GA. (2007). Effect of lead exposure on liver lipid peroxidative and antioxidant defense 

systems of protein-undernourished rats. Biol Trace Elem Res 116: 219-225. 

http://dx.doi.org/10.1007/BF02685932.

Ademuyiwa, O; Ugbaja, RN; Idumebor, F; Adebawo, O. (2005a). Plasma lipid profiles and risk of cardiovascular 

disease in occupational lead exposure in Abeokuta, Nigeria. Lipids Health Dis 4: 19. 

http://dx.doi.org/10.1186/1476-511X-4-19.

Ademuyiwa, O; Ugbaja, RN; Ojo, DA; Owoigbe, AO; Adeokun, SE. (2005b). Reversal of aminolevulinic acid 

dehydratase (ALAD) inhibition and reduction of erythrocyte protoporphyrin levels by Vitamin C in 

occupational lead exposure in Abeokuta, Nigeria. Environ Toxicol Pharmacol 20: 404-411. 

http://dx.doi.org/10.1016/j.etap.2005.04.002.

Ademuyiwa, O; Agarwal, R; Chandra, R; Behari, JR. (2009). Lead-induced phospholipidosis and cholesterogenesis 

in rat tissues. Chem Biol Interact 179: 314-320. http://dx.doi.org/10.1016/j.cbi.2008.10.057.

Adonaylo, VN; Oteiza, PI. (1999). Pb2+ promotes lipid oxidation and alterations in membrane physical properties. 

Toxicology 132: 19-32. http://dx.doi.org/10.1016/S0300-483X(98)00134-6.
February 2012 5-540 Draft – Do Not Cite or Quote

Afeiche, M; Peterson, KE; Sánchez, BN; Cantonwine, D; Lamadrid-Figueroa, H; Schnaas, L; Ettinger, AS; 

Hernández-Avila, M; Hu, H; Téllez-Rojo, MM. (2011). Prenatal lead exposure and weight of 0 to 5 yearold children in Mexico City. Environ Health Perspect 119: 1436-1441. 

http://dx.doi.org/10.1289/ehp.1003184.

Agarwal, S; Roy, S; Ray, A; Mazumder, S; Bhattacharya, S. (2009). Arsenic trioxide and lead acetate induce 

apoptosis in adult rat hepatic stem cells. Cell Biol Toxicol 25: 403-413. http://dx.doi.org/10.1007/s10565-

008-9094-6.

Aguilar, RP; Genta, S; Sanchez, S. (2008). Renal gangliosides are involved in lead intoxication. J Appl Toxicol 28: 

122-131. http://dx.doi.org/10.1002/jat.1256.

Ahamed, M; Verma, S; Kumar, A; Siddiqui, MK. (2005). Environmental exposure to lead and its correlation with 

biochemical indices in children. Sci Total Environ 346: 48-55. 

http://dx.doi.org/10.1016/j.scitotenv.2004.12.019.

Ahamed, M; Verma, S; Kumar, A; Siddiqui, MK. (2006). Delta-aminolevulinic acid dehydratase inhibition and 

oxidative stress in relation to blood lead among urban adolescents. Hum Exp Toxicol 25: 547-553. 

http://dx.doi.org/10.1191/0960327106het657oa.

Ahamed, M; Singh, S; Behari, JR; Kumar, A; Siddiqui, MK. (2007). Interaction of lead with some essential trace 

metals in the blood of anemic children from Lucknow, India. Clin Chim Acta 377: 92-97. 

http://dx.doi.org/10.1016/j.cca.2006.08.032.

Ahamed, M; Siddiqui, MK. (2007). Low level lead exposure and oxidative stress: Current opinions. Clin Chim Acta 

383: 57-64. http://dx.doi.org/10.1016/j.cca.2007.04.024.

Ahamed, M; Fareed, M; Kumar, A; Siddiqui, WA; Siddiqui, MKJ. (2008). Oxidative stress and neurological 

disorders in relation to blood lead levels in children. Redox Rep 13: 117-122. 

http://dx.doi.org/10.1179/135100008X259213.

Ahamed, M; Mehrotra, PK; Kumar, P; Siddiqui, MKJ. (2009). Placental lead-induced oxidative stress and preterm 

delivery. Environ Toxicol Pharmacol 27: 70-74. http://dx.doi.org/10.1016/j.etap.2008.08.013.

Akel, A; Wagner, CA; Kovacikova, J; Kasinathan, RS; Kiedaisch, V; Koka, S; Alper, SL; Bernhardt, I; Wieder, T; 

Huber, SM; Lang, F. (2007). Enhanced suicidal death of erythrocytes from gene-targeted mice lacking the 

Cl-/HCO3- exchanger AE1. Am J Physiol Cell Physiol 292: C1759-C1767. 

http://dx.doi.org/10.1152/ajpcell.00158.2006.

Akesson, A; Lundh, T; Vahter, M; Bjellerup, P; Lidfeldt, J; Nerbrand, C; Goran, S; Stromberg, U; Skerfving, S.

(2005). Tubular and glomerular kidney effects in Swedish women with low environmental cadmium 

exposure. Environ Health Perspect 113: 1627-1631. http://dx.doi.org/10.1289/ehp.8033.

Akesson, A; Bjellerup, P; Lundh, T; Lidfeldt, J; Nerbrand, C; Samsioe, G; Skerfving, S; Vahter, M. (2006). 

Cadmium-induced effects on bone in a population-based study of women. Environ Health Perspect 114: 

830-834. http://dx.doi.org/10.1289/ehp.8763.

Al-Saleh, I; Nester, M; DeVol, E; Shinwari, N; Munchari, L; Al-Shahria, S. (2001). Relationships between blood 

lead concentrations, intelligence, and academic achievement of Saudi Arabian schoolgirls. Int J Hyg 

Environ Health 204: 165-174. http://dx.doi.org/10.1078/1438-4639-00091.

Al-Saleh, I; Coskun, S; Mashhour, A; Shinwari, N; El-Doush, I; Billedo, G; Jaroudi, K; Al-Shahrani, A; Al-Kabra, 

M; El Din Mohamed, G. (2008a). Exposure to heavy metals (lead, cadmium and mercury) and its effect on 

the outcome of in-vitro fertilization treatment. Int J Hyg Environ Health 211: 560-579. 

http://dx.doi.org/10.1016/j.ijheh.2007.09.005.

Al-Saleh, I; Shinwari, N; Nester, M; Mashhour, A; Moncari, L; Mohamed, GED; Rabah, A. (2008b). Longitudinal 

study of prenatal and postnatal lead exposure and early cognitive development in Al-Kharj, Saudi Arabia: 

A preliminary results of cord blood lead levels. J Trop Pediatr 54: 300-307. 

http://dx.doi.org/10.1093/tropej/fmn019.

Alasia, DD; Emem-Chioma, PC; Wokoma, FS. (2010). Occupational and environmental lead exposure in Port 

Harcourt, Nigeria: Analysis of its association with renal function indices. Niger J Med 19: 407-414. 
February 2012 5-541 Draft – Do Not Cite or Quote

Alatise, OI; Schrauzer, GN. (2010). Lead exposure: A contributing cause of the current breast cancer epidemic in 

Nigerian women. Biol Trace Elem Res 136: 127-139. http://dx.doi.org/10.1007/s12011-010-8608-2.

Alber, SA; Strupp, BJ. (1996). An in-depth analysis of lead effects in a delayed spatial alternation task: assessment 

of mnemonic effects, side bias, and proactive interference. Neurotoxicol Teratol 18: 3-15. 

http://dx.doi.org/10.1016/0892-0362(95)02026-8.

Alexander, KR; Fishman, GA. (1984). Supernormal scotopic ERG in cone dystrophy. Br J Ophthalmol 68: 69-78. 

Alfano, DP; Petit, TL. (1981). Behavioral effects of postnatal lead exposure: possible relationship to hippocampal 

dysfunction. Behav Neural Biol 32: 319-333. 

Alghazal, MA; Lenártová, V; Holovská, K; Sobeková, A; Falis, M; Legáth, J. (2008a). Activities of antioxidant and 

detoxifying enzymes in rats after lead exposure. Acta Vet (Beogr) 77: 347-354. 

http://dx.doi.org/10.2754/avb200877030347.

Alghazal, MA; Sutiakova, I; Kovalkovicova, N; Legath, J; Falis, M; Pistl, J; Sabo, R; Benova, K; Sabova, L; Vaczi, 

P. (2008b). Induction of micronuclei in rat bone marrow after chronic exposure to lead acetate trihydrate. 

Toxicol Ind Health 24: 587-593. http://dx.doi.org/10.1177/0748233708100089.

Aligne, CA; Auinger, P; Byrd, RS; Weitzman, M. (2000). Risk factors for pediatric asthma: Contributions of 

poverty, race, and urban residence. Am J Respir Crit Care Med 162: 873-877. 

Alimonti, A; Bocca, B; Lamazza, A; Forte, G; Rahimi, S; Mattei, D; Fiori, E; Iacomino, M; Schillaci, A; De Masi, 

E; Pino, A. (2008). A study on metals content in patients with colorectal polyps. J Toxicol Environ Health 

A 71: 342-347. http://dx.doi.org/10.1080/15287390701839133.

Alinovi, R; Scotti, E; Andreoli, R; De Palma, G; Goldoni, M; Apostoli, P; Mutti, A. (2005). [Neuroendocrine and 

renal effects of inorganic lead]. G Ital Med Lav Ergon 27: 33-38. 

Allouche, L; Hamadouche, M; Touabti, A. (2009). Chronic effects of low lead levels on sperm quality, 

gonadotropins and testosterone in albino rats. Exp Toxicol Pathol 61: 503-510. 

http://dx.doi.org/10.1016/j.etp.2008.12.003.

Alomran, AH; Shleamoon, MN. (1988). The influence of chronic lead exposure on lymphocyte proliferative 

response and immunoglobulin levels in storage battery workers. J Biol Sci 19: 575-585. 

Altmann, L; Weinsberg, F; Sveinsson, K; Lilienthal, H; Wiegand, H; Winneke, G. (1993). Impairment of long-term 

potentiation and learning following chronic lead exposure. Toxicol Lett 66: 105-112. 

http://dx.doi.org/10.1016/0378-4274(93)90085-C.

Altmann, P; Maruna, RF; Maruna, H; Michalica, W; Wagner, G. (1981). [Lead detoxication effect of a combined 

calcium phosphate and ascorbic acid therapy in pregnant women with increased lead burden (author's 

transl)]. Wiener medizinische Wochenschrift 131: 311-314. 

Alvarez, J; Garcia-Sancho, J; Herreros, B. (1986). Inhibition of Ca2+-dependent K+ channels by lead in one-step 

inside-out vesicles from human red cell membranes. Biochim Biophys Acta 857: 291-294. 

http://dx.doi.org/10.1016/0005-2736(86)90359-7.

Amici, A; Emanuelli, M; Magni, G; Raffaelli, N; Ruggieri, S. (1997). Pyrimidine nucleotidases from human 

erythrocyte possess phosphotransferase activities specific for pyrimidine nucleotides. FEBS Lett 419: 263-

267. http://dx.doi.org/10.1016/S0014-5793(97)01464-6.

Andersen, ZJ; Wahlin, P; Raaschou-Nielsen, O; Scheike, T; Loft, S. (2007). Ambient particle source apportionment 

and daily hospital admissions among children and elderly in Copenhagen. J Expo Sci Environ Epidemiol 

17: 625-636. 

Andreae, MO. (1983). Biotransformation of arsenic in the marine environment. In WH Lederer; RJ Fensterheim 

(Eds.), Arsenic: Industrial, biomedical and environmental perspectives: Proceedings of the Arsenic 

Symposium (pp. 378-391). New York, NY: Van Nostrand Reinhold. 

Anetor, JI; Adeniyi, FAA. (1998). Decreased immune status in Nigerian workers occupationally exposed to lead. 

Afr J Med Med Sci 28: 169-172. 
February 2012 5-542 Draft – Do Not Cite or Quote

Annesi-Maesano, I; Pollitt, R; King, G; Bousquet, J; Hellier, G; Sahuquillo, J; Huel, G. (2003). In utero exposure to 

lead and cord blood total IgE: Is there a connection? Allergy 58: 589-594. http://dx.doi.org/10.1034/j.1398-

9995.2003.00111.x.

Antonowicz, J; Andrzejak, R; Smolik, R. (1990). Influence of heavy metal mixtures on erythrocyte metabolism. Int 

Arch Occup Environ Health 62: 195-198. http://dx.doi.org/10.1007/BF00379431.

Antonyuk, SV; Strange, RW; Marklund, SL; Hasnain, SS. (2009). The structure of human extracellular copper-zinc 

superoxide dismutase at 1.7 A resolution: Insights into heparin and collagen binding. J Mol Biol 338: 310-

326. http://dx.doi.org/10.1016/j.jmb.2009.03.026.

Anwar, WA; Kamal, AAM. (1988). Cytogenetic effects in a group of traffic policemen in Cairo. Mutat Res Lett 

208: 225-231. http://dx.doi.org/10.1016/0165-7992(88)90065-6.

Apostoli, P; Romeo, L; De Matteis, MC; Menegazzi, M; Faggionato, G; Vettore, L. (1988). Effects of lead on red 

blood cell membrane proteins. Int Arch Occup Environ Health 61: 71-75. 

http://dx.doi.org/10.1007/BF00381610.

Apostoli, P; Huard, C; Chaumontet, C; Martel, P; Allesio, L; Mazzoleni, G. (2000). Effects of four inorganic lead 

compounds on the proliferation and junctional coupling of cultured REL liver cells. Am J Ind Med 38: 340-

348. 

Aquilonius, SM; Hartvig, P. (1986). A Swedish county with unexpectedly high utilization of anti-parkinsonian 

drugs. Acta Neurol Scand 74: 379-382. http://dx.doi.org/10.1111/j.1600-0404.1986.tb03529.x.

Ari, E; Kaya, Y; Demir, H; Asicioglu, E; Keskin, S. (2011). The Correlation of Serum Trace Elements and Heavy 

Metals with Carotid Artery Atherosclerosis in Maintenance Hemodialysis Patients. Biol Trace Elem Res 

144: 351-359. http://dx.doi.org/10.1007/s12011-011-9103-0.

Ariza, ME; Williams, MV. (1996). Mutagenesis of AS52 cells by low concentrations of lead(II) and mercury(II). 

Environ Mol Mutagen 27: 30-33. 

Ariza, ME; Bijur, GN; Williams, MV. (1998). Lead and mercury mutagenesis: Role of H2O2, superoxide dismutase, 

and xanthine oxidase. Environ Mol Mutagen 31: 352-361. http://dx.doi.org/10.1002/(SICI)1098-

2280(1998)31:4<352::AID-EM8>3.0.CO;2-K.

Ariza, ME; Williams, MV. (1999). Lead and mercury mutagenesis: Type of mutation dependent upon metal 

concentration. J Biochem Mol Toxicol 13: 107-112. http://dx.doi.org/10.1002/(SICI)1099-

0461(1999)13:2<107::AID-JBT6>3.0.CO;2-0.

Arnold, WJ; Kelley, WN. (1978). Adenine phosphoribosyltransferase. Methods Enzymol 51: 568-574. 

http://dx.doi.org/10.1016/S0076-6879(78)51079-3.

Arora, M; Chan, SW; Ryan, CG; Kennedy, BJ; Walker, DM. (2005). Spatial distribution of lead in enamel and 

coronal dentine of wistar rats. Biol Trace Elem Res 105: 159-170. http://dx.doi.org/10.1385/BTER:105:1-

3:159.

Arora, M; Weuve, J; Weisskopf, MG; Sparrow, D; Nie, HL; Garcia, RI; Hu, H. (2009). Cumulative lead exposure 

and tooth loss in men: The normative aging study. Environ Health Perspect 117: 1531-1534. 

http://dx.doi.org/10.1289/ehp.0900739.

Arrieta, O; Palencia, G; Garcia-Arenas, G; Morales-Espinosa, D; Hernandez-Pedro, N; Sotelo, J. (2005). Prolonged 

exposure to lead lowers the threshold of pentylenetetrazole-induced seizures in rats. Epilepsia 46: 1599-

1602. http://dx.doi.org/10.1111/j.1528-1167.2005.00267.x.

Aslani, MR; Najarnezhad, V; Mohri, M. (2010). Individual and combined effect of meso-2,3-dimercaptosuccinic 

acid and allicin on blood and tissue lead content in mice. Planta Med 76: 241-244. 

http://dx.doi.org/10.1055/s-0029-1186141.

Astrin, KH; Bishop, DF; Wetmur, JG; Kaul, B; Davidow, B; Desnick, RJ. (1987). δ-aminolevulinic acid dehydratase 

isozymes and lead toxicity. Ann N Y Acad Sci 514: 23-29. http://dx.doi.org/10.1111/j.1749-

6632.1987.tb48757.x.
February 2012 5-543 Draft – Do Not Cite or Quote

Atabek, ME; Kurtoglu, S; Pirgon, O; Uzum, K; Saraymen, R. (2007). Relation of in utero lead exposure with 

insulin-like growth factor-I levels and neonatal anthropometric parameters. Int J Hyg Environ Health 210: 

91-95. http://dx.doi.org/10.1016/j.ijheh.2006.06.007.

Atchison, WD. (2003). Effects of toxic environmental contaminants on voltage-gated calcium channel function: 

From past to present. J Bioenerg Biomembr 35: 507-532. 

http://dx.doi.org/10.1023/B:JOBB.0000008023.11211.13.

ATS (American Thoracic Society). (2000). What constitutes an adverse health effect of air pollution? This official 

statement of the American Thoracic Society was adopted by the ATS Board of Directors, July 1999. Am J 

Respir Crit Care Med 161: 665-673. 

Attri, J; Dhawan, V; Mahmood, S; Pandhi, P; Parwana, HK; Nath, R. (2003). Effect of vitamin C supplementation 

on oxidative DNA damage in an experimental model of lead-induced hypertension. Ann Nutr Metab 47: 

294-301. http://dx.doi.org/10.1159/000072402.

Audesirk, G; Audesirk, T. (1991). Effects of inorganic lead on voltage-sensitive calcium channels in N1E-115 

neuroblastoma cells. Neurotoxicology 12: 519-528. 

Audesirk, G; Audesirk, T. (1993). The effects of inorganic lead on voltage-sensitive calcium channels differ among 

cell types and among channel subtypes. Neurotoxicology 142: 259-266. 

Ayres, AJ. (1964). Tactile functions. Their relation to hyperactive and perceptual motor behavior. Am J Occup Ther 

18: 6-11. 

Azar, A; Trochimowicz, HJ; Maxfield, ME. (1973). Review of lead studies in animals carried out at Haskell 

Laboratory: Two year feeding study and response to hemorrhage study. In D Barth; A Berlin; R Engel; P 

Recht; J Smeets (Eds.), Environmental health aspects of lead: Proceedings of an international symposium 

(pp. 199-210). Amsterdam, The Netherlands: Commission of the European Communities. 

Baccarelli, A; Bollati, V. (2009). Epigenetics and environmental chemicals. Curr Opin Pediatr 21: 243-251. 

http://dx.doi.org/10.1097/MOP.0b013e32832925cc.

Badavi, M; Mehrgerdi, FZ; Sarkaki, A; Naseri, MK; Dianat, M. (2008). Effect of grape seed extract on lead induced 

hypertension and heart rate in rat. Pak J Biol Sci 11: 882-887. 

Bagchi, D; Preuss, HG. (2005). Effects of acute and chronic oral exposure of lead on blood pressure and bone 

mineral density in rats. J Inorg Biochem 99: 1155-1164. http://dx.doi.org/10.1016/j.jinorgbio.2005.02.011.

Baghurst, PA; McMichael, AJ; Wigg, NR; Vimpani, GV; Robertson, EF; Roberts, RJ; Tong, SL. (1992). 

Environmental exposure to lead and children's intelligence at the age of seven years: The Port Pirie cohort 

study. N Engl J Med 327: 1279-1284. 

Baker, EL; Landrigan, PJ; Barbour, AG; Cox, DH; Folland, DS; Ligo, RN; Throckmorton, J. (1979). Occupational 

lead poisoning in the United States: Clinical and biochemical findings related to blood lead levels. Occup 

Environ Med 36: 314-322. http://dx.doi.org/10.1136/oem.36.4.314.

Baker, EL; Feldman, RG; White, RA; Harley, JP; Niles, CA; Dinse, GE; Berkey, CS. (1984). Occupational lead 

neurotoxicity: A behavioural and electrophysiological evaluation: Study design and year one results. Occup 

Environ Med 41: 352-361. http://dx.doi.org/10.1136/oem.41.3.352.

Baker, EL; White, RF; Pothier, LJ; Berkey, CS; Dinse, GE; Travers, PH; Harley, JP; Feldman, RG. (1985). 

Occupational lead neurotoxicity: Improvement in behavioural effects after reduction of exposure. Br J Ind 

Med 42: 507-516. http://dx.doi.org/10.1136/oem.42.8.507.

Balbuena, P; Li, W; Ehrich, M. (2011). Assessments of tight junction proteins occludin, claudin 5 and scaffold 

proteins ZO1 and ZO2 in endothelial cells of the rat blood-brain barrier: cellular responses to 

neurotoxicants malathion and lead acetate. Neurotoxicology 32: 58-67. 

http://dx.doi.org/10.1016/j.neuro.2010.10.004.

Ball, GV; Sorensen, LB. (1969). Pathogenesis of hyperuricemia in saturnine gout. N Engl J Med 280: 1199-1202. 

http://dx.doi.org/10.1056/nejm196905292802203.
February 2012 5-544 Draft – Do Not Cite or Quote

Bandeen-Roche, K; Glass, TA; Bolla, KI; Todd, AC; Schwartz, BS. (2009). Cumulative lead dose and cognitive 

function in older adults. Epidemiology 20: 831-839. http://dx.doi.org/10.1097/EDE.0b013e3181b5f100.

Bao, QS; Lu, CY; Song, H; Wang, M; Ling, WH; Chen, WQ; Deng, XQ; Hao, YT; Rao, SQ. (2009). Behavioural 

development of school-aged children who live around a multi-metal sulphide mine in Guangdong province, 

China: A cross-sectional study. BMC Public Health 9: 217. http://dx.doi.org/10.1186/1471-2458-9-217.

Baranek, GT; Berkson, G. (1994). Tactile defensiveness in children with developmental disabilities: Responsiveness 

and habituation. J Autism Dev Disord 24: 457-471. http://dx.doi.org/10.1007/BF02172128.

Baranowska-Bosiacka, I; Hlynczak, AJ. (2003). The effect of lead ions on the energy metabolism of human 

erythrocytes in vitro. Comp Biochem Physiol C Toxicol Pharmacol 134: 403-416. 

http://dx.doi.org/10.1016/S1532-0456(03)00008-5.

Baranowska-Bosiacka, I; Hlynczak, AJ. (2004). Effect of lead ions on rat erythrocyte purine content. Biol Trace 

Elem Res 100: 259-273. http://dx.doi.org/10.1385/BTER:100:3:259.

Baranowska-Bosiacka, I; Dziedziejko, V; Safranow, K; Gutowska, I; Marchlewicz, M; Dołegowska, B; Rać, ME; 

Wiszniewska, B; Chlubek, D. (2009). Inhibition of erythrocyte phosphoribosyltransferases (APRT and 

HPRT) by Pb2+: A potential mechanism of lead toxicity. Toxicology 259: 77-83. 

http://dx.doi.org/10.1016/j.tox.2009.02.005.

Baranowska-Bosiacka, I; Gutowska, I; Marchetti, C; Rutkowska, M; Marchlewicz, M; Kolasa, A; Prokopowicz, A; 

Wiernicki, I; Piotrowska, K; Baśkiewicz, M; Safranow, K; Wiszniewska, B; Chlubek, D. (2011a). Altered 

energy status of primary cerebellar granule neuronal cultures from rats exposed to lead in the pre- and 

neonatal period. Toxicology 280: 24-32. http://dx.doi.org/10.1016/j.tox.2010.11.004.

Baranowska-Bosiacka, I; Dąbrowska-Bouta, B; Strużyńska, L. (2011b). Regional changes in purines and selected 

purinergic receptors in immature rat brain exposed to lead. Toxicology 279: 100-107. 

http://dx.doi.org/10.1016/j.tox.2010.09.016.

Barbeito, AG; Martinez-Palma, L; Vargas, MR; Pehar, M; Mañay, N; Beckman, JS; Barbeito, L; Cassina, P. (2010). 

Lead exposure stimulates VEGF expression in the spinal cord and extends survival in a mouse model of 

ALS. Neurobiol Dis 37: 574-580. http://dx.doi.org/10.1016/j.nbd.2009.11.007.

Barbosa, F; Sandrim, VC; Uzuelli, JA; Gerlach, RF; Tanus-Santos, JE. (2006a). eNOS genotype-dependent 

correlation between whole blood lead and plasma nitric oxide products concentrations. Nitric Oxide 14: 58-

64. http://dx.doi.org/10.1016/j.niox.2005.09.007.

Barker, DJ; Osmond, C; Golding, J; Kuh, D; Wadsworth, ME. (1989). Growth in utero, blood pressure in childhood 

and adult life, and mortality from cardiovascular disease. BMJ 298: 564-567. 

Barker, DJP; Bagby, SP. (2005). Developmental antecedents of cardiovascular disease: a historical perspective. J 

Am Soc Nephrol 16: 2537-2544. http://dx.doi.org/10.1681/ASN.2005020160.

Baronas, ET; Lee, JW; Alden, C; Hsieh, FY. (2007). Biomarkers to monitor drug-induced phospholipidosis. Toxicol 

Appl Pharmacol 218: 72-78. http://dx.doi.org/10.1016/j.taap.2006.10.015.

Basha, MR; Wei, W; Brydie, M; Razmiafshari, M; Zawia, NH. (2003). Lead-induced developmental perturbations 

in hippocampal Sp1 DNA-binding are prevented by zinc supplementation: In vivo evidence for Pb and Zn 

competition. Int J Dev Neurosci 21: 1-12. http://dx.doi.org/10.1016/S0736-5748(02)00137-5.

Basha, MR; Murali, M; Siddiqi, HK; Ghosal, K; Siddiqi, OK; Lashuel, HA; Ge, YW; Lahiri, DK; Zawia, NH.

(2005). Lead (Pb) exposure and its effect on APP proteolysis and A beta aggregation. FASEB J 19: 2083-

2084. http://dx.doi.org/10.1096/fj.05-4375fje.

Batalha, JR; Saldiva P, H; Clarke, RW; Coull, BA; Stearns, RC; Lawrence, J; Murthy, GG; Koutrakis, P; Godleski, 

JJ. (2002). Concentrated ambient air particles induce vasoconstriction of small pulmonary arteries in rats. 

Environ Health Perspect 110: 1191-1197. 

Batuman, V; Landy, E; Maesaka, JK; Wedeen, RP. (1983). Contribution of lead to hypertension with renal 

impairment. N Engl J Med 309: 17-21. http://dx.doi.org/10.1056/NEJM198307073090104.

Batuman, V. (1993). Lead nephropathy, gout, and hypertension. Am J Med Sci 305: 241-247. 
February 2012 5-545 Draft – Do Not Cite or Quote

Bayer, SA. (1989). Cellular aspects of brain development. Neurotoxicology 10: 307-320. 

Bear, MF; Malenka, RC. (1994). Synaptic plasticity: LTP and LTD. Curr Opin Neurobiol 4: 389-399. 

http://dx.doi.org/10.1016/0959-4388(94)90101-5.

Beaudin, SA; Stangle, DE; Smith, DR; Levitsky, DA; Strupp, BJ. (2007). Succimer chelation normalizes reactivity 

to reward omission and errors in lead-exposed rats. Neurotoxicol Teratol 29: 188-202. 

http://dx.doi.org/10.1016/j.ntt.2006.11.004.

Bechara, EJH; Medeiros, MHG; Monteiro, HP; Hermes-Lima, M; Pereira, B; Demasi, M; Costa, CA; Abdalla, DSP; 

Onuki, J; Wendel, CMA; Mascio, PD. (1993). A free radical hypothesis of lead poisoning and inborn 

porphyrias associated with 5-aminolevulinic acid overload. Quimica Nova 16: 385-392. 

Behl, M; Zhang, Y; Zheng, W. (2009). Involvement of insulin-degrading enzyme in the clearance of beta-amyloid at 

the blood-CSF barrier: Consequences of lead exposure. Cerebrospinal Fluid Research 6: 11. 

http://dx.doi.org/10.1186/1743-8454-6-11.

Bell, M; Ebisu, K; Peng, R; Samet, J; Dominici, F. (2009). Hospital admissions and chemical composition of fine 

particle air pollution. Am J Respir Crit Care Med 179: 1115-1120. http://dx.doi.org/10.1164/rccm.200808-

1240OC.

Belles-Isles, M; Ayotte, P; Dewailly, E; Weber, JP; Roy, R. (2002). Cord blood lymphocyte functions in newborns 

from a remote maritime population exposed to organochlorines and methylmercury. J Toxicol Environ 

Health A 65: 165-182. http://dx.doi.org/10.1080/152873902753396794.

Bellinger, D; Needleman, HL; Bromfield, R; Mintz, M. (1984). A followup study of the academic attainment and 

classroom behavior of children with elevated dentine lead levels. Biol Trace Elem Res 6: 207-223. 

http://dx.doi.org/10.1007/BF02917507.

Bellinger, D; Leviton, A; Waternaux, C; Needleman, H; Rabinowitz, M. (1987). Longitudinal analyses of prenatal 

and postnatal lead exposure and early cognitive development. N Engl J Med 316: 1037-1043. 

http://dx.doi.org/10.1056/NEJM198704233161701.

Bellinger, D; Leviton, A; Sloman, J. (1990). Antecedents and correlates of improved cognitive performance in 

children exposed in utero to low levels of lead. Environ Health Perspect 89: 5-11. 

Bellinger, D; Sloman, J; Leviton, A; Rabinowitz, M; Needleman, HL; Waternaux, C. (1991). Low-level lead 

exposure and children's cognitive function in the preschool years. Pediatrics 87: 219-227. 

Bellinger, D; Stiles, KM; Needleman, HL. (1992). Low-level lead exposure, intelligence and academic achievement: 

A long-term follow-up study. Pediatrics 90: 855-861. 

Bellinger, D; Stiles, KM. (1993). Epidemiologic approaches to assessing the developmental toxicity of lead. 

Neurotoxicology 14: 151-160. 

Bellinger, D; Hu, H; Titlebaum, L; Needleman, HL. (1994a). Attentional correlates of dentin and bone lead levels in 

adolescents. Arch Environ Health 49: 98-105. http://dx.doi.org/10.1080/00039896.1994.9937461.

Bellinger, D; Leviton, A; Allred, E; Rabinowitz, M. (1994b). Pre- and postnatal lead exposure and behavior 

problems in school-aged children. Environ Res 66: 12-30. http://dx.doi.org/10.1006/enrs.1994.1041.

Bellinger, D; Needleman, HL. (2003). Intellectual impairment and blood lead levels [letter] [Letter]. N Engl J Med 

349: 500. http://dx.doi.org/10.1056/NEJM200307313490515.

Bellinger, D; Rappaport, L. (2002). Developmental assessment and interventions. In B Harvey (Ed.), Managing 

elevated blood lead levels among young children: recommendations from the Advisory Committee on 

Childhood Lead Poisoning Prevention (pp. 79-95). Atlanta, GA: Centers for Disease Control. 

http://www.cdc.gov/nceh/lead/.

Bellinger, DC. (2004a). Assessing environmental neurotoxicant exposures and child neurobehavior: Confounded by 

confounding? Epidemiology 15: 383-384. http://dx.doi.org/10.1097/01.ede.0000129525.15064.a4.
February 2012 5-546 Draft – Do Not Cite or Quote

Bellinger, DC. (2004b). What is an adverse effect? A possible resolution of clinical and epidemiological 

perspectives on neurobehavioral toxicity. Environ Res 95: 394-405. 

http://dx.doi.org/10.1016/j.envres.2003.07.013.

Bellinger, DC. (2007). Interpretation of small effect sizes in occupational and environmental neurotoxicology: 

Individual versus population risk. Neurotoxicology 28: 245-251. 

http://dx.doi.org/10.1016/j.neuro.2006.05.009.

Bellinger, DC. (2008). Email from David Bellinger, Harvard Medical School, to Jee-Young Kim, EPA. SUBJECT: 

Request for additional information on Bellinger and Needleman (2003). (EPA-HQ-OAR-2006-0735-5156). 

Bellinger, DC. http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OAR-2006-0735-5156.

Belloni-Olivi, L; Annadata, M; Goldstein, GW; Bressler, JP. (1996). Phosphorylation of membrane proteins in 

erythrocytes treated with lead. Biochemistry 315: 401-406. 

Bener, A; Obineche, E; Gillett, M; Pasha, MAH; Bishawi, B. (2001a). Association between blood levels of lead, 

blood pressure and risk of diabetes and heart disease in workers. Int Arch Occup Environ Health 74: 375-

378. http://dx.doi.org/10.1007/s004200100231.

Bener, A; Almehdi, AM; Alwash, R; Al-Neamy, FR. (2001b). A pilot survey of blood lead levels in various types of 

workers in the United Arab Emirates. Environ Int 27: 311-314. http://dx.doi.org/10.1016/S0160-

4120(01)00061-7.

Bennet, C; Bettaiya, R; Rajanna, S; Baker, L; Yallapragada, PR; Brice, JJ; White, SL; Bokara, KK. (2007). Region 

specific increase in the antioxidant enzymes and lipid peroxidation products in the brain of rats exposed to 

lead. Free Radic Res 41: 267-273. http://dx.doi.org/10.1080/10715760600889855.

Berenson, GS. (2002). Childhood risk factors predict adult risk associated with subclinical cardiovascular disease: 

The Bogalusa Heart Study. Am J Cardiol 90: 3L-7L. http://dx.doi.org/10.1016/S0002-9149(02)02953-3.

Bergdahl, IA; Grubb, A; Schutz, A; Desnick, RJ; Wetmur, JG; Sassa, S; Skerfving, S. (1997a). Lead binding to 

delta-aminolevulinic acid dehydratase (ALAD) in human erythrocytes. Basic Clin Pharmacol Toxicol 81: 

153-158. http://dx.doi.org/10.1111/j.1600-0773.1997.tb02061.x.

Bergdahl, IA; Schutz, A; Gerhardsson, L; Jensen, A; Skerfving, S. (1997b). Lead concentrations in human plasma, 

urine and whole blood. Scand J Work Environ Health 23: 359-363. 

Bergdahl, IA; Sheveleva, M; Schutz, A; Artamonova, VG; Skerfving, S. (1998). Plasma and blood lead in humans: 

Capacity-limited binding to delta-aminolevulinic acid dehydratase and other lead-binding components. 

Toxicol Sci 46: 247-253. http://dx.doi.org/10.1093/toxsci/46.2.247.

Bergeret, A; Pouget, E; Tedone, R; Meygert, T; Cadot, R; Descotes, J. (1990). Neutrophil functions in lead-exposed 

workers. Hum Exp Toxicol 9: 231-233. http://dx.doi.org/10.1177/096032719000900405.

Berkowitz, Z; Price-Green, P; Bove, FJ; Kaye, WE. (2006). Lead exposure and birth outcomes in five communities 

in Shoshone County, Idaho. Int J Hyg Environ Health 209: 123-132. 

http://dx.doi.org/10.1016/j.ijheh.2005.11.001.

Berrahal, AA; Nehdi, A; Hajjaji, N; Gharbi, N; El-Fazâa, S. (2007). Antioxidant enzymes activities and bilirubin 

level in adult rat treated with lead. C R Biol 330: 581-588. http://dx.doi.org/10.1016/j.crvi.2007.05.007.

Berrahal, AA; Lasram, M; El Elj, N; Kerkeni, A; Gharbi, N; El-Fazâa, S. (2011). Effect of age-dependent exposure 

to lead on hepatotoxicity and nephrotoxicity in male rats. Environ Toxicol 26: 68-78. 

http://dx.doi.org/10.1002/tox.20530.

Berry, WD, Jr; Moriarty, CM; Lau, YS. (2002). Lead attenuation of episodic growth hormone secretion in male rats. 

Int J Toxicol 21: 93-98. 

Bhattacharya, A; Shukla, R; Dietrich, K; Bornschein, R; Berger, O. (1995). Effect of early lead exposure on 

children's postural balance. Dev Med Child Neurol 37: 861-878. 

Bhattacharya, A; Shukla, R; Dietrich, KN; Bornschein, RL. (2006). Effect of early lead exposure on the maturation 

of children's postural balance: A longitudinal study. Neurotoxicol Teratol 3: 376-385. 

http://dx.doi.org/10.1016/j.ntt.2006.02.003.
February 2012 5-547 Draft – Do Not Cite or Quote

Bhatti, P; Stewart, PA; Hutchinson, A; Rothman, N; Linet, MS; Inskip, PD; Rajaraman, P. (2009). Lead exposure, 

polymorphisms in genes related to oxidative stress, and risk of adult brain tumors. Cancer Epidemiol 

Biomarkers Prev 18: 1841-1848. http://dx.doi.org/10.1158/1055-9965.EPI-09-0197.

Bielarczyk, H; Tian, X; Suszkiw, JB. (1996). Cholinergic denervation-like changes in rat hippocampus following 

developmental lead exposure. Brain Res 708: 108-115. http://dx.doi.org/10.1016/0006-8993(95)01315-6.

Bihaqi, SW; Huang, H; Wu, J; Zawia, NH. (2011). Infant exposure to lead (Pb) and epigenetic modifications in the 

aging primate brain: Implications for Alzheimer's disease. J Alzheimers Dis 27: 819-833. 

http://dx.doi.org/10.3233/JAD-2011-111013.

Bilban, M. (1998). Influence of the work environment in a Pb-Zn mine on the incidence of cytogenetic damage in 

miners. Am J Ind Med 34: 455-463. 

Bishayi, B; Sengupta, M. (2006). Synergism in immunotoxicological effects due to repeated combined 

administration of arsenic and lead in mice. Int Immunopharmacol 6: 454-464. 

http://dx.doi.org/10.1016/j.intimp.2005.09.011.

Biswas, NM; Ghosh, PK. (2006). Protection of adrenal and male gonadal functions by androgen in lead-treated rats. 

Kathmandu Univ Med J 4: 218-221. 

Bitto, E; Bingman, CA; Wesenberg, GE; McCoy, JG; Phillips, GN, Jr. (2006). Structure of pyrimidine 5'-

nucleotidase type 1: Insight into mechanism of action and inhibition during lead poisoning. J Biol Chem 

281: 20521-20529. http://dx.doi.org/10.1074/jbc.M602000200.

Blackman, SS, Jr. (1936). Intranuclear inclusion bodies in the kidney and liver caused by lead poisoning. Johns 

Hopkins Med J 58: 384-402. 

Bleecker, ML; Lindgren, KN; Ford, DP. (1997). Differential contribution of current and cumulative indices of lead 

dose to neuropsychological performance by age. Neurology 48: 639-645. 

Bleecker, ML; Ford, DP; Vaughan, CG; Walsh, KS; Lindgren, KN. (2007a). The association of lead exposure and 

motor performance mediated by cerebral white matter change. Neurotoxicology 28: 318-323. 

http://dx.doi.org/10.1016/j.neuro.2006.04.008.

Bleecker, ML; Ford, DP; Celio, MA; Vaughan, CG; Lindgren, KN. (2007b). Impact of cognitive reserve on the 

relationship of lead exposure and neurobehavioral performance. Neurology 69: 470-476. 

http://dx.doi.org/10.1212/01.wnl.0000266628.43760.8c.

Bloom, MS; Parsons, PJ; Steuerwald, AJ; Schisterman, EF; Browne, RW; Kim, K; Coccaro, GA; Conti, GC; 

Narayan, N; Fujimoto, VY. (2010). Toxic trace metals and human oocytes during in vitro fertilization 

(IVF). Reprod Toxicol 29: 298-305. http://dx.doi.org/10.1016/j.reprotox.2010.01.003.

Bloom, MS; Louis, GM; Sundaram, R; Kostyniak, PJ; Jain, J. (2011a). Associations between blood metals and 

fecundity among women residing in New York State. Reprod Toxicol 31: 158-163. 

http://dx.doi.org/10.1016/j.reprotox.2010.09.013.

Bloom, MS; Parsons, PJ; Kim, D; Steuerwald, AJ; Vaccari, S; Cheng, G; Fujimoto, VY. (2011b). Toxic trace metals 

and embryo quality indicators during in vitro fertilization (IVF). Reprod Toxicol 31: 164-170. 

http://dx.doi.org/10.1016/j.reprotox.2010.11.011.

Boerrigter, G; Burnett, JC, Jr. (2009). Soluble guanylate cyclase: Not a dull enzyme. Circulation 119: 2752-2754. 

http://dx.doi.org/10.1161/CIRCULATIONAHA.109.860288.

Boillee, S; Yamanaka, K; Lobsiger, CS; Copeland, NG; Jenkins, NA; Kassiotis, G; Kollias, G; Cleveland, DW.

(2006). Onset and progression in inherited ALS determined by motor neurons and microglia. Science 312: 

1389-1392. http://dx.doi.org/10.1126/science.1123511.

Bokara, KK; Blaylock, I; Denise, SB; Bettaiya, R; Rajanna, S; Yallapragada, PR. (2009). Influence of lead acetate 

on glutathione and its related enzymes in different regions of rat brain. J Appl Toxicol 29: 452-458. 

http://dx.doi.org/10.1002/jat.1423.
February 2012 5-548 Draft – Do Not Cite or Quote

Bolin, CM; Basha, R; Cox, D; Zawia, NH; Maloney, B; Lahiri, DK; Cardozo-Pelaez, F. (2006). Exposure to lead 

(Pb) and the developmental origin of oxidative DNA damage in the aging brain. FASEB J 20: 788-790. 

http://dx.doi.org/10.1096/fj.05-5091fje.

Bollati, V; Baccarelli, A; Hou, L; Bonzini, M; Fustinoni, S; Cavallo, D; Byun, HM; Jiang, J; Marinelli, B; Pesatori, 

AC; Bertazzi, PA; Yang, AS. (2007). Changes in DNA methylation patterns in subjects exposed to lowdose benzene. Cancer Res 67: 876-880. http://dx.doi.org/10.1158/0008-5472.CAN-06-2995.

Bonacker, D; Stoiber, T; Bohm, KJ; Prots, I; Wang, M; Unger, E; Thier, R; Bolt, HM; Degen, GH. (2005). 

Genotoxicity of inorganic lead salts and disturbance of microtubule function. Environ Mol Mutagen 45: 

346-353. http://dx.doi.org/10.1002/em.20100.

Borgstahl, GE; Parge, HE; Hickey, MJ; Beyer, WF, Jr; Hallewell, RA; Tainer, JA. (1992). The structure of human 

mitochondrial manganese superoxide dismutase reveals a novel tetrameric interface of two 4-helix bundles. 

Cell 71: 107-118. http://dx.doi.org/10.1016/0092-8674(92)90270-M.

Boscolo, P; Di Gioacchino, M; Sabbioni, E; Benvenuti, F; Conti, P; Reale, M; Bavazzano, P; Giuliano, G. (1999). 

Expression of lymphocyte subpopulations, cytokine serum levels, and blood and urinary trace elements in 

asymptomatic atopic men exposed to an urban environment. Int Arch Occup Environ Health 72: 26-32. 

http://dx.doi.org/10.1007/s004200050330.

Boscolo, P; De Gioacchino, M; Sabbioni, E; Di Giacomo, F; Reale, M; Volpe, AR; Di Sciascio, MB; Conti, P; 

Giuliano, G. (2000). Lymphocyte subpopulations, cytokines and trace elements in asymptomatic atopic 

women exposed to an urban environment. Life Sci 67: 1119-1126. http://dx.doi.org/10.1016/S0024-

3205(00)00712-8.

Bouchard, MF; Bellinger, DC; Weuve, J; Matthews-Bellinger, J; Gilman, SE; Wright, RO; Schwartz, J; Weisskopf, 

MG. (2009). Blood lead levels and major depressive disorder, panic disorder, and generalized anxiety 

disorder in US young adults. Arch Gen Psychiatry 66: 1313-1319. 

http://dx.doi.org/10.1001/archgenpsychiatry.2009.164.

Boudene, C; Despaux-Pages, N; Comoy, E; Bohuon, C. (1984). Immunological and enzymatic studies of 

erythrocytic delta-aminolevulinate dehydratase: Comparison of results obtained in normal and leadexposed subjects. Int Arch Occup Environ Health 55: 87-96. http://dx.doi.org/10.1007/BF00378071.

Bound, JP; Harvey, PW; Francis, BJ; Awwad, F; Gatrell, AC. (1997). Involvement of deprivation and 

environmental lead in neural tube defects: A matched case-control study. Arch Dis Child 76: 107-112. 

http://dx.doi.org/10.1136/adc.76.2.107.

Bouton, CM; Frelin, LP; Forde, CE; Arnold Godwin, H; Pevsner, J. (2001). Synaptotagmin I is a molecular target 

for lead. J Neurochem 76: 1724-1735. http://dx.doi.org/10.1046/j.1471-4159.2001.00168.x.

Bowers, TS; Beck, BD. (2006). What is the meaning of non-linear dose-response relations between blood lead 

concentrations and IQ? Neurotoxicology 27: 520-524. http://dx.doi.org/10.1016/j.neuro.2006.02.001.

Bradberry, S; Vale, A. (2009). A comparison of sodium calcium edetate (edetate calcium disodium) and succimer 

(DMSA) in the treatment of inorganic lead poisoning. Clin Toxicol 47: 841-858. 

http://dx.doi.org/10.3109/15563650903321064.

Bragadin, M; Marton, D; Murgia, M; Rizzoli, V; Scutari, G; Deana, R. (1998). Interactions of trialkyllead 

compounds with rat liver mitochondria. J Inorg Biochem 69: 259-262. http://dx.doi.org/10.1016/S0162-

0134(97)10036-8.

Bragadin, M; Marton, D; Manente, S. (2007). Trialkyllead compounds induce the opening of the MTP pore in rat 

liver mitochondria. J Inorg Biochem 101: 876-878. http://dx.doi.org/10.1016/j.jinorgbio.2007.01.016.

Bratton, GR; Hiney, JK; Dees, WL. (1994). Lead (Pb) alters the norepinephrine-induced secretion of luteinizing 

hormone releasing hormone from the medium eminence of adult male rats in vitro. Life Sci 55: 563-571. 

http://dx.doi.org/10.1016/0024-3205(94)00482-X.

Braun, JM; Kahn, RS; Froehlich, T; Auinger, P; Lanphear, BP. (2006). Exposures to environmental toxicants and 

attention deficit hyperactivity disorder in U.S. children. Environ Health Perspect 114: 1904-1909. 

http://dx.doi.org/10.1289/ehp.9478.
February 2012 5-549 Draft – Do Not Cite or Quote

Braun, JM; Froehlich, TE; Daniels, JL; Dietrich, KN; Hornung, R; Auinger, P; Lanphear, BP. (2008). Association of 

environmental toxicants and conduct disorder in U.S. children: NHANES 2001-2004. Environ Health 

Perspect 116: 956-962. http://dx.doi.org/10.1289/ehp.11177.

Bravo, Y; Quiroz, Y; Ferrebuz, A; Vaziri, ND; Rodriguez-Iturbe, B. (2007). Mycophenolate mofetil administration 

reduces renal inflammation, oxidative stress, and arterial pressure in rats with lead-induced hypertension. 

Am J Physiol Renal Physiol 293: F616-F623. http://dx.doi.org/10.1152/ajprenal.00507.2006.

Brender, JD; Suarez, L; Felkner, M; Gilani, Z; Stinchcomb, D; Moody, K; Henry, J; Hendricks, K. (2006). Maternal 

exposure to arsenic, cadmium, lead, and mercury and neural tube defects in offspring. Environ Res 101: 

132-139. http://dx.doi.org/10.1016/j.envres.2005.08.003.

Brockel, BJ; Cory-Slechta, DA. (1998). Lead, attention, and impulsive behavior: Changes in a fixed-ratio waitingfor-reward paradigm. Pharmacol Biochem Behav 60: 545-552. http://dx.doi.org/10.1016/S0091-

3057(98)00023-9.

Brockel, BJ; Cory-Slechta, DA. (1999). The effects of postweaning low-level Pb exposure on sustained attention: a 

study of target densities, stimulus presentation rate, and stimulus predictability. Neurotoxicology 20: 921-

933. 

Brondum, J. (2007). Environmental exposures and ADHD [Letter]. Environ Health Perspect 115: A398; author 

reply A399. http://dx.doi.org/10.1289/ehp.10274.

Brondum, J. (2011). ADHD, lead, and PCBs: Appropriate comparison studies [Letter]. Environ Health Perspect 119: 

a282-a282. http://dx.doi.org/10.1289/ehp.1103513.

Brubaker, CJ; Schmithorst, VJ; Haynes, EN; Dietrich, KN; Egelhoff, JC; Lindquist, DM; Lanphear, BP; Cecil, KM.

(2009). Altered myelination and axonal integrity in adults with childhood lead exposure: A diffusion tensor 

imaging study. Neurotoxicology 30: 867-875. http://dx.doi.org/10.1016/j.neuro.2009.07.007.

Brubaker, CJ; Dietrich, KN; Lanphear, BP; Cecil, KM. (2010). The influence of age of lead exposure on adult gray 

matter volume. Neurotoxicology 31: 259-266. http://dx.doi.org/10.1016/j.neuro.2010.03.004.

Bryant-Stephens, T. (2009). Asthma disparities in urban environments. J Allergy Clin Immunol 123: 1199-1206. 

http://dx.doi.org/10.1016/j.jaci.2009.04.030.

Buchanan, LH; Counter, SA; Ortega, F. (2011). Environmental lead exposure and otoacoustic emissions in Andean 

children. J Toxicol Environ Health A 74: 1280-1293. http://dx.doi.org/10.1080/15287394.2011.587106.

Budtz-Jorgensen, E; Keiding, N; Grandjean, P. (2001). Benchmark dose calculation from epidemiological data. 

Biometrics 57: 698-706. http://dx.doi.org/10.1111/j.0006-341X.2001.00698.x.

Bunn, TL; Marsh, JA; Dietert, RR. (2000). Gender differences in developmental immunotoxicity to lead in a 

chicken: Analysis following a single early low-level exposure in ovo. J Toxicol Environ Health A 61: 677-

693. http://dx.doi.org/10.1080/00984100050195152.

Bunn, TL; Ladics, GS; Holsapple, MP; Dietert, RR. (2001a). Developmental immunotoxicology assessment in the 

rat: Age, gender and strain comparisons after exposure to Pb. Toxicol Mech Meth 11: 41-58. 

http://dx.doi.org/10.1080/105172301300055151.

Bunn, TL; Parsons, PJ; Kao, E; Dietert, RR. (2001b). Exposure to lead during critical windows of embryonic 

development: Differential immunotoxic outcome based on stage of exposure and gender. Toxicol Sci 64: 

57-66. http://dx.doi.org/10.1093/toxsci/64.1.57.

Bunn, TL; Parsons, PJ; Kao, E; Dietert, RR. (2001c). Gender-based profiles of developmental immunotoxicity to 

lead in the rat: Assessment in juveniles and adults. J Toxicol Environ Health A 64: 223-240. 

http://dx.doi.org/10.1080/15287390152543708.

Burger, J; Gochfeld, M. (2005). Effects of lead on learning in herring gulls: An avian wildlife model for 

neurobehavioral deficits. Neurotoxicology 26: 615-624. http://dx.doi.org/10.1016/j.neuro.2005.01.005.

Burns, JM; Baghurst, PA; Sawyer, MG; McMichael, AJ; Tong, SL. (1999). Lifetime low-level exposure to 

environmental lead and children's emotional and behavioral development at ages 11-13 years: The Port 

Pirie cohort study. Am J Epidemiol 149: 740-749. 
February 2012 5-550 Draft – Do Not Cite or Quote

Bussolaro, D; Neto, FF; Gargioni, R; Fernandes, LC; Randi, MAF; Pelletier, E; Oliveira Ribeiro, CA. (2008). The 

immune response of peritoneal macrophages due to exposure to inorganic lead in the house mouse Mus 

musculus. Toxicol In Vitro 22: 254-260. http://dx.doi.org/10.1016/j.tiv.2007.09.003.

Cabaravdic, M; Mijanovic, M; Kusturica, J; Cabaravdic, A. (2010). Occupational exposure of workers at gas station 

to inorganic lead. Med Arh 64: 107-109. 

Cabb, EE; Gorospe, EC; Rothweiler, AM; Gerstenberger, SL. (2008). Toxic remedy: A case of a 3-year-old child 

with lead colic treated with lead monoxide (Greta). Clin Pediatr (Phila) 47: 77-79. 

http://dx.doi.org/10.1177/0009922807304385.

Caffo, B; Chen, S; Stewart, W; Bolla, K; Yousem, D; Davatzikos, C; Schwartz, BS. (2008). Are brain volumes 

based on magnetic resonance imaging mediators of the associations of cumulative lead dose with cognitive 

function? Am J Epidemiol 167: 429-437. http://dx.doi.org/10.1093/aje/kwm326.

Cai, MY; Arenaz, P. (1998). Antimutagenic effect of crown ethers on heavy metal-induced sister chromatid 

exchanges. Mutagenesis 13: 27-32. http://dx.doi.org/10.1093/mutage/13.1.27.

Calderon-Salinas, JV; Quintanar-Escorza, MA; Hernandez-Luna, CE; Gonzalez-Martinez, MT. (1999a). Effect of 

lead on the calcium transport in human erythrocyte. Hum Exp Toxicol 18: 146-153. 

http://dx.doi.org/10.1177/09603271990180030319901800303.

Calderon-Salinas, JV; Quintanar-Escorcia, MA; Gonzalez-Martinez, MT; Hernandez-Luna, CE. (1999b). Lead and 

calcium transport in human erythrocyte. Hum Exp Toxicol 18: 327-332. 

http://dx.doi.org/10.1191/096032799678840138.

Calvo, FB; Santos, D; Rodrigues, CJ; Krug, FJ; Marumo, JT; Schor, N; Bellini, MH. (2009). Variation in the 

distribution of trace elements in renal cell carcinoma. Biol Trace Elem Res 130: 107-113. 

http://dx.doi.org/10.1007/s12011-009-8325-x.

Camoratto, AM; White, LM; Lau, YS; Ware, GO; Berry, WD; Moriarty, CM. (1993). Effect of exposure to low 

level lead on growth and growth hormone release in rats. Toxicology 83: 101-114. 

http://dx.doi.org/10.1016/0300-483X(93)90095-A.

Campagna, D; Huel, G; Hellier, G; Girard, F; Sahuquillo, J; Fagot-Campagna, A; Godin, J; Blot, P. (2000). Negative 

relationships between erythrocyte Ca-pump activity and lead levels in mothers and newborns. Life Sci 68: 

203-215. http://dx.doi.org/10.1016/S0024-3205(00)00928-0.

Campbell, JR; Auinger, P. (2007). The association between blood lead levels and osteoporosis among adults: 

Results from the Third National Health and Nutrition Examination Survey (NHANES III). Environ Health 

Perspect 115: 1018-1022. http://dx.doi.org/10.1289/ehp.9716.

Camyre, E; Wise, SS; Milligan, P; Gordon, N; Goodale, B; Stackpole, M; Patzlaff, N; Aboueissa, AM; Wise, JP.

(2007). Ku80 deficiency does not affect particulate chromate-induced chromosome damage and 

cytotoxicity in chinese hamster ovary cells. Toxicol Sci 97: 348-354. 

http://dx.doi.org/10.1093/toxsci/kfm045.

Can, S; Bagci, C; Ozaslan, M; Bozkurt, AI; Cengiz, B; Cakmak, EA; Kocabas, R; Karadag, E; Tarakcioglu, M.

(2008). Occupational lead exposure effect on liver functions and biochemical parameters. Acta Physiol 

Hung 95: 395-403. http://dx.doi.org/10.1556/APhysiol.95.2008.4.6.

Canfield, R. (2008). Email from Dr. Richard Canfield, Cornell University, to Jee-Young Kim, USEPA Subject: 

Request for additional information on Canfield et al. (2003). (EPA-HQ-OAR-2006-0735-5811). Canfield, 

R. http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OAR-2006-0735-5811.

Canfield, RL; Henderson, CR, Jr; Cory-Slechta, DA; Cox, C; Jusko, TA; Lanphear, BP. (2003a). Intellectual 

impairment in children with blood lead concentrations below 10 micrograms per deciliter. N Engl J Med 

348: 1517-1526. http://dx.doi.org/10.1056/NEJMoa022848.

Canfield, RL; Kreher, DA; Cornwell, C; Henderson, CR, Jr. (2003b). Low-level lead exposure, executive 

functioning, and learning in early childhood. Child Neuropsychol 9: 35-53. 

http://dx.doi.org/10.1076/chin.9.1.35.14496.
February 2012 5-551 Draft – Do Not Cite or Quote

Canfield, RL; Gendle, MH; Cory-Slechta, DA. (2004). Impaired neuropsychological functioning in lead-exposed 

children. Dev Neuropsychol 26: 513-540. http://dx.doi.org/10.1207/s15326942dn2601_8.

Cantarow, A; Trumper, M. (1944). Lead poisoning. Hagerstown, MD: Williams & Wilkins Co. 

Cantonwine, D; Hu, H; Sánchez, BN; Lamadrid-Figueroa, H; Smith, D; Ettinger, AS; Mercado-García, A; 

Hernández-Avila, M; Wright, RO; Téllez-Rojo, MM. (2010a). Critical windows of fetal lead exposure: 

Adverse impacts on length of gestation and risk of premature delivery. J Occup Environ Med 52: 1106-

1111. http://dx.doi.org/10.1097/JOM.0b013e3181f86fee.

Cantonwine, D; Hu, H; Tellez-Rojo, MM; Sanchez, BN; Lamadrid-Figueroa, H; Ettinger, AS; Mercado-Garcia, A; 

Hernandez-Avila, M; Wright, RO. (2010b). HFE gene variants modify the association between maternal 

lead burden and infant birthweight: A prospective birth cohort study in Mexico City, Mexico. Environ 

Health Global Access Sci Source 9: 43. http://dx.doi.org/10.1186/1476-069X-9-43.

Cao, SQ; Chen, ZY; Liu, GQ; Jiang, L; Yuan, HB; Ren, G; Bian, XH; Jian, HY; Ma, XL. (2009). The Arabidopsis 

Ethylene-Insensitive 2 gene is required for lead resistance. Plant Physiol Biochem 47: 308-312. 

http://dx.doi.org/10.1016/j.plaphy.2008.12.013.

Cao, XJ; Huang, SH; Wang, M; Chen, JT; Ruan, DY. (2008). S-adenosyl-L-methionine improves impaired 

hippocampal long-term potentiation and water maze performance induced by developmental lead exposure 

in rats. Eur J Pharmacol 595: 30-34. http://dx.doi.org/10.1016/j.ejphar.2008.07.061.

Carafoli, E. (2005). The Theodor Bucher Lecture - Calcium - a universal carrier of biological signals. Delivered on 3 

July 2003 at the Special FEBS Meeting in Brussels. FEBS J 272: 1073-1089. 

http://dx.doi.org/10.1111/j.1742-4658.2005.04546.x.

Cardenas, A; Roels, H; Bernard, AM; Barbon, R; Buchet, JP; Lauwerys, RR; Rosello, J; Ramis, I; Mutti, A; 

Franchini, I; Fels, LM; Stolte, H; De Broe, ME; Nuyts, GD; Taylor, SA; Price, RG. (1993). Markers of 

early renal changes induced by industrial pollutants: II Application to workers exposed to lead. Br J Ind 

Med 50: 28-36. http://dx.doi.org/10.1136/oem.50.1.28.

Carey, JB; Allshire, A; Van Pelt, FN. (2006). Immune modulation by cadmium and lead in the acute reporter 

antigen-popliteal lymph node assay. Toxicol Sci 91: 113-122. http://dx.doi.org/10.1093/toxsci/kfj142.

Carmignani, M; Boscolo, P; Poma, A; Volpe, AR. (1999). Kininergic system and arterial hypertension following 

chronic exposure to inorganic lead. Int Immunopharmacol 44: 105-110. http://dx.doi.org/10.1016/S0162-

3109(99)00115-0.

Carmignani, M; Volpe, AR; Boscolo, P; Qiao, N; MDi, G; Grilli, A; Felaco, M. (2000). Catcholamine and nitric 

oxide systems as targets of chronic lead exposure in inducing selective functional impairment. Life Sci 68: 

401-415. http://dx.doi.org/10.1016/S0024-3205(00)00954-1.

Carmona, ER; Creus, A; Marcos, R. (2011). Genotoxicity testing of two lead-compounds in somatic cells of 

Drosophila melanogaster. Mutat Res 724: 35-40. http://dx.doi.org/10.1016/j.mrgentox.2011.05.008.

Carreras, HA; Wannaz, ED; Pignata, ML. (2009). Assessment of human health risk related to metals by the use of 

biomonitors in the province of Cordoba, Argentina. Environ Pollut 157: 117-122. 

http://dx.doi.org/10.1016/j.envpol.2008.07.018.

Casadoa, MF; Cecchinia, AL; Simãoa, ANC; Oliveiraa, RD; Cecchini, R. (2007). Free radical-mediated prehemolytic injury in human red blood cells subjected to lead acetate as evaluated by chemiluminescence. 

Food Chem Toxicol 45: 945-952. http://dx.doi.org/10.1016/j.fct.2006.12.001.

Caylak, E; Aytekin, M; Halifeoglu, I. (2008). Antioxidant effects of methionine, alpha-lipoic acid, N-acetylcysteine 

and homocysteine on lead-induced oxidative stress to erythrocytes in rats. Exp Toxicol Pathol 60: 289-294. 

http://dx.doi.org/10.1016/j.etp.2007.11.004.

Cecil, K; Yuan, W; Holland, S; Wessel, S; Dietrich, K; Ris, D; Lanphear, B. (2005). The influence of childhood lead 

exposure on language function in young adults: An FMRI study [Abstract]. Proc Int Soc Magn Reson Med 

13: 1441. 
February 2012 5-552 Draft – Do Not Cite or Quote

Cecil, KM; Brubaker, CJ; Adler, CM; Dietrich, KN; Altaye, M; Egelhoff, JC; Wessel, S; Elangovan, I; Hornung, R; 

Jarvis, K; Lanphear, BP. (2008). Decreased brain volume in adults with childhood lead exposure. PLoS 

Med 5: e112. http://dx.doi.org/10.1371/journal.pmed.0050112.

Cecil, KM; Dietrich, KN; Altaye, M; Egelhoff, JC; Lindquist, DM; Brubaker, CJ; Lanphear, BP. (2011). Proton 

magnetic resonance spectroscopy in adults with childhood lead exposure. Environ Health Perspect 119: 

403-408. http://dx.doi.org/10.1289/ehp.1002176.

Celik, A; Ogenler, O; Comelekoglu, U. (2005). The evaluation of micronucleus frequency by acridine orange 

fluorescent staining in peripheral blood of rats treated with lead acetate. Mutagenesis 20: 411-415. 

http://dx.doi.org/10.1093/mutage/gei055.

Cerulli, N; Campanella, L; Grossi, R; Politi, L; Scandurra, R; Soda, G; Gallo, F; Damiani, S; Alimonti, A; Petrucci, 

F; Caroli, S. (2006). Determination of Cd, Cu, Pb and Zn in neoplastic kidneys and in renal tissue of 

fetuses, newborns and corpses. J Trace Elem Med Biol 20: 171-179. 

http://dx.doi.org/10.1016/j.jtemb.2006.03.002.

Cezard, C; Demarquilly, C; Boniface, M; Haguenor, JM. (1992). Influence of the degree of exposure to lead on 

relations between alcohol consumption and the biological indices of lead exposure: Epidemiological study 

in a lead acid battery factory. Occup Environ Med 49: 645-647. http://dx.doi.org/10.1136/oem.49.9.645.

Chai, S; Webb, RC. (1988). Effects of lead on vascular reactivity. Environ Health Perspect 78: 85-89. 

Chakraborty, I; Sharma, A; Talukder, G. (1987). Antagonistic and synergistic effects of lead and selenium in Rattus 

norvegicus. Toxicol Lett 37: 21-26. http://dx.doi.org/10.1016/0378-4274(87)90162-7.

Chancellor, AM; Slattery, JM; Fraser, H; Warlow, CP. (1993). Risk factors for motor neuron disease: A case-control 

study based on patients from the Scottish Motor Neuron Disease Register. J Neurol Neurosurg Psychiatry 

56: 1200-1206. http://dx.doi.org/10.1136/jnnp.56.11.1200.

Chandramouli, K; Steer, CD; Ellis, M; Emond, AM. (2009). Effects of early childhood lead exposure on academic 

performance and behaviour of school age children. Arch Dis Child 94: 844-848. 

http://dx.doi.org/10.1136/adc.2008.149955.

Chang, HR; Chen, SS; Tsao, DA; Cheng, JT; Ho, CK; Yu, HS. (1997). Change of cardiac "beta"-adrenoceptors in 

lead-exposed rats. Toxicology 123: 27-32. http://dx.doi.org/10.1016/S0300-483X(97)00100-5.

Chang, HR; Tsao, DA; Yu, HS; Ho, CK. (2005). The change of "beta"-adrenergic system after cessation of lead 

exposure. Toxicology 207: 73-80. http://dx.doi.org/10.1016/j.tox.2004.08.018.

Chang, MK; Raggatt, LJ; Alexander, KA; Kuliwaba, JS; Fazzalari, NL; Schroder, K; Maylin, ER; Ripoll, VM; 

Hume, DA; Pettit, AR. (2008a). Osteal tissue macrophages are intercalated throughout human and mouse 

bone lining tissues and regulate osteoblast function in vitro and in vivo. J Immunol 181: 1232-1244. 

Chang, SH; Cheng, BH; Lee, SL; Chuang, HY; Yang, CY; Sung, FC; Wu, TN. (2006). Low blood lead 

concentration in association with infertility in women. Environ Res 101: 380-386. 

http://dx.doi.org/10.1016/j.envres.2005.10.004.

Chang, WC; Chang, CC; Wang, YS; Weng, WT; Yoshioka, T; Juo, SH. (2011). Involvement of the epidermal 

growth factor receptor in Pb(2+)-induced activation of cPLA(2)/COX-2 genes and PGE(2) production in 

vascular smooth muscle cells. Toxicology 279: 45-53. http://dx.doi.org/10.1016/j.tox.2010.09.004.

Chang, YF; Teng, HC; Cheng, SY; Wang, CT; Chiou, SH; Kao, LS; Kao, FJ; Chiou, A; Yang, DM. (2008b). OrailSTIM1 formed store-operated Ca[sub]2+[/sub] channels (SOCs) as the molecular components needed for 

Pb[sub]2+[/sub] entry in living cells. Toxicol Appl Pharmacol 227: 430-439. 

http://dx.doi.org/10.1016/j.taap.2007.11.025.

Chao, SL; Moss, JM; Harry, GJ. (2007). Lead-induced alterations of apoptosis and neurotrophic factor mRNA in the 

developing rat cortex, hippocampus, and cerebellum. J Biochem Mol Toxicol 21: 265-272. 

http://dx.doi.org/10.1002/jbt.20191.

Chaurasia, SS; Kar, A. (1997). Protective effects of vitamin E against lead-induced deterioration of membrane 

associated type-I iodothyronine 5'-monodeiodinase (5'D-I) activity in male mice. Toxicology 124: 203-209. 

http://dx.doi.org/10.1016/S0300-483X(97)00155-8.
February 2012 5-553 Draft – Do Not Cite or Quote

Chaurasia, SS; Gupta, P; Maiti, PK; Kar, A. (1998). Possible involvement of lipid peroxidation in the inhibition of 

type I iodothyronine 5'-monodeiodinase activity by lead in chicken liver. J Appl Toxicol 18: 299-300. 

Chen, A; Dietrich, KN; Ware, JH; Radcliffe, J; Rogan, WJ. (2005). IQ and blood lead from 2 to 7 years of age: Are 

the effects in older children the residual of high blood lead concentrations in 2-year-olds? Environ Health 

Perspect 113: 597-601. http://dx.doi.org/10.1289/ehp.7625.

Chen, A; Cai, B; Dietrich, KN; Radcliffe, J; Rogan, WJ. (2007). Lead exposure, IQ, and behavior in urban 5- to 7-

year-olds: Does lead affect behavior only by lowering IQ? Pediatrics 119: e650-e658. 

http://dx.doi.org/10.1542/peds.2006-1973.

Chen, B; Lamberts, LV; Behets, GJ; Zhao, TT; Zhou, MX; Liu, G; Hou, XH; Guan, GJ; D'Haese, PC. (2009a). 

Selenium, lead, and cadmium levels in renal failure patients in China. Biol Trace Elem Res 131: 1-12. 

http://dx.doi.org/10.1007/s12011-009-8340-y.

Chen, HH; Chan, MH. (2002). Developmental lead exposure differentially alters the susceptibility to 

chemoconvulsants in rats. Toxicology 173: 249-257. http://dx.doi.org/10.1016/S0300-483X(02)00032-X.

Chen, HI; Chiu, YW; Hsu, YK; Li, WF; Chen, YC; Chuang, HY. (2010a). The association of metallothionein-4 

gene polymorphism and renal function in long-term lead-exposed workers. Biol Trace Elem Res 137: 55-

62. http://dx.doi.org/10.1007/s12011-009-8564-x.

Chen, J; Mercer, G; Roth, SR; Abraham, L; Lutz, P; Ercal, N; Neal, RE. (2011). Sub-chronic lead exposure alters 

kidney proteome profiles. Hum Exp Toxicol 30: 1616-1625. http://dx.doi.org/10.1177/0960327110396521.

Chen, PC; Pan, IJ; Wang, JD. (2006a). Parental exposure to lead and small for gestational age births. Am J Ind Med 

49: 417-422. http://dx.doi.org/10.1002/ajim.20313.

Chen, S; Golemboski, KA; Sanders, FS; Dietert, RR. (1999). Persistent effect of in utero meso-2,3-

dimercaptosuccinic acid (DMSA) on immune function and lead-induced immunotoxicity. Toxicology 132: 

67-79. http://dx.doi.org/10.1016/S0300-483X(98)00139-5.

Chen, S; Golemboski, KA; Piepenbrink, M; Dietert, RR. (2004). Developmental immunotoxicity of lead in the rat: 

Influence of maternal diet. J Toxicol Environ Health A 67: 495-511. 

http://dx.doi.org/10.1080/15287390490276520.

Chen, XK; Yang, Q; Smith, G; Krewski, D; Walker, M; Wen, SW. (2006c). Environmental lead level and 

pregnancy-induced hypertension. Environ Res 100: 424-430. 

http://dx.doi.org/10.1016/j.envres.2005.07.006.

Chen, Y; Zhao, JX; Liu, JW; Cui, J; Li, L; Tian, W. (2008c). Lack of association of delta-aminolevulinic acid 

dehydratase genotype with blood lead levels in environmentally exposed children of Uygur and Han 

populations. Acta Paediatr 97: 1717-1720. http://dx.doi.org/10.1111/j.1651-2227.2008.01003.x.

Cheng, Y; Schwartz, J; Vokonas, PS; Weiss, ST; Aro, A; Hu, H. (1998). Electrocardiographic conduction 

disturbances in association with low-level lead exposure (the Normative Aging Study). Am J Cardiol 82: 

594-599. http://dx.doi.org/10.1016/S0002-9149(98)00402-0.

Cheng, Y; Schwartz, J; Sparrow, D; Aro, A; Weiss, ST; Hu, H. (2001). Bone lead and blood lead levels in relation to 

baseline blood pressure and the prospective development of hypertension: The Normative Aging Study. 

Am J Epidemiol 153: 164-171. http://dx.doi.org/10.1093/aje/153.2.164.

Cheng, YJ; Yang, BC; Hsieh, WC; Huang, BM; Liu, MY. (2002). Enhancement of TNF-alpha expression does not 

trigger apoptosis upon exposure of glial cells to lead and lipopolysaccharide. Toxicology 178: 183-191. 

http://dx.doi.org/10.1016/S0300-483X(02)00225-1.

Cheng, YJ; Yang, BC; Liu, MY. (2006). Lead increases lipopolysaccharide-induced liver injury through tumor 

necrosis factor-alpha overexpression by monocytes/macrophages: Role of protein kinase C and p42/44 

mitogen-activated protein kinase. Environ Health Perspect 114: 507-513. 

http://dx.doi.org/10.1289/ehp.8550.

Chetty, CS; Reddy, GR; Murthy, KS; Johnson, J; Sajwan, K; Desaiah, D. (2001). Perinatal lead exposure alters the 

expression of neuronal nitric oxide synthase in rat brain. Int J Toxicol 20: 113-120. 

http://dx.doi.org/10.1080/109158101317097692.
February 2012 5-554 Draft – Do Not Cite or Quote

Chetty, CS; Vemuri, MC; Campbell, K; Suresh, C. (2005). Lead-induced cell death of human neuroblastoma cells 

involves GSH deprivation. Cell Mol Biol Lett 10: 413-423. 

Chia, SE; Zhou, H; Tham, MT; Yap, E; Dong, NV; Tu, NH; Chia, KS. (2005). Possible influence of "delta"-

aminolevulinic acid dehydratase polymorphism and susceptibility to renal toxicity of lead: A study of a 

Vietnamese population. Environ Health Perspect 113: 1313-1317. http://dx.doi.org/10.1289/ehp.7904.

Chia, SE; Zhou, HJ; Yap, E; Tham, MT; Dong, NV; Hong Tu, NT; Chia, KS. (2006). Association of renal function 

and delta-aminolevulinic acid dehydratase polymorphism among Vietnamese and Singapore workers 

exposed to inorganic lead. Occup Environ Med 63: 180-186. http://dx.doi.org/10.1136/oem.2005.021154.

Chia, SE; Zhou, HJ; Theng, TM; Yap, E. (2007). Possibilities of newer ALAD polymorphism influencing human 

susceptibility to effects of inorganic lead on the neurobehavioral functions. Neurotoxicology 28: 312-317. 

http://dx.doi.org/10.1016/j.neuro.2006.04.003.

Chiodo, LM; Jacobson, SW; Jacobson, JL. (2004). Neurodevelopmental effects of postnatal lead exposure at very 

low levels. Neurotoxicol Teratol 26: 359-371. http://dx.doi.org/10.1016/j.ntt.2004.01.010.

Chiodo, LM; Covington, C; Sokol, RJ; Hannigan, JH; Jannise, J; Ager, J; Greenwald, M; Delaney-Black, V. (2007). 

Blood lead levels and specific attention effects in young children. Neurotoxicol Teratol 29: 538-546. 

http://dx.doi.org/10.1016/j.ntt.2007.04.001.

Chiu, TY; Teng, HC; Huang, PC; Kao, FJ; Yang, DM. (2009). Dominant role of Orai1 with STIM1 on the cytosolic 

entry and cytotoxicity of lead ions. Toxicol Sci 110: 353-362. http://dx.doi.org/10.1093/toxsci/kfp099.

Cho, SC; Kim, BN; Hong, YC; Shin, MS; Yoo, HJ; Kim, JW; Bhang, SY; Cho, IH; Kim, HW. (2010). Effect of 

environmental exposure to lead and tobacco smoke on inattentive and hyperactive symptoms and 

neurocognitive performance in children. J Child Psychol Psychiatry 51: 1050-1057. 

http://dx.doi.org/10.1111/j.1469-7610.2010.02250.x.

Chobanian, AV; Bakris, GL; Black, HR; Cushman, WC; Green, LA; Izzo, JL, Jr; Jones, DW; Materson, BJ; Oparil, 

S; Wright, JT, Jr; Roccella, EJ. (2003). Seventh report of the Joint National Committee on Prevention, 

Detection, Evaluation, and Treatment of High Blood Pressure. Hypertension 42: 1206-1252. 

http://dx.doi.org/10.1161/01.HYP.0000107251.49515.c2.

Choie, DD; Richter, GW. (1972). Lead poisoning: Rapid formation of intranuclear inclusions. Science 177: 1194-

1195. http://dx.doi.org/10.1126/science.177.4055.1194.

Choie, DD; Richter, GW; Young, LB. (1975). Biogenesis of intranuclear lead-protein inclusions in mouse kidney. 

Pathol Res Pract 155: 197-203. 

Chou, YH; Woon, PY; Huang, WC; Shiurba, R; Tsai, YT; Wang, YS; Hsieh, TJ; Chang, WC; Chuang, HY; Chang, 

WC. (2011). Divalent lead cations induce cyclooxygenase-2 gene expression by epidermal growth factor 

receptor/nuclear factor-kappa B signaling in A431carcinoma cells. Toxicol Lett 203: 147-153. 

http://dx.doi.org/10.1016/j.toxlet.2011.03.017.

Chuang, HY; Schwartz, J; Gonzales-Cossio, T; Lugo, MC; Palazuelos, E; Aro, A; Hu, H; Hernandez-Avila, M.

(2001). Interrelations of lead levels in bone, venous blood, and umbilical cord blood with exogenous lead 

exposure through maternal plasma lead in peripartum women. Environ Health Perspect 109: 527-532. 

http://dx.doi.org/10.1289/ehp.01109527.

Chuang, HY; Kuo, CH; Chiu, YW; Ho, CK; Chen, CJ; Wu, TN. (2007). A case-control study on the relationship of 

hearing function and blood concentrations of lead, manganese, arsenic, and selenium. Sci Total Environ 

387: 79-85. http://dx.doi.org/10.1016/j.scitotenv.2007.07.032.

Church, HJ; Day, JP; Braithwaite, RA; Brown, SS. (1993a). Binding of lead to a metallothionein-like protein in 

human erythrocytes. J Inorg Biochem 49: 55-68. http://dx.doi.org/10.1016/0162-0134(93)80048-E.

Church, HJ; Day, JP; Braithwaite, RA; Brown, SS. (1993b). The speciation of lead in erythrocytes in relation to lead 

toxicity: Case studies of two lead-exposed workers. Neurotoxicology 14: 359-364.

Ciubar, R; Ciofrangeanu, C; Mitran, V; Cimpean, A; Stanescu, A; Iordachescu, D. (2007). The effect of lead ions on 

human erythrocytes in vitro. Rev Chim 58: 895-897. 
February 2012 5-555 Draft – Do Not Cite or Quote

Clark, CS; Bornschein, RL; Succop, P; Que Hee, SS; Hammond, PB; Peace, B. (1985). Condition and type of 

housing as an indicator of potential environmental lead exposure and pediatric blood lead levels. Environ 

Res 38: 46-53. http://dx.doi.org/10.1016/0013-9351(85)90071-4.

Claus Henn, B; Schnaas, L; Ettinger, AS; Schwartz, J; Lamadrid-Figueroa, H; Hernández-Avila, M; 

Amarasiriwardena, C; Hu, H; Bellinger, DC; Wright, RO; Téllez-Rojo, MM. (2011). Associations of Early 

Childhood Manganese and Lead Co-exposure with Neurodevelopment. Environ Health Perspect 120: 126-

131. http://dx.doi.org/10.1289/ehp.1003300.

Cline, HT; Witte, S; Jones, KW. (1996). Low lead levels stunt neuronal growth in a reversible manner. PNAS 93: 

9915-9920. 

Coban, TA; Senturk, M; Ciftci, M; Kufrevioglu, OI. (2007). Effects of some metal ions on human erythrocyte 

glutathione reductase: an in vitro study. Protein Pept Lett 14: 1027-1030. 

http://dx.doi.org/10.2174/092986607782541060.

Cocco, P; Fadda, D; Atzeri, S; Avataneo, G; Meloni, M; Flore, C. (2007). Causes of death among lead smelters in 

relation to the glucose-6-phosphate dehydrogenase polymorphism. Occup Environ Med 64: 414-416. 

http://dx.doi.org/10.1136/oem.2006.028779.

Cockcroft, DW; Gault, MH. (1976). Prediction of creatinine clearance from serum creatinine. Nephron 16: 31-41. 

Cohen, BS; Li, W; Xiong, JQ; Lippmann, M. (2000). Detecting H+ in ultrafine ambient aerosol using iron nano-film 

detectors and scanning probe microscopy. Appl Occup Environ Hyg 15: 80-89. 

http://dx.doi.org/10.1080/104732200301881.

Cohen, N; Modai, D; Golik, A; Weissgarten, J; Peller, S; Katz, A; Averbukh, Z; Shaked, U. (1989). Increased 

concanavalin A-induced suppressor cell activity in humans with occupational lead exposure. Environ Res 

48: 1-6. http://dx.doi.org/10.1016/S0013-9351(89)80079-9.

Columbano, A; Ledda-Columbano, GM; Coni, P; Pani, P. (1987). Failure of mitogen-induced cell proliferation to 

achieve initiation of rat liver carcinogenesis. Carcinogenesis 8: 345-347. 

http://dx.doi.org/10.1093/carcin/8.2.345.

Columbano, A; Ledda-Columbano, GM; Ennas, MG; Curto, M; Chelo, A; Pani, P. (1990). Cell proliferation and 

promotion of rat liver carcinogenesis: Different effect of hepatic regeneration and mitogen induced 

hyperplasia on the development of enzyme-altered foci. Carcinogenesis 11: 771-776. 

http://dx.doi.org/10.1093/carcin/11.5.771.

Columbano, A; Endoh, T; Denda, A; Noguchi, O; Nakae, D; Hasegawa, K; Ledda-Columbano, GM; Zedda, AI; 

Konishi, Y. (1996). Effects of cell proliferation and cell death (apoptosis and necrosis) on the early stages 

of rat hepatocarcinogenesis. Carcinogenesis 17: 395-400. http://dx.doi.org/10.1093/carcin/17.3.395.

Comper, WD; Russo, LM. (2009). The glomerular filter: An imperfect barrier is required for perfect renal function. 

Curr Opin Nephrol Hypertens 18: 336-342. http://dx.doi.org/10.1097/Mnh.0b013e32832cb96a.

Coni, P; Pichiri-Coni, G; Ledda-Columbano, GM; Semple, E; Rajalakshmi, S; Rao, PM; Sarma, DSR; Columbano, 

A. (1992). Stimulation of DNA synthesis by rat plasma following in vivo treatment with three liver 

mitogens. Cancer Lett 61: 233-238. http://dx.doi.org/10.1016/0304-3835(92)90293-5.

Conroy, MC; Adkinson, NF; lichtenstein, LM. (1977). Measurement of IgE on human basophils: Relation to serum 

IgE and anti-IgE-induced histamine release. J Immunol 118: 1317-1321. 

Conterato, GM; Bulcão, RP; Sobieski, R; Moro, AM; Charão, MF; Freitas, FA; Almeida, FL; Moreira, AP; Roehrs, 

M; Tonello, R; Batista, BL; Grotto, D; Barbosa, F; Garcia, SC; Emanuelli, T. (In Press) Blood thioredoxin 

reductase activity, oxidative stress and hematological parameters in painters and battery workers: 

Relationship with lead and cadmium levels in blood. J Appl Toxicol. http://dx.doi.org/10.1002/jat.1731.

Conterato, GMM; Augusti, PR; Somacal, S; Einsfeld, L; Sobieski, R; Torres, JRV; Emanuelli, T. (2007). Effect of 

lead acetate on cytosolic thioredoxin reductase activity and oxidative stress parameters in rat kidneys. Basic 

Clin Pharmacol Toxicol 101: 96-100. http://dx.doi.org/10.1111/j.1742-7843.2007.00084.x.
February 2012 5-556 Draft – Do Not Cite or Quote

Coon, S; Stark, A; Peterson, E; Gloi, A; Kortsha, G; Pounds, J; Chettle, D; Gorell, J. (2006). Whole-body lifetime 

occupational lead exposure and risk of Parkinson's disease. Environ Health Perspect 114: 1872-1876. 

http://dx.doi.org/10.1289/ehp.9102.

Cooper, GP; Manalis, RS. (1984). Interactions of lead and cadmium on acetylcholine release at the frog 

neuromuscular junction. Toxicol Appl Pharmacol 74: 411-416. http://dx.doi.org/10.1016/0041-

008X(84)90294-1.

Coratelli, P; Giannattasio, M; Lomonte, C; Marzolla, R; Rana, F; L'Abbate, N. (1988). Enzymuria to detect tubular 

injury in workers exposed to lead: A 12-month follow-up. In C Bianchi; V Bocci; FA Carone; R Rabkin 

(Eds.), Kidney and proteins in health and disease: Fifth international symposium in health and disease; July 

1987; Montecatini Terme, Italy (pp. 207-211). Basel, Switzerland: S. Karger. 

Coria, C; Cabello, A; Tassara, E; Lopez, E; Rosales, H; Perez, M; Zavala, C; Munoz, P; Orellana, G; Inostroza, MI; 

Contreras, L; Kirsten, L. (2009). [Long term consequences among children exposed to lead poisoning]. Rev 

Med Chil 137: 1037-1044. http://dx.doi.org/10.4067/S0034-98872009000800006.

Coria, F; Berciano, MT; Berciano, J; Lafarga, M. (1984). Axon membrane remodeling in the lead-induced 

demyelinating neuropathy of the rat. Brain Res 291: 369-372. http://dx.doi.org/10.1016/0006-

8993(84)91271-x.

Corongiu, FP; Milia, A. (1982). Rise of hepatic glutathione concentration induced in rats by chronic lead nitrate 

treatment. Its role in aflatoxin B1 intoxication. Res Comm Chem Pathol Pharmacol 38: 97-112. 

Cory-Slechta, DA. (1990). Exposure duration modifies the effects of low level lead on fixed-interval performance. 

Neurotoxicology 11: 427-441. 

Cory-Slechta, DA; Pokora, MJ; Widzowski, DV. (1991). Behavioral manifestations of prolonged lead exposure 

initiated at different stages of the life cycle: II. Delayed spatial alternation. Neurotoxicology 12: 761-776. 

Cory-Slechta, DA; Pokora, MJ; Widzowski, DV. (1992). Postnatal lead exposure induces supersensitivity to the 

stimulus properties of D2D3 agonist. Brain Res 598: 162-172. http://dx.doi.org/10.1016/0006-

8993(92)90180-H.

Cory-Slechta, DA; O'Mara, DJ; Brockel, BJ. (1998). Nucleus accumbens dopaminergic medication of fixed interval 

schedule-controlled behavior and its modulation by low-level lead exposure. J Pharmacol Exp Ther 286: 

794-805. 

Cory-Slechta, DA; Virgolini, MB; Thiruchelvam, M; Weston, DD; Bauter, MR. (2004). Maternal stress modulates 

the effects of developmental lead exposure. Environ Health Perspect 112: 717-730. 

http://dx.doi.org/10.1289/ehp.6481.

Cory-Slechta, DA; Virgolini, MB; Rossi-George, A; Thiruchelvam, M; Lisek, R; Weston, D. (2008). Lifetime 

consequences of combined maternal lead and stress. Basic Clin Pharmacol Toxicol 102: 218-227. 

http://dx.doi.org/10.1111/j.1742-7843.2007.00189.x.

Cory-Slechta, DA; Virgolini, MB; Rossi-George, A; Weston, D; Thiruchelvam, M. (2009). Experimental 

manipulations blunt time-induced changes in brain monoamine levels and completely reverse stress, but not 

Pb +/- stress-related modifications to these trajectories. Behav Brain Res 205: 76-87.

http://dx.doi.org/10.1016/j.bbr.2009.06.040.

Cory-Slechta, DA; Stern, S; Weston, D; Allen, JL; Liu, S. (2010). Enhanced learning deficits in female rats 

following lifetime pb exposure combined with prenatal stress. Toxicol Sci 117: 427-438. 

http://dx.doi.org/10.1093/toxsci/kfq221.

Cory-Slechta, DA. (1994). Neurotoxicant-induced changes in schedule-controlled behavior. In LW Chang (Ed.), 

Principles of neurotoxicology (pp. 313-344). New York, NY: Marcel Dekker, Inc. 

Costa, LG; Fox, DA. (1983). A selective decrease of cholinergic muscarinic receptors in the visual cortex of adult 

rats following developmental lead exposure. Brain Res 276: 259-266. http://dx.doi.org/10.1016/0006-

8993(83)90733-3.
February 2012 5-557 Draft – Do Not Cite or Quote

Costa, M; Zhitkovich, A; Gargas, M; Paustenbach, D; Finley, B; Kuykendall, J; Billings, R; Carlson, TJ; 

Wetterhahn, K; Xu, J; Patierno, S; Bogdanffy, M. (1996). Interlaboratory validation of a new assay for 

DNA-protein crosslinks. Mutat Res Genet Toxicol 369: 13-21. http://dx.doi.org/10.1016/S0165-

1218(96)90043-9.

Counter, SA; Buchanan, LH; Ortega, F; Rifai, N; Shannon, MW. (2007). Comparative analysis of zinc 

protoporphyrin and blood lead levels in lead-exposed Andean children. Clin Biochem 40: 787-792. 

http://dx.doi.org/10.1016/j.clinbiochem.2007.03.003.

Counter, SA; Buchanan, LH; Ortega, F. (2008). Zinc protoporphyrin levels, blood lead levels and neurocognitive 

deficits in Andean children with chronic lead exposure. Clin Biochem 41: 41-47. 

http://dx.doi.org/10.1016/j.clinbiochem.2007.10.002.

Counter, SA; Buchanan, LH; Ortega, F. (2009). Neurocognitive screening of lead-exposed andean adolescents and 

young adults. J Toxicol Environ Health A 72: 625-632. http://dx.doi.org/10.1080/15287390902769410.

Counter, SA; Buchanan, LH; Ortega, F; van der Velde, J; Borg, E. (2011). Assessment of auditory brainstem 

function in lead-exposed children using stapedius muscle reflexes. J Neurol Sci 306: 29-37. 

http://dx.doi.org/10.1016/j.jns.2011.04.003.

Courtois, E; Marques, M; Barrientos, A; Casado, S; Lopez-Farre, A. (2003). Lead-induced downregulation of 

soluble guanylage cyclase in isolated rat aortic segments mediated by reactive oxygen species and 

cyclooxygenase-2. J Am Soc Nephrol 14: 1464-1470. http://dx.doi.org/10.1097/01.ASN.0000064947.1499.

Coyle, JT; Tsai, G. (2004). The NMDA receptor glycine modulatory site: A therapeutic target for improving 

cognition and reducing negative symptoms in schizophrenia. Psychopharmacology 174: 32-38. 

http://dx.doi.org/10.1007/s00213-003-1709-2.

Croes, K; Baeyens, W; Bruckers, L; Den Hond, E; Koppen, G; Nelen, V; Van de Mieroop, E; Keune, H; Dhooge, 

W; Schoeters, G; Van Larebeke, N. (2009). Hormone levels and sexual development in Flemish 

adolescents residing in areas differing in pollution pressure. Int J Hyg Environ Health 212: 612-625. 

http://dx.doi.org/10.1016/j.ijheh.2009.05.002.

Crofton, KM; Taylor, DH; Bull, RJ; Sivulka, DJ; Lutkenhoff, SD. (1980). Developmental delays in exploration and 

locomotor activity in male rats exposed to low level lead. Life Sci 26: 823-831. 

http://dx.doi.org/10.1016/0024-3205(80)90289-1.

Crooks, DR; Ghosh, MC; Haller, RG; Tong, WH; Rouault, TA. (2010). Posttranslational stability of the heme 

biosynthetic enzyme ferrochelatase is dependent on iron availability and intact iron-sulfur cluster assembly 

machinery. Blood 115: 860-869. http://dx.doi.org/10.1182/blood-2009-09-243105.

Daggett, DA; Oberley, TD; Nelson, SA; Wright, LS; Kornguth, SE; Siegel, FL. (1998). Effects of lead on rat kidney 

and liver: GST expression and oxidative stress. Toxicology 128: 191-206. http://dx.doi.org/10.1016/S0300-

483X(98)00080-8.

Danadevi, K; Rozati, R; Saleha Banu, B; Hanumanth Rao, P; Grover, P. (2003). DNA damage in workers exposed 

to lead using comet assay. Toxicology 187: 183-193. http://dx.doi.org/10.1016/S0300-483X(03)00054-4.

Daniels, SR. (2011). International differences in secular trends in childhood blood pressure: A puzzle to be solved 

[Editorial]. Circulation 124: 378-380. http://dx.doi.org/10.1161/CIRCULATIONAHA.111.042747.

Davenport, A; Murcutt, G; Whiting, S. (2009). Cross-sectional audit of blood lead levels in regular outpatient 

haemodialysis patients dialysing in north London. Nephrology 14: 476-481. 

http://dx.doi.org/10.1111/j.1440-1797.2009.01087.x.

David, O; Clark, J; Voeller, K. (1972). Lead and hyperactivity. Lancet 300: 900-903. 

http://dx.doi.org/10.1016/S0140-6736(72)92534-2.

Davidson, R; MacKinnon, JG. (1981). Several tests for model specification in the presence of alternative 

hypotheses. Econometrica 49: 781-793. 

Davis, JM. (1982). Ethological approaches to behavioral toxicology. In CL Mitchell (Ed.), Nervous system 

toxicology (pp. 29-44). New York, NY: Raven Press. 
February 2012 5-558 Draft – Do Not Cite or Quote

De, M; Ghosh, S; Palit, S; Ghosh, A; Talukder, G; Sharma, A. (1995). Clastogenic effects in human samples 

following prolonged exposure in metal industry. Bull Environ Contam Toxicol 54: 357-362. 

http://dx.doi.org/10.1007/BF00195105.

de Restrepo, HG; Sicard, D; Torres, MM. (2000). DNA damage and repair in cells of lead exposed people. Am J Ind 

Med 38: 330-334. http://dx.doi.org/10.1002/1097-0274(200009)38:3<330::AID-AJIM13>3.0.CO;2-Z.

De Burbure, C; Buchet, JP; Leroyer, A; Nisse, C; Haguenoer, JM; Mutti, A; Smerhovsky, Z; Cikrt, M; TrczinkaOchocka, M; Razniewska, G; Jakubowski, M; Bernard, A. (2006). Renal and neurologic effects of 

cadmium, lead, mercury, and arsenic in children: Evidence of early effects and multiple interactions at 

environmental exposure levels. Environ Health Perspect 114: 584-590. http://dx.doi.org/10.1289/ehp.8202.

De Coster, S; Koppen, G; Bracke, M; Schroijen, C; Den Hond, E; Nelen, V; Van de Mieroop, E; Bruckers, L; Bilau, 

M; Baeyens, W; Schoeters, G; van Larebeke, N. (2008). Pollutant effects on genotoxic parameters and 

tumor-associated protein levels in adults: A cross sectional study. Environ Health Global Access Sci Source 

7: 26. http://dx.doi.org/10.1186/1476-069X-7-26.

De Keyser, J; Mostert, JP; Koch, MW. (2008). Dysfunctional astrocytes as key players in the pathogenesis of central 

nervous system disorders [Review]. J Neurol Sci 267: 3-16. http://dx.doi.org/10.1016/j.jns.2007.08.044.

De Palma, G; Goldoni, M; Catalani, S; Carbognani, P; Poli, D; Mozzoni, P; Acampa, O; Internullo, E; Rusca, M; 

Apostoli, P. (2008). Metallic elements in pulmonary biopsies from lung cancer and control subjects. Ateneo 

Parmense 1 79: 43-51. 

de Souza Lisboa, SF; Goncalves, G; Komatsu, F; Queiroz, CAS; Almeida, AA; Moreira, EG. (2005). 

Developmental lead exposure induces depressive-like behavior in female rats. Drug Chem Toxicol 28: 67-

77. http://dx.doi.org/10.1081/dct-200039696.

Dearth, RK; Hiney, JK; Srivastava, V; Burdick, SB; Bratton, GR; Dees, WL. (2002). Effects of lead (Pb) exposure 

during gestation and lactation on female pubertal development in the rat. Reprod Toxicol 16: 343-352. 

http://dx.doi.org/10.1016/S0890-6238(02)00037-0.

Degawa, M; Arai, H; Miura, S; Hashimoto, Y. (1993). Preferential inhibitions of hepatic P450IA2 expression and 

induction by lead nitrate in the rat. Carcinogenesis 14: 1091-1094. 

http://dx.doi.org/10.1093/carcin/14.6.1091.

Dehlink, E; Baker, AH; Yen, E; Nurko, S; Fiebiger, E. (2010). Relationships between levels of serum IgE, cellbound IgE, and IgE-receptors on peripheral blood cells in a pediatric population. PLoS ONE 5: e12204. 

http://dx.doi.org/10.1371/journal.pone.0012204.

Dehpour, AR; Essalat, M; Ala, S; Ghazi-Khansari, M; Ghafourifar, P. (1999). Increase by NO synthase inhibitor of 

lead-induced release of N-acetyl-beta-D-glucosaminidase from perfused rat kidney. Toxicology 132: 119-

125. http://dx.doi.org/10.1016/S0300-483X(98)00143-7.

Dempster, EL; Pidsley, R; Schalkwyk, LC; Owens, S; Georgiades, A; Kane, F; Kalidindi, S; Picchioni, M; 

Kravariti, E; Toulopoulou, T; Murray, RM; Mill, J. (2011). Disease-associated epigenetic changes in 

monozygotic twins discordant for schizophrenia and bipolar disorder. Hum Mol Genet 20: 4786-4796. 

http://dx.doi.org/10.1093/hmg/ddr416.

Den Hond, E, .; Dhooge, W, .; Bruckers, L, .; Schoeters, G, .; Nelen, V, .; van de Mieroop, E, .; Koppen, G, .; Bilau, 

M, .; Schroijen, C, .; Keune, H, .; Baeyens, W, .; van Larebeke, N, . (2011). Internal exposure to pollutants 

and sexual maturation in Flemish adolescents. J Expo Sci Environ Epidemiol 21: 224-233. 

http://dx.doi.org/10.1038/jes.2010.2.

Deng, H; Callender, R; Schramm, VL; Grubmeyer, C. (2010). Pyrophosphate activation in hypoxanthine--guanine 

phosphoribosyltransferase with transition state analogue. Biochemistry 49: 2705-2714. 

http://dx.doi.org/10.1021/bi100012u.

Deng, W; Poretz, RD. (2002). Protein kinase C activation is required for the lead-induced inhibition of proliferation 

and differentiation of cultured oligodendroglial progenitor cells. Brain Res 929: 87-95. 

http://dx.doi.org/10.1016/S0006-8993(01)03385-6.
February 2012 5-559 Draft – Do Not Cite or Quote

Denham, M; Schell, LM; Deane, G; Gallo, MV; Ravenscroft, J; DeCaprio, AP. (2005). Relationship of lead, 

mercury, mirex, dichlorodiphenyldichloroethylene, hexachlorobenzene, and polychlorinated biphenyls to 

timing of menarche among Akwesasne Mohawk girls. Pediatrics 115: e127-e134. 

http://dx.doi.org/10.1542/peds.2004-1161.

Dentener, MA; Greve, JW; Maessen, JG; Buurman, WA. (1989). Role of tumour necrosis factor in the enhanced 

sensitivity of mice to endotoxin after exposure to lead. Immunopharmacol Immunotoxicol 11: 321-334. 

http://dx.doi.org/10.3109/08923978909005373.

Despres, C; Beuter, A; Richer, F; Poitras, K; Veilleux, A; Ayotte, P; Dewailly, E; Saint-Amour, D; Muckle, G.

(2005). Neuromotor functions in Inuit preschool children exposed to Pb, PCBs, and Hg. Neurotoxicol 

Teratol 27: 245-257. http://dx.doi.org/10.1016/j.ntt.2004.12.001.

Devarajan, P. (2007). Emerging biomarkers of acute kidney injury. Contrib Nephrol 156: 203-212. 

http://dx.doi.org/10.1159/000102085.

Deveci, E. (2006). Ultrastructural effects of lead acetate on brain of rats. Toxicol Ind Health 22: 419-422. 

http://dx.doi.org/10.1177/07482337060220100101.

Devi, CB; Reddy, GH; Prasanthi, RPJ; Chetty, CS; Reddy, GR. (2005). Developmental lead exposure alters 

mitochondrial monoamine oxidase and synaptosomal catecholamine levels in rat brain. Int J Dev Neurosci 

23: 375-381. http://dx.doi.org/10.1016/j.ijdevneu.2004.11.003.

Devi, KD; Banu, BS; Grover, P; Jamil, K. (2000). Genotoxic effect of lead nitrate on mice using SCGE (Comet 

assay). Toxicology 145: 195-201. http://dx.doi.org/10.1016/S0300-483X(00)00154-2.

Dhir, H; Roy, AK; Sharma, A; Talukder, G. (1990). Modification of clastogenicity of lead and aluminium in mouse 

bone marrow cells by dietary ingestion of Phyllanthus emblica fruit extract. Mutat Res Genet Toxicol 241: 

305-312. http://dx.doi.org/10.1016/0165-1218(90)90029-2.

Dhir, H; Ghosh, S; Sharma, A; Talukder, G. (1992a). Interaction between two group IV metals--lead and zirconium-

-in bone marrow cells of Mus musculus in vivo. Biometals 5: 81-86. 

http://dx.doi.org/10.1007/BF01062218.

Dhir, H; Sharma, A; Talukder, G. (1992b). Modifying effect of iron on lead-induced clastogenicity in mouse bone 

marrow cells. Biol Trace Elem Res 34: 279-286. http://dx.doi.org/10.1007/BF02783683.

Dhir, H; Roy, AK; Sharma, A. (1993). Relative efficiency of Phyllanthus emblica fruit extract and ascorbic acid in 

modifying lead and aluminium-induced sister-chromatid exchanges in mouse bone marrow. Environ Mol 

Mutagen 21: 229-236. http://dx.doi.org/10.1002/em.2850210305.

Di Lorenzo, L; Silvestroni, A; Martino, MG; Gagliardi, T; Corfiati, M; Soleo, L. (2006). Evaluation of peripheral 

blood neutrophil leucocytes in lead-exposed workers. Int Arch Occup Environ Health 79: 491-498. 

http://dx.doi.org/10.1007/s00420-005-0073-4.

Di Lorenzo, L; Vacca, A; Corfiati, M; Lovreglio, P; Soleo, L. (2007). Evaluation of tumor necrosis factor-alpha and 

granulocyte colony-stimulating factor serum levels in lead-exposed smoker workers. Int J Immunopathol 

Pharmacol 20: 239-247. 

Dietert, RR; Piepenbrink, MS. (2006). Perinatal immunotoxicity: Why adult exposure assessment fails to predict 

risk. Environ Health Perspect 114: 477-483. http://dx.doi.org/10.1289/ehp.8566.

Dietert, RR. (2011). Role of developmental immunotoxicity and immune dysfunction in chronic disease and cancer. 

Reprod Toxicol 31: 319-326. http://dx.doi.org/10.1016/j.reprotox.2010.09.006.

Dietert, RR; McCabe, MJ, Jr. (2007). Lead immunotoxicity. In RW Luebke; RV House; I Kimber (Eds.), 

Immunotoxicology and immunopharmacology (3rd ed., pp. 207-223). Boca Raton, FL: CRC Press. 

Dietrich, KN; Krafft, KM; Bier, M; Succop, PA; Berger, O; Bornschein, RL. (1986). Early effects of fetal lead 

exposure: Neurobehavioral findings at 6 months. International Journal for Biosocial Research 8: 151-168. 

Dietrich, KN; Krafft, KM; Bornschein, RL; Hammond, PB; Berger, O; Succop, PA; Bier, M. (1987a). Low-level 

fetal lead exposure effect on neurobehavioral development in early infancy. Pediatrics 80: 721-730. 
February 2012 5-560 Draft – Do Not Cite or Quote

Dietrich, KN; Succop, PA; Berger, OG; Hammond, PB; Bornschein, RL. (1991). Lead exposure and the cognitive 

development of urban preschool children: The Cincinnati Lead Study cohort at age 4 years. Neurotoxicol 

Teratol 13: 203-211. http://dx.doi.org/10.1016/0892-0362(91)90012-L.

Dietrich, KN; Succop, PA; Berger, OG; Keith, RW. (1992). Lead exposure and the central auditory processing 

abilities and cognitive development of urban children: The Cincinnati Lead Study cohort at age 5 years. 

Neurotoxicol Teratol 14: 51-56. http://dx.doi.org/10.1016/0892-0362(92)90028-9.

Dietrich, KN; Berger, OG; Succop, PA; Hammond, PB; Bornschein, RL. (1993a). The developmental consequences 

of low to moderate prenatal and postnatal lead exposure: Intellectual attainment in the Cincinnati Lead 

Study Cohort following school entry. Neurotoxicol Teratol 15: 37-44. http://dx.doi.org/10.1016/0892-

0362(93)90043-N.

Dietrich, KN; Berger, OG; Succop, PA. (1993b). Lead exposure and the motor developmental status of urban sixyear-old children in the Cincinnati Prospective Study. Pediatrics 91: 301-307. 

Dietrich, KN; Ris, MD; Succop, PA; Berger, OG; Bornschein, RL. (2001). Early exposure to lead and juvenile 

delinquency. Neurotoxicol Teratol 23: 511-518. http://dx.doi.org/10.1016/S0892-0362(01)00184-2.

Dietrich, KN; Krafft, KM; Shukla, R; Bornschein, RL; Succop, PA. (1987b). The neurobehavioral effects of early 

lead exposure. In SR Schroeder (Ed.), Toxic substances and mental retardation: Neurobehavioral 

toxicology and teratology (pp. 71-95). Washington, DC: American Association on Mental Deficiency. 

Ding, Y; Gonick, HC; Vaziri, ND. (2000). Lead promotes hydroxyl radical generation and lipid peroxidation in 

cultured aortic endothelial cells. Am J Hypertens 13: 552-555. http://dx.doi.org/10.1016/S0895-

7061(99)00226-5.

Ding, Y; Gonick, HC; Vaziri, ND; Liang, K; Wei, L. (2001). Lead-induced hypertension. III. Increased hydroxyl 

radical production. Am J Hypertens 14: 169-173. http://dx.doi.org/10.1016/S0895-7061(00)01248-6.

Diouf, A; Garcon, G; Diop, Y; Ndiaye, B; Thiaw, C; Fall, M; Kane-Barry, O; Ba, D; Haguenoer, JM; Shirali, P.

(2006). Environmental lead exposure and its relationship to traffic density among Senegalese children: A 

cross-sectional study. Hum Exp Toxicol 25: 637-644. http://dx.doi.org/10.1177/0960327106074591.

Dogru, MI; Dogru, AK; Gul, M; Esrefoglu, M; Yurekli, M; Erdogan, S; Ates, B. (2008). The effect of 

adrenomedullin on rats exposed to lead. J Appl Toxicol 28: 140-146. http://dx.doi.org/10.1002/jat.1259.

Dogu, O; Louis, ED; Tamer, L; Unal, O; Yilmaz, A; Kaleagasi, H. (2007). Elevated blood lead concentrations in 

essential tremor: A case-control study in Mersin, Turkey. Environ Health Perspect 115: 1564-1568. 

http://dx.doi.org/10.1289/ehp.10352.

Dolinoy, DC; Jirtle, RL. (2008). Environmental epigenomics in human health and disease. Environ Mol Mutagen 

49: 4-8. http://dx.doi.org/10.1002/em.20366.

Donaldson, WE; Knowles, SO. (1993). Is lead toxicosis a reflection of altered fatty acid composition of membranes? 

[Review]. Comp Biochem Physiol C Toxicol Pharmacol 104: 377-379. http://dx.doi.org/10.1016/0742-

8413(93)90003-4.

Dong, S; Liang, D; An, N; Jia, L; Shan, Y; Chen, C; Sun, K; Niu, F; Li, H; Fu, S. (2009). The role of MAPK and 

FAS death receptor pathways in testicular germ cell apoptosis induced by lead. Acta Biochim Biophys Sin 

41: 800-807. http://dx.doi.org/10.1093/abbs/gmp069.

Dorman, RV; Freeman, EJ. (2002). Lead-dependent effects on arachidonic acid accumulation and the proliferation 

of vascular smooth muscle. J Biochem Mol Toxicol 16: 245-253. http://dx.doi.org/10.1002/jbt.10045.

Dorsey, CD; Lee, BK; Bolla, KI; Weaver, VM; Lee, SS; Lee, GS; Todd, AC; Shi, W; Schwartz, BS. (2006). 

Comparison of patella lead with blood lead and tibia lead and their associations with neurobehavioral test 

scores. J Occup Environ Med 48: 489-496. http://dx.doi.org/10.1097/01.jom.0000199678.86629.3b.

Dou, C; Zhang, J. (2011). Effects of lead on neurogenesis during zebrafish embryonic brain development. J Hazard 

Mater 194: 277-282. http://dx.doi.org/10.1016/j.jhazmat.2011.07.106.

Dowd, JB; Aiello, AE. (2009). Socioeconomic differentials in immune response. Epidemiology 20: 902-908. 

http://dx.doi.org/10.1097/EDE.0b013e3181bb5302.
February 2012 5-561 Draft – Do Not Cite or Quote

Dowd, TL; Rosen, JF; Gupta, RK. (1990). 31P NMR and saturation transfer studies of the effect of PB2+ on 

cultured osteoblastic bone cells. J Biol Chem 265: 20833-20838. 

Dowd, TL; Gupta, RK. (1991). 19F-NMR study of the effect of lead on intracellular free calcium in human platelets. 

Biochim Biophys Acta 1092: 341-346. http://dx.doi.org/10.1016/S0167-4889(97)90010-0.

Dowd, TL; Rosen, JF; Gundberg, CM; Gupta, RK. (1994). The displacement of calcium from osteocalcin at 

submicromolar concentrations of free lead. Biochim Biophys Acta 1226: 131-137. 

http://dx.doi.org/10.1016/0925-4439(94)90020-5.

Dowd, TL; Rosen, JF; Mints, L; Gundberg, CM. (2001). The effect of Pb2+ on the structure and hydroxyapatite 

binding properties of osteocalcin. Biochim Biophys Acta 1535: 153-163. http://dx.doi.org/10.1016/S0925-

4439(00)00094-6.

DS, H; Geraldine, M; T, V. (2009). Influence of minerals on lead-induced alterations in liver function in rats

exposed to long-term lead exposure. J Hazard Mater 166: 1410-1414. 

http://dx.doi.org/10.1016/j.jhazmat.2008.12.070.

Dumitrescu, E; Trif, A; Argherie, D. (2007). Consequences of chronic exposure of female rats to lead acetate on 

ovarian and uterine histoarhitectonic. LII: 198-205. 

Dumitrescu, E; Trif, A; Florin, M. (2008a). The consequences of female rats chronic exposure to lead acetate in the 

biomarkers emphasizing the hormonal disrupting potential of the reproductive function for in vivo 

evaluation. Bull Univ Agric Sci Vet Med Cluj-Napoca Vet Med 65: 382-387. 

Dumitrescu, E; Trif, A; Petrovici, S. (2008b). Lead acetate impact on some markers of female reproductive system 

performances (litter size, sex ratio) and physical development (vaginal opening) in rats. Bull Univ Agric 

Sci Vet Med Cluj-Napoca Vet Med 65: 283-287. 

Dundar, B; Oktem, F; Arslan, MK; Delibas, N; Baykal, B; Arslan, C; Gultepe, M; Ilhan, IE. (2006). The effect of 

long-term low-dose lead exposure on thyroid function in adolescents. Environ Res 101: 140-145. 

http://dx.doi.org/10.1016/j.envres.2005.10.002.

Dursun, N; Arifoglu, C; Suer, C; Keskinol, L. (2005). Blood pressure relationship to nitric oxide, lipid peroxidation, 

renal function, and renal blood flow in rats exposed to low lead levels. Biol Trace Elem Res 104: 141-149. 

http://dx.doi.org/10.1385/BTER:104:2:141.

DuVal, G; Fowler, BA. (1989). Preliminary purification and characterization studies of a low molecular weight, 

high affinity cytosolic lead-binding protein in rat brain. Biochem Biophys Res Commun 159: 177-184. 

http://dx.doi.org/10.1016/0006-291X(89)92420-0.

Duvall, RM; Norris, GA; Dailey, LA; Burke, JM; McGee, JK; Gilmour, MI; Gordon, T; Devlin, RB. (2008). Source 

apportionment of particulate matter in the U.S. and associations with lung inflammatory markers. Inhal 

Toxicol 20: 671-683. http://dx.doi.org/10.1080/08958370801935117.

Duydu, Y; Suzen, HS; Aydin, A; Cander, O; Uysal, H; Isimer, A; Vural, N. (2001). Correlation between lead 

exposure indicators and sister chromatid exchange (SCE) frequencies in lymphocytes from inorganic lead 

exposed workers. Arch Environ Contam Toxicol 41: 241-246. http://dx.doi.org/10.1007/s002440010244.

Duydu, Y; Dur, A; Suzen, HS. (2005). Evaluation of increased proportion of cells with unusually high sister 

chromatid exchange counts as a cytogenetic biomarker for lead exposure. Biol Trace Elem Res 104: 121-

129. http://dx.doi.org/10.1385/BTER:104:2:121.

Dyatlov, VA; Platoshin, AV; Lawrence, DA; Carpenter, DO. (1998). Lead potentiates cytokine- and glutamatemediated increases in permeability of the blood-brain barrier. Neurotoxicology 19: 283-291. 

Dyatlov, VA; Lawrence, DA. (2002). Neonatal lead exposure potentiates sickness behavior induced by Listeria 

monocytogenes infection of mice. Brain Behav Immun 16: 477-492. 

http://dx.doi.org/10.1006/brbi.2001.0641.

Eder, K; Reichlmayr-Lais, AM; Kirchgessner, M. (1990). Activity of Na-K-ATPase and Ca-Mg-ATPase in red 

blood cell membranes of lead-depleted rats. J Trace Elem Med Biol 4: 21-24. 
February 2012 5-562 Draft – Do Not Cite or Quote

Egle, PM; Shelton, KR. (1986). Chronic lead intoxication causes a brain-specific nuclear protein to accumulate in 

the nuclei of cells lining kidney tubules. J Biol Chem 261: 2294-2298. 

Ehrlich, R; Robins, T; Jordaan, E; Miller, S; Mbuli, S; Selby, P; Wynchank, S; Cantrell, A; De Broe, M; D'Haese, P; 

Todd, A; Landrigan, P. (1998). Lead absorption and renal dysfunction in a South African battery factory. 

Occup Environ Med 55: 453-460. http://dx.doi.org/10.1136/oem.55.7.453.

Ekinci, D; Beydemir, S; Küfrevioğlu, OI. (2007). In vitro inhibitory effects of some heavy metals on human 

erythrocyte carbonic anhydrases. J Enzyme Inhib Med Chem 22: 745-750. 

http://dx.doi.org/10.1080/14756360601176048.

El-Ashmawy, IM; Ashry, KM; El-Nahas, AF; Salama, OM. (2006). Protection by turmeric and myrrh against liver 

oxidative damage and genotoxicity induced by lead acetate in mice. Basic Clin Pharmacol Toxicol 98: 32-

37. http://dx.doi.org/10.1111/j.1742-7843.2006.pto_228.x.

El-Fawal, HAN; Waterman, SJ; De Feo, A; Shamy, MY. (1999). Neuroimmunotoxicology: Humoral assessment of 

neurotoxicity and autoimmune mechanisms. Environ Health Perspect 107: 767-775. 

El-Ghor, AA; Noshy, MM; Eid, JI. (In Press) Lead acetate and arsenic trioxide induce instability of microsatellites 

at three different fragile sites (6q21, 9q32-9q33 and 15p14) within the genome of the rat. Mutat Res. 

http://dx.doi.org/10.1016/j.mrgentox.2011.10.003.

El-Nekeety, AA; El-Kady, AA; Soliman, MS; Hassan, NS; Abdel-Wahhab, MA. (2009). Protective effect of 

Aquilegia vulgaris (L.) against lead acetate-induced oxidative stress in rats. Food Chem Toxicol 47: 2209-

2215. http://dx.doi.org/10.1016/j.fct.2009.06.019.

El-Neweshy, MS; El-Sayed, YS. (2011). Influence of vitamin C supplementation on lead-induced histopathological 

alterations in male rats. Exp Toxicol Pathol 63: 221-227. http://dx.doi.org/10.1016/j.etp.2009.12.003.

El-Sokkary, GH; Abdel-Rahman, GH; Kamel, ES. (2005). Melatonin protects against lead-induced hepatic and renal 

toxicity in male rats. Toxicology 213: 25-33. http://dx.doi.org/10.1016/j.tox.2005.05.003.

El Shafai, A; Zohdy, N; El Mulla, K; Hassan, M; Morad, N. (2011). Light and electron microscopic study of the 

toxic effect of prolonged lead exposure on the seminiferous tubules of albino rats and the possible 

protective effect of ascorbic acid. Food Chem Toxicol 49: 734-743. 

http://dx.doi.org/10.1016/j.fct.2010.11.033.

Elinder, CG; Friberg, L; Lind, B; Nilsson, B; Svartengren, M; Övermark, I. (1986). Decreased blood lead levels in 

residents of Stockholm for the period 1980-1984. Scand J Work Environ Health 12: 114-120. 

Elmarsafawy, SF; Jain, NB; Schwartz, J; Sparrow, D; Nie, HL; Hu, H. (2006). Dietary calcium as a potential 

modifier of the relationship of lead burden to blood pressure. Epidemiology 17: 531-537. 

http://dx.doi.org/10.1097/01.ede.0000231285.86968.2b.

Emmerson, BT. (1965). The renal excretion of urate in chronic lead nephropathy. Intern Med J 14: 295-303. 

Emmerson, BT; Ravenscroft, PJ. (1975). Abnormal renal urate homeostasis in systemic disorders. Nephron 14: 62-

80. http://dx.doi.org/10.1159/000180436.

Ercal, N; Treeratphan, P; Hammond, TC; Matthews, RH; Grannemann, NH; Spitz, DR. (1996). In vivo indices of 

oxidative stress in lead-exposed C57BL/6 mice are reduced by treatment with meso-2,3-dimercaptosuccinic 

acid or N-acetylcysteine. Free Radic Biol Med 21: 157-161. http://dx.doi.org/10.1016/0891-

5849(96)00020-2.

Ergurhan-Ilhan, I; Cadir, B; Koyuncu-Arslan, M; Gultepe, FM; Ozkan, G. (2008). Level of oxidative stress and 

damage in erythrocytes in apprentices indirectly exposed to lead. Pediatrics International 50: 45-50. 

http://dx.doi.org/10.1111/j.1442-200X.2007.02442.x.

Erie, JC; Good, JA; Butz, JA. (2009). Excess Lead in the Neural Retina in Age-Related Macular Degeneration. Am 

J Ophthalmol 148: 890-894. http://dx.doi.org/10.1016/j.ajo.2009.07.001.

Ernhart, CB; Wolf, AW; Kennard, MJ; Erhard, P; Filipovich, HF; Sokol, RJ. (1986). Intrauterine exposure to low 

levels of lead: the status of the neonate. Arch Environ Health 41: 287-291. 
February 2012 5-563 Draft – Do Not Cite or Quote

Ernhart, CB; Morrow-Tlucak, M; Marler, MR; Wolf, AW. (1987). Low level lead exposure in the prenatal and early 

preschool periods: Early preschool development. Neurotoxicol Teratol 9: 259-270. 

http://dx.doi.org/10.1016/0892-0362(87)90011-0.

Ernhart, CB; Morrow-Tlucak, M; Wolf, AW. (1988). Low level lead exposure and intelligence in the preschool 

years. Sci Total Environ 71: 453-459. 

Ernhart, CB; Morrow-Tlucak, M; Wolf, AW; Super, D; Drotar, D. (1989). Low level lead exposure in the prenatal 

and early preschool periods: intelligence prior to school entry. Neurotoxicol Teratol 11: 161-170. 

Ettinger, AS; Tellez-Rojo, MM; Amarasiriwardena, C; Bellinger, D; Peterson, K; Schwartz, J; Hu, H; HernandezAvila, M. (2004a). Effect of breast milk lead on infant blood lead levels at 1 month of age. Environ Health 

Perspect 112: 1381-1385. 

Ettinger, AS; Lamadrid-Figueroa, H; Téllez-Rojo, MM; Mercado-García, A; Peterson, KE; Schwartz, J; Hu, H; 

Hernández-Avila, M. (2009). Effect of calcium supplementation on blood lead levels in pregnancy: A 

randomized placebo-controlled trial. Environ Health Perspect 117: 26-31. 

http://dx.doi.org/10.1289/ehp.11868.

Eum, KD; Nie, LH; Schwartz, J; Vokonas, PS; Sparrow, D; Hu, H; Weisskopf, MG. (2011). Prospective cohort 

study of lead exposure and electrocardiographic conduction disturbances in the Department of Veterans 

Affairs Normative Aging Study. Environ Health Perspect 119: 490-494. 

http://dx.doi.org/10.1289/ehp.1003279.

Evans, M; Fored, CM; Nise, G; Bellocco, R; Nyren, O; Elinder, CG. (2010). Occupational lead exposure and severe 

CKD: A population-based case-control and prospective observational cohort study in Sweden. Am J 

Kidney Dis 55: 497-506. http://dx.doi.org/10.1053/j.ajkd.2009.11.012.

Ewers, U; Stiller-Winkler, R; Idel, H. (1982). Serum immunoglobulin, complement C3, and salivary IgA levels in 

lead workers. Environ Res 29: 351-357. http://dx.doi.org/10.1016/0013-9351(82)90036-6.

Factor-Litvak, P; Graziano, JH; Kline, JK; Popovac, D; Mehmeti, A; Ahmedi, G; Shrout, P; Murphy, MJ; Gashi, E; 

Haxhiu, R; Rajovic, L; Nenezic, DU; Stein, ZA. (1991). A prospective study of birthweight and length of 

gestation in a population surrounding a lead smelter in Kosovo, Yugoslavia. Int J Epidemiol 20: 722-728. 

http://dx.doi.org/10.1093/ije/20.3.722.

Factor-Litvak, P; Kline, JK; Popovac, D; Hadzialjevic, S; Lekic, V; Preteni-Rexhepi, E; Capuni-Paracka, S; 

Slavkovich, V; Graziano, J. (1996). Blood lead and blood pressure in young children. Epidemiology 7: 

633-637. 

Factor-Litvak, P; Wasserman, G; Kline, JK; Graziano, J. (1999). The Yugoslavia prospective study of 

environmental lead exposure. Environ Health Perspect 107: 9-15. http://dx.doi.org/10.1289/ehp.991079.

Fadrowski, JJ; Navas-Acien, A; Tellez-Plaza, M; Guallar, E; Weaver, VM; Furth, SL. (2010). Blood lead level and 

kidney function in US adolescents: The Third National Health and Nutrition Examination Survey. Arch 

Intern Med 170: 75-82. http://dx.doi.org/10.1001/archinternmed.2009.417.

Fahmy, MA. (1999). Lead acetate genotoxicity in mice. Cytologia (Tokyo) 64: 357-365. 

Faikoglu, R; Savan, K; Utku, C; Takar, N; Zebitay, AG. (2006). Significance of maternal plasma lead level in early 

pregnancy loss. J Environ Sci Health A Tox Hazard Subst Environ Eng 41: 501-506. 

http://dx.doi.org/10.1080/10934520500428435.

Faith, RE; Luster, MI; Kimmel, CA. (1979). Effect of chronic developmental lead exposure on cell-mediated 

immune functions. Clin Exp Immunol 35: 413-420. 

Fan, G; Feng, C; Li, Y; Wang, CH; Yan, J; Li, W; Feng, JG; Shi, XL; Bi, YY. (2009a). Selection of nutrients for 

prevention or amelioration of lead-induced learning and memory impairment in rats. Ann Occup Hyg 53: 

341-351. http://dx.doi.org/10.1093/annhyg/mep019.

Fan, G; Feng, C; Wu, F; Ye, W; Lin, F; Wang, C; Yan, J; Zhu, G; Xiao, Y; Bi, Y. (2010). Methionine choline 

reverses lead-induced cognitive and N-methyl-d-aspartate receptor subunit 1 deficits. Toxicology 272: 23-

31. http://dx.doi.org/10.1016/j.tox.2010.03.018.
February 2012 5-564 Draft – Do Not Cite or Quote

Fan, Y; Yamada, T; Shimizu, T; Nanashima, N; Akita, M; Suto, K; Tsuchida, S. (2009b). Ferritin expression in rat 

hepatocytes and Kupffer cells after lead nitrate treatment. Toxicol Pathol 37: 209-217. 

http://dx.doi.org/10.1177/0192623308328544.

Fang, F; Kwee, LC; Allen, KD; Umbach, DM; Ye, W; Watson, M; Keller, J; Oddone, EZ; Sandler, DP; Schmidt, S; 

Kamel, F. (2010). Association between blood lead and the risk of amyotrophic lateral sclerosis. Am J 

Epidemiol 171: 1126-1133. http://dx.doi.org/10.1093/aje/kwq06.

Farant, JP; Wigfield, DC. (1987). Interaction of divalent metal ions with normal and lead-inhibited human 

erythrocytic porphobilinogen synthase in vitro. Toxicol Appl Pharmacol 89: 9-18. 

http://dx.doi.org/10.1016/0041-008X(87)90171-2.

Farant, JP; Wigfield, DC. (1990). The effects of copper, zinc, mercury, and cadmium on rabbit erythrocytic 

porphobilinogen synthase in vivo. J Anal Toxicol 14: 222-226. 

Faraone, SV; Doyle, AE. (2001). The nature and heritability of attention-deficit/hyperactivity disorder. Child 

Adolesc Psychiatr Clin N Am 10: 299-316, viii-ix. 

Farmand, F; Ehdaie, A; Roberts, CK; Sindhu, RK. (2005). Lead-induced dysregulation of superoxide dismutases, 

catalase, glutathione peroxidase, and guanylate cyclase. Environ Res 98: 33-39. 

http://dx.doi.org/10.1016/j.envres.2004.05.016.

Farrer, DG; Hueber, SM; McCabe, MJ, Jr. (2005). Lead enhances CD4+ T cell proliferation indirectly by targeting 

antigen presenting cells and modulating antigen-specific interactions. Toxicol Appl Pharmacol 207: 125-

137. http://dx.doi.org/10.1016/j.taap.2004.12.017.

Farrer, DG; Hueber, S; Laiosa, MD; Eckles, KG; McCabe, MJ, Jr. (2008). Reduction of myeloid suppressor cell 

derived nitric oxide provides a mechanistic basis of lead enhancement of alloreactive CD4(+) T cell 

proliferation. Toxicol Appl Pharmacol 229: 135-145. http://dx.doi.org/10.1016/j.taap.2007.12.011.

Favalli, L; Chiari, MC; Piccinini, F; Rozza, A. (1977). Experimental investigations on the contraction induced by 

lead in arterial smooth muscle. Acta Pharmacol Toxicol 41: 412-420. http://dx.doi.org/10.1111/j.1600-

0773.1977.tb03362.x.

Fazli-Tabaei, S; Fahim, M; Khoshbaten, A. (2006). Acute lead exposure and contraction of rat isolated aorta induced 

by D1-dopaminergic and alpha-adrenergic drugs. Arch Iran Med 9: 119-123. 

Fehlau, R; Grygorczyk, R; Fuhrmann, GF; Schwarz, W. (1989). Modulation of the Ca2+- or Pb2+-activated K+-

selective channels in human red cells. II. Parallelisms to modulation of the activity of a membrane-bound 

oxidoreductase. Biochim Biophys Acta 978: 37-42. http://dx.doi.org/10.1016/0005-2736(89)90495-1.

Feinberg, AP. (2007). Phenotypic plasticity and the epigenetics of human disease. Nature 447: 433-440. 

http://dx.doi.org/10.1038/nature05919.

Fels, LM; Wunsch, M; Baranowski, J; Norska-Borowka, I; Price, RG; Taylor, SA; Patel, S; De Broe, M; Elsevier, 

MM; Lauwerys, R; Roels, H; Bernard, A; Mutti, A; Gelpi, E; Rosello, J; Stolte, H. (1998). Adverse effects 

of chronic low level lead exposure on kidney function: A risk group study in children. Nephrol Dial 

Transplant 13: 2248-2256. http://dx.doi.org/10.1093/ndt/13.9.2248.

Ferguson, C; Kern, M; Audesirk, G. (2000). Nanomolar concentrations of inorganic lead increase Ca2+ efflux and 

decrease intracellular free Ca2+ ion concentrations in cultured rat hippocampal neurons by a calmodulindependent mechanism. Neurotoxicology 21: 365-378. 

Ferguson, MA; Vaidya, VS; Bonventre, JV. (2008). Biomarkers of nephrotoxic acute kidney injury. Toxicology 

245: 182-193. http://dx.doi.org/10.1016/j.tox.2007.12.024.

Fergusson, DM; Fergusson, JE; Horwood, LJ; Kinzett, NG. (1988a). A longitudinal study of dentine lead levels, 

intelligence, school performance and behaviour Part I Dentine lead levels and exposure to environmental 

risk factors. J Child Psychol Psychiatry 29: 781-792. 

Fergusson, DM; Fergusson, JE; Horwood, LJ; Kinzett, NG. (1988b). A longitudinal study of dentine lead levels, 

intelligence, school performance and behaviour Part II Dentine lead and cognitive ability. J Child Psychol 

Psychiatry 29: 793-809. 
February 2012 5-565 Draft – Do Not Cite or Quote

Fergusson, DM; Boden, JM; Horwood, LJ. (2008). Dentine lead levels in childhood and criminal behaviour in late 

adolescence and early adulthood. J Epidemiol Community Health 62: 1045-1050. 

http://dx.doi.org/10.1136/jech.2007.072827.

Fernandez-Cabezudo, MJ; Ali, SAE; Ullah, A; Hasan, MY; Kosanovic, M; Fahim, MA; Adem, A; al-Ramadi, BK.

(2007). Pronounced susceptibility to infection by Salmonella enterica serovar Typhimurium in mice 

chronically exposed to lead correlates with a shift to Th2-type immune responses. Toxicol Appl Pharmacol 

218: 215-226. http://dx.doi.org/10.1016/j.taap.2006.11.018.

Finkelstein, Y; Markowitz, ME; Rosen, JF. (1998). Low-level lead-induced neurotoxicity in children: An update on 

central nervous system effects. Brain Res 27: 168-176. http://dx.doi.org/10.1016/S0165-0173(98)00011-3.

Fiorim, J; Ribeiro Júnior, RF; Silveira, EA; Padilha, AS; Vescovi, MV; de Jesus, HC; Stefanon, I; Salaices, M; 

Vassallo, DV. (2011). Low-level lead exposure increases systolic arterial pressure and endothelium-derived 

vasodilator factors in rat aortas. PLoS ONE 6: e17117. http://dx.doi.org/10.1371/journal.pone.0017117.

Fischbein, A; Tsang, P; Luo, JCJ; Roboz, JP; Jiang, JD; Bekesi, JG. (1993). Phenotypic aberrations of the CD3+ and 

CD4+ cells and functional impairments of lymphocytes at low-level occupational exposure to lead. 66: 

163-168. http://dx.doi.org/10.1006/clin.1993.1020.

Flegal, AR; Smith, DR. (1992). Lead levels in preindustrial humans [Letter]. N Engl J Med 326: 1293-1294. 

Flohe, SB; Bruggemann, J; Herder, C; Goebel, C; Kolb, H. (2002). Enhanced proinflammatory response to 

endotoxin after priming of macrophages with lead ions. J Lepid Soc 71: 417-424. 

Flora, SJS; Singh, S; Tandon, SK. (1989). Thiamine and zinc in prevention or therapy of lead intoxication. J Int Med 

Res 17: 68-75. 

Flora, SJS; Saxena, G; Gautam, P; Kaur, P; Gill, KD. (2007). Response of lead-induced oxidative stress and 

alterations in biogenic amines in different rat brain regions to combined administration of DMSA and 

MiADMSA. Chem Biol Interact 170: 209-220. http://dx.doi.org/10.1016/j.cbi.2007.08.003.

Foglieni, C; Fulgenzi, A; Ticozzi, P; Pellegatta, F; Sciorati, C; Belloni, D; Ferrero, E; Ferrero, ME. (2006). 

Protective effect of EDTA preadministration on renal ischemia. BMC Nephrol 7: 5. 

http://dx.doi.org/10.1186/1471-2369-7-5.

Föller, M; Huber, SM; Lang, F. (2008). Erythrocyte programmed cell death. IUBMB Life 60: 661-668. 

http://dx.doi.org/10.1002/iub.106.

Fonte, R; Agosti, A; Scafa, F; Candura, SM. (2007). Anaemia and abdominal pain due to occupational lead 

poisoning. Haematologica 92: e13-e14. http://dx.doi.org/10.3324/haematol.10951.

Forst, LS; Freels, S; Persky, V. (1997). Occupational lead exposure and hearing loss. J Occup Environ Med 39: 658-

660. 

Fortier, M; Omara, F; Bernier, J; Brousseau, P; Fournier, M. (2008). Effects of physiological concentrations of 

heavy metals both individually and in mixtures on the viability and function of peripheral blood human 

leukocytes in vitro. J Toxicol Environ Health A 71: 1327-1337. 

http://dx.doi.org/10.1080/15287390802240918.

Fortoul, TI; Saldivar, L; Espejel-Mayab, G; Bazarro, P; Mussali-Galante, P; Avila-Casado, MD; Colin-Barenque, L; 

Avila-Costa, MR. (2005). Inhalation of cadmium, lead or its mixture effects on the bronchiolar structure 

and its relation with metal tissue concentrations. Environ Toxicol Pharmacol 19: 329-334. 

http://dx.doi.org/10.1016/j.etap.2004.08.007.

Fortune, T; Lurie, DI. (2009). Chronic low-level lead exposure affects the monoaminergic system in the mouse 

superior olivary complex. J Comp Neurol 513: 542-558. http://dx.doi.org/10.1002/cne.21978.

Foster, WG. (1992). Reproductive toxicity of chronic lead exposure in the female cynomolgus monkey. Reprod 

Toxicol 6: 123-131. 

Foster, WG; McMahon, A; YoungLai, EV; Hughes, EG; Rice, DC. (1993). Reproductive endocrine effects of 

chronic lead exposure in the male cynomolgus monkey. Reprod Toxicol 7: 203-209. 
February 2012 5-566 Draft – Do Not Cite or Quote

Fowler, BA; Kimmel, CA; Woods, JS; McConnell, EE; Grant, LD. (1980). Chronic low-level lead toxicity in the 

rat: III An integrated assessment of long-term toxicity with special reference to the kidney. Toxicol Appl

Pharmacol 56: 59-77. http://dx.doi.org/10.1016/0041-008X(80)90131-3.

Fowler, BA; DuVal, G. (1991). Effects of lead on the kidney: Roles of high-affinity lead-binding proteins. Environ 

Health Perspect 91: 77-80. http://dx.doi.org/10.1289/ehp.919177.

Fowler, BA; Kahng, MW; Smith, DR; Conner, EA; Laughlin, NK. (1993). Implications of lead binding proteins for 

risk assessment of lead exposure. J Expo Sci Environ Epidemiol 3: 441-448. 

Fox, DA; Sillman, AJ. (1979). Heavy metals affect rod, but not cone, photoreceptors. Science 206: 78-80. 

http://dx.doi.org/10.1126/science.314667.

Fox, DA; Farber, DB. (1988). Rods are selectively altered by lead: I. Electrophysiology and biochemistry. Exp Eye 

Res 46: 597-611. http://dx.doi.org/10.1016/S0014-4835(88)80016-2.

Fox, DA; Chu, LWF. (1988). Rods are selectively altered by lead: II. Ultrastructure and quantitative histology. Exp 

Eye Res 46: 613-625. http://dx.doi.org/10.1016/S0014-4835(88)80017-4.

Fox, DA; Katz, LM; Farber, DB. (1991). Low level developmental lead exposure decreases the sensitivity, 

amplitude and temporal resolution of rods. Neurotoxicology 12: 641-654. 

Fox, DA; Katz, LM. (1992). Developmental lead exposure selectively alters the scotopic ERG component of dark 

and light adaptation and increases rod calcium content. Vision Res 32: 249-255. 

Fox, DA; Campbell, ML; Blocker, YS. (1997). Functional alterations and apoptotic cell death in the retina following 

developmental or adult lead exposure. Neurotoxicology 18: 645-664. 

Fox, DA; Kala, SV; Hamilton, WR; Johnson, JE; O'Callaghan, JA. (2008). Low-level human equivalent gestational 

lead exposure produces supernormal scotopic electroretinograms, increased retinal neurogenesis, and 

decreased retinal dopamine utilization in rats. Environ Health Perspect 116: 618-625. 

http://dx.doi.org/10.1289/ehp.11268.

Fox, DA; Opanashuk, L; Zharkovsky, A; Weiss, B. (2010). Gene-chemical interactions in the developing 

mammalian nervous system: Effects on proliferation, neurogenesis and differentiation. Neurotoxicology 

31: 589-597. http://dx.doi.org/10.1016/j.neuro.2010.03.007.

Fracasso, ME; Perbellini, L; Solda, S; Talamini, G; Franceschetti, P. (2002). Lead induced DNA strand breaks in 

lymphocytes of exposed workers: Role of reactive oxygen species and protein kinase C. Mutat Res Genet 

Toxicol Environ Mutagen 515: 159-169. http://dx.doi.org/10.1016/S1383-5718(02)00012-8.

Francis, DD; Diorio, J; Plotsky, PM; Meaney, MJ. (2002). Environmental enrichment reverses the effects of 

maternal separation on stress reactivity. J Neurosci 22: 7840-7843. 

Fraser, S; Muckle, G; Despres, C. (2006). The relationship between lead exposure, motor function and behaviour in 

Inuit preschool children. Neurotoxicol Teratol 28: 18-27. http://dx.doi.org/10.1016/j.ntt.2005.10.008.

Fried, LF. (2009). Creatinine and cystatin C: What are the values? Kidney Int 75: 578-580. 

http://dx.doi.org/10.1038/Ki.2008.688.

Froehlich, TE; Lanphear, BP; Dietrich, KN; Cory-Slechta, DA; Wang, N; Kahn, RS. (2007). Interactive effects of a 

DRD4 polymorphism, lead and sex on executive functions in children. Biol Psychiatry 62: 243-249. 

http://dx.doi.org/10.1016/j.biopsych.2006.09.039.

Froehlich, TE; Lanphear, BP; Auinger, P; Hornung, R; Epstein, JN; Braun, J; Kahn, RS. (2009). Association of 

tobacco and lead exposures with attention-deficit/hyperactivity disorder. Pediatrics 124: E1054-E1063. 

http://dx.doi.org/10.1542/peds.2009-0738.

Fujita, H; Orii, Y; Sano, S. (1981). Evidence of increased synthesis of delta-aminolevulinic acid dehydratase in 

experimental lead-poisoned rats. Biochim Biophys Acta 678: 39-50. http://dx.doi.org/10.1016/0304-

4165(81)90045-3.
February 2012 5-567 Draft – Do Not Cite or Quote

Fujita, H; Sato, K; Sano, S. (1982). Increase in the amount of erythrocyte delta-aminolevulinic acid dehydratase in 

workers with moderate lead exposure. Int Arch Occup Environ Health 50: 287-297. 

http://dx.doi.org/10.1007/BF00378090.

Fujiwara, Y; Kaji, T; Sakurai, S; Sakamoto, M; Kozuka, H. (1997). Inhibitory effect of lead on the repair of 

wounded monolayers of cultured vascular endothelial cells. Toxicology 117: 193-198. 

http://dx.doi.org/10.1016/S0300-483X(96)03575-5.

Fukumoto, K; Karai, I; Horiguchi, S. (1983). Effect of lead on erythrocyte membranes. Occup Environ Med 40: 

220-223. http://dx.doi.org/10.1136/oem.40.2.220.

Fuller, GM; Zhang, Z. (2001). Transcriptional control mechanism of fibrinogen gene expression. Ann N Y Acad Sci 

936: 469-479. http://dx.doi.org/10.1111/j.1749-6632.2001.tb03534.x.

Gao, A; Lu, XT; Li, QY; Tian, L. (2010a). Effect of the delta-aminolevulinic acid dehydratase gene polymorphism 

on renal and neurobehavioral function in workers exposed to lead in China. Sci Total Environ 408: 4052-

4055. http://dx.doi.org/10.1016/j.scitotenv.2010.04.024.

Gao, D; Kasten-Jolly, J; Lawrence, DA. (2006). The paradoxical effects of lead in interferon-gamma knockout 

BALB/c mice. Toxicol Sci 89: 444-453. http://dx.doi.org/10.1093/toxsci/kfj043.

Gao, D; Mondal, TK; Lawrence, DA. (2007). Lead effects on development and function of bone marrow-derived 

dendritic cells promote Th2 immune responses. Toxicol Appl Pharmacol 222: 69-79. 

http://dx.doi.org/10.1016/j.taap.2007.04.001.

Gao, D; Lawrence, DA. (2010). Dendritic cells in immunotoxicity testing. Methods Mol Biol 598: 259-281. 

http://dx.doi.org/10.1007/978-1-60761-401-2_19.

Gao, SJ; Jin, YL; Unverzagt, FW; Ma, F; Hall, KS; Murrell, JR; Cheng, YB; Shen, JZ; Ying, B; Ji, RD; Matesan, J; 

Liang, C; Hendrie, HC. (2008). Trace element levels and cognitive function in rural elderly Chinese. J 

Gerontol A Biol Sci Med Sci 63: 635-641. 

Garavan, H; Morgan, RE; Levitsky, DA; Hermer-Vazquez, L; Strupp, BJ. (2000). Enduring effects of early lead 

exposure: Evidence for a specific deficit in associative ability. Neurotoxicol Teratol 22: 151-164. 

García-Lestón, J; Roma-Torres, J; Vilares, M; Pinto, R; Cunha, LM; Prista, J; Teixeira, JP; Mayan, O; Pásaro, E; 

Méndez, J; Laffon, B. (2011). Biomonitoring of a population of Portuguese workers exposed to lead. Mutat 

Res 721: 81-88. http://dx.doi.org/10.1016/j.mrgentox.2011.01.001.

Garcon, G; Leleu, B; Marez, T; Zerimech, F; Jean-Marie, HD; Daniel, FB; Shirali, P. (2007). Biomonitoring of the 

adverse effects induced by the chronic exposure to lead and cadmium on kidney function: Usefulness of 

alpha-glutathione S-transferase. Sci Total Environ 377: 165-172. 

http://dx.doi.org/10.1016/j.scitotenv.2007.02.002.

Gargioni, R; Neto, FF; Buchi, DF; Randi, MAF; Franco, CRC; Paludo, KS; Pelletier, E; Ferraro, MVM; Cestari, 

MM; Bussolaro, D; Ribeiro, CAO. (2006). Cell death and DNA damage in peritoneal macrophages of mice 

(Mus musculus) exposed to inorganic lead. Cell Biol Int 30: 615-623. 

http://dx.doi.org/10.1016/j.cellbi.2006.03.010.

Garza, A; Vega, R; Soto, E. (2006). Cellular mechanisms of lead neurotoxicity. Med Sci Monit 12: RA57-RA65. 

Gastaldo, J; Viau, M; Bencokova, Z; Joubert, A; Charvet, AM; Balosso, J; Foray, N. (2007). Lead contamination 

results in late and slowly repairable DNA double-strand breaks and impacts upon the ATM-dependent 

signaling pathways. Toxicol Lett 173: 201-214. http://dx.doi.org/10.1016/j.toxlet.2007.08.003.

Gautam, P; Flora, SJ. (2010). Oral supplementation of gossypin during lead exposure protects alteration in heme 

synthesis pathway and brain oxidative stress in rats. Nutrition 26: 563-570. 

http://dx.doi.org/10.1016/j.jhazmat.2008.12.070.

Gedeon, Y; Ramesh, GT; Wellman, PJ; Jadhav, AL. (2001). Changes in mesocorticolimbic dopamine and D1/D2 

receptor levels after low level lead exposure: A time course study. Toxicol Lett 123: 217-226. 

http://dx.doi.org/10.1016/S0378-4274(01)00408-8.
February 2012 5-568 Draft – Do Not Cite or Quote

Gent, JF; Koutrakis, P; Belanger, K; Triche, E; Holford, TR; Bracken, MB; Leaderer, BP. (2009). Symptoms and 

medication use in children with asthma and traffic-related sources of fine particle pollution. Environ Health 

Perspect 117: 1168-1174. http://dx.doi.org/10.1289/ehp.0800335.

Gerr, F; Letz, R; Stokes, L; Chettle, D; McNeill, F; Kaye, W. (2002). Association between bone lead concentration 

and blood pressure among young adults. Am J Ind Med 42: 98-106. 

Ghering, AB; Jenkins, LMM; Schenck, BL; Deo, S; Mayer, RA; Pikaart, MJ; Omichinski, JG; Godwin, HA. (2005). 

Spectroscopic and functional determination of the interaction of Pb2+ with GATA proteins. J Am Chem 

Soc 127: 3751-3759. http://dx.doi.org/10.1021/ja0464544.

Ghosh-Narang, J; Jones, TM; Menke, A; Todd, AC; Muntner, P; Batuman, V. (2007). Parathyroid hormone status 

does not influence blood and bone lead levels in dialysis patients. Am J Med Sci 334: 415-420. 

http://dx.doi.org/10.1097/MAJ.0b013e318068b237.

Giddabasappa, A; Hamilton, WR; Chaney, S; Xiao, W; Johnson, JE; Mukherjee, S; Fox, DA. (2011). Low-level 

gestational lead exposure increases retinal progenitor cell proliferation and rod photoreceptor and bipolar 

cell neurogenesis in mice. Environ Health Perspect 119: 71-77. http://dx.doi.org/10.1289/ehp.1002524.

Giedd, JN; Lalonde, FM; Celano, MJ; White, SL; Wallace, GL; Lee, NR; Lenroot, RK. (2009). Anatomical brain 

magnetic resonance imaging of typically developing children and adolescents. J Am Acad Child Adolesc 

Psychiatry 48: 465-470. http://dx.doi.org/10.1097/CHI.0b013e31819f2715.

Gilbert, ME; Mack, CM; Lasley, SM. (1999). Chronic developmental lead exposure and hippocampal long-term 

potentiation: biphasic dose-response relationship. Neurotoxicology 20: 71-82. 

Gilbert, ME; Kelly, ME; Samsam, TE; Goodman, JH. (2005). Chronic developmental lead exposure reduces 

neurogenesis in adult rat hippocampus but does not impair spatial learning. Toxicol Sci 86: 365-374. 

Gilbert, SG; Rice, DC. (1987). Low-level lifetime lead exposure produces behavioral toxicity (spatial discrimination 

reversal) in adult monkeys. Toxicol Appl Pharmacol 91: 484-490. 

Gilsanz, V; Chalfant, J; Kalkwarf, H; Zemel, B; Lappe, J; Oberfield, S; Shepherd, J; Wren, T; Winer, K. (2011). 

Age at onset of puberty predicts bone mass in young adulthood. J Pediatr 158: 100-105, 105.e101-102. 

http://dx.doi.org/10.1016/j.jpeds.2010.06.054.

Giridhar, J; Isom, GE. (1990). Interaction of lead acetate with atrial natriuretic factor in rats. Life Sci 46: 569-576. 

http://dx.doi.org/10.1016/0024-3205(90)90124-A.

Gittleman, R; Eskenazi, B. (1983). Lead and hyperactivity revisited: An investigation of non-disadvantaged 

children. Arch Gen Psychiatry 40: 827-833. http://dx.doi.org/10.1001/archpsyc.1983.01790070017002.

Glahn, F; Schmidt-Heck, W; Zellmer, S; Guthke, R; Wiese, J; Golka, K; Hergenroder, R; Degen, GH; Lehmann, T; 

Hermes, M; Schormann, W; Brulport, M; Bauer, A; Bedawy, E; Gebhardt, R; Hengstler, JG; Foth, H.

(2008). Cadmium, cobalt and lead cause stress response, cell cycle deregulation and increased steroid as 

well as xenobiotic metabolism in primary normal human bronchial epithelial cells which is coordinated by 

at least nine transcription factors. Arch Toxicol 82: 513-524. http://dx.doi.org/10.1007/s00204-008-0331-9.

Glass, TA; Bandeen-Roche, K; McAtee, M; Bolla, K; Todd, AC; Schwartz, BS. (2009). Neighborhood psychosocial 

hazards and the association of cumulative lead dose with cognitive function in older adults. Am J 

Epidemiol 169: 683-692. http://dx.doi.org/10.1093/aje/kwn390.

Glenn, BS; Bandeen-Roche, K; Lee, BK; Weaver, VM; Todd, AC; Schwartz, BS. (2006). Changes in systolic blood 

pressure associated with lead in blood and bone. Epidemiology 17: 538-544. 

http://dx.doi.org/10.1097/01.ede.0000231284.19078.4b.

Godleski, JJ; Clarke, RW; Coull, BA; Saldiva, PHN; Jiang, NF; Lawrence, J; Koutrakis, P. (2002). Composition of 

inhaled urban air particles determines acute pulmonary responses. Ann Occup Hyg 46: 419-424. 

Goering, PL; Fowler, BA. (1984). Regulation of lead inhibition of delta-aminolevulinic acid dehydratase by a low 

molecular weight, high affinity renal lead-binding protein. J Pharmacol Exp Ther 231: 66-71. 

Goering, PL; Fowler, BA. (1985). Mechanism of renal lead-binding protein reversal of delta-aminolevulinic acid 

dehydratase inhibition by lead. J Pharmacol Exp Ther 234: 365-371. 
February 2012 5-569 Draft – Do Not Cite or Quote

Goering, PL; Mistry, P; Fowler, BA. (1986). A low molecular weight lead-binding protein in brain attenuates lead 

inhibition of delta-aminolevulinic acid dehydratase: Comparison with a renal lead-binding protein. J 

Pharmacol Exp Ther 237: 220-225. 

Goering, PL; Fowler, BA. (1987a). Kidney zinc-thionein regulation of delta-aminolevulinic acid dehydratase 

inhibition by lead. Arch Biochem Biophys 253: 48-55. http://dx.doi.org/10.1016/0003-9861(87)90635-7.

Goering, PL; Fowler, BA. (1987b). Regulatory roles of high-affinity metal-binding proteins in mediating lead 

effects on delta-aminolevulinic acid dehydratase. Ann N Y Acad Sci 514: 235-247. 

http://dx.doi.org/10.1111/j.1749-6632.1987.tb48778.x.

Golabek, T; Darewicz, B; Borawska, M; Markiewicz, R; Socha, K; Kudelski, J. (2009). Lead concentration in the 

bladder tissue and blood of patients with bladder cancer. Scand J Urol Nephrol 43: 467-470. 

http://dx.doi.org/10.3109/00365590903198991.

Goldstein, GW. (1993). Evidence that lead acts as a calcium substitute in second messenger metabolism. 

Neurotoxicology 14: 97-101. 

Gollenberg, AL; Hediger, ML; Lee, PA; Himes, JH; Buck Louis, GM. (2010). Association between lead and 

cadmium and reproductive hormones in peripubertal U.S. girls. Environ Health Perspect 118: 1782-1787. 

http://dx.doi.org/10.1289/ehp.1001943.

Gomaa, A; Hu, H; Bellinger, D; Schwartz, J; Tsaih, SW; Gonzalez-Cossio, T; Schnaas, L; Peterson, K; Aro, A; 

Hernandez-Avila, M. (2002). Maternal bone lead as an independent risk factor for fetal neurotoxicity: A 

prospective study. Pediatrics 110: 110-118. 

Gong, Z; Evans, HL. (1997). Effect of chelation with meso-dimercaptosuccinic acid (DMSA) before and after the 

appearance of lead-induced neurotoxicity in the rat. Toxicol Appl Pharmacol 144: 205-214. 

http://dx.doi.org/10.1006/taap.1997.8118.

Gonick, HC; Cohen, AH; Ren, Q; Saldanha, LF; Khalil-Manesh, F; Anzalone, J; Sun, YY. (1996). Effect of 2,3-

dimercaptosuccinic acid on nephrosclerosis in the Dahl rat I role of reactive oxygen species. Kidney Int 50: 

1572-1581. http://dx.doi.org/10.1038/ki.1996.473.

Gonick, HC; Ding, Y; Bondy, SC; Ni, Z; Vaziri, ND. (1997). Lead-induced hypertension: Interplay of nitric oxide 

and reactive oxygen species. Hypertension 30: 1487-1492. 

Gonick, HC; Ding, Y; Vaziri, ND. (1998). Effect of low lead exposure on eicosanoid excretion in rats. 

Prostaglandins Other Lipid Mediat 55: 77-82. http://dx.doi.org/10.1016/S0090-6980(98)00010-0.

Gonick, HC; Khalil-Manesh, F; Raghavan, SRV. (1985). Characterization of human erythrocyte lead-binding 

protein. In TD Lekkas (Ed.), Heavy metals in the environment: Athens, September 1985 (pp. 313-316). 

Edinburgh, UK: CEP Consultants. 

Gorell, JM; Johnson, CC; Rybicki, BA; Peterson, EL; Kortsha, GX; Brown, GG; Richardson, RJ. (1997). 

Occupational exposures to metals as risk factors for Parkinson's disease. Neurology 48: 650-658. 

Goswami, K; Gachhui, R; Bandopadhyay, A. (2005). Hepatorenal dysfunctions in lead pollution. J Environ Eng Sci 

47: 75-80. 

Governa, M; Valentino, M; Visona, I. (1987). In vitro impairment of human granulocyte functions by lead. Arch 

Toxicol 59: 421-425. http://dx.doi.org/10.1007/BF00316208.

Goyer, RA. (1968). The renal tubule in lead poisoning: I. mMitochondrial swelling and aminoaciduria. Lab Invest 

19: 71-77. 

Goyer, RA; Krall, A; Kimball, JP. (1968). The renal tubule in lead poisoning: II. In vitro studies of mitochondrial 

structure and function. Lab Invest 19: 78-83. 

Goyer, RA; May, P; Cates, MM; Krigman, MR. (1970a). Lead and protein content of isolated intranuclear inclusion 

bodies from kidneys of lead-poisoned rats. Lab Invest 22: 245-251. 

Goyer, RA; Leonard, DL; Moore, JF; Rhyne, B; Krigman, MR. (1970b). Lead dosage and the role of the 

intranuclear inclusion body: An experimental study. Arch Environ Occup Health 20: 705-711. 
February 2012 5-570 Draft – Do Not Cite or Quote

Goyer, RA; Rhyne, BC. (1973). Pathological effects of lead. Int Rev Exp Pathol 12: 1-77. 

Goyer, RA; Cherian, MG; Delaquerriere-Richardson, L. (1978). Renal effects of repeated administration of calcium 

disodium ethylenediaminetetraacetate during excessive exposure to lead in rats. J Environ Pathol Toxicol 

Oncol 1: 403-410. 

Grabowska, M; Guminska, M. (1996). The effect of lead on lactate formation, ATP level and membrane ATPase 

activities in human erythrocytes in vitro. Int J Occup Med Environ Health 9: 265-274. 

Gracia, CR; Sammel, MD; Coutifaris, C; Guzick, DS; Barnhart, KT. (2005). Occupational exposures and male 

infertility. Am J Epidemiol 162: 729-733. http://dx.doi.org/10.1093/aje/kwi269.

Graf, U; Würgler, FE. (1986). The present status of validation of the wing spot test in Drosophila. Prog Clin Biol 

Res 209B: 391-398. 

Gramigni, E; Tadini-Buoninsegni, F; Bartolommei, G; Santini, G; Chelazzi, G; Moncelli, MR. (2009). Inhibitory 

effect of Pb2+ on the transport cycle of the Na+,K+-ATPase. Chem Res Toxicol 22: 1699-1704. 

http://dx.doi.org/10.1021/tx9001786.

Grant, LD; Kimmel, CA; West, GL; Martinez-Vargas, CM; Howard, JL. (1980). Chronic low-level lead toxicity in 

the rat: II Effects on postnatal physical and behavioral development. Toxicol Appl Pharmacol 56: 42-58. 

http://dx.doi.org/10.1016/0041-008X(80)90130-1.

Greene, T; Ernhart, CB. (1993). Dentine lead and intelligence prior to school entry: A statistical sensitivity analysis. 

J Clin Epidemiol 46: 323-339. http://dx.doi.org/10.1016/0895-4356(93)90147-S.

Grizzo, LT; Cordellini, S. (2008). Perinatal lead exposure affects nitric oxide and cyclooxygenase pathways in aorta 

of weaned rats. Toxicol Sci 103: 207-214. http://dx.doi.org/10.1093/toxsci/kfn018.

Grlickova-Duzevik, E; Wise, SS; Munroe, RC; Thompson, WD; Wise, JP, Sr. (2006). XRCC1 protects against 

particulate chromate-induced chromosome damage and cytotoxicity in Chinese hamster ovary cells. 

Toxicol Sci 92: 409-415. http://dx.doi.org/10.1093/toxsci/kfl021.

Grover, P; Rekhadevi, PV; Danadevi, K; Vuyyuri, SB; Mahboob, M; Rahman, MF. (2010). Genotoxicity evaluation 

in workers occupationally exposed to lead. Int J Hyg Environ Health 213: 99-106. 

http://dx.doi.org/10.1016/j.ijheh.2010.01.005.

Guguchkova, PT, s. (1972). [Electroretino- and electrooculographic studies in persons occupationally exposed to 

lead for a long period of time]. Vestn Oftalmol 1: 60-65. 

Guilarte, TR; McGlothan, JL. (1998). Hippocampal NMDA receptor mRNA undergoes subunit specific changes 

during developmental lead exposure. Brain Res 790: 98-107. http://dx.doi.org/10.1016/S0006-

8993(98)00054-7.

Guilarte, TR; Toscano, CD; McGlothan, JL; Weaver, SA. (2003). Environmental enrichment reverses cognitive and 

molecular deficits induced by developmental lead exposure. Ann Neurol 53: 50-56. 

http://dx.doi.org/10.1002/ana.10399.

Gulson, B; Mahaffey, KR; Jameson, CW; Patison, N; Law, AJ; Mizon, KJ; Korsch, MJ; Pederson, D. (1999). 

Impact of diet on lead in blood and urine in female adults and relevance to mobilization of lead from bone 

stores. Environ Health Perspect 107: 257-263. http://dx.doi.org/10.1289/ehp.99107257.

Gulson, B; Mizon, KJ; Korsch, MJ; Palmer, JM; Donnelly, JB. (2003). Mobilization of lead from human bone tissue 

during pregnancy and lactation: A summary of long-term research. Sci Total Environ 303: 79-104. 

http://dx.doi.org/10.1016/S0048-9697(02)00355-8.

Gulson, B; Mizon, KJ; Palmer, JM; Korsch, MJ; Taylor, AJ; Mahaffey, KR. (2004a). Blood lead changes during 

pregnancy and postpartum with calcium supplementation. Environ Health Perspect 112: 1499-1507. 

http://dx.doi.org/10.1289/ehp.6548.

Gump, BB; Stewart, P; Reihman, J; Lonky, E; Darvill, T; Matthews, KA; Parsons, PJ. (2005). Prenatal and early 

childhood blood lead levels and cardiovascular functioning in 9 1/2 year old children. Neurotoxicol Teratol 

27: 655-665. http://dx.doi.org/10.1016/j.ntt.2005.04.002.
February 2012 5-571 Draft – Do Not Cite or Quote

Gump, BB; Reihman, J; Stewart, P; Lonky, E; Darvill, T; Matthews, KA. (2007). Blood lead (Pb) levels: A potential 

environmental mechanism explaining the relation between socioeconomic status and cardiovascular 

reactivity in children. Health Psychol 26: 296-304. http://dx.doi.org/10.1037/0278-6133.26.3.296.

Gump, BB; Stewart, P; Reihman, J; Lonky, E; Darvill, T; Parsons, PJ; Granger, DA. (2008). Low-level prenatal and 

postnatal blood lead exposure and adrenocortical responses to acute stress in children. Environ Health 

Perspect 116: 249-255. http://dx.doi.org/10.1289/ehp.10391.

Gump, BB; Reihman, J; Stewart, P; Lonky, E; Granger, DA; Matthews, KA. (2009). Blood lead (Pb) levels: Further 

evidence for an environmental mechanism explaining the association between socioeconomic status and 

psychophysiological dysregulation in children. Health Psychol 28: 614-620. 

http://dx.doi.org/10.1037/a0015611.

Gump, BB; Mackenzie, JA; Bendinskas, K; Morgan, R; Dumas, AK; Palmer, CD; Parsons, PJ. (2011). Low-level 

Pb and cardiovascular responses to acute stress in children: the role of cardiac autonomic regulation. 

Neurotoxicol Teratol 33: 212-219. http://dx.doi.org/10.1016/j.ntt.2010.10.001.

Gundacker, C; Fröhlich, S; Graf-Rohrmeister, K; Eibenberger, B; Jessenig, V; Gicic, D; Prinz, S; Wittmann, KJ; 

Zeisler, H; Vallant, B; Pollak, A; Husslein, P. (2010). Perinatal lead and mercury exposure in Austria. Sci 

Total Environ 408: 5744-5749. http://dx.doi.org/10.1016/j.scitotenv.2010.07.079.

Guo, TL; Mudzinski, SP; Lawrence, DA. (1996a). The heavy metal lead modulates the expression of both TNF-

"alpha" and TNF-"alpha" receptors in lipopolysaccharide-activated human peripheral blood mononuclear 

cells. J Leukoc Biol 59: 932-939. 

Guo, TL; Mudzinski, SP; Lawrence, DA. (1996b). Regulation of HLA-DR and invariant chain expression by human 

peripheral blood mononuclear cells with lead, interferon-"gamma", or interleukin-4. Cell Immunol 171: 1-

9. http://dx.doi.org/10.1006/cimm.1996.0166.

Gustafson, A; Hedner, P; Schutz, A; Skerfving, S. (1989). Occupational lead exposure and pituitary function. Int 

Arch Occup Environ Health 61: 277-281. http://dx.doi.org/10.1007/BF00381426.

Gustavsson, P; Plato, N; Hallqvist, J; Hogstedt, C; Lewne, M; Reuterwall, C; Scheele, P. (2001). A populationbased case-referent study of myocardial infarction and occupational exposure to motor exhaust, other 

combustion products, organic solvents, lead, and dynamite stockholm heart epidemiology program 

(SHEEP) study group. Epidemiology 12: 222-228. 

Habermann, E; Crowell, K; Janicki, P. (1983). Lead and other metals can substitute for Ca2+ in calmodulin. Arch 

Toxicol 54: 61-70. http://dx.doi.org/10.1007/BF00277816.

Hage, FG; Szalai, AJ. (2007). C-reactive protein gene polymorphisms, C-reactive protein blood levels, and 

cardiovascular disease risk. J Am Coll Cardiol 50: 1115-1122. http://dx.doi.org/10.1016/j.jacc.2007.06.012.

HaMai, D; Bondy, SC. (2004). Oxidative basis of manganese neurotoxicity. Ann N Y Acad Sci 1012: 129-141. 

http://dx.doi.org/10.1196/annals.1306.010.

Hamed, EA; Meki, AR; Abd El-Mottaleb, NA. (2010). Protective effect of green tea on lead-induced oxidative 

damage in rat's blood and brain tissue homogenates. J Physiol Biochem 66: 143-151. 

http://dx.doi.org/10.1007/s13105-010-0019-5.

Hamidinia, SA; Erdahl, WL; Chapman, CJ; Steinbaugh, GE; Taylor, RW; Pfeiffer, DR. (2006). Monensin improves 

the effectiveness of meso-dimercaptosuccinate when used to treat lead intoxication in rats. Environ Health 

Perspect 114: 484-493. http://dx.doi.org/10.1289/ehp.8279.

Hanas, JS; Rodgers, JS; Bantle, JA; Cheng, YG. (1999). Lead inhibition of DNA-binding mechanism of 

Cys(2)His(2) zinc finger proteins. Mol Pharmacol 56: 982-988. 

Hara, N; Yamada, K; Terashima, M; Osago, H; Shimoyama, M; Tsuchiya, M. (2003). Molecular identification of 

human glutamine- and ammonia-dependent NAD synthetases: Carbon-nitrogen hydrolase domain confers 

glutamine dependency. J Biol Chem 278: 10914-10921. http://dx.doi.org/10.1074/jbc.M209203200.

Harry, GJ; Schmitt, TJ; Gong, A; Brown, H; Zawia, N; Evans, HL. (1996). Lead-induced alterations of glial 

fibrillary acidic protein (GFAP) in the developing rat brain. Toxicol Appl Pharmacol 139: 84-93. 

http://dx.doi.org/10.1006/taap.1996.0145.
February 2012 5-572 Draft – Do Not Cite or Quote

Hartwig, A; Schlepegrell, R; Beyersmann, D. (1990). Indirect mechanism of lead-induced genotoxicity in cultured 

mammalian cells. Mutat Res Genet Toxicol 241: 75-82. http://dx.doi.org/10.1016/0165-1218(90)90110-N.

Hashemzadeh-Gargari, H; Guilarte, TR. (1999). Divalent cations modulate N-methyl-D-aspartate receptor function 

at the glycine site. J Pharmacol Exp Ther 290: 1356-1362. 

Hashmi, NS; Kachru, DN; Khandelwal, S; Tandon, SK. (1989). Interrelationship between iron deficiency and lead 

intoxication (part 2). Biol Trace Elem Res 22: 299-307. http://dx.doi.org/10.1007/BF02916618.

Hauser, R; Sergeyev, O; Korrick, S; Lee, MM; Revich, B; Gitin, E; Burns, JS; Williams, PL. (2008). Association of 

blood lead levels with onset of puberty in Russian boys. Environ Health Perspect 116: 976-980. 

http://dx.doi.org/10.1289/ehp.10516.

Hausmann, OV; Gentinetta, T; Fux, M; Ducrest, S; Pichler, WJ; Dahinden, CA. (2011). Robust expression of CCR3 

as a single basophil selection marker in flow cytometry. Allergy 66: 85-91. 

http://dx.doi.org/10.1111/j.1398-9995.2010.02431.x.

He, LH; Poblenz, AT; Medrano, CJ; Fox, DA. (2000). Lead and calcium produce rod photoreceptor cell apoptosis 

by opening the mitochondrial permeability transition pore. J Biol Chem 275: 12175-12184. 

http://dx.doi.org/10.1074/jbc.275.16.12175.

He, S; Dong, W; Deng, Q; Weng, S; Sun, W. (2003). Seeing more clearly: Recent advances in understanding retinal 

circuitry. Science 302: 408-411. http://dx.doi.org/10.1126/science.1085457.

He, T; Hirsch, HVB; Ruden, DM; Lnenicka, GA. (2009). Chronic lead exposure alters presynaptic calcium 

regulation and synaptic facilitation in Drosophila larvae. Neurotoxicology 30: 777-784. 

http://dx.doi.org/10.1016/j.neuro.2009.08.007.

Hechtenberg, S; Beyersmann, D. (1991). Inhibition of sarcoplasmic reticulum Ca2+-ATPase activity by cadmium, 

lead and mercury. Enzyme 45: 109-115. 

Hegazy, RM; Hamdy, R; Kamel, HF. (2011). Modulation of ige levels in lead exposed children by parental cigarette 

smoking, Qualyobia Governate, Egypt. Int J Pharma Bio Sci 2: 272-385. 

Heiman, AS; Tonner, LE. (1995). The acute effect of lead acetate on glucocorticoid regulation of tyrosine 

aminotransferase in hepatoma cells. Toxicology 100: 57-68. http://dx.doi.org/10.1016/0300-

483X(95)03061-J.

Hellstrom, L; Elinder, CG; Dahlberg, B; Lundberg, M; Jarup, L; Persson, B; Axelson, O. (2001). Cadmium 

exposure and end-stage renal disease. Am J Kidney Dis 38: 1001-1008. 

http://dx.doi.org/10.1053/ajkd.2001.28589.

Hemdan, NYA; Emmrich, F; Adham, K; Wichmann, G; Lehmann, I; El-Massry, A; Ghoneim, H; Lehmann, J; Sack, 

U. (2005). Dose-dependent modulation of the in vitro cytokine production of human immune competent 

cells by lead salts. Toxicol Sci 86: 75-83. http://dx.doi.org/10.1093/toxsci/kfi177.

Hengstler, JG; Bolm-Audorff, U; Faldum, A; Janssen, K; Reifenrath, M; Gotte, W; Jung, D; Mayer-Popken, O; 

Fuchs, J; Gebhard, S; Bienfait, HG; Schlink, K; Dietrich, C; Faust, D; Epe, B; Oesch, F. (2003). 

Occupational exposure to heavy metals: DNA damage induction and DNA repair inhibition prove coexposures to cadmium, cobalt and lead as more dangerous than hitherto expected. Carcinogenesis 24: 63-

73. http://dx.doi.org/10.1093/carcin/24.1.63.

Heo, Y; Parsons, PJ; Lawrence, DA. (1996). Lead differentially modifies cytokine production in vitro and in vivo. 

Toxicol Appl Pharmacol 138: 149-157. http://dx.doi.org/10.1006/taap.1996.0108.

Heo, Y; Lee, BK; Ahn, KD; Lawrence, DA. (2004). Serum IgE elevation correlates with blood lead levels in battery 

manufacturing workers. Hum Exp Toxicol 23: 209-213. http://dx.doi.org/10.1191/0960327104ht442oa.

Heo, Y; Mondal, TK; Gao, DH; Kasten-Jolly, J; Kishikawa, H; Lawrence, DA. (2007). Posttranscriptional inhibition 

of interferon-gamma production by lead. Toxicol Sci 96: 92-100. http://dx.doi.org/10.1093/toxsci/kfl182.

Hermes-Lima, M; Pereira, B; Bechara, EJH. (1991). Are free radicals involved in lead poisoning? Xenobiotica 21: 

1085-1090. http://dx.doi.org/10.3109/00498259109039548.
February 2012 5-573 Draft – Do Not Cite or Quote

Hernandez-Avila, M; Smith, D; Meneses, F; Sanin, LH; Hu, H. (1998). The influence of bone and blood lead on 

plasma lead levels in environmentally exposed adults. Environ Health Perspect 106: 473-477. 

http://dx.doi.org/10.1289/ehp.98106473.

Hernández-Franco, P; Silva, M; Valverde, M; Rojas, E. (2011). Induction of oxidative stress by low doses of lead in 

human hepatic cell line WRL-68. Biometals 24: 951-958. http://dx.doi.org/10.1007/s10534-011-9453-2.

Hernandez-Ochoa, I; Sanchez-Gutierrez, M; Solis-Heredia, MJ; Quintanilla-Vega, B. (2006). Spermatozoa nucleus 

takes up lead during the epididymal maturation altering chromatin condensation. Reprod Toxicol 21: 171-

178. http://dx.doi.org/10.1016/j.reprotox.2005.07.015.

Hernandez-Serrato, MI; Fortoul, TI; Rojas-Martinez, R; Mendoza-Alvarado, LR; Canales-Trevino, L; BochichioRiccardelli, T; Avila-Costa, MR; Olaiz-Fernandez, G. (2006). Lead blood concentrations and renal function 

evaluation: Study in an exposed Mexican population. Environ Res 100: 227-231. 

http://dx.doi.org/10.1016/j.envres.2005.03.004.

Herold, C. (2010). NMDA and D2-like receptors modulate cognitive flexibility in a color discrimination reversal 

task in pigeons. Behav Neurosci 124: 381-390. http://dx.doi.org/10.1037/a0019504.

Heydari, A; Norouzzadeh, A; Khoshbaten, A; Asgari, A; Ghasemi, A; Najafi, S; Badalzadeh, R. (2006). Effects of 

short-term and subchronic lead poisoning on nitric oxide metabolites and vascular responsiveness in rat. 

Toxicol Lett 166: 88-94. http://dx.doi.org/10.1016/j.toxlet.2006.05.014.

Hilson, JA; Strupp, BJ. (1997). Analyses of response patterns clarify lead effects in olfactory reversal and 

extradimensional shift tasks: Assessment of inhibitory control, associative ability, and memory. Behav 

Neurosci 111: 532-542. 

Holmes, AL; Wise, SS; Sandwick, SJ; Lingle, WL; Negron, VC; Thompson, WD; Wise, JP, Sr. (2006a). Chronic 

exposure to lead chromate causes centrosome abnormalities and aneuploidy in human lung cells. Cancer 

Res 66: 4041-4048. http://dx.doi.org/10.1158/0008-5472.can-05-3312.

Holmes, AL; Wise, SS; Sandwick, SJ; Wise, JP. (2006b). The clastogenic effects of chronic exposure to particulate 

and soluble Cr(VI) in human lung cells. Mutat Res Genet Toxicol Environ Mutagen 610: 8-13. 

http://dx.doi.org/10.1016/j.mrgentox.2006.06.006.

Holtzman, D; Olson, JE; DeVries, C; Bensch, K. (1987). Lead toxicity in primary cultured cerebral astrocytes and 

cerebellar granular neurons. Toxicol Appl Pharmacol 89: 211-225. http://dx.doi.org/10.1016/0041-

008X(87)90042-1.

Hon, KL; Ching, GK; Hung, EC; Leung, TF. (2009). Serum lead levels in childhood eczema. Clin Exp Dermatol 34: 

e508-e509. http://dx.doi.org/10.1111/j.1365-2230.2009.03596.x.

Hon, KL; Wang, SS; Hung, EC; Lam, HS; Lui, HH; Chow, CM; Ching, GK; Fok, TF; Ng, PC; Leung, TF. (2010). 

Serum levels of heavy metals in childhood eczema and skin diseases: Friends or foes. Pediatric Allergy and 

Immunology 21: 831-836. http://dx.doi.org/10.1111/j.1399-3038.2010.01022.x.

Hong, YC; Hwang, SS; Kim, JH; Lee, KH; Lee, HJ; Yu, SD; Kim, DS. (2007b). Metals in particulate pollutants 

affect peak expiratory flow of schoolchildren. Environ Health Perspect 115: 430-434. 

http://dx.doi.org/10.1289/ehp.9531.

Hornung, R; Lanphear, B; Kietrich, K. (2006). Response to: "What is the meaning of non-linear dose-response 

relationships between blood lead concentration and IQ?" [letter] [Letter]. Neurotoxicology 27: 635. 

Hornung, R. (2008). Email from Richard Hornung, Cincinnati Children's Hospital Medical Center (CCHMC) to JeeYoung Kim, USEPA Subject: Request for additional information on Lanphear et al. (2005). (EPA-HQOAR-2006-0735-5814). Hornung, R. http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OAR2006-0735-5814.

Hornung, RW; Lanphear, BP; Dietrich, KN. (2009). Age of greatest susceptibility to childhood lead exposure: A 

new statistical approach. Environ Health Perspect 117: 1309-1312. http://dx.doi.org/10.1289/ehp.0800426.

Hotz, P; Buchet, JP; Bernard, A; Lison, D; Lauwerys, R. (1999). Renal effects of low-level environmental cadmium 

exposure: 5-year follow-up of a subcohort from the Cadmibel study. Lancet 354: 1508-1513. 

http://dx.doi.org/10.1016/S0140-6736(99)91145-5.
February 2012 5-574 Draft – Do Not Cite or Quote

Howard, JK. (1974). Human erythrocyte glutathione reductase and glucose 6-phosphate dehydrogenase activities in 

normal subjects and in persons exposed to lead. Clin Sci Mol Med 47: 515-520. 

Hsiao, CL; Wu, KH; Wan, KS. (2011). Effects of environmental lead exposure on T-helper cell-specific cytokines in 

children. J Immunotoxicol 8: 284-287. http://dx.doi.org/10.3109/1547691X.2011.592162.

Hsiao, CY; Wu, HD; Lai, JS; Kuo, HW. (2001). A longitudinal study of the effects of long-term exposure to lead 

among lead battery factory workers in Taiwan (1989-1999). Sci Total Environ 279: 151-158. 

http://dx.doi.org/10.1016/S0048-9697(01)00762-8.

Hsieh, SJ; Chiu, YW; Li, WF; Wu, CH; Chen, HI; Chuang, HY. (2009a). Increased concentrations of serum inhibin 

B among male workers with long-term moderate lead exposure. Sci Total Environ 407: 2603-2607. 

http://dx.doi.org/10.1016/j.scitotenv.2008.12.056.

Hsieh, TJ; Chen, YC; Li, CW; Liu, GC; Chiu, YW; Chuang, HY. (2009b). A proton magnetic resonance 

spectroscopy study of the chronic lead effect on the basal ganglion and frontal and occipital lobes in 

middle-age adults. Environ Health Perspect 117: 941-945. http://dx.doi.org/10.1289/ehp.0800187.

Hsu, CW; Lin, JL; Lin-Tan, DT; Yen, TH; Huang, WH; Ho, TC; Huang, YL; Yeh, LM; Huang, LM. (2009a). 

Association of environmental cadmium exposure with inflammation and malnutrition in maintenance 

haemodialysis patients. Nephrol Dial Transplant 24: 1282-1288. http://dx.doi.org/10.1093/ndt/gfn602.

Hsu, JM. (1981). Lead toxicity as related to glutathione metabolism. J Nutr 111: 26-33. 

Hsu, PC; Liu, MY; Hsu, CC; Chen, LY; Guo, YL. (1998a). Effects of vitamin E and/or C on reactive oxygen 

species-related lead toxicity in the rat sperm. Toxicology 128: 169-179. http://dx.doi.org/10.1016/S0300-

483X(98)00068-7.

Hsu, PC; Hsu, CC; Liu, MY; Chen, LY; Guo, YL. (1998b). Lead-induced changes in spermatozoa function and 

metabolism. J Toxicol Environ Health A 55: 45-64. http://dx.doi.org/10.1080/009841098158610.

Hsu, PC; Chang, HY; Guo, YL; Liu, YC; Shih, TS. (2009b). Effect of smoking on blood lead levels in workers and 

role of reactive oxygen species in lead-induced sperm chromatin DNA damage. Fertil Steril 91: 1096-1103. 

http://dx.doi.org/10.1016/j.fertnstert.2008.01.005.

Hu, H. (1991). A 50-year follow-up of childhood plumbism: Hypertension, renal function, and hemoglobin levels 

among survivors. Am J Dis Child 145: 681-687. 

Hu, H; Payton, M; Korrick, S; Aro, A; Sparrow, D; Weiss, ST; Rotnitzky, A. (1996a). Determinants of bone and 

blood lead levels among community-exposed middle-aged to elderly men: The normative aging study. Am 

J Epidemiol 144: 749-759. 

Hu, H; Aro, A; Payton, M; Korrick, S; Sparrow. (1996b). The relationship of bone and blood lead to hypertension: 

The Normative Aging Study. JAMA 275: 1171-1176. 

http://dx.doi.org/10.1001/jama.1996.03530390037031.

Hu, H; Hernandez-Avila, M. (2002). Lead, bones, women, and pregnancy: The poison within? Am J Epidemiol 156: 

1088-1091. http://dx.doi.org/10.1093/aje/kwf164.

Hu, H; Tellez-Rojo, MM; Bellinger, D; Smith, D; Ettinger, AS; Lamadrid-Figueroa, H; Schwartz, J; Schnaas, L; 

Mercado-Garca, A; Hernandez-Avila, M. (2006). Fetal lead exposure at each stage of pregnancy as a 

predictor of infant mental development. Environ Health Perspect 114: 1730-1735. 

http://dx.doi.org/10.1289/ehp.9067.

Hu, P; Wang, M; Chen, WH; Liu, J; Chen, L; Yin, ST; Yong, W; Chen, JT; Wang, HL; Ruan, DY. (2008a). 

Quercetin relieves chronic lead exposure-induced impairment of synaptic plasticity in rat dentate gyrus in 

vivo. Naunyn Schmiedebergs Arch Pharmacol 378: 43-51. http://dx.doi.org/10.1007/s00210-008-0301-z.

Hu, QS; Fu, HJ; Ren, TL; Wang, SY; Zhou, W; Song, H; Han, YF; Dong, SZ. (2008b). Maternal low-level lead 

exposure reduces the expression of PSA-NCAM and the activity of sialyltransferase in the hippocampi of 

neonatal rat pups. Neurotoxicology 29: 675-681. http://dx.doi.org/10.1016/j.neuro.2008.04.002.
February 2012 5-575 Draft – Do Not Cite or Quote

Huang, BM; Lai, HY; Liu, MY. (2002). Concentration dependency in lead-inhibited steroidogenesis in MA-10 

mouse Leydig tumor cells. J Toxicol Environ Health A 65: 557-567. 

http://dx.doi.org/10.1080/15287390252808000.

Huang, BM; Liu, MY. (2004). Inhibitory actions of lead on steroidogenesis in MA-10 mouse Leydig tumor cells. 

Sys Biol Reprod Med 50: 5-9. http://dx.doi.org/10.1080/01485010490250434.

Huang, H; Bihaqi, SW; Cui, L; Zawia, NH. (2011a). In vitro Pb exposure disturbs the balance between Aβ 

production and elimination: the role of AβPP and neprilysin. Neurotoxicology 32: 300-306. 

http://dx.doi.org/10.1016/j.neuro.2011.02.001.

Huang, J; Wu, J; Li, T; Song, X; Zhang, B; Zhang, P; Zheng, X. (2011b). Effect of exposure to trace elements in the 

soil on the prevalence of neural tube defects in a high-risk area of China. Biomed Environ Sci 24: 94-101. 

http://dx.doi.org/10.3967/0895-3988.2011.02.002.

Huang, M; Krepkiy, D; Hu, W; Petering, DH. (2004). Zn-, Cd-, and Pb-transcription factor IIIA: Properties, DNA 

binding, and comparison with TFIIIA-finger 3 metal complexes. J Inorg Biochem 98: 775-785. 

http://dx.doi.org/10.1016/j.jinorgbio.2004.01.014.

Huang, XP; Feng, ZY; Zhai, WL; Xu, JH. (1988). Chromosomal aberrations and sister chromatid exchanges in 

workers exposed to lead. Biomed Environ Sci 1: 382-387. 

Hudson, CA; Cao, L; Kasten-Jolly, J; Kirkwood, JN; Lawrence, DA. (2003). Susceptibility of lupus-prone NZM 

mouse strains to lead exacerbation of systemic lupus erythematosus symptoms. J Toxicol Environ Health A 

66: 895-918. http://dx.doi.org/10.1080/15287390306456.

Huel, G; Sahuquillo, J; Debotte, G; Oury, JF; Takser, L. (2008). Hair mercury negatively correlates with calcium 

pump activity in human term newborns and their mothers at delivery. Environ Health Perspect 116: 263-

267. http://dx.doi.org/10.1289/ehp.10381.

Huo, X; Peng, L; Xu, XJ; Zheng, LK; Qiu, B; Qi, ZL; Zhang, B; Han, D; Piao, ZX. (2007). Elevated blood lead 

levels of children in Guiyu, an electronic waste recycling town in China. Environ Health Perspect 115: 

1113-1117. http://dx.doi.org/10.1289/ehp.9697.

Huseman, CA; Moriarty, CM; Angle, CR. (1987). Childhood lead toxicity and impaired release of thyrotropinstimulating hormone. Environ Res 42: 524-533. http://dx.doi.org/10.1016/S0013-9351(87)80219-0.

Huseman, CA; Varma, MM; Angle, CR. (1992). Neuroendocrine effects of toxic and low blood lead levels in 

children. Pediatrics 90: 186-189. 

Hwang, KY; Lee, BK; Bressler, JP; Bolla, KI; Stewart, WF; Schwartz, BS. (2002). Protein kinase C activity and the 

relations between blood lead and neurobehavioral function in lead workers. Environ Health Perspect 110: 

133-138. http://dx.doi.org/10.1289/ehp.02110133.

Hwang, YH; Chiang, HY; Yen-Jean, MC; Wang, JD. (2009). The association between low levels of lead in blood 

and occupational noise-induced hearing loss in steel workers. Sci Total Environ 408: 43-49. 

http://dx.doi.org/10.1016/j.scitotenv.2009.09.016.

Hwua, YS; Yang, JL. (1998). Effect of 3-aminotriazole on anchorage independence and mutagenicity in cadmiumand lead-treated diploid human fibroblasts. Carcinogenesis 19: 881-888. 

http://dx.doi.org/10.1093/carcin/19.5.881.

IARC (International Agency for Research on Cancer). (2006a). Inorganic and organic lead compounds. Lyon, 

France. http://monographs.iarc.fr/ENG/Monographs/vol87/index.php.

Iavicoli, I; Sgambato, A; Carelli, G; Ardito, R; Cittadini, A; Castellino, N. (2001). Lead-related effects on rat 

fibroblasts. Mol Cell Biochem 222: 35-40. http://dx.doi.org/10.1023/A:1017930401906.

Iavicoli, I; Carelli, G; III, SE; Castellino, N; Calabrese, EJ. (2004). Effects of low doses of dietary lead on puberty 

onset in female mice. Reprod Toxicol 19: 35-41. 

Iavicoli, I; Carelli, G; Stanek, EJ, III; Castellino, N; Calabrese, EJ. (2006a). Below background levels of blood lead 

impact cytokine levels in male and female mice. Toxicol Appl Pharmacol 210: 94-99. 

http://dx.doi.org/10.1016/j.taap.2005.09.016.
February 2012 5-576 Draft – Do Not Cite or Quote

Iavicoli, I; Carelli, G; Stanek, EJ; Castellino, N; Li, Z; Calabrese, EJ. (2006b). Low doses of dietary lead are 

associated with a profound reduction in the time to the onset of puberty in female mice. Reprod Toxicol 22: 

586-590. http://dx.doi.org/10.1016/j.reprotox.2006.03.016.

Iavicoli, I; Carelli, G. (2007). The effects of lead on the hematological parameters of a second and third generation 

of Swiss mice. Toxicol Lett 172: S116-S116. http://dx.doi.org/10.1016/j.toxlet.2007.05.309.

Ignasiak, Z; Slawinska, T; Rozek, K; Little, BB; Malina, RM. (2006). Lead and growth status of schoolchildren 

living in the copper basin of south-western Poland: Differential effects on bone growth. Ann Hum Biol 33: 

401-414. http://dx.doi.org/10.1080/03014460600730752.

Iijima, K; Otake, T; Yoshinaga, J; Ikegami, M; Suzuki, E; Naruse, H; Yamanaka, T; Shibuya, N; Yasumizu, T; 

Kato, N. (2007). Cadmium, lead, and selenium in cord blood and thyroid hormone status of newborns. Biol 

Trace Elem Res 119: 10-18. http://dx.doi.org/10.1007/s12011-007-0057-1.

Ikebuchi, H; Teshima, R; Suzuki, K; Terao, T; Yamane, Y. (1986). Simultaneous induction of Pb-metallothioneinlike protein and Zn-thionein in the liver of rats given lead acetate. Biochemistry 233: 541-546. 

Ikonomidou, C; Bosch, F; Miksa, M; Bittigau, P; Vöckler, J; Dikranian, K; Tenkova, TI; Stefovska, V; Turski, L; 

Olney, JW. (1999). Blockade of NMDA receptors and apoptotic neurodegeneration in the developing brain. 

Science 283: 70-74. http://dx.doi.org/10.1126/science.283.5398.70.

Ingelsson, E; Gona, P; Larson, MG; Lloyd-Jones, DM; Kannel, WB; Vasan, RS; Levy, D. (2008). Altered blood 

pressure progression in the community and its relation to clinical events. Arch Intern Med 168: 1450-1457. 

http://dx.doi.org/10.1001/archinte.168.13.1450.

Inglis, JA; Henderson, DA; Emmerson, BT. (1978). The pathology and pathogenesis of chronic lead nephropathy 

occurring in Queensland. J Pathol 124: 65-76. http://dx.doi.org/10.1002/path.1711240202.

Institoris, L; Kovacs, D; Kecskemeti-Kovacs, I; Lukacs, A; Szabo, A; Lengyel, Z; Papp, A; Nagymajtenyi, L; Desi, 

I. (2006). Immunotoxicological investigation of subacute combined exposure with low doses of PB, HG 

and CD in rats. Acta Biol Hung 57: 433-439. http://dx.doi.org/10.1556/ABiol.57.2006.4.5.

Ionescu, JG; Novotny, J; Stejskal, V; Latsch, A; Blaurock-Busch, E; Eisenmann-Klein, M. (2007). Breast tumours 

strongly accumulate transition metals. Maedica 2: 5-9. 

Iranpour, R; Besharati, AA; Nasseri, F; Hashemipour, M; Balali-Mood, M; Kelishadi, R. (2007). Comparison of 

blood lead levels of mothers and cord blood in intrauterine growth retarded neonates and normal term 

neonates. Saudi Med J 28: 877-880. 

Irgens, A; Kruger, K; Skorve, AH; Irgens, LM. (1998). Reproductive outcome in offspring of parents occupationally 

exposed to lead in Norway. Am J Ind Med 34: 431-437. 

Iwata, T; Yano, E; Karita, K; Dakeishi, M; Murata, K. (2005). Critical dose of lead affecting postural balance in 

workers. Am J Ind Med 48: 319-325. http://dx.doi.org/10.1002/ajim.20220.

Jaako-Movits, K; Zharkovsky, T; Romantchik, O; Jurgenson, M; Merisalu, E; Heidmets, LT; Zharkovsky, A.

(2005). Developmental lead exposure impairs contextual fear conditioning and reduces adult hippocampal 

neurogenesis in the rat brain. Int J Dev Neurosci 23: 627-635. 

http://dx.doi.org/10.1016/j.ijdevneu.2005.07.005.

Jabeen, R; Tahir, M; Waqas, S. (2010). Teratogenic effects of lead acetate on kidney. J Ayub Med Coll Abbottabad 

22: 76-79. 

Jackson, LW; Howards, PP; Wactawski-Wende, J; Schisterman, EF. (2011). The association between cadmium, lead 

and mercury blood levels and reproductive hormones among healthy, premenopausal women. Hum Reprod 

26: 2887-2895. http://dx.doi.org/10.1093/humrep/der250.

Jacobsen, C; Hartvigsen, K; Thomsen, MK; Hansen, LF; Lund, P; Skibsted, LH; Hølmer, G; Adler-Nissen, J; 

Meyer, AS. (2001). Lipid oxidation in fish oil enriched mayonnaise: Calcium disodium 

ethylenediaminetetraacetate, but not gallic acid, strongly inhibited oxidative deterioration. J Agric Food 

Chem 49: 1009-1019. http://dx.doi.org/10.1021/jf000729r.
February 2012 5-577 Draft – Do Not Cite or Quote

Jadhav, SH; Sarkar, SN; Ram, GC; Tripathi, HC. (2007). Immunosuppressive effect of subchronic exposure to a 

mixture of eight heavy metals, found as groundwater contaminants in different areas of india, through 

drinking water in male rats. Arch Environ Contam Toxicol 53: 450-458. http://dx.doi.org/10.1007/s00244-

006-0177-1.

Jagetia, GC; Aruna, R. (1998). Effect of various concentrations of lead nitrate on the induction of micronuclei in 

mouse bone marrow. Mutat Res Genet Toxicol Environ Mutagen 415: 131-137. 

http://dx.doi.org/10.1016/S1383-5718(98)00052-7.

Jain, NB; Potula, V; Schwartz, J; Vokonas, PS; Sparrow, D; Wright, RO; Nie, H; Hu, H. (2007). Lead levels and 

ischemic heart disease in a prospective study of middle-aged and elderly men: The VA Normative Aging 

Study. Environ Health Perspect 115: 871-875. http://dx.doi.org/10.1289/ehp.9629.

Jamieson, JA; Shuhyta, JN; Taylor, CG. (2007). Lead does not affect transcription of intestinal zinc-binding proteins 

in growing rats. Exp Biol Med 232: 744-753. 

Jamieson, JA; Stringer, DM; Zahradka, P; Taylor, CG. (2008). Dietary zinc attenuates renal lead deposition but 

metallothionein is not directly involved. Biometals 21: 29-40. http://dx.doi.org/10.1007/s10534-007-9090-

y.

Jang, HO; Kim, JS; Kwon, WC; Kim, JK; Ko, MS; Kim, DH; Kim, WI; Jeon, YC; Chung, IK; Shin, SH; Chung, J; 

Bae, MK; Yun, I. (2008). The effect of lead on calcium release activated calcium influx in primary cultures 

of human osteoblast-like cells. Arch Pharm Res 31: 188-194. http://dx.doi.org/10.1007/s12272-001-1140-3.

Jang, WH; Lim, KM; Kim, K; Noh, JY; Kang, S; Chang, YK; Chung, JH. (2011). Low level of lead can induce 

phosphatidylserine exposure and erythrophagocytosis: A new mechanism underlying lead-associated 

anemia. Toxicol Sci 122: 177-184. http://dx.doi.org/10.1093/toxsci/kfr079.

Janjua, NZ; Delzell, E; Larson, RR; Meleth, S; Kristensen, S; Kabagambe, E; Sathiakumar, N. (2009). Determinants 

of low birth weight in urban Pakistan. Public Health Nutr 12: 789-798. 

http://dx.doi.org/10.1017/S1368980008002942.

Jayakumar, T; Sridhar, MP; Bharathprasad, TR; Ilayaraja, M; Govindasamy, S; Balasubramanian, MP. (2009). 

Experimental studies of Achyranthes aspera (L) preventing nephrotoxicity induced by lead in albino rats. J 

Health Sci 55: 701-708. http://dx.doi.org/10.1248/jhs.55.701.

Jedrychowski, W; Perera, F; Jankowski, J; Rauh, V; Flak, E; Caldwell, KL; Jones, RL; Pac, A; Lisowska-Miszczyk, 

I. (2008). Prenatal low-level lead exposure and developmental delay of infants at age 6 months (Krakow 

inner city study). Int J Hyg Environ Health 211: 345-351. http://dx.doi.org/10.1016/j.ijheh.2007.07.023.

Jedrychowski, W; Perera, F; Jankowski, J; Mrozek-Budzyn, D; Mroz, E; Flak, E; Edwards, S; Skarupa, A; 

Lisowska-Miszczyk, I. (2009a). Gender specific differences in neurodevelopmental effects of prenatal 

exposure to very low-lead levels: The prospective cohort study in three-year olds. Early Hum Dev 85: 503-

510. http://dx.doi.org/10.1016/j.earlhumdev.2009.04.006.

Jedrychowski, W; Perera, FP; Jankowski, J; Mrozek-Budzyn, D; Mroz, E; Flak, E; Edwards, S; Skarupa, A; 

Lisowska-Miszczyk, I. (2009b). Very low prenatal exposure to lead and mental development of children in 

infancy and early childhood: Krakow prospective cohort study. Neuroepidemiology 32: 270-278. 

http://dx.doi.org/10.1159/000203075.

Jedrychowski, W; Perera, F; Maugeri, U; Rembiasz, M; Flak, E; Mroz, E; Majewska, R; Zembala, M. (2011). 

Intrauterine exposure to lead may enhance sensitization to common inhalant allergens in early childhood: A 

prospective prebirth cohort study. Environ Res 111: 119-124. 

http://dx.doi.org/10.1016/j.envres.2010.11.002.

Jehan, ZS; Motlag, DB. (1995). Metal induced changes in the erythrocyte membrane of rats. Toxicol Lett 78: 127-

133. http://dx.doi.org/10.1016/0378-4274(94)03245-3.

Jelliffe-Pawlowski, LL; Miles, SQ; Courtney, JG; Materna, B; Charlton, V. (2006). Effect of magnitude and timing 

of maternal pregnancy blood lead (Pb) levels on birth outcomes. J Perinatol 26: 154-162. 

http://dx.doi.org/10.1038/sj.jp.7211453.
February 2012 5-578 Draft – Do Not Cite or Quote

Jett, DA; Kuhlmann, AC; Farmer, SJ; Guilarte, TR. (1997). Age-dependent effects of developmental lead exposure 

on performance in the Morris water maze. Pharmacol Biochem Behav 57: 271-279. 

http://dx.doi.org/10.1016/S0091-3057(96)00350-4.

Jiang, YM; Long, LL; Zhu, XY; Zheng, H; Fu, X; Ou, SY; Wei, DL; Zhou, HL; Zheng, W. (2008). Evidence for 

altered hippocampal volume and brain metabolites in workers occupationally exposed to lead: A study by 

magnetic resonance imaging and 1H magnetic resonance spectroscopy. Toxicol Lett 181: 118-125. 

http://dx.doi.org/10.1016/j.toxlet.2008.07.009.

Jin, C; Li, Y; Li, YL; Zou, Y; Zhang, GL; Normura, M; Zhu, GY. (2008). Blood lead: Its effect on trace element 

levels and iron structure in hemoglobin. Nucl Instrum Methods Phys Res B 266: 3607-3613. 

http://dx.doi.org/10.1016/j.nimb.2008.05.087.

Jin, YP; Liao, YJ; Lu, CW; Li, GX; Yu, F; Zhi, XP; Xu, JH; Liu, SX; Liu, MM; Yang, J. (2006). Health effects in 

children aged 3-6 years induced by environmental lead exposure. Ecotoxicol Environ Saf 63: 313-317. 

http://dx.doi.org/10.1016/j.ecoenv.2005.05.011.

Johnson, B, . L.; Elbatawi, M, .; Xintaras, C, .; Baker, E, . L., J.,r.; Hanninen, H, .; Seppalainen, A, . M. (Eds.). 

(1987). Prevention of neurotoxic illness in working populations. New York: John Willey & Sons. 

Johnson, RJ; Kang, DH; Feig, D; Kivlighn, S; Kanellis, J; Watanabe, S; Tuttle, KR; Rodriguez-Iturbe, B; HerreraAcosta, J; Mazzali, M. (2003). Is there a pathogenetic role for uric acid in hypertension and cardiovascular 

and renal disease? Hypertension 41: 1183-1190. http://dx.doi.org/10.1161/01.HYP.0000069700.62727.C5.

Jomova, K; Valko, M. (2011). Advances in metal-induced oxidative stress and human disease. Toxicology 283: 65-

87. http://dx.doi.org/10.1016/j.tox.2011.03.001.

Jomova, K; Jenisova, Z; Feszterova, M; Baros, S; Liska, J; Hudecova, D; Rhodes, CJ; Valko, M. (2011). Arsenic: 

toxicity, oxidative stress and human disease. J Appl Toxicol 31: 95-107. http://dx.doi.org/10.1002/jat.1649.

Jones, AYM; Lam, PKW; Dean, E. (2006). Respiratory health of bus drivers in Hong Kong. Int Arch Occup 

Environ Health 79: 414-418. http://dx.doi.org/10.1007/s00420-005-0061-8.

Jones, AYM; Lam, PK; Gohel, MD. (2008). Respiratory health of road-side vendors in a large industrialized city. 

Environ Sci Pollut Res Int 15: 150-154. http://dx.doi.org/10.1065/espr2006.12.368.

Jones, EA; Wright, JM; Rice, G; Buckley, BT; Magsumbol, MS; Barr, DB; Williams, BL. (2010). Metal exposures 

in an inner-city neonatal population. Environ Int 36: 649-654. 

http://dx.doi.org/10.1016/j.envint.2010.04.007.

Jones, SR; Atkin, P; Holroyd, C; Lutman, E; Batlle, JVI; Wakeford, R; Walker, P. (2007). Lung cancer mortality at 

a UK tin smelter. Occup Med (Lond) 57: 238-245. http://dx.doi.org/10.1093/occmed/kql153.

Joseph, CLM; Havstad, S; Ownby, DR; Peterson, EL; Maliarik, M; McCabe, MJ; Barone, C; Johnson, CC. (2005). 

Blood lead levels and risk of asthma. Environ Health Perspect 113: 900-904. 

http://dx.doi.org/10.1289/ehp.7453.

Jurczuk, M; Moniuszko-Jakoniuk, J; Brzóska, MM. (2006). Involvement of some low-molecular thiols in the 

peroxidative mechanisms of lead and ethanol action on rat liver and kidney. Toxicology 219: 11-21. 

http://dx.doi.org/10.1016/j.tox.2005.10.022.

Jurczuk, M; Brzoska, MM; Moniuszko-Jakoniuk, J. (2007). Hepatic and renal concentrations of vitamins E and C in 

lead- and ethanol-exposed rats. An assessment of their involvement in the mechanisms of peroxidative 

damage. Food Chem Toxicol 45: 1478-1486. http://dx.doi.org/10.1016/j.fct.2007.02.007.

Jusko, TA; Henderson, CR; Lanphear, BP; Cory-Slechta, DA; Parsons, PJ; Canfield, RL. (2008). Blood lead 

concentrations < 10 microg/dL and child intelligence at 6 years of age. Environ Health Perspect 116: 243-

248. http://dx.doi.org/10.1289/ehp.10424.

Kaczynska, K; Walski, M; Szereda-Przestaszewska, M. (2011). Ultrastructural changes in lung tissue after acute 

lead intoxication in the rat. J Electron Microsc (Tokyo) 60: 289-294. 

http://dx.doi.org/10.1093/jmicro/dfr035.
February 2012 5-579 Draft – Do Not Cite or Quote

Kaji, T; Suzuki, M; Yamamoto, C; Mishima, A; Sakamoto, M; Kozuka, H. (1995). Severe damage of cultured 

vascular endothelial cell monolayer after simultaneous exposure to cadmium and lead. Arch Environ 

Contam Toxicol 28: 168-172. http://dx.doi.org/10.1007/BF00217612.

Kamal, M; Fathy, MM; Taher, E; Hasan, M; Tolba, M. (2011). Assessment of the role of paraoxonase gene 

polymorphism (Q192R) and paraoxonase activity in the susceptibility to atherosclerosis among leadexposed workers. Ann Saudi Med 31: 481-487. http://dx.doi.org/10.4103/0256-4947.84625.

Kamel, F; Umbach, DM; Munsat, TL; Shefner, JM; Hu, H; Sandler, DP. (2002). Lead exposure and amyotrophic 

lateral sclerosis. Epidemiology 13: 311-319. 

Kamel, F; Umbach, DM; Hu, H; Munsat, TL; Shefner, JM; Taylor, JA; Sandler, DP. (2005). Lead exposure as a risk 

factor for amyotrophic lateral sclerosis. Neurodegener Dis 2: 195-201. 

http://dx.doi.org/10.1159/000089625.

Kamel, F; Umbach, DM; Stallone, L; Richards, M; Hu, H; Sandler, DP. (2008). Association of lead exposure with 

survival in amyotrophic lateral sclerosis. Environ Health Perspect 116: 943-947. 

http://dx.doi.org/10.1289/ehp.11193.

Kanduc, D; Rossiello, MR; Aresta, A; Cavazza, C; Quagliariello, E; Farber, E. (1991). Transitory DNA 

hypomethylation during liver cell proliferation induced by a single dose of lead nitrate. Arch Biochem 

Biophys 286: 212-216. http://dx.doi.org/10.1016/0003-9861(91)90030-M.

Kannel, WB. (2000a). Elevated systolic blood pressure as a cardiovascular risk factor. Am J Cardiol 85: 251-255. 

http://dx.doi.org/10.1016/S0002-9149(99)00635-9.

Kannel, WB. (2000b). Risk stratification in hypertension: New insights from the Framingham Study. Am J 

Hypertens 13: 3S-10S. http://dx.doi.org/10.1016/S0895-7061(99)00252-6.

Kapuku, GK; Harshfield, GA; Davis, HC; Treiber, FA. (2006). Early markers of cardiovascular disease. Vascul 

Pharmacol 45: 277-280. http://dx.doi.org/10.1016/j.vph.2006.08.009.

Karaca, T; Simsek, N. (2007). Effects of spirulina on the number of ovary mast cells in lead-induced toxicity in rats. 

Phytother Res 21: 44-46. http://dx.doi.org/10.1002/ptr.2015.

Karimooy, HN; Mood, MB; Hosseini, M; Shadmanfar, S. (2010). Effects of occupational lead exposure on renal and 

nervous system of workers of traditional tile factories in Mashhad (northeast of Iran). Toxicol Ind Health 

26: 633-638. http://dx.doi.org/10.1177/0748233710377774.

Karita, K; Yano, E; Dakeishi, M; Iwata, T; Murata, K. (2005). Benchmark dose of lead inducing anemia at the 

workplace. Risk Anal 25: 957-962. http://dx.doi.org/10.1111/j.1539-6924.2005.00652.x.

Karmaus, W; Brooks, KR; Nebe, T; Witten, J; Obi-Osius, N; Kruse, H. (2005). Immune function biomarkers in 

children exposed to lead and organochlorine compounds: A cross-sectional study. Environ Health Global 

Access Sci Source 4: 1-10. http://dx.doi.org/10.1186/1476-069X-4-5.

Kasperczyk, A; Kasperczyk, S; Horak, S; Ostalowska, A; Grucka-Mamczar, E; Romuk, E; Olejek, A; Birkner, E.

(2008). Assessment of semen function and lipid peroxidation among lead exposed men. Toxicol Appl 

Pharmacol 228: 378-384. http://dx.doi.org/10.1016/j.taap.2007.12.024.

Kasperczyk, S; Kasperczyk, J; Ostałowska, A; Zalejska-Fiolka, J; Wielkoszyński, T; Swietochowska, E; Birkner, E.

(2009). The role of the antioxidant enzymes in erythrocytes in the development of arterial hypertension 

among humans exposed to lead. Biol Trace Elem Res 130: 95-106. http://dx.doi.org/10.1007/s12011-009-

8323-z.

Kasprzak, KS; Hoover, KL; Poirier, LA. (1985). Effects of dietary calcium acetate on lead subacetate 

carcinogenicity in kidneys of male Sprague- Dawley rats. Carcinogenesis 6: 279-282. 

http://dx.doi.org/10.1093/carcin/6.2.279.

Kasten-Jolly, J; Heo, Y; Lawrence, DA. (2010). Impact of developmental lead exposure on splenic factors. Toxicol 

Appl Pharmacol 247: 105-115. http://dx.doi.org/10.1016/j.taap.2010.06.003.

Kaufman, AS. (2001). Do low levels of lead produce IQ loss in children? A careful examination of the literature. 

Arch Clin Neuropsychol 16: 303-341. 
February 2012 5-580 Draft – Do Not Cite or Quote

Kazi, TG; Jalbani, N; Kazi, N; Jamali, MK; Arain, MB; Afridi, HI; Kandhro, A; Pirzado, Z. (2008). Evaluation of 

toxic metals in blood and urine samples of chronic renal failure patients, before and after dialysis. Ren Fail 

30: 737-745. http://dx.doi.org/10.1080/08860220802212999.

Ke, H. (2004). Implications of PDE4 structure on inhibitor selectivity across PDE families. Int J Impot Res 16: S24-

S27. http://dx.doi.org/10.1038/sj.ijir.3901211.

Kemp, FW; Neti, P; Howell, RW; Wenger, P; Louria, DB; Bogden, JD. (2007). Elevated blood lead concentrations 

and vitamin D deficiency in winter and summer in young urban children. Environ Health Perspect 115: 

630-635. http://dx.doi.org/10.1289/ehp.9389.

Kempe, DA; Lang, PA; Eisele, K; Klarl, BA; Wieder, T; Huber, SM; Duranton, C; Lang, F. (2005). Stimulation of 

erythrocyte phosphatidylserine exposure by lead ions. Am J Physiol 288: C396-C402. 

http://dx.doi.org/10.1152/ajpcell.00115.2004.

Kempermann, G; Krebs, J; Fabel, K. (2008). The contribution of failing adult hippocampal neurogenesis to 

psychiatric disorders. Curr Opin Psychiatry 21: 290-295. 

http://dx.doi.org/10.1097/YCO.0b013e3282fad375.

Kerkvliet, NI; Baecher-Steppan, L. (1982). Immunotoxicology studies on lead: Effects of exposure on tumor growth

and cell-mediated immunity after syngeneic or allogeneic stimulation. Int Immunopharmacol 4: 213-224. 

http://dx.doi.org/10.1016/0162-3109(82)90003-0.

Kermani, S; Karbalaie, K; Madani, SH; Jahangirnejad, AA; Eslaminejad, MB; Nasr-Esfahani, MH; Baharvand, H.

(2008). Effect of lead on proliferation and neural differentiation of mouse bone marrow-mesenchymal stem 

cells. Toxicol In Vitro 22: 995-1001. http://dx.doi.org/10.1016/j.tiv.2008.02.009.

Kern, M; Wisniewski, M; Cabell, L; Audesirk, G. (2000). Inorganic lead and calcium interact positively in 

activation of calmodulin. Neurotoxicology 21: 353-363. 

Kern, M; Audesirk, G. (2000). Stimulatory and inhibitory effects of inorganic lead on calcineurin. Toxicology 150: 

171-178. http://dx.doi.org/10.1016/S0300-483X(00)00258-4.

Khaĭrullina, AY; Ol’shanskaya, TV; Filimonenko, DS; Yasinski, VM; Slobozhanina, EI; Kozlova, NM. (2008). 

Study of optical and nanostructural metal-induced changes in erythrocyte membranes by scattering and 

atomic force microscopy. Optics and Spectroscopy 105: 154-160. 

http://dx.doi.org/10.1134/S0030400X08070242.

Khalil-Manesh, F; Gonick, HC; Cohen, AH; Alinovi, R; Bergamaschi, E; Mutti, A; Rosen, VJ. (1992a). 

Experimental model of lead nephropathy. I. Continuous high-dose lead administration. Kidney Int 41: 

1192-1203. http://dx.doi.org/10.1038/ki.1992.181.

Khalil-Manesh, F; Gonick, HC; Cohen, A; Bergamaschi, E; Mutti, A. (1992b). Experimental model of lead 

nephropathy: II. Effect of removal from lead exposure and chelation treatment with dimercaptosuccinic 

acid (DMSA). Environ Res 58: 35-54. http://dx.doi.org/10.1016/S0013-9351(05)80203-8.

Khalil-Manesh, F; Gonick, HC; Cohen, AH. (1993a). Experimental model of lead nephropathy III Continuous lowlevel lead administration. Arch Environ Occup Health 48: 271-278. 

http://dx.doi.org/10.1080/00039896.1993.9940372.

Khalil-Manesh, F; Gonick, HC; Weiler, EWJ; Prins, B; Weber, MA; Purdy, RE. (1993b). Lead-induced 

hypertension: Possible role of endothelial factors. Am J Hypertens 6: 723-729. 

Khalil-Manesh, F; Gonick, HC; Weiler, EWJ; Prins, B; Weber, MA; Purdy, R; Ren, Q. (1994). Effect of chelation 

treatment with dimercaptosuccinic acid (DMSA) on lead-related blood pressure changes. Environ Res 65: 

86-99. http://dx.doi.org/10.1006/enrs.1994.1023.

Khalil, N; Cauley, JA; Wilson, JW; Talbott, EO; Morrow, L; Hochberg, MC; Hillier, TA; Muldoon, SB; Cummings, 

SR. (2008). Relationship of blood lead levels to incident nonspine fractures and falls in older women: The 

study of osteoporotic fractures. J Bone Miner Res 23: 1417-1425. http://dx.doi.org/10.1359/jbmr.080404.

Khalil, N; Wilson, JW; Talbott, EO; Morrow, LA; Hochberg, MC; Hillier, TA; Muldoon, SB; Cummings, SR; 

Cauley, JA. (2009a). Association of blood lead concentrations with mortality in older women: a prospective 

cohort study. Environ Health Global Access Sci Source 8: 15. http://dx.doi.org/10.1186/1476-069x-8-15.
February 2012 5-581 Draft – Do Not Cite or Quote

Khalil, N; Morrow, LA; Needleman, H; Talbott, EO; Wilson, JW; Cauley, JA. (2009b). Association of cumulative 

lead and neurocognitive function in an occupational cohort. Neuropsychology 23: 10-19. 

http://dx.doi.org/10.1037/a0013757.

Khalil, N. (2010). Re: Blood lead mortality study: Khalil et al., 2009. Khalil, N. 

http://www.regulations.gov/#!documentDetail;D=EPA-HQ-ORD-2011-0051-0006.

Khan, D; Qayyum, S; Saleem, S; Ansari, W; Khan, F. (2010a). Lead exposure and its adverse health effects among 

occupational worker’s children. Toxicol Ind Health 26: 497. http://dx.doi.org/10.1177/0748233710373085.

Khan, DA; Qayyum, S; Saleem, S; Khan, FA. (2008). Lead-induced oxidative stress adversely affects health of the 

occupational workers. Toxicol Ind Health 24: 611-618. http://dx.doi.org/10.1177/0748233708098127.

Khan, MI; Ahmad, I; Mahdi, AA; Akhtar, MJ; Islam, N; Ashquin, M; Venkatesh, T. (2010b). Elevated blood lead 

levels and cytogenetic markers in buccal epithelial cells of painters in India: Genotoxicity in painters 

exposed to lead containing paints. Environ Sci Pollut Res Int 17: 1347-1354. 

http://dx.doi.org/10.1007/s11356-010-0319-x.

Khan, MI; Islam, N; Sahasrabuddhe, AA; Mahdi, AA; Siddiqui, H; Ashquin, M; Ahmad, I. (2011). Ubiquitous 

hazardous metal lead induces TNF-α in human phagocytic THP-1 cells: Primary role of ERK 1/2. J Hazard 

Mater 189: 255-264. http://dx.doi.org/10.1016/j.jhazmat.2011.02.027.

Kharoubi, O; Slimani, M; Aoues, A; Seddik, L. (2008a). Prophylactic effects of Wormwood on lipid peroxidation in 

an animal model of lead intoxication. Indian J Nephrol 18: 51-57. http://dx.doi.org/10.4103/0971-

4065.42333.

Kharoubi, O; Slimani, M; Krouf, D; Seddik, L; Aoues, A. (2008b). Role of wormwood (Artemisia absinthium) 

extract on oxidative stress in ameliorating lead induced haematotoxicity. African Journal of Traditional, 

Complementary and Alternative Medicines 5: 263-270. 

Khorasani, G; Shokrzade, M; Salehifar, E; Asadi, M; Shabankhani, B; Rezaeinejad, S. (2008). The comparison of 

lead and zinc plasma levels in gastric cancer patients with healthy volunteers. Res J Biol Sci 3: 631-634. 

http://dx.doi.org/10.3923/rjbsci.2008.631.634.

Khotimchenko, MY; Kolenchenko, EA. (2007). Efficiency of low-esterified pectin in toxic damage to the liver 

inflicted by lead treatment. Bull Exp Biol Med 144: 60-62. http://dx.doi.org/10.1007/s10517-007-0254-0.

Kibayashi, K; Nakao, KI; Taki, T; Koyama, H. (2010). Changes in the brain after intracerebral implantation of metal 

lead in a rat model. J Neurotrauma 27: 1925-1934. http://dx.doi.org/10.1089/neu.2010.1379.

Kim, D; Lawrence, DA. (2000). Immunotoxic effects of inorganic lead on host resistance of mice with different 

circling behavior preferences. Brain Behav Immun 14: 305-317. http://dx.doi.org/10.1006/brbi.2000.0609.

Kim, HK; Yoon, EK; Jang, J; Hwang, M; Kim, J; Ha, JH; Jang, DD; Yoo, TM; Park, KL. (2009a). Assessment of 

heavy metal exposure via the intake of oriental medicines in Korea. J Toxicol Environ Health A 72: 1336-

1342. http://dx.doi.org/10.1080/15287390903212485.

Kim, HS; Lee, SS; Lee, GS; Hwangbo, Y; Ahn, KD; Lee, BK. (2004). The protective effect of delta-aminolevulinic 

acid dehydratase 1-2 and 2-2 isozymes against blood lead with higher hematologic parameters. Environ 

Health Perspect 112: 538-541. http://dx.doi.org/10.1289/ehp.6464.

Kim, JH; Lee, KH; Yoo, DH; Kang, D; Cho, SH; Hong, YC. (2007). GSTM1 and TNF-alpha gene polymorphisms 

and relations between blood lead and inflammatory markers in a non-occupational population. Mutat Res 

Genet Toxicol Environ Mutagen 629: 32-39. http://dx.doi.org/10.1016/j.mrgentox.2007.01.004.

Kim, R; Rotnitsky, A; Sparrow, D; Weiss, ST; Wager, C; Hu, H. (1996). A longitudinal study of low-level lead 

exposure and impairment of renal function The Normative Aging Study. JAMA 275: 1177-1181. 

http://dx.doi.org/10.1001/jama.1996.03530390043032.

Kim, Y; Kim, BN; Hong, YC; Shin, MS; Yoo, HJ; Kim, JW; Bhang, SY; Cho, SC. (2009b). Co-exposure to 

environmental lead and manganese affects the intelligence of school-aged children. Neurotoxicology 30: 

564-571. http://dx.doi.org/10.1016/j.neuro.2009.03.012.
February 2012 5-582 Draft – Do Not Cite or Quote

Kimber, I; Stonard, MD; Gidlow, DA; Niewola, Z. (1986). Influence of chronic low-level exposure to lead on 

plasma immunoglobulin concentration and cellular immune function in man. Int Arch Occup Environ 

Health 57: 117-125. http://dx.doi.org/10.1007/BF00381379.

Király, E; Jones, DG. (1982). Dendritic spine changes in rat hippocampal pyramidal cells after postnatal lead 

treatment: A golgi study. Exp Neurol 77: 236-239. http://dx.doi.org/10.1016/0014-4886(82)90158-3.

Kiran Kumar, B; Prabhakara Rao, Y; Noble, T; Weddington, K; McDowell, VP; Rajanna, S; Bettaiya, R. (2009). 

Lead-induced alteration of apoptotic proteins in different regions of adult rat brain. Toxicol Lett 184: 56-

60. http://dx.doi.org/10.1016/j.toxlet.2008.10.023.

Kirberger, M; Yang, JJ. (2008). Structural differences between Pb2+- and Ca2+-binding sites in proteins: 

Implications with respect to toxicity. J Inorg Biochem 102: 1901-1909. 

http://dx.doi.org/10.1016/j.jinorgbio.2008.06.014.

Kishi, R; Ikeda, T; Miyake, H; Uchino, E; Tsuzuki, T; Inoue, K. (1983). Effects of low lead exposure on neurobehavioral function in the rat. Arch Environ Occup Health 38: 25-33. 

Kishimoto, T; Oguri, T; Ueda, D; Tada, M. (1995). Effect of lead on tube formation by cultured human vascular 

endothelial cells. Arch Toxicol 69: 718-721. http://dx.doi.org/10.1007/s002040050238.

Kiziler, AR; Aydemir, B; Onaran, I; Alici, B; Ozkara, H; Gulyasar, T; Akyolcu, MC. (2007). High levels of 

cadmium and lead in seminal fluid and blood of smoking men are associated with high oxidative stress and 

damage in infertile subjects. Biol Trace Elem Res 120: 82-91. http://dx.doi.org/10.1007/s12011-007-8020-

8.

Klann, E; Shelton, KR. (1989). The effect of lead on the metabolism of a nuclear matrix protein which becomes 

prominent in lead-induced intranuclear inclusion bodies. J Biol Chem 264: 16969-16972. 

Klein, D; Y-JY, W; Kamyab, S; Okuda, H; Sokol, RZ. (1994). Effects of toxic levels of lead on gene regulation in 

the male axis: increase in messenger ribonucleic acids and intracellular stores of gonadotrophs within the 

central nervous system. Biol Reprod 50: 802-811. http://dx.doi.org/10.1095/biolreprod50.4.802.

Knowles, SO; Donaldson, WE. (1990). Dietary modification of lead toxicity: Effects on fatty acid and eicosanoid 

metabolism in chicks. Comp Biochem Physiol C Toxicol Pharmacol 95: 99-104. 

http://dx.doi.org/10.1016/0742-8413(90)90088-Q.

Knowles, SO; Donaldson, WE. (1997). Lead disrupts eicosanoid metabolism, macrophage function, and disease 

resistance in birds. Biol Trace Elem Res 60: 13-26. http://dx.doi.org/10.1007/BF02783306.

Kojima, M; Sekikawa, K; Nemoto, K; Degawa, M. (2005). Tumor necrosis factor-alpha-independent 

downregulation of hepatic cholesterol 7 alpha-hydroxylase gene in mice treated with lead nitrate. Toxicol 

Sci 87: 537-542. http://dx.doi.org/10.1093/toxsci/kfi267.

Kojima, M; Degawa, M. (2006). Gender-related difference in altered gene expression of a sterol regulatory element 

binding protein, SREBP-2, by lead nitrate in rats: Correlation with development of hypercholesterolemia. J 

Appl Toxicol 26: 381-384. http://dx.doi.org/10.1002/jat.1138.

Koka, S; Huber, SM; Boini, KM; Lang, C; Foller, M; Lang, F. (2007). Lead decreases parasitemia and enhances 

survival of Plasmodium berghei-infected mice. Biochem Biophys Res Commun 363: 484-489. 

http://dx.doi.org/10.1016/j.bbrc.2007.08.173.

Kolesarova, A; Roychoudhury, S; Slivkova, J; Sirotkin, A; Capcarova, M; Massanyi, P. (2010). In vitro study on the 

effects of lead and mercury on porcine ovarian granulosa cells. J Environ Sci Health A Tox Hazard Subst 

Environ Eng 45: 320-331. http://dx.doi.org/10.1080/10934520903467907.

Koller, LD; Kerkvliet, NI; Exon, JH. (1985). Neoplasia induced in male rats fed lead acetate, ethyl urea, and sodium 

nitrite. Toxicol Pathol 13: 50-57. http://dx.doi.org/10.1177/019262338501300107.

Korashy, HM; El-Kadi, AOS. (2004). Differential effects of mercury, lead and copper on the constitutive and 

inducible expression of aryl hydrocarbon receptor (AHR)-regulated genes in cultured hepatoma Hepa 

1c1c7 cells. Toxicology 201: 153-172. http://dx.doi.org/10.1016/j.tox.2004.04.011.
February 2012 5-583 Draft – Do Not Cite or Quote

Korashy, HM; El-Kadi, AOS. (2008). NF-kappa B and AP-1 are key signaling pathways in the modulation of 

NAD(P)H: Quinone oxidoreductase 1 gene by mercury, lead, and copper. J Biochem Mol Toxicol 22: 274-

283. http://dx.doi.org/10.1002/jbt.20238.

Korashy, HM; Ei-Kadi, AOS. (2008). The role of redox-sensitive transcription factors NF-kappa B and AP-1 in the 

modulation of the Cyp1a1 gene by mercury, lead, and copper. Free Radic Biol Med 44: 795-806. 

http://dx.doi.org/10.1016/j.freeradbiomed.2007.11.003.

Kordas, K; Lopez, P; Rosado, JL; Vargas, GG; Rico, JA; Ronquillo, D; Cebrian, ME; Stoltzfus, RJ. (2004). Blood 

lead, anemia, and short stature are independently associated with cognitive performance in Mexican school 

children. J Nutr 134: 363-371. 

Kordas, K; Canfield, RL; Lopez, P; Rosado, JL; Vargas, GG; Cebrian, ME; Rico, JA; Ronquillo, D; Stoltzfus, RJ.

(2006). Deficits in cognitive function and achievement in Mexican first-graders with low blood lead 

concentrations. Environ Res 100: 371-386. http://dx.doi.org/10.1016/j.envres.2005.07.007.

Kordas, K; Casavantes, KA; Mendoza, C; Lopez, P; Ronquillo, D; Rosado, JL; Vargas, GG; Stoltzfus, RJ. (2007). 

The association between lead and micronutrient status, and children's sleep, classroom behavior, and 

activity. Arch Environ Occup Health 62: 105-112. http://dx.doi.org/10.3200/AEOH.62.2.105-112.

Kordas, K; Ettinger, AS; Lamadrid-Figueroa, H; Tellez-Rojo, MM; Hernandez-Avila, M; Hu, H; Wright, RO.

(2009). Methylenetetrahydrofolate reductase (MTHFR) C677T, A1298C and G1793A genotypes, and the 

relationship between maternal folate intake, tibia lead and infant size at birth. Br J Nutr 102: 907-914. 

http://dx.doi.org/10.1017/s0007114509318280.

Kordas, K; Ettinger, AS; Bellinger, DC; Schnaas, L; Téllez Rojo, MM; Hernández-Avila, M; Hu, H; Wright, RO.

(2011). A dopamine receptor (DRD2) but not dopamine transporter (DAT1) gene polymorphism is 

associated with neurocognitive development of Mexican preschool children with lead exposure. J Pediatr 

159: 638-643. http://dx.doi.org/10.1016/j.jpeds.2011.03.043.

Kottgen, A; Selvin, E; Stevens, LA; Levey, AS; Van Lente, F; Coresh, J. (2008). Serum cystatin C in the United 

States: The Third National Health and Nutrition Examination Survey (NHANES III). Am J Kidney Dis 51: 

385-394. http://dx.doi.org/10.1053/j.ajkd.2007.11.019.

Krieg, EF, Jr; Chrislip, DW; Crespo, CJ; Brightwell, WS; Ehrenberg, RL; Otto, DA. (2005). The relationship 

between blood lead levels and neurobehavioral test performance in NHANES III and related occupational 

studies. Public Health Rep 120: 240-251. 

Krieg, EF, Jr. (2007). The relationships between blood lead levels and serum follicle stimulating hormone and 

luteinizing hormone in the third national health and nutrition examination survey. Environ Res 104: 374-

382. http://dx.doi.org/10.1016/j.envres.2006.09.009.

Krieg, EF, Jr; Butler, MA. (2009). Blood lead, serum homocysteine, and neurobehavioral test performance in the 

third National Health and Nutrition Examination Survey. Neurotoxicology 30: 281-289. 

http://dx.doi.org/10.1016/j.neuro.2008.12.014.

Krieg, EF, Jr; Butler, MA; Chang, MH; Liu, TB; Yesupriya, A; Lindegren, ML; Dowling, N. (2009). Lead and 

cognitive function in ALAD genotypes in the Third National Health and Nutrition Examination Survey. 

Neurotoxicol Teratol 31: 364-371. http://dx.doi.org/10.1016/j.ntt.2009.08.003.

Krieg, EF, Jr; Butler, MA; M-h, C; Liu, T; Yesupriya, A; Dowling, N; Lindegren, ML. (2010). Lead and cognitive 

function in VDR genotypes in the Third National Health and Nutrition Examination Survey. Neurotoxicol 

Teratol 32: 262-272. http://dx.doi.org/10.1016/j.ntt.2009.12.004.

Krishnamoorthy, MS; Parthiban, N; Muthu, P; Paul, V; Balagopal, G; Kumaravel, TS. (1993). Effect of acute 

pretreatment of lead on picrotoxin-induced convulsions in rats. J Appl Toxicol 13: 155-159. 

Kristal-Boneh, E; Coller, D; Froom, P; Harari, G; Ribak, J. (1999). The association between occupational lead 

exposure and serum cholesterol and lipoprotein levels. Am J Public Health 89: 1083-1087. 

Kuhlmann, AC; McGlothan, JL; Guilarte, TR. (1997). Developmental lead exposure causes spatial learning deficits 

in adult rats. Neurosci Lett 233: 101-104. http://dx.doi.org/10.1016/S0304-3940(97)00633-2.
February 2012 5-584 Draft – Do Not Cite or Quote

Kuo, CY; Wong, RH; Lin, JY; Lai, JC; Lee, H. (2006). Accumulation of chromium and nickel metals in lung tumors 

from lung cancer patients in Taiwan. J Toxicol Environ Health A 69: 1337-1344. 

http://dx.doi.org/10.1080/15287390500360398.

Kuruvilla, A; Pillay, VV; Adhikari, P; Venkatesh, T; Chakrapani, M; Rao, HTJ; Bastia, BK; Rajeev, A; Saralaya, 

KM; Rai, M. (2006). Clinical manifestations of lead workers of Mangalore, India. Toxicol Ind Health 22: 

405-413. http://dx.doi.org/10.1177/0748233706074174.

Laden, F; Neas, LM; Dockery, DW; Schwartz, J. (2000). Association of fine particulate matter from different 

sources with daily mortality in six US cities. Environ Health Perspect 108: 941-947. 

http://dx.doi.org/10.1289/ehp.00108941.

Lai, CC; Lin, HH; Chen, CW; Chen, SH; Chiu, TH. (2002). Excitatory action of lead on rat sympathetic 

preganglionic neurons in vitro and in vivo. Life Sci 71: 1035-1045. http://dx.doi.org/10.1016/S0024-

3205(02)01789-7.

Lai, LH; Chou, SY; Wu, FY; Chen, JJH; Kuo, HW. (2008a). Renal dysfunction and hyperuricemia with low blood 

lead levels and ethnicity in community-based study. Sci Total Environ 401: 39-43. 

http://dx.doi.org/10.1016/j.scitotenv.2008.04.004.

Lal, B; Murthy, RC; Anand, M; Chandra, SV; Kumar, R; Tripathi, O; Srimal, RC. (1991). Cardiotoxicity and 

hypertension in rats after oral lead exposure. Drug Chem Toxicol 14: 305-318. 

http://dx.doi.org/10.3109/01480549109002192.

Lamadrid-Figueroa, H; Téllez-Rojo, MM; Hernández-Avila, M; Trejo-Valdivia, B; Solano-González, M; MercadoGarcia, A; Smith, D; Hu, H; Wright, RO. (2007). Association between the plasma/whole blood lead ratio 

and history of spontaneous abortion: A nested cross-sectional study. BMC Pregnancy Childbirth 7: 22. 

http://dx.doi.org/10.1186/1471-2393-7-22.

Lamb, MR; Janevic, T; Liu, X; Cooper, T; Kline, J; Factor-Litvak, P. (2008). Environmental lead exposure, 

maternal thyroid function, and childhood growth. Environ Res 106: 195-202. 

http://dx.doi.org/10.1016/j.envres.2007.09.012.

Landrigan, PJ; Suk, WA; Amler, RW. (1999). Chemical wastes, children's health, and the Superfund Basic Research 

Program. Environ Health Perspect 107: 423-427. 

Lang, F; Gulbins, E; Lerche, H; Huber, SM; Kempe, DS; Foller, M. (2008). Eryptosis, a window to systemic 

disease. Cell Physiol Biochem 22: 373-380. 

Lanphear, BP; Dietrich, K; Auinger, P; Cox, C. (2000). Cognitive deficits associated with blood lead concentrations 

<10 microg/dL in US children and adolescents. Public Health Rep 115: 521-529. 

Lanphear, BP; Hornung, R; Khoury, J; Yolton, K; Baghurst, P; Bellinger, DC; Canfield, RL; Dietrich, KN; 

Bornschein, R; Greene, T; Rothenberg, SJ; Needleman, HL; Schnaas, L; Wasserman, G; Graziano, J; 

Roberts, R. (2005). Low-level environmental lead exposure and children's intellectual function: An 

international pooled analysis. Environ Health Perspect 113: 894-899. 

Lara-Tejero, M; Pamer, EG. (2004). T cell responses to Listeria monocytogenes. Curr Opin Microbiol 7: 45-50. 

http://dx.doi.org/10.1016/j.mib.2003.12.002.

Laschi-Loquerie, A; Decotes, J; Tachon, P; Evreux, JC. (1984). Influence of lead acetate on hypersensitivity: 

Experimental study. J Immunopharm 6: 87-93. http://dx.doi.org/10.3109/08923978409026461.

Lasley, SM; Gilbert, ME. (2002). Rat hippocampal glutamate and GABA release exhibit biphasic effects as a 

function of chronic lead exposure level. Toxicol Sci 66: 139-147. http://dx.doi.org/10.1093/toxsci/66.1.139.

Laties, AM; Zrenner, E. (2002). Viagra® (sildenafil citrate) and ophthalmology. Prog Retin Eye Res 21: 485-506. 

http://dx.doi.org/10.1016/s1350-9462(02)00013-7.

Lau, YS; Camoratto, AM; White, LM; Moriarty, CM. (1991). Effect of lead on TRH and GRF binding in rat anterior 

pituitary membranes. Toxicology 68: 169-179. http://dx.doi.org/10.1016/0300-483X(91)90019-W.

Laughlin, NK; Luck, ML; Lasky, RE. (2008). Postnatal lead effects on the development of visual spatial acuity in 

rhesus monkeys (Macaca mulatta). Dev Psychobiol 50: 608-614. http://dx.doi.org/10.1002/dev.20315.
February 2012 5-585 Draft – Do Not Cite or Quote

Laughlin, NK; Luck, ML; Lasky, RE. (2009). Early lead exposure effects on an auditory threshold task in the rhesus 

monkey (Macaca mulatta). Dev Psychobiol 51: 289-300. http://dx.doi.org/10.1002/dev.20364.

Laviola, G; Rea, M; Moreley-Fletcher, S; Di Carlo, S; Bacosi, A; De Simone, R; Bertini, M; Pacifici, R. (2004). 

Beneficial effects of enriched environment on adolescent rats from stressed pregnancies. Eur J Neurosci 20: 

1655-1664. http://dx.doi.org/10.1111/j.1460-9568.2004.03597.x.

Laviola, G; Hannan, AJ; Marci, S; Solinas, M; Jaber, M. (2008). Effects of enriched environment on animal models 

of neurodegenerative diseases and psychiatric disorders. Neurobiol Dis 31: 159-168. 

http://dx.doi.org/10.1016/j.nbd.2008.05.001.

Lawton, LJ; Donaldson, WE. (1991). Lead-induced tissue fatty acid alterations and lipid peroxidation. Biol Trace 

Elem Res 28: 83-97. http://dx.doi.org/10.1007/BF02863075.

Lazarus, JH. (2005). Thyroid disorders associated with pregnancy: etiology, diagnosis, and management. Treat 

Endocrinol 4: 31-41. 

Leasure, JL; Giddabasappa, A; Chaney, S; Johnson, JE; Pothakos, K; Lau, YS; Fox, DA. (2008). Low-level human 

equivalent gestational lead exposure produces sex-specific motor and coordination abnormalities and lateonset obesity in year-old mice. Environ Health Perspect 116: 355-361. http://dx.doi.org/10.1289/ehp.10862.

Ledda-Columbano, GM; Coni, O; Curto, M; Giacomini, O; Faa, G; Sarma, DSR; Columbano, A. (1992). Mitogeninduced liver hyperplasia does not substitute for compensatory regeneration during promotion of chemical 

hepatocarcinogenesis. Carcinogenesis 13: 379-383. http://dx.doi.org/10.1093/carcin/13.3.379.

Lee, BK; Lee, GS; Stewart, WF; Ahn, KD; Simon, D; Kelsey, KT; Todd, AC; Schwartz, BS. (2001a). Associations 

of blood pressure and hypertension with lead dose measures and polymorphisms in the vitamin D receptor 

and delta-aminolevulinic acid dehydratase genes. Environ Health Perspect 109: 383-389. 

Lee, DH; Blomhoff, R; Jacobs, DR, Jr. (2004). Is serum gamma glutamyltransferase a marker of oxidative stress? 

Free Radic Res 38: 535-539. http://dx.doi.org/10.1080/10715760410001694026.

Lee, DH; Lim, JS; Song, K; Boo, Y; Jacobs, DR. (2006a). Graded associations of blood lead and urinary cadmium 

concentrations with oxidative-stress-related markers in the US population: Results from the Third National 

Health and Nutrition Examination Survey. Environ Health Perspect 114: 350-354. 

http://dx.doi.org/10.1289/ehp.8518.

Lee, JE; Chen, S; Golemboski, KA; Parsons, PJ; Dietert, RR. (2001b). Developmental windows of differential leadinduced immunotoxicity in chickens. Toxicology 156: 161-170. http://dx.doi.org/10.1016/S0300-

483X(00)00350-4.

Lee, JJ; Battles, AH. (1994). Lead toxicity via arachidonate signal transduction to growth responses in the splenic 

macrophage. Environ Res 67: 209-219. http://dx.doi.org/10.1006/enrs.1994.1075.

Lee, MK; Cho, SY; Kim, DJ; Jang, JY; Shin, KH; Park, SA; Park, EM; Lee, JS; Choi, MS; Kim, MJ. (2005). Duzhong (Eucommia ulmoides Oliv.) cortex water extract alters heme biosynthesis and erythrocyte 

antioxidant defense system in lead-administered rats. J Med Food 8: 86-92. 

http://dx.doi.org/10.1089/jmf.2005.8.86.

Lee, MY; Shin, JH; Han, HS; Chung, JH. (2006b). In vivo effects of lead on erythrocytes following chronic 

exposure through drinking water. Arch Pharm Res 29: 1158-1163. 

Lee, TH; Tseng, MC; Chen, CJ; Lin, JL. (2009). Association of high body lead store with severe intracranial carotid 

atherosclerosis. Neurotoxicology 30: 876-880. http://dx.doi.org/10.1016/j.neuro.2009.07.004.

Legare, ME; Barhoumi, R; Burghardt, RC; Tiffany-Castiglioni, E. (1993). Low-level lead exposure in cultured 

astroglia: Identification of cellular targets with vital fluorescent probes. Neurotoxicology 14: 267-272. 

Lenroot, RK; Giedd, JN. (2006). Brain development in children and adolescents: insights from anatomical magnetic 

resonance imaging. Neurosci Biobehav Rev 30: 718-729. 

http://dx.doi.org/10.1016/j.neubiorev.2006.06.001.
February 2012 5-586 Draft – Do Not Cite or Quote

Leret, ML; Garcia-Uceda, F; Antonio, MT. (2002). Effects of maternal lead administration on monoaminergic, 

GABAergic and glutamatergic systems. Brain Res Bull 58: 469-473. http://dx.doi.org/10.1016/S0361-

9230(02)00819-5.

Leseney, AM; Dème, D; Legué, O; Ohayon, R; Chanson, P; Sales, JP; Carvalho, DP; Dupuy, C; Virion, A. (1999). 

Biochemical characterization of a Ca2+/NAD(P)H-dependent H2O2 generator in human thyroid tissue. 

Biochimie 81: 373-380. http://dx.doi.org/10.1016/S0300-9084(99)80084-4.

Levey, AS; Bosc, JP; Lewis, JB; Greene, T; Rogers, N; Roth, D. (1999). A more accurate method to estimate 

glomerular filtration rate from serum creatinine: A new prediction equation. Ann Intern Med 130: 461-470. 

Levey, AS; Greene, T; Kusek, JW; Beck, G. (2000). A simplified equation to predict glomerular filtration rate from 

serum creatinine [Abstract]. J Am Soc Nephrol 11: A0828. 

Levey, AS; Stevens, LA; Schmid, CH; Zhang, YP; Castro, AF; Feldman, HI; Kusek, JW; Eggers, P; Van Lente, F; 

Greene, T; Coresh, J. (2009). A new equation to estimate glomerular filtration rate. Ann Intern Med 150: 

604-612. 

Levin, ED; Bowman, RE. (1986). Effects of the dopamine D-2 receptor agonist, LY 171555, on radial arm maze 

performance in rats. Pharmacol Biochem Behav 25: 1117-1119. 

Levin, ED; Bowman, RE; Wegert, S; Vuchetich, J. (1987). Psychopharmacological investigations of a lead-induced 

long-term cognitive deficit in monkeys. Psychopharmacology 91: 334-341. 

Leviton, A; Bellinger, D; Allred, EN; Rabinowitz, M; Needleman, H; Schoenbaum, S. (1993). Pre- and postnatal 

low-level lead exposure and children's dysfunction in school. Environ Res 60: 30-43. 

http://dx.doi.org/10.1006/enrs.1993.1003.

Lewis, M; Pitts, D. (2004). Inorganic lead exposure in the rat activates striatal cFOS expression at lower blood 

levels and inhibits amphetamine-induced cFOS expression at higher blood levels. J Pharmacol Exp Ther 

310: 815-820. http://dx.doi.org/10.1124/jpet.103.063941.

Li, JP; Wang, CY; Tang, YA; Lin, YW; Yang, JL. (2008a). Role of extracellular signal-regulated kinase (ERK) 

signaling in nucleotide excision repair and genotoxicity in response to As(III) and Pb(II). Pure Appl Chem 

80: 2735-2750. http://dx.doi.org/10.1351/pac200880122735.

Li, N; Yu, ZL; Wang, L; Zheng, YT; Jia, JX; Wang, Q; Zhu, MJ; Liu, XH; Xia, X; Li, WJ. (2009c). Early-life lead 

exposure affects the activity of TNF-alpha and expression of SNARE complex in hippocampus of mouse 

pups. Biol Trace Elem Res 132: 227-238. http://dx.doi.org/10.1007/s12011-009-8551-2.

Li, N; Yu, ZL; Wang, L; Zheng, YT; Jia, JX; Wang, Q; Zhu, MJ; Liu, XL; Xia, X; Li, WJ. (2010b). Increased tau 

phosphorylation and beta amyloid in the hipocampus of mouse pups by early life lead exposure. Acta Biol 

Hung 61: 123-134. http://dx.doi.org/10.1556/ABiol.61.2010.2.1.

Li, S; Chen, W; Srinivasan, SR; Bond, MG; Tang, R; Urbina, EM; Berenson, GS. (2003). Childhood cardiovascular 

risk factors and carotid vascular changes in adulthood: The Bogalusa Heart Study. JAMA 290: 2271-2276. 

http://dx.doi.org/10.1001/jama.290.17.2271.

Li, S; Zhao, Z; Zhou, X; Liu, S. (2008c). The effect of lead on intracellular Ca<sup>2+</sup> in mouse 

lymphocytes. Toxicol In Vitro 22: 1815-1819. http://dx.doi.org/10.1016/j.tiv.2008.08.005.

Li, TY; Zhang, X; Wei, XP; Liu, YF; Qu, P; Liu, YX; Chen, J. (2011). [Impact of antioxidant vitamins and heavy 

metal levels at birth on neurodevelopment of children assessed at two years of age]. Zhonghua Er Ke Za 

Zhi 49: 439-444. 

Li, Y; Xu, XJ; Wu, KS; Chen, GJ; Liu, JX; Chen, SJ; Gu, CW; Zhang, B; Zheng, LK; Zheng, MH; Huo, X. (2008d). 

Monitoring of lead load and its effect on neonatal behavioral neurological assessment scores in Guiyu, an 

electronic waste recycling town in China. J Environ Monit 10: 1233-1238. 

http://dx.doi.org/10.1039/b804959a.

Li Chen, T; Wise, SS; Kraus, S; Shaffiey, F; Levine, KM; Thompson, WD; Romano, T; O'Hara, T; Wise, JP, Sr.

(2009). Particulate hexavalent chromium is cytotoxic and genotoxic to the North Atlantic right whale 

(Eubalaena glacialis) lung and skin fibroblasts. Environ Mol Mutagen 50: 387-393. 

http://dx.doi.org/10.1002/em.20471.
February 2012 5-587 Draft – Do Not Cite or Quote

Lidsky, TI; Schneider, JS. (2003). Lead neurotoxicity in children: basic mechanisms and clinical correlates. Brain 

126: 5-19. http://dx.doi.org/10.1093/brain/awg014.

Lidsky, TI; Schneider, JS. (2004). Lead and public health: Review of recent findings, re-evaluation of clinical risks. 

J Environ Monit 6: 36N-42N. http://dx.doi.org/10.1039/B403259b.

Lilienthal, H; Lenaerts, C; Winneke, G; Hennekes, R. (1988). Alteration of the visual evoked potential and the 

electroretinogram in lead-treated monkeys. Neurotoxicol Teratol 10: 417-422. 

http://dx.doi.org/10.1016/0892-0362(88)90002-5.

Lilienthal, H; Kohler, K; Turfeld, M; Winneke, G. (1994). Persistent increases in scotopic b-wave amplitudes after 

lead exposure in monkeys. Exp Eye Res 59: 203-209. http://dx.doi.org/10.1006/exer.1994.1098.

Lilis, R; Fischbein, A; Eisinger, J; Blumberg, WE; Diamond, S; Anderson, HA; Rom, W; Rice, C; Sarkozi, L; Kon, 

S; Selikoff, IJ. (1977). Prevalence of lead disease among secondary lead smelter workers and biological 

indicators of lead exposure. Environ Res 14: 255-285. http://dx.doi.org/10.1016/0013-9351(77)90037-8.

Lim, SY; Doherty, JD; McBride, K; Miller-Ihli, NJ; Carmona, GN; Stark, KD; Salem, N. (2005). Lead exposure and 

(n-3) fatty acid deficiency during rat neonatal development affect subsequent spatial task performance and 

olfactory discrimination. J Nutr 135: 1019-1026. 

Lin-Tan, DT; Lin, JL; Wang, LH; Wang, LM; Huang, LM; Liu, L; Huang, JY; Huang, YL. (2007a). Fasting glucose 

levels in predicting 1-year all-cause mortality in patients who do not have diabetes and are on maintenance 

hemodialysis. J Am Soc Nephrol 18: 2385-2391. http://dx.doi.org/10.1681/Asn.2006121409.

Lin-Tan, DT; Lin, JL; Yen, TH; Chen, KH; Huang, YL. (2007b). Long-term outcome of repeated lead chelation 

therapy in progressive non-diabetic chronic kidney diseases. Nephrol Dial Transplant 22: 2924-2931. 

http://dx.doi.org/10.1093/ndt/gfm342.

Lin, CN; Wang, LH; Shen, KH. (2009). Determining urinary trace elements (Cu, Zn, Pb, As, and Se) in patients 

with bladder cancer. J Clin Lab Anal 23: 192-195. http://dx.doi.org/10.1002/jcla.20318.

Lin, JL; Yu, CC; Lin-Tan, DT; Ho, HH. (2001). Lead chelation therapy and urate excretion in patients with chronic 

renal diseases and gout. Kidney Int 60: 266-271. http://dx.doi.org/10.1046/j.1523-1755.2001.00795.x.

Lin, JL; Lin-Tan, DT; Hsu, KH; Yu, CC. (2003). Environmental lead exposure and progression of chronic renal 

diseases in patients without diabetes. N Engl J Med 348: 277-286. 

http://dx.doi.org/10.1097/01.ASN.0000118529.01681.

Lin, JL; Lin-Tan, DT; Yu, CC; Li, YJ; Huang, YY; Li, KL. (2006a). Environmental exposure to lead and 

progressive diabetic nephropathy in patients with type II diabetes. Kidney Int 69: 2049-2056. 

http://dx.doi.org/10.1038/sj.ki.5001505.

Lin, JL; Lin-Tan, DT; Li, YJ; Chen, KH; Huang, YL. (2006b). Low-level environmental exposure to lead and 

progressive chronic kidney diseases. Am J Med 119: 1-9. http://dx.doi.org/10.1016/j.amjmed.2006.01.005.

Lin, JL; Lin-Tan, DT; Yen, TH; Hsu, CW; Jenq, CC; Chen, KH; Hsu, KH; Huang, YL. (2008). Blood lead levels, 

malnutrition, inflammation, and mortality in patients with diabetes treated by long-term hemodialysis. Am 

J Kidney Dis 51: 107-115. http://dx.doi.org/10.1053/j.ajkd.2007.10.002.

Lin, JL; Lin-Tan, DT; Chen, KH; Hsu, CW; Yen, TH; Huang, WH; Huang, YL. (2010). Blood lead levels 

association with 18-month all-cause mortality in patients with chronic peritoneal dialysis. Nephrol Dial 

Transplant 25: 1627-1633. http://dx.doi.org/10.1093/ndt/gfp663.

Lin, JL; Lin-Tan, DT; Hsu, CW; Yen, TH; Chen, KH; Hsu, HH; Ho, TC; Hsu, KH. (2011). Association of blood 

lead levels with mortality in patients on maintenance hemodialysis. Am J Med 124: 350-358. 

http://dx.doi.org/10.1016/j.amjmed.2010.10.022.

Lin, RH; Lee, CH; Chen, WK; Lin-Shiau, SY. (1994). Studies on cytotoxic and genotoxic effects of cadmium nitrate 

and lead nitrate in Chinese hamster ovary cells. Environ Mol Mutagen 23: 143-149. 

http://dx.doi.org/10.1002/em.2850230212.

Lin, TA; Tai-yi, J. (2007). Benchmark dose approach for renal dysfunction in workers exposed to lead. Environ 

Toxicol 22: 229-233. http://dx.doi.org/10.1002/tox.20260.
February 2012 5-588 Draft – Do Not Cite or Quote

Lind, SE; Park, JS; Drexler, JW. (2009). Pyrithione and 8-hydroxyquinolines transport lead across erythrocyte 

membranes. Transl Res 154: 153-159. http://dx.doi.org/10.1016/j.trsl.2009.06.002.

Lison, D; Raguzzi, F; Lauwerys, R. (1990). Comparison of the effects of auranofin, heavy metals and retinoids on 

protein kinase C in vitro and on a protein kinase C mediated response in macrophages. Basic Clin 

Pharmacol Toxicol 67: 239-242. http://dx.doi.org/10.1111/j.1600-0773.1990.tb00820.x.

Little, BB; Spalding, S; Walsh, B; Keyes, DC; Wainer, J; Pickens, S; Royster, M; Villanacci, J; Gratton, T. (2009). 

Blood lead levels and growth status among African-American and Hispanic children in Dallas, Texas-1980 

and 2002: Dallas Lead Project II. Ann Hum Biol 36: 331-341. 

http://dx.doi.org/10.1080/03014460902806615.

Liu, HT; Niu, RY; Wang, JM; He, Y; Wang, JD. (2008). Changes caused by fluoride and lead in energy metabolic 

enzyme activities in the reproductive system of male offspring rats. Fluoride 41: 184-191. 

Liu, J; Kershaw, WC; Klaassen, CD. (1991). The protective effect of metallothionein on the toxicity of various 

metals in rat primary hepatocyte culture. Toxicol Appl Pharmacol 107: 27-34. 

http://dx.doi.org/10.1016/0041-008X(91)90327-B.

Liu, J; Han, D; Li, Y; Zheng, L; Gu, C; Piao, Z; Au, WW; Xu, X; Huo, X. (2010b). Lead affects apoptosis and 

related gene XIAP and Smac expression in the hippocampus of developing rats. Neurochem Res 35: 473-

479. http://dx.doi.org/10.1007/s11064-009-0083-9.

Liu, J; Xu, X; Wu, K; Piao, Z; Huang, J; Guo, Y; Li, W; Zhang, Y; Chen, A; Huo, X. (2011a). Association between 

lead exposure from electronic waste recycling and child temperament alterations. Neurotoxicology 32: 458-

464. http://dx.doi.org/10.1016/j.neuro.2011.03.012.

Liu, L; Wong, T; Pozza, M; Lingenhoehl, K; Wang, Y; Sheng, M; Auberson, Y; Wang, Y. (2004). Role of NMDA 

receptor subtypes in governing the direction of hippocampal synaptic plasticity. Science 304: 1021-1024. 

http://dx.doi.org/10.1126/science.1096615.

Liu, MY; Hsieh, WC; Yang, BC. (2000). In vitro aberrant gene expression as the indicator of lead-induced

neurotoxicity in U-373MG cells. Toxicology 147: 59-64. http://dx.doi.org/10.1016/S0300-483X(00)00186-

4.

Llanos, MN; Ronco, AM. (2009). Fetal growth restriction is related to placental levels of cadmium, lead and arsenic 

but not with antioxidant activities. Reprod Toxicol 27: 88-92. 

http://dx.doi.org/10.1016/j.reprotox.2008.11.057.

Lodi, S; Sharma, V; Kansal, L. (2011). The protective effect of Rubia cordifolia against lead nitrate-induced

immune response impairment and kidney oxidative damage. Indian J Pharmacol 43: 441-444. 

http://dx.doi.org/10.4103/0253-7613.83118.

Lohmann, C; Bonhoeffer, T. (2008). A role for local calcium signaling in rapid synaptic partner selection by 

dendritic filopodia. Neuron 59: 253-260. http://dx.doi.org/10.1016/j.neuron.2008.05.025.

Lolin, Y; O'Gorman, P. (1988). An intra-erythrocyctic low molecular weight lead-binding protein in acute and 

chronic lead exposure and its possible protective role in lead toxicity. Ann Clin Biochem 25: 688-697. 

Long, GJ; Rosen, JF; Pounds, JG. (1990). Cellular lead toxicity and metabolism in primary and clonal osteoblastic 

bone cells. Toxicol Appl Pharmacol 102: 346-361. 

Long, GJ; Pounds, JG; Rosen, JF. (1992). Lead intoxication alters basal and parathyroid hormone-regulated cellular 

calcium homeostasis in rat osteosarcoma (ROS 17/28) cells. Calcif Tissue Int 50: 451-458. 

http://dx.doi.org/10.1007/BF00296777.

Long, GJ; Rosen, JF. (1992). Lead perturbs epidermal growth factor (EGF) modulation of intracellular calcium 

metabolism and collagen synthesis in clonal rat osteoblastic (ROS 17/28) cells. Toxicol Appl Pharmacol 

114: 63-70. http://dx.doi.org/10.1016/0041-008X(92)90097-C.

Long, GJ; Rosen, JF; Schanne, FAX. (1994). Lead activation of protein kinase C from rat brain: Determination of 

free calcium, lead, and zinc by 19F NMR. J Biol Chem 269: 834-837. 
February 2012 5-589 Draft – Do Not Cite or Quote

Long, Y, .; Li, Q, .; Li, J, .; Cui, Z, . (2011). Molecular analysis, developmental function and heavy metal-induced 

expression of ABCC5 in zebrafish. Comp Biochem Physiol B Biochem Mol Biol 158: 46-55. 

http://dx.doi.org/10.1016/j.cbpb.2010.09.005.

López-Larrubia, P; Cauli, O. (2011). Alterations of apparent diffusion coefficient (ADC) in the brain of rats 

chronically exposed to lead acetate. Toxicology 281: 1-6. http://dx.doi.org/10.1016/j.tox.2010.12.009.

Lopez, CM; Pineiro, AE; Nunez, N; Avagnina, AM; Villaamil, EC; Roses, OE. (2000). Thyroid hormone changes in 

males exposed to lead in the Buenos Aires area (Argentina). Pharmacol Res 42: 599-602. 

http://dx.doi.org/10.1006/phrs.2000.0734.

Louis, ED; Jurewicz, EC; Applegate, L; Factor-Litvak, P; Parides, M; Andrews, L; Slavkovich, V; Graziano, JH; 

Carroll, S; Todd, A. (2003). Association between essential tremor and blood lead concentration. Environ 

Health Perspect 111: 1707-1711. 

Louis, ED; Applegate, L; Graziano, JH; Parides, M; Slavkovich, V; Bhat, HK. (2005). Interaction between blood 

lead concentration and delta-amino-levulinic acid dehydratase gene polymorphisms increases the odds of 

essential tremor. Mov Disord 20: 1170-1177. http://dx.doi.org/10.1002/mds.20565.

Lu, H; Guizzetti, M; Costa, LG. (2002). Inorganic lead activates the mitogen-activated protein kinase kinasemitogen-activated protein kinase-p90RSK signaling pathway in human astrocytoma cells via a protein 

kinase C-dependent mechanism. J Pharmacol Exp Ther 300: 818-823. 

Lundstrom, NG; Englyst, V; Gerhardsson, L; Jin, T; Nordberg, G. (2006). Lung cancer development in primary 

smelter workers: A nested case-referent study. J Occup Environ Med 48: 376-380. 

http://dx.doi.org/10.1097/01.jom.0000201556.95982.95.

Lurie, DI; Brooks, DM; Gray, LC. (2006). The effect of lead on the avian auditory brainstem. Neurotoxicology 27: 

108-117. http://dx.doi.org/10.1016/j.neuro.2005.07.005.

Lustberg, M; Silbergeld, E. (2002). Blood lead levels and mortality. Arch Intern Med 162: 2443-2449. 

Lutz, PM; Wilson, TJ; Ireland, AL; Gorman, JS; Gale, NL; Johnson, JC; Hewett, JE. (1999). Elevated 

immunoglobulin E (IgE) levels in children with exposure to environmental lead. Toxicology 134: 63-78. 

http://dx.doi.org/10.1016/S0300-483X(99)00036-0.

Lynes, MA; Fontenot, AP; Lawrence, DA; Rosenspire, AJ; Pollard, KM. (2006). Gene expression influences on 

metal immunomodulation. Toxicol Appl Pharmacol 210: 9-16. 

http://dx.doi.org/10.1016/j.taap.2005.04.021.

Ma, T; Chen, H; Ho, I. (1999). Effects of chronic lead (Pb) exposure on neurobehavioral function and dopaminergic 

neurotransmitter receptors in rats. Toxicol Lett 105: 111-121. http://dx.doi.org/10.1016/S0378-

4274(98)00388-9.

MacDonald, JF; Jackson, MF; Beazely, MA. (2006). Hippocampal long-term synaptic plasticity and signal 

amplification of NMDA receptors. Crit Rev Neurobiol 18: 71-84. 

Machida, M; Sun, SJ; Oguma, E; Kayama, F. (2009). High bone matrix turnover predicts blood levels of lead among 

perimenopausal women. Environ Res 109: 880-886. http://dx.doi.org/10.1016/j.envres.2009.06.005.

Maciejczyk, P; Chen, LC. (2005). Effects of subchronic exposures to concentrated ambient particles (CAPs) in 

mice: VIII. Source-related daily variations in in vitro responses to CAPs. Inhal Toxicol 17: 243-253. 

http://dx.doi.org/10.1080/08958370590912914.

Madhavi, D; Devi, KR; Rao, KK; Reddy, PP. (2007). Modulating effect of Phyllanthus fruit extract against lead 

genotoxicity in germ cells of mice. J Environ Biol 28: 115-117. 

Maeda, K; Sugino, H; Hirose, T; Kitagawa, H; Nagai, T; Mizoguchi, H; Takuma, K; Yamada, K. (2007). Clozapine 

prevents a decrease in neurogenesis in mice repeatedly treated with phencyclidine. J Pharmacol Sci 103: 

299-308. http://dx.doi.org/10.1254/jphs.FP0061424.

Maes, M. (2011). Depression is an inflammatory disease, but cell-mediated immune activation is the key component 

of depression. Prog Neuropsychopharmacol Biol Psychiatry 35: 664-675. 

http://dx.doi.org/10.1016/j.pnpbp.2010.06.014.
February 2012 5-590 Draft – Do Not Cite or Quote

Magalhaes, C; Socodato, R; Paes-de-Carvalho, R. (2006). Nitric oxide regulates the proliferation of chick embryo 

retina cells by a cyclic GMP-independent mechanism. Int J Dev Neurosci 24: 53-60. 

http://dx.doi.org/10.1016/j.ijdevneu.2005.10.004.

Magyar, JS; Weng, TC; Stern, CM; Dye, DF; Rous, BW; Payne, JC; Bridgewater, BM; Mijovilovich, A; Parkin, G; 

Zaleski, JM; Penner-Hahn, JE; Godwin, HA. (2005). Reexamination of lead(II) coordination preferences in 

sulfur-rich sites: Implications for a critical mechanism of lead poisoning. J Am Chem Soc 127: 9495-9505. 

http://dx.doi.org/10.1021/ja0424530.

Mahaffey, KR; Fowler, BA. (1977). Effects of concurrent administration of lead, cadmium, and arsenic in the rat. 

Environ Health Perspect 19: 165-171. http://dx.doi.org/10.1289/ehp.7719165.

Mahaffey, KR; Capar, SG; Gladen, BC; Fowler, BA. (1981). Concurrent exposure to lead, cadmium, and arsenic. 

Effects on toxicity and tissue metal concentrations in the rat. Transl Res 98: 463-481. 

Mahram, M; Mousavinasab, N; Dinmohammadi, H; Soroush, S; Sarkhosh, F. (2007). Effect of living in lead mining 

area on growth. Indian J Pediatr 74: 555-559. http://dx.doi.org/10.1007/s12098-007-0107-x.

Maitani, T; Watahiki, A; Suzuki, KT. (1986). Induction of metallothionein after lead administration by three 

injection routes in mice. Toxicol Appl Pharmacol 83: 211-217. http://dx.doi.org/10.1016/0041-

008X(86)90298-X.

Maizlish, NA; Parra, G; Feo, O. (1995). Neurobehavioural evaluation of Venezuelan workers exposed to inorganic 

lead. Occup Environ Med 52: 408-414. http://dx.doi.org/10.1136/oem.52.6.408.

Malveaux, FJ; Conroy, MC; Adkinson, NF; Lichtenstein, LM. (1978). IgE receptors on human basophils. 

Relationship to serum IgE concentration. J Clin Invest 62: 176-181. http://dx.doi.org/10.1172/JCI109103.

Malvezzi, CK; Moreira, EG; Vassilieff, I; Vassilieff, VS; Cordellini, S. (2001). Effect of L-arginine, 

dimercaptosuccinic acid (DMSA) and the association of L-arginine and DMSA on tissue lead mobilization 

and blood pressure level in plumbism. Braz J Med Biol Res 34: 1341-1346. 

http://dx.doi.org/10.1590/S0100-879X2001001000016.

Marchlewicz, M; Baranowska-Bosiacka, I; Kolasa, A; Kondarewicz, A; Chlubek, D; Wiszniewska, B. (2009). 

Disturbances of energetic metabolism in rat epididymal epithelial cells as a consequence of chronic lead 

intoxication. Biometals 22: 877-887. http://dx.doi.org/10.1007/s10534-009-9238-z.

Marcus, DK; Fulton, JJ; Clarke, EJ. (2010). Lead and conduct problems: A meta-analysis. J Clin Child Adolesc 

Psychol 39: 234-241. http://dx.doi.org/10.1080/15374411003591455.

Maric, C; Hall, JE. (2011). Obesity, metabolic syndrome and diabetic nephropathy. Contrib Nephrol 170: 28-35. 

http://dx.doi.org/10.1159/000324941.

Markovac, J; Goldstein, GW. (1988a). Lead activates protein kinase C in immature rat brain microvessels. Toxicol 

Appl Pharmacol 96: 14-23. http://dx.doi.org/10.1016/0041-008X(88)90242-6.

Markovac, J; Goldstein, GW. (1988b). Picomolar concentrations of lead stimulate brain protein kinase C. Nature 

334: 71-73. http://dx.doi.org/10.1038/334071a0.

Marques, CC; Nunes, AC; Pinheiro, T; Lopes, PA; Santos, MC; Viegas-Crespo, AM; Ramalhinho, MG; Mathias, 

ML. (2006). An assessment of time-dependent effects of lead exposure in algerian mice (Mus spretus) 

using different methodological approaches. Biol Trace Elem Res 109: 75-90. 

http://dx.doi.org/10.1385/BTER:109:1:075.

Marques, M; Millas, I; Jimenez, A; Garcia-Colis, E; Rodriguez-Feo, JA; Velasco, S; Barrientos, A; Casado, S; 

Lopez-Farre, A. (2001). Alteration of the soluble guanylate cyclase system in the vascular wall of leadinduced hypertension in rats. J Am Soc Nephrol 12: 2594-2600. 

Martin, D; Glass, TA; Bandeen-Roche, K; Todd, AC; Shi, WP; Schwartz, BS. (2006). Association of blood lead and 

tibia lead with blood pressure and hypertension in a community sample of older adults. Am J Epidemiol 

163: 467-478. http://dx.doi.org/10.1093/aje/kwj060.
February 2012 5-591 Draft – Do Not Cite or Quote

Martinez, T; Lartigue, J; Juárez, F; Avila-perez, P; Márquez, C; Zarazua, G; Tejeda, S. (2007). 206Pb/207Pb ratios 

in dry deposit samples from the Metropolitan Zone of Mexico Valley. Journal of Radioanal Chem 273: 

577-582. http://dx.doi.org/10.1007/s10967-007-0913-7.

Mas-Oliva, J. (1989). Effect of lead on the erythrocyte (Ca2+,Mg2+)-ATPase activity: Calmodulin involvement. 

Mol Cell Biochem 89: 87-93. http://dx.doi.org/10.1007/BF00228283.

Massadeh, AM; Al-Safi, SA; Momani, IF; Al-Mahmoud, M; Alkofahi, AS. (2007). 115. Biol Trace Elem Res 115: 

157-167. http://dx.doi.org/10.1007/BF02686027.

Massanyi, P; Lukac, N; Makarevich, AV; Chrenek, P; Forgacs, Z; Zakrzewski, M; Stawarz, R; Toman, R; Lazor, P; 

Flesarova, S. (2007). Lead-induced alterations in rat kidneys and testes in vivo. J Environ Sci Health A Tox 

Hazard Subst Environ Eng 42: 671-676. http://dx.doi.org/10.1080/10934520701244474.

Massó-González, EL; Antonio-García, MT. (2009). Natural antioxidants protect against lead-induced damage during 

pregnancy and lactation in rat's pups. Ecotoxicol Environ Saf 72: 2137-2142. 

http://dx.doi.org/10.1016/j.ecoenv.2009.03.013.

Massó, EL; Corredor, L; Antonio, MT. (2007). Oxidative damage in liver after perinatal intoxication with lead 

and/or cadmium. J Trace Elem Med Biol 21: 210-216. http://dx.doi.org/10.1016/j.jtemb.2007.03.002.

Matović, V; Buha, A; Bulat, Z; Dukić-Ćosić, D. (2011). Cadmium toxicity revisited: Focus on oxidative stress 

induction and interactions with zinc and magnesium [Review]. Arh Hig Rada Toksikol 62: 65-76. 

http://dx.doi.org/10.2478/10004-1254-62-2011-2075.

Mauel, J; Ransijn, A; Buchmuller-Rouiller, Y. (1989). Lead inhibits intracellular killing of Leishmania parasites and 

extracellular cytolysis of target cells by macrophages exposed to macrophage activating factor. J Lepid Soc 

45: 401-409. 

Mazumdar, M; Bellinger, DC; Gregas, M; Abanilla, K; Bacic, J; Needleman, HL. (2011). Low-level environmental 

lead exposure in childhood and adult intellectual function: A follow-up study. Environ Health Global 

Access Sci Source 10: 24. http://dx.doi.org/10.1186/1476-069X-10-24.

McCabe, MJ; Lawrence, DA. (1991). Lead, a major environmental pollutant, is immunomodulatory by its 

differential effects on CD4+ T cell subsets. Toxicol Appl Pharmacol 111: 13-23. 

http://dx.doi.org/10.1016/0041-008X(91)90129-3.

McCabe, MJ, Jr; Singh, KP; Reiners, JJ, Jr. (1999). Lead intoxication impairs the generation of a delayed type 

hypersensitivity response. Toxicology 139: 255-264. http://dx.doi.org/10.1016/S0300-483X(99)00147-X.

McElroy, JA; Shafer, MM; Gangnon, RE; Crouch, LA; Newcomb, PA. (2008). Urinary lead exposure and breast 

cancer risk in a population-based case-control study. Cancer Epidemiol Biomarkers Prev 17: 2311-2317. 

http://dx.doi.org/10.1158/1055-9965.epi-08-0263.

McGrath, J; Saha, S; Chant, D; Welham, J. (2008). Schizophrenia: A concise overview of incidence, prevalence, and 

mortality. Epidemiol Rev 30: 67-76. http://dx.doi.org/10.1093/epirev/mxn001.

McLachlin, JR; Goyer, RA; Cherian, MG. (1980). Formation of lead-induced inclusion bodies in primary rat kidney 

epithelial cell cultures: Effect of actinomycin D and cycloheximide. Toxicol Appl Pharmacol 56: 418-431. 

http://dx.doi.org/10.1016/0041-008X(80)90076-9.

McNeill, DR; Wong, HK; Narayana, A; Wilson, DM. (2007). Lead promotes abasic site accumulation and comutagenesis in mammalian cells by inhibiting the major abasic endonuclease Ape1. Mol Carcinog 46: 91-

99. http://dx.doi.org/10.1002/mc.20196.

Meeker, JD; Rossano, MG; Protas, B; Diamond, MP; Puscheck, E; Daly, D; Paneth, N; Wirth, JJ. (2008). Cadmium, 

lead, and other metals in relation to semen quality: Human evidence for molybdenum as a male 

reproductive toxicant. Environ Health Perspect 116: 1473-1479. http://dx.doi.org/10.1289/ehp.11490.

Meeker, JD; Rossano, MG; Protas, B; Padmanahban, V; Diamond, MP; Puscheck, E; Daly, D; Paneth, N; Wirth, JJ.

(2010). Environmental exposure to metals and male reproductive hormones: Circulating testosterone is 

inversely associated with blood molybdenum. Fertil Steril 93: 130-140. 

http://dx.doi.org/10.1016/j.fertnstert.2008.09.044.
February 2012 5-592 Draft – Do Not Cite or Quote

Méndez-Armenta, M; Nava-Ruiz, C; Fernández-Valverde, F; Sánchez-García, A; Rios, C. (2011). Histochemical 

changes in muscle of rats exposed subchronically to low doses of heavy metals. Environ Toxicol 

Pharmacol 32: 107-112. http://dx.doi.org/10.1016/j.etap.2011.03.001.

Méndez-Gómez, J; García-Vargas, GG; López-Carrillo, L; Calderón-Aranda, ES; Gómez, A; Vera, E; Valverde, M; 

Cebrián, ME; Rojas, E. (2008). Genotoxic effects of environmental exposure to arsenic and lead on 

children in region Lagunera, Mexico. Ann N Y Acad Sci 1140: 358-367. 

http://dx.doi.org/10.1196/annals.1454.027.

Mendiola, J; Moreno, JM; Roca, M; Vergara-Juárez, N; Martínez-García, MJ; García-Sánchez, A; Elvira-Rendueles, 

B; Moreno-Grau, S; López-Espín, JJ; Ten, J; Bernabeu, R; Torres-Cantero, AM. (2011). Relationships 

between heavy metal concentrations in three different body fluids and male reproductive parameters: A 

pilot study. Environ Health Global Access Sci Source 10: 6. http://dx.doi.org/10.1186/1476-069X-10-6.

Mendy, A; Gasana, J; Vieira, ER. (In Press) Urinary heavy metals and associated medical conditions in the US adult 

population. Int J Environ Health Res. http://dx.doi.org/10.1080/09603123.2011.605877.

Menegazzi, M; Carcereri-De Prati, A; Suzuki, H; Shinozuka, H; Pibiri, M; Piga, R; Columbano, A; LeddaColumbano, GM. (1997). Liver cell proliferation induced by nafenopin and cyproterone acetate is not 

associated with increases in activation of transcription factors NF-kappaB and AP-1 or with expression of 

tumor necrosis factor alpha. Hepatology 25: 585-592. http://dx.doi.org/10.1002/hep.510250316.

Meng, XM; Zhu, DM; Ruan, DY; She, JQ; Luo, L. (2005). Effects of chronic lead exposure on H MRS of 

hippocampus and frontal lobes in children. Neurology 64: 1644-1647. 

Menke, A; Muntner, P; Batuman, V; Silbergeld, EK; Guallar, E. (2006). Blood lead below 0.48 micromol/L (10 

microg/dL) and mortality among US adults. Circulation 114: 1388-1394. 

http://dx.doi.org/10.1161/circulationaha.106.628321.

Mesdaghinia, A; Yazdanpanah, H; Seddighi, M; Banafshe, HR; Heydari, A. (2010). Effect of short-term lead 

exposure on PTZ-induced seizure threshold in mice. Toxicol Lett 199: 6-9. 

http://dx.doi.org/10.1016/j.toxlet.2010.07.012.

Messerschmidt, A, .; Huber, R, .; Wieghardt, K, .; Poulos, T, . (Eds.). (2001). Handbook of metalloproteins. 

Chichester, UK: Wiley. http://dx.doi.org/10.1002/0470028637.

Mielzyńska, D; Siwińska, E; Kapka, L; Szyfter, K; Knudsen, LE; Merlo, DF. (2006). The influence of 

environmental exposure to complex mixtures including PAHs and lead on genotoxic effects in children 

living in Upper Silesia, Poland. Mutagenesis 21: 295-304. http://dx.doi.org/10.1093/mutage/gel037.

Miller, TE; Golemboski, KA; Ha, RS; Bunn, T; Sanders, FS; Dietert, RR. (1998). Developmental exposure to lead 

causes persistent immunotoxicity in Fischer 344 rats. Toxicol Sci 42: 129-135. 

http://dx.doi.org/10.1006/toxs.1998.2424.

Min, JY; Min, KB; Cho, SI; Kim, R; Sakong, J; Paek, D. (2007). Neurobehavioral function in children with low 

blood lead concentrations. Neurotoxicology 28: 421-425. http://dx.doi.org/10.1016/j.neuro.2006.3.007.

Min, JY; Min, KB; Kim, R; Cho, SI; Paek, D. (2008a). Blood lead levels and increased bronchial responsiveness. 

Biol Trace Elem Res 123: 41-46. http://dx.doi.org/10.1007/s12011-008-8099-6.

Min, KB; Min, JY; Cho, SI; Kim, R; Kim, H; Paek, D. (2008b). Relationship between low blood lead levels and 

growth in children of white-collar civil servants in Korea. Int J Hyg Environ Health 211: 82-87. 

http://dx.doi.org/10.1016/j.ijheh.2007.03.003.

Min, MYO; Singer, LT; Kirchner, HL; Minnes, S; Short, E; Hussain, Z; Nelson, S. (2009). Cognitive development 

and low-level lead exposure in poly-drug exposed children. Neurotoxicol Teratol 31: 225-231. 

http://dx.doi.org/10.1016/j.ntt.2009.03.002.

Minozzo, R; Deimling, LI; Gigante, LP; Santos-Mello, R. (2004). Micronuclei in peripheral blood lymphocytes of 

workers exposed to lead. Mutat Res Genet Toxicol Environ Mutagen 565: 53-60. 

http://dx.doi.org/10.1016/j.mrgentox.2004.09.003.
February 2012 5-593 Draft – Do Not Cite or Quote

Minozzo, R; Deimling, LI; Santos-Mello, R. (2010). Cytokinesis-blocked micronucleus cytome and comet assays in 

peripheral blood lymphocytes of workers exposed to lead considering folate and vitamin B12 status. Mutat 

Res Genet Toxicol Environ Mutagen 697: 24-32. http://dx.doi.org/10.1016/j.mrgentox.2010.01.009.

Miranda, ML; Kim, D; Galeano, M; Paul, CJ; Hull, AP; Morgan, SP. (2007b). The relationship between early 

childhood blood lead levels and performance on end of grade tests. Environ Health Perspect 115: 1242-

1247. http://dx.doi.org/10.1289/ehp.9994.

Miranda, ML; Kim, D; Reiter, J; Overstreet Galeano, MA; Maxson, P. (2009). Environmental contributors to the 

achievement gap. Neurotoxicology 30: 1019-1024. http://dx.doi.org/10.1016/j.neuro.2009.07.012.

Miranda, ML; Edwards, SE; Swamy, GK; Paul, CJ; Neelon, B. (2010). Blood lead levels among pregnant women: 

Historical versus contemporaneous exposures. Int J Environ Res Public Health 7: 1508-1519. 

http://dx.doi.org/10.3390/ijerph7041508.

Mishra, KP; Singh, VK; Rani, R; Yadav, VS; Chandran, V; Srivastava, SP; Seth, PK. (2003). Effect of lead 

exposure on the immune response of some occupationally exposed individuals. Toxicology 188: 251-259. 

http://dx.doi.org/10.1016/S0300-483X(03)00091-X.

Mishra, KP; Chauhan, UK; Naik, S. (2006a). Effect of lead exposure on serum immunoglobulins and reactive 

nitrogen and oxygen intermediate. Hum Exp Toxicol 25: 661-665. 

http://dx.doi.org/10.1177/0960327106070453.

Mishra, KP. (2009). Lead exposure and its impact on immune system: A review [Review]. Toxicol In Vitro 23: 969-

972. http://dx.doi.org/10.1016/j.tiv.2009.06.014.

Mishra, KP; Rani, R; Yadav, VS; Naik, S. (2010). Effect of lead exposure on lymphocyte subsets and activation 

markers. Immunopharmacol Immunotoxicol 32: 446-449. http://dx.doi.org/10.3109/08923970903503668.

Misonou, H; Morishima-Kawashima, M; Ihara, Y. (2000). Oxidative stress induces intracellular accumulation of 

amyloid beta-protein (Abeta) in human neuroblastoma cells. Biochemistry 39: 6951-6959. 

http://dx.doi.org/10.1021/bi000169p.

Mistry, P; Lucier, GW; Fowler, BA. (1985). High-affinity lead binding proteins in rat kidney cytosol mediate cellfree nuclear translocation of lead. J Pharmacol Exp Ther 232: 462-469. 

Mistry, P; Mastri, C; Fowler, BA. (1986). Influence of metal ions on renal cytosolic lead-binding proteins and 

nuclear uptake of lead in the kidney. Biochem Pharmacol 35: 711-713. http://dx.doi.org/10.1016/0006-

2952(86)90371-0.

Moffitt, TE; Caspi, A; Rutter, M; Silva, PA. (2001). Sex differences in antisocial behavior: Conduct disorder, 

delinquency, and violence in the Dunedin Longitudinal Study. Cambridge, UK: University of Cambridge 

Press. 

Mohammad, A; Ali, N; Reza, B; Ali, K. (2010). Effect of ascorbic acid supplementation on nitric oxide metabolites 

and systolic blood pressure in rats exposed to lead. Indian J Pharmacol 42: 78-81. 

http://dx.doi.org/10.4103/0253-7613.64501.

Mohammad, IK; Mahdi, AA; Raviraja, A; Najmul, I; Iqbal, A; Thuppil, V. (2008). Oxidative stress in painters 

exposed to low lead levels. Arh Hig Rada Toksikol 59: 161-169. http://dx.doi.org/10.2478/10004-1254-59-

2008-1883.

Molero, L; Carrasco, C; Marques, M; Vaziri, ND; Mateos-Caceres, PJ; Casado, S; Macaya, C; Barrientos, A; LopezFarre, AJ. (2006). Involvement of endothelium and endothelin-1 in lead-induced smooth muscle cell 

dysfunction in rats. Kidney Int 69: 685-690. http://dx.doi.org/10.1038/sj.ki.5000103.

Molina, RM; Phattanarudee, S; Kim, J; Thompson, K; Wessling-Resnick, M; Maher, TJ; Brain, JD. (2011). 

Ingestion of Mn and Pb by rats during and after pregnancy alters iron metabolism and behavior in 

offspring. Neurotoxicology 32: 413-422. http://dx.doi.org/10.1016/j.neuro.2011.03.010.

Monsell, S. (2003). Task switching. Trends Cogn Sci 7: 134-140. http://dx.doi.org/10.1016/S1364-6613(03)00028-

7.
February 2012 5-594 Draft – Do Not Cite or Quote

Monteiro, HP; Abdalla, DSP; Faljoni-Alàrio, A; Bechara, EJH. (1986). Generation of active oxygen species during 

coupled autoxidation of oxyhemoglobin and delta-aminolevulinic acid. Biochim Biophys Acta 881: 100-

106. http://dx.doi.org/10.1016/0304-4165(86)90102-9.

Monteiro, HP; Abdalla, DSP; Augusto, O; Bechara, EJH. (1989). Free radical generation during deltaaminolevulinic acid autoxidation: Induction by hemoglobin and connections with porphyrinpathies. Arch 

Biochem Biophys 271: 206-216. http://dx.doi.org/10.1016/0003-9861(89)90271-3.

Monteiro, HP; Bechara, EJH; Abdalla, DSP. (1991). Free radicals involvement in neurological porphyrias and lead 

poisoning. Mol Cell Biochem 103: 73-83. http://dx.doi.org/10.1007/BF00229595.

Montenegro, MF; Barbosa, F, Jr; Sandrim, VC; Gerlach, RF; Tanus-Santos, JE. (2006). A polymorphism in the 

delta-aminolevulinic acid dehydratase gene modifies plasma/whole blood lead ratio. Arch Toxicol 80: 394-

398. http://dx.doi.org/10.1007/s00204-005-0056-y.

Moore, JF; Goyer, RA; Wilson, M. (1973). Lead-induced inclusion bodies: Solubility, amino acid content, and 

relationship to residual acidic nuclear proteins. Lab Invest 29: 488-494. 

Moore, JF; Goyer, RA. (1974). Lead-induced inclusion bodies: Composition and probable role in lead metabolism. 

Environ Health Perspect 7: 121-127. http://dx.doi.org/10.1289/ehp.747121.

Moorhouse, SR; Carden, S; Drewitt, PN; Eley, BP; Hargreaves, RJ; Pelling, D. (1988). The effect of chronic low 

level lead exposure on blood-brain barrier function in the developing rat. Biochem Pharmacol 37: 4539-

4547. http://dx.doi.org/10.1016/0006-2952(88)90670-3.

Moorman, WJ; Skaggs, SR; Clark, JC; Turner, TW; Sharpnack, DD; Murrell, JA; Simon, SD; Chapin, RE; 

Schrader, SM. (1998). Male reproductive effects of lead, including species extrapolation for the rabbit 

model. Reprod Toxicol 12: 333-346. http://dx.doi.org/10.1016/S0890-6238(98)00010-0.

Moreley-Fletcher, S; Rea, M; Maccari, S; Laviola, G. (2003). Environmental enrichment during adolescence 

reverses the effects of prenatal stress on play behaviour and HPA axis reactivity in rats. Eur J Neurosci 18: 

3367-3374. http://dx.doi.org/10.1111/j.1460-9568.2003.03070.x.

Morita, Y; Sakai, T; Araki, S; Araki, T; Masuyama, Y. (1997). Nicotinamide adenine dinucleotide synthetase 

activity in erythrocytes as a tool for the biological monitoring of lead exposure. Int Arch Occup Environ 

Health 70: 195-198. http://dx.doi.org/10.1007/s004200050206.

Morris, S; van Aardt, WJ; Ahern, MD. (2005). The effect of lead on the metabolic and energetic status of the Yabby, 

Cherax destructor, during environmental hypoxia. Aquat Toxicol 75: 16-31. 

http://dx.doi.org/10.1016/j.aquatox.2005.07.001.

Mosad, SM; Ghanem, AA; El-Fallal, HM; El-Kannishy, AM; El Baiomy, AA; Al-Diasty, AM; Arafa, LF. (2010). 

Lens cadmium, lead, and serum vitamins C, E, and beta carotene in cataractous smoking patients. Curr Eye 

Res 35: 23-30. http://dx.doi.org/10.3109/02713680903362880.

Moser, R; Oberley, TD; Daggett, DA; Friedman, AL; Johnson, JA; Siegel, FL. (1995). Effects of lead administration 

on developing rat kidney: I Glutathione S-transferase isoenzymes. Toxicol Appl Pharmacol 131: 85-93. 

http://dx.doi.org/10.1006/taap.1995.1050.

Mudipalli, A. (2007). Lead hepatotoxicity & potential health effects. Indian J Med Res 126: 518-527. 

Muldoon, SB; Cauley, JA; Kuller, LH; Morrow, L; Needleman, HL; Scott, J; Hooper, FJ. (1996). Effects of blood 

lead levels on cognitive function of older women. Neuroepidemiology 15: 62-72. 

http://dx.doi.org/10.1159/000109891.

Muller, S; Gillert, KE; Krause, C; Gross, U; L'Age-Stehr, J; Diamantstein, T. (1977). Suppression of delayed type 

hypersensitivity of mice by lead. Cell Mol Life Sci 33: 667-668. http://dx.doi.org/10.1007/BF01946564.

Müller, YM; Kobus, K; Schatz, JC; Ammar, D; Nazari, EM. (In Press) Prenatal lead acetate exposure induces 

apoptosis and changes GFAP expression during spinal cord development. Ecotoxicol Environ Saf. 

http://dx.doi.org/10.1016/j.ecoenv.2011.08.004.
February 2012 5-595 Draft – Do Not Cite or Quote

Muntner, P; He, J; Vupputuri, S; Coresh, J; Batuman, V. (2003). Blood lead and chronic kidney disease in the 

general United States population: Results from NHANES III. Kidney Int 63: 1044-1050. 

http://dx.doi.org/10.1046/j.1523-1755.2003.00812.x.

Muntner, P; Menke, A; DeSalvo, KB; Rabito, FA; Batuman, V. (2005). Continued decline in blood lead levels 

among adults in the United States - The National Health and Nutrition Examination Surveys. Arch Intern 

Med 165: 2155-2161. 

Muntner, P; Menke, A; Batuman, V; Rabito, FA; He, J; Todd, AC. (2007). Association of tibia lead and blood lead 

with end-stage renal disease: A pilot study of African-Americans. Environ Res 104: 396-401. 

http://dx.doi.org/10.1016/j.envres.2007.04.001.

Murakami, K; Feng, G; Chen, SG. (1993). Inhibition of brain protein kinase C subtypes by lead. J Pharmacol Exp 

Ther 264: 757-761. 

Murata, K; Iwata, T; Dakeishi, M; Karita, K. (2009). Lead toxicity: Does the critical level of lead resulting in 

adverse effects differ between adults and children? J Occup Health 51: 1-12. 

http://dx.doi.org/10.1539/joh.K8003.

Naha, N; Chowdhury, AR. (2006). Inorganic lead exposure in battery and paint factory: Effect on human sperm 

structure and functional activity. J UOEH 28: 157-171. 

Naha, N; Manna, B. (2007). Mechanism of lead induced effects on human spermatozoa after occupational exposure. 

Kathmandu Univ Med J 5: 85-94. 

Naicker, N; Norris, SA; Mathee, A; Becker, P; Richter, L. (2010). Lead exposure is associated with a delay in the 

onset of puberty in South African adolescent females: Findings from the birth to twenty cohort. Sci Total 

Environ 408: 4949-4954. http://dx.doi.org/10.1016/j.scitotenv.2010.07.037.

Nakagawa, K. (1989). Hepatic glutathione metabolism in mice acutely treated with lead acetate. Jpn J Pharmacol 51: 

173-179. http://dx.doi.org/10.1254/jjp.51.173.

Nakagawa, K. (1991). Decreased glutathione S-transferase activity in mice livers by acute treatment with lead, 

independent of alteration in glutathione content. Toxicol Lett 56: 13-17. http://dx.doi.org/10.1016/0378-

4274(91)90085-K.

Nakajima, T; Deguchi, T; Kagawa, K; Hikita, H; Ueda, K; Katagishi, T; Ohkawara, T; Kakusui, M; Kimura, H; 

Okanoue, T; Kashima, K; Ashihara, T. (1995). Identification of apoptotic hepatocytes in situ in rat liver 

after lead nitrate administration. J Gastroenterol 30: 725-730. http://dx.doi.org/10.1007/BF02349638.

Nakhoul, F; Kayne, LH; Brautbar, N; Hu, MS; Mcdonough, A; Eggena, P; Golub, MS; Berger, M; Chang, CT; 

Jamgotchian, N; Lee, DBN. (1992). Rapid hypertensinogenic effect of lead: studies in the spontaneously

hypertensive rat. Toxicol Ind Health 8: 89-102. 

Nampoothiri, LP; Gupta, S. (2006). Simultaneous effect of lead and cadmium on granulosa cells: A cellular model 

for ovarian toxicity. Reprod Toxicol 21: 179-185. http://dx.doi.org/10.1016/j.reprotox.2005.07.010.

Nampoothiri, LP; Agarwal, A; Gupta, S. (2007). Effect of co-exposure to lead and cadmium on antioxidant status in 

rat ovarian granulose cells. Arch Toxicol 81: 145-150. http://dx.doi.org/10.1007/s00204-006-0133-x.

Nampoothiri, LP; Gupta, S. (2008). Biochemical effects of gestational coexposure to lead and cadmium on 

reproductive performance, placenta, and ovary. J Biochem Mol Toxicol 22: 337-344. 

http://dx.doi.org/10.1002/jbt.20246.

Nandi, S; Gupta, PS; Selvaraju, S; Roy, SC; Ravindra, JP. (2010). Effects of exposure to heavy metals on viability, 

maturation, fertilization, and embryonic development of buffalo (Bubalus bubalis) oocytes in vitro. Arch 

Environ Contam Toxicol 58: 194-204. http://dx.doi.org/10.1007/s00244-009-9342-7.

Narayana, K; Al-Bader, M. (2011). Ultrastructural and DNA damaging effects of lead nitrate in the liver. Exp 

Toxicol Pathol 63: 43-51. http://dx.doi.org/10.1016/j.etp.2009.09.007.

Nation, JR; Gleaves, DH. (2001). Low-level lead exposure and intelligence in children. Arch Clin Neuropsychol 16: 

375-388. 
February 2012 5-596 Draft – Do Not Cite or Quote

National Kidney Foundation. (2002). K/DOQI clinical practice guidelines for chronic kidney disease: Evaluation, 

classification, and stratification, Part 4: Definition and stages of chronic kidney disease. Am J Kidney Dis 

39: S46-S75. http://dx.doi.org/10.1053/ajkd.2002.30943.

Nava-Hernandez, MP; Hauad-Marroquin, LA; Bassol-Mayagoitia, S; Garcia-Arenas, G; Mercado-Hernandez, R; 

Echavarri-Guzman, MA; Cerda-Flores, RM. (2009). Lead-, cadmium-, and arsenic-induced DNA damage 

in rat germinal cells. DNA Cell Biol 28: 241-248. http://dx.doi.org/10.1089/dna.2009.0860.

Navarro-Moreno, LG; Quintanar-Escorza, MA; Gonzalez, S; Mondragon, R; Cerbon-Solorzano, J; Valdes, J; 

Calderon-Salinas, JV. (2009). Effects of lead intoxication on intercellular junctions and biochemical 

alterations of the renal proximal tubule cells. Toxicol In Vitro 23: 1298-1304. 

http://dx.doi.org/10.1016/j.tiv.2009.07.020.

Navas-Acien, A; Selvin, E; Sharrett, AR; Calderon-Aranda, E; Silbergeld, E; Guallar, E. (2004). Lead, cadmium, 

smoking, and increased risk of peripheral arterial disease. Circulation 109: 3196-3201. 

http://dx.doi.org/10.1161/01.CIR.0000130848.18636.B2.

Navas-Acien, A; Silbergeld, EK; Sharrett, AR; Calderon-Aranda, E; Selvin, E; Guallar, E. (2005). Metals in urine 

and peripheral arterial disease. Environ Health Perspect 113: 164-169. http://dx.doi.org/10.1289/ehp.7329.

Navas-Acien, A; Schwartz, BS; Rothenberg, SJ; Hu, H; Silbergeld, EK; Guallar, E. (2008). Bone lead levels and 

blood pressure endpoints: A meta-analysis. Epidemiology 19: 496-504. 

http://dx.doi.org/10.1097/EDE.0b013e31816a2400.

Navas-Acien, A; Tellez-Plaza, M; Guallar, E; Muntner, P; Silbergeld, E; Jaar, B; Weaver, V. (2009). Blood 

cadmium and lead and chronic kidney disease in US adults: A joint analysis. Am J Epidemiol 170: 1156-

1164. http://dx.doi.org/10.1093/aje/kwp248.

Naves, M; Díaz-López, JB; Gómez, C; Rodríguez-Rebollar, A; Cannata-Andía, JB. (2005). Determinants of 

incidence of osteoporotic fractures in the female Spanish population older than 50. Osteoporos Int 16: 

2013-2017. http://dx.doi.org/10.1007/s00198-005-1983-4.

Nawrot, TS; Thijs, L; Den Hond, EM; Roels, HA; Staessen, JA. (2002). An epidemiological re-appraisal of the 

association between blood pressure and blood lead: A meta-analysis. J Hum Hypertens 16: 123-131. 

http://dx.doi.org/10.1038/sj/jhh/1001300.

Nayak, BN; Ray, M; Persaud, TVN. (1989a). Maternal and fetal chromosomal aberrations in mice following 

prenatal exposure to subembryotoxic doses of lead nitrate. Acta Anat 135: 185-188. 

http://dx.doi.org/10.1159/000146751.

Nayak, BN; Ray, M; Persaud, TVN; Nigli, M. (1989b). Relationship of embryotoxicity to genotoxicity of lead 

nitrate in mice. Exp Pathol 36: 65-73. 

Neal, A; Stansfield, K; Worley, P; Thompson, R; Guilarte, T. (2010a). Lead exposure during synaptogenesis alters 

vesicular proteins and impairs vesicular release: Potential role of NMDA receptor-dependent BDNF 

signaling. Toxicol Sci 116: 249-263. http://dx.doi.org/10.1093/toxsci/kfq111.

Neal, RE; Lin, C; Isom, R; Vaishnav, K; Zigler, JS. (2010b). Opacification of lenses cultured in the presence of Pb. 

Mol Vis 16: 2137-2145. 

Neaton, JD; Kuller, L; Stamler, J; Wentworth, DN. (1995). Impact of systolic and diastolic blood pressure on 

cardiovascular mortality. In JH Laragh; BM Brenner (Eds.), Hypertension: Pathophysiology, diagnosis, and 

management (2nd ed., pp. 127-144). New York, NY: Raven Press Ltd. 

Needleman, HL; Gunnoe, C; Leviton, A; Reed, R; Peresie, H; Maher, C; Barrett, P. (1979). Deficits in psychologic 

and classroom performance of children with elevated dentine lead levels. N Engl J Med 300: 689-695. 

Needleman, HL; Leviton, A; Bellinger, D. (1982). Lead-associated intellectual deficit [letter] [Letter]. N Engl J Med 

306: 367. 

Needleman, HL; Schell, A; Bellinger, D; Leviton, A; Allred, EN. (1990). The long-term effects of exposure to low 

doses of lead in childhood: An 11-year follow-up report. N Engl J Med 322: 83-88. 
February 2012 5-597 Draft – Do Not Cite or Quote

Needleman, HL; Gatsonis, CA. (1990). Low-level lead exposure and the IQ of children: A meta-analysis of modern 

studies. JAMA 263: 673-678. http://dx.doi.org/10.1001/jama.1990.03440050067035.

Needleman, HL; Riess, JA; Tobin, MJ; Biesecker, GE; Greenhouse, JB. (1996). Bone lead levels and delinquent 

behavior. JAMA 275: 363-369. http://dx.doi.org/10.1001/jama.1996.03530290033034.

Needleman, HL; Bellinger, D. (2001). Studies of lead exposure and the developing central nervous system: A reply 

to Kaufman. Arch Clin Neuropsychol 16: 359-374. http://dx.doi.org/10.1016/S0887-6177(00)00087-1.

Needleman, HL; McFarland, C; Ness, RB; Fienberg, SE; Tobin, MJ. (2002). Bone lead levels in adjudicated 

delinquents: A case control study. Neurotoxicol Teratol 24: 711-717. http://dx.doi.org/10.1016/S0892-

0362(02)00269-6.

Nehez, M; Lorencz, R; Desi, I. (2000). Simultaneous action of cypermethrin and two environmental pollutant 

metals, cadmium and lead, on bone marrow cell chromosomes of rats in subchronic administration. 

Ecotoxicol Environ Saf 45: 55-60. http://dx.doi.org/10.1006/eesa.1999.1831.

Nelson, AE; Chaudhary, S; Kraus, VB; Fang, F; Chen, JC; Schwartz, TA; Shi, XA; Renner, JB; Stabler, TV; 

Helmick, CG; Caldwell, K; Robin Poole, A; Jordan, JM. (In Press) Whole blood lead levels are associated 

with biomarkers of joint tissue metabolism in African American and white men and women: The Johnston 

County Osteoarthritis Project. Environ Res. http://dx.doi.org/10.1016/j.envres.2011.08.002.

Nelson, MM; Espy, KA. (2009). Low-Level Lead Exposure and Contingency-Based Responding in Preschoolers: 

An Exploratory Study. Dev Neuropsychol 34: 494-506. http://dx.doi.org/10.1080/87565640902964565.

Nemoto, K; Ito, S; Yoshida, C; Miyata, M; Kojima, M; Degawa, M. (2011). Hepatic expression of spermatogenic 

genes and their transiently remarkable downregulations in Wistar-Kyoto rats in response to lead-nitrate 

administration: strain-difference in the gene expression patterns. J Toxicol Sci 36: 357-364. 

Nemsadze, K; Sanikidze, T; Ratiani, L; Gabunia, L; Sharashenidze, T. (2009). Mechanisms of lead-induced 

poisoning. Georgian Med News92-96. 

Nenov, VD; Taal, MW; Sakharova, OV; Brenner, BM. (2000). Multi-hit nature of chronic renal disease. Curr Opin 

Nephrol Hypertens 9: 85-97. 

Nestmann, ER; Zhang, BW. (2007). Chromosome aberration test of Pigment Yellow 34 (lead chromate) in Chinese 

hamster ovary cells. Mutat Res Genet Toxicol Environ Mutagen 633: 126-132. 

http://dx.doi.org/10.1016/j.mrgentox.2007.05.012.

Neuberger, JS; Hu, SC; Drake, KD; Jim, R. (2009). Potential health impacts of heavy-metal exposure at the Tar 

Creek Superfund site, Ottawa County, Oklahoma. Environ Geochem Health 31: 47-59. 

http://dx.doi.org/10.1007/s10653-008-9154-0.

Newland, MC; Yezhou, S; Logdberg, B; Berlin, M. (1994). Prolonged behavioral effects of in utero exposure to lead 

or methyl mercury: Reduced sensitivity to changes in reinforcement contingencies during behavioral 

transitions and in steady state. Toxicol Appl Pharmacol 126: 6-15. 

http://dx.doi.org/10.1006/taap.1994.1084.

Ni, Z; Hou, S; Barton, CH; Vaziri, ND. (2004). Lead exposure raises superoxide and hydrogen peroxide in human 

endothelial and vascular smooth muscle cells. Kidney Int 66: 2329-2336. http://dx.doi.org/10.1111/j.1523-

1755.2004.66032.x.

Nicolescu, R; Petcu, C; Cordeanu, A; Fabritius, K; Schlumpf, M; Krebs, R; Kramer, U; Winneke, G. (2010). 

Environmental exposure to lead, but not other neurotoxic metals, relates to core elements of ADHD in 

Romanian children: Performance and questionnaire data. Environ Res 110: 476-483. 

http://dx.doi.org/10.1016/j.envres.2010.04.002.

Nigg, J; Knottnerus, G; Martel, M; Nikolas, M; Cavanagh, K; Karmaus, W; Rappley, M. (2008). Low blood lead 

levels associated with clinically diagnosed attention-deficit/hyperactivity disorder and mediated by weak 

cognitive control. Biol Psychiatry 63: 325-331. http://dx.doi.org/10.1016/j.biopsych.2007.07.013.

Nikolova, P; Kavaldzhieva, B. (1991). The effect of certain heavy metals (Mn and Pb) on parameters of erythrocyte 

energy metabolism. J Hyg Epidemiol Microbiol Immunol 35: 361-365. 
February 2012 5-598 Draft – Do Not Cite or Quote

Niu, RY; Sun, ZL; Cheng, ZT; Li, ZG; Wang, JD. (2009). Decreased learning ability and low hippocampus 

glutamate in offspring rats exposed to fluoride and lead. Environ Toxicol Pharmacol 28: 254-258. 

http://dx.doi.org/10.1016/j.etap.2009.04.012.

Nordberg, M; Winblad, B; Fratiglioni, L; Basun, H. (2000). Lead concentrations in elderly urban people related to 

blood pressure and mental performance: Results from a population-based study. Am J Ind Med 38: 290-

294. http://dx.doi.org/10.1002/1097-0274(200009)38:3<290::AID-AJIM7>3.0.CO;2-T.

Novak, J; Banks, RO. (1995). Lead and nickel alter the cardiorenal actions of endothelin in the rat. Proc Soc Exp 

Biol Med 208: 191-198. 

Nowak, P; Szczerbak, G; Nitka, D; Kostrzewa, RM; Sitkiewicz, T; Brus, R. (2008). Effect of prenatal lead exposure 

on nigrostriatal neurotransmission and hydroxyl radical formation in rat neostriatum: Dopaminergicnitrergic interaction. Toxicology 246: 83-89. http://dx.doi.org/10.1016/j.tox.2007.12.026.

NRC (National Research Council). (2000). Toxicological effects of methylmercury. Washington, DC: National 

Academy Press. http://books.nap.edu/catalog.php?record_id=9899.

Nriagu, J; Afeiche, M; Linder, A; Arowolo, T; Ana, G; Sridhar, MKC; Oloruntoba, EO; Obi, E; Ebenebe, JC; 

Orisakwe, OE; Adesina, A. (2008). Lead poisoning associated with malaria in children of urban areas of 

Nigeria. Int J Hyg Environ Health 211: 591-605. http://dx.doi.org/10.1016/j.ijheh.2008.05.001.

NTP (National Toxicology Program). (2004). Eleventh report on carcinogens: Lead (CAS no 7439-92-1) and lead 

compounds. Research Triangle Park, NC: NTP. 

http://ntp.niehs.nih.gov/ntp/roc/eleventh/profiles/s101lead.pdf.

Nuyts, GD; Van Vlem, E; Thys, J; De Leersnijder, D; D'Haese, PC; Elseviers, MM; De Broe, ME. (1995). New 

occupational risk factors for chronic renal failure. Lancet 346: 7-11. http://dx.doi.org/10.1016/S0140-

6736(95)92648-8.

Oberley, TD; Friedman, AL; Moser, R; Siegel, FL. (1995). Effects of lead administration on developing rat kidney: 

II Functional, morphologic, and immunohistochemical studies. Toxicol Appl Pharmacol 131: 94-107. 

http://dx.doi.org/10.1006/taap.1995.1051.

Oberto, A; Marks, N; Evans, HL; Guidotti, A. (1996). Lead (Pb+2) promotes apoptosis in newborn rat cerebellar 

neurons: Pathological implications. J Pharmacol Exp Ther 279: 435-442. 

Obhodas, J, Tucak-Zorić, S., Kutle, A., Valković, V. (2007). Indications for synergetic and antagonistic effects 

between trace elements in the environment to human health. Coll Antropol 31: 209-219. 

Oishi, H; Nakashima, M; Totoki, T; Tomokuni, K. (1996). Chronic lead exposure may inhibit endotheliumdependent hyperpolarizing factor in rats. J Cardiovasc Pharmacol 28: 558-563. 

http://dx.doi.org/10.1097/00005344-199610000-00013.

Olaleye, SB; Adaramoye, OA; Erigbali, PP; Adeniyi, OS. (2007). Lead exposure increases oxidative stress in the 

gastric mucosa of HCl/ethanol-exposed rats. World J Gastroenterol 13: 5121-5126. 

Olewińska, E; Kasperczyk, A; Kapka, L; Kozłowska, A; Pawlas, N; Dobrakowski, M; Birkner, E; Kasperczyk, S.

(2010). Level of DNA damage in lead-exposed workers. Ann Agric Environ Med 17: 231-236. 

Oliveira, H; Spano, M; Santos, C; Pereira, MDL. (2009). Lead chloride affects sperm motility and acrosome 

reaction in mice: Lead affects mice sperm motility and acrosome reaction. Cell Biol Toxicol 25: 341-353. 

http://dx.doi.org/10.1007/s10565-008-9088-4.

Olivero-Verbel, J; Duarte, D; Echenique, M; Guette, J; Johnson-Restrepo, B; Parsons, PJ. (2007). Blood lead levels 

in children aged 5-9 years living in Cartagena, Colombia. Sci Total Environ 372: 707-716. 

http://dx.doi.org/10.1016/j.scitotenv.2006.10.025.

Olszewski, J; Latusinski, J; Kita, A; Pietkiewicz, P; Starostecka, B; Majak, J. (2006). Comparative assessment of 

aluminum and lead concentrations in serum and tissue bioptates in patients with laryngeal papilloma or 

cancer. B-ENT 2: 47-49. 

Ong, CN; Lee, WR. (1980a). Distribution of lead-203 in human peripheral blood in vitro. Br J Ind Med 37: 78-84. 

http://dx.doi.org/10.1136/oem.37.1.78.
February 2012 5-599 Draft – Do Not Cite or Quote

Ong, CN; Lee, WR. (1980b). High affinity of lead for fetal haemoglobin. Br J Ind Med 37: 292-298. 

http://dx.doi.org/10.1136/oem.37.3.292.

Onuegbu, AJ; Olisekodiaka, MJ; Nwaba, EI; Adeyeye, AD; Akinola, FFS. (2011). Assessment of some renal indices 

in people occupationally exposed to lead. Toxicol Ind Health 27: 475-479. 

http://dx.doi.org/10.1177/0748233710390020.

Opler, MGA; Brown, AS; Graziano, J; Desai, M; Zheng, W; Schaefer, C; Factor-Litvak, P; Susser, ES. (2004). 

Prenatal lead exposure, delta-aminolevulinic acid, and schizophrenia. Environ Health Perspect 112: 548-

552. http://dx.doi.org/10.1289/ehp.10464.

Opler, MGA; Buka, SL; Groeger, J; McKeague, I; Wei, C; Factor-Litvak, P; Bresnahan, M; Graziano, J; Goldstein, 

JM; Seidman, LJ; Brown, AS; Susser, ES. (2008). Prenatal exposure to lead, delta-aminolevulinic acid, and 

schizophrenia: Further evidence. Environ Health Perspect 116: 1586-1590. 

http://dx.doi.org/10.1289/ehp.10464.

Orisakwe, OE; Nwachukwu, E; Osadolor, HB; Afonne, OJ; Okocha, CE. (2007). Liver and kidney function tests 

amongst paint factory workers in Nkpor, Nigeria. Toxicol Ind Health 23: 161-165. 

http://dx.doi.org/10.1177/0748233707081908.

Örün, E; Yalçın, SS; Aykut, O; Orhan, G; Morgil, GK; Yurdakök, K; Uzun, R. (2011). Breast milk lead and 

cadmium levels from suburban areas of Ankara. Sci Total Environ 409: 2467-2472. 

http://dx.doi.org/10.1016/j.scitotenv.2011.02.035.

Oskarsson, A; Squibb, KS; Fowler, BA. (1982). Intracellular binding of lead in the kidney: The partial isolation and 

characterization of postmitochondrial lead binding components. Biochem Biophys Res Commun 104: 290-

298. http://dx.doi.org/10.1016/0006-291X(82)91973-8.

Oskarsson, A; Fowler, BA. (1985). Effects of lead inclusion bodies on subcellular distribution of lead in rat kidney: 

The relationship to mitochondrial function. Exp Mol Pathol 43: 397-408. http://dx.doi.org/10.1016/0014-

4800(85)90076-0.

Osman, K; Pawlas, K; Schutz, A; Gazdzik, M; Sokal, JA; Vahter, M. (1999). Lead exposure and hearing effects in 

children in Katowice, Poland. Environ Res 80: 1-8. http://dx.doi.org/10.1006/enrs.1998.3886.

Osterode, W; Ulberth, F. (2000). Increased concentration of arachidonic acid in erythrocyte membranes in 

chronically lead-exposed men. J Toxicol Environ Health A 59: 87-95. 

http://dx.doi.org/10.1080/009841000156998.

Ostro, B; Feng, WY; Broadwin, R; Green, S; Lipsett, M. (2007). The effects of components of fine particulate air 

pollution on mortality in California: Results from CALFINE. Environ Health Perspect 115: 13-19. 

http://dx.doi.org/10.1289/ehp.9281.

Oteiza, PI; Kleinman, CG; Demasi, M; Bechara, EJH. (1995). 5-Aminolevulinic acid induces iron release from 

ferritin. Arch Biochem Biophys 316: 607-611. http://dx.doi.org/10.1006/abbi.1995.1080.

Otto, DA; Baumann, SB; Robinson, GS; Schroeder, SR; Kleinbaum, DG; Barton, CN; Mushak, P. (1985b). 

Auditory and visual evoked potentials in children with undue lead absorption [Abstract]. Toxicologist 5: 

81. 

Otto, DA; Fox, DA. (1993). Auditory and visual dysfunction following lead exposure. Neurotoxicology 14: 191-

207. 

Overmann, SR. (1977). Behavioral effects of asymptomatic lead exposure during neonatal development in rats. 

Toxicol Appl Pharmacol 41: 459-471. http://dx.doi.org/10.1016/S0041-008X(77)80002-1.

Ozsoy, SY; Ozsoy, B; Ozyildiz, Z; Aytekin, I. (2010). Protective effect of L-carnitine on experimental lead toxicity 

in rats: A clinical, histopathological and immunohistochemical study. Biotech Histochem 86: 436-443. 

http://dx.doi.org/10.3109/10520295.2010.529825.

Pace, BM; Lawrence, DA; Behr, MJ; Parsons, PJ; Dias, JA. (2005). Neonatal lead exposure changes quality of 

sperm and number of macrophages in testes of BALB/c mice. Toxicology 210: 247-256. 

http://dx.doi.org/10.1016/j.tox.2005.02.004.
February 2012 5-600 Draft – Do Not Cite or Quote

Pace, TWW; Miller, AH. (2009). Cytokines and glucocorticoid receptor signaling: Relevance to major depression. 

Ann N Y Acad Sci 1179: 86-105. http://dx.doi.org/10.1111/j.1749-6632.2009.04984.x.

Paglia, DE; Valentine, WN. (1975). Characteristics of a pyrimidine-specific 5'-nucleotidase in human erythrocytes. J 

Biol Chem 250: 7973-7979. 

Palaniappan, K; Roy, A; Balakrishnan, K; Gopalakrishnan, L; Mukherjee, B; Hu, H; Bellinger, DC. (2011). Lead 

exposure and visual-motor abilities in children from Chennai, India. Neurotoxicology 32: 465-470. 

http://dx.doi.org/10.1016/j.neuro.2011.03.011.

Palus, J; Rydzynski, K; Dziubaltowska, E; Wyszynska, K; Natarajan, AT; Nilsson, R. (2003). Genotoxic effects of 

occupational exposure to lead and cadmium. Mutat Res Genet Toxicol Environ Mutagen 540: 19-28. 

http://dx.doi.org/10.1016/S1383-5718(03)00167-0.

Pan, SY; Morrison, H; Gibbons, L; Zhou, J; Wen, SW; DesMeules, M; Mao, Y. (2011). Breast cancer risk 

associated with residential proximity to industrial plants in Canada. J Occup Environ Med 53: 522-529. 

http://dx.doi.org/10.1097/JOM.0b013e318216d0b3.

Pandya, C; Pillai, P; Nampoothiri, LP; Bhatt, N; Gupta, S; Gupta, S. (In Press) Effect of lead and cadmium coexposure on testicular steroid metabolism and antioxidant system of adult male rats. Andrologia. 

http://dx.doi.org/10.1111/j.1439-0272.2010.01137.x.

Pandya, CD; Pillai, PP; Gupta, SS. (2010). Lead and cadmium co-exposure mediated toxic insults on hepatic steroid 

metabolism and antioxidant system of adult male rats. Biol Trace Elem Res 134: 307-317. 

http://dx.doi.org/10.1007/s12011-009-8479-6.

Park, SK; Schwartz, J; Weisskopf, M; Sparrow, D; Vokonas, PS; Wright, RO; Coull, B; Nie, H; Hu, H. (2006). 

Low-level lead exposure, metabolic syndrome, and heart rate variability: The VA Normative Aging Study. 

Environ Health Perspect 114: 1718-1724. 

Park, SK; Mukherjee, B; Xia, X; Sparrow, D; Weisskopf, MG; Nie, H; Hu, H. (2009a). Bone lead level prediction 

models and their application to examine the relationship of lead exposure and hypertension in the third 

National Health and Nutrition Examination Survey. J Occup Environ Med 51: 1422-1436. 

http://dx.doi.org/10.1097/JOM.0b013e3181bf6c8d.

Park, SK; Hu, H; Wright, RO; Schwartz, J; Cheng, Y; Sparrow, D; Vokonas, PS; Weisskopf, MG. (2009b). Iron 

metabolism genes, low-level lead exposure, and QT interval. Environ Health Perspect 117: 80-85. 

http://dx.doi.org/10.1289/ehp.11559.

Park, SK; Elmarsafawy, S; Mukherjee, B; Spiro, A, III; Vokonas, PS; Nie, H; Weisskopf, MG; Schwartz, J; Hu, H.

(2010). Cumulative lead exposure and age-related hearing loss: The VA Normative Aging Study. Hear Res 

269: 48-55. http://dx.doi.org/10.1016/j.heares.2010.07.004.

Parkinson, DK; Ryan, C; Bromet, EJ; Connell, MM. (1986). A psychiatric epidemiologic study of occupational lead 

exposure. Am J Epidemiol 123: 261-269. 

Parr, DR; Harris, EJ. (1976). The effect of lead on the calcium-handling capacity of rat heart mitochondria. 

Biochemistry 158: 289-294. 

Pasha, Q; Malik, SA; Iqbal, J; Shah, MH. (2007). Characterization and distribution of the selected metals in the 

scalp hair of cancer patients in comparison with normal donors. Biol Trace Elem Res 118: 207-216. 

http://dx.doi.org/10.1007/s12011-007-0035-7.

Pasha, Q; Malik, SA; Iqbal, J; Shaheen, N; Shah, MH. (2008a). Comparative distribution of the scalp hair trace 

metal contents in the benign tumour patients and normal donors. Environ Monit Assess 147: 377-388. 

http://dx.doi.org/10.1007/s10661-007-0127-z.

Pasha, Q; Malik, SA; Iqbal, J; Shaheen, N; Shah, MH. (2008b). Comparative evaluation of trace metal distribution 

and correlation in human malignant and benign breast tissues. Biol Trace Elem Res 125: 30-40. 

http://dx.doi.org/10.1007/s12011-008-8158-z.

Pasha, Q; Malik, SA; Shah, MH. (2008c). Statistical analysis of trace metals in the plasma of cancer patients versus 

controls. J Hazard Mater 153: 1215-1221. http://dx.doi.org/10.1016/j.jhazmat.2007.09.115.
February 2012 5-601 Draft – Do Not Cite or Quote

Pasha, Q; Malik, SA; Shaheen, N; Shah, MH. (2010). Investigation of trace metals in the blood plasma and scalp 

hair of gastrointestinal cancer patients in comparison with controls. Clin Chim Acta 411: 531-539. 

http://dx.doi.org/10.1016/j.cca.2010.01.010.

Pasha Shaik, A; Sankar, S; Reddy, SC; Das, PG; Jamil, K. (2006). Lead-induced genotoxicity in lymphocytes from 

peripheral blood samples of humans: In vitro studies. Drug Chem Toxicol 29: 111-124. 

http://dx.doi.org/10.1080/01480540500408739.

Pastor-Barriuso, R; Banegas, JR; Damián, J; Appel, LJ; Guallar, E. (2003). Systolic blood pressure, diastolic blood 

pressure, and pulse pressure: An evaluation of their joint effect on mortality. Ann Intern Med 139: 731-739. 

Patel, AB; Prabhu, AS. (2009). Determinants of lead level in umbilical cord blood. Indian Pediatr 46: 791-793. 

Patil, AJ; Bhagwat, VR; Patil, JA; Dongre, NN; Ambekar, JG; Das, KK. (2006a). Biochemical aspects of lead 

exposure in silver jewelry workers in western Maharashtra (India). J Basic Clin Physiol Pharmacol 17: 213-

229. 

Patil, AJ; Bhagwat, VR; Patil, JA; Dongre, NN; Ambekar, JG; Jailkhani, R; Das, KK. (2006b). Effect of lead (Pb) 

exposure on the activity of superoxide dismutase and catalase in battery manufacturing workers (BMW) of 

Western Maharashtra (India) with reference to heme biosynthesis. Int J Environ Res Public Health 3: 329-

337. 

Patil, AJ; Bhagwat, VR; Patil, JA; Dongre, NN; Ambekar, JG; Das, KK. (2007). Occupational lead exposure in 

battery manufacturing workers, silver jewelry workers, and spray painters in western Maharashtra (India): 

Effect on liver and kidney function. J Basic Clin Physiol Pharmacol 18: 87-100. 

Pavanello, S; Bollati, V; Pesatori, AC; Kapka, L; Bolognesi, C; Bertazzi, PA; Baccarelli, A. (2009). Global and 

gene-specific promoter methylation changes are related to anti-B[a]PDE-DNA adduct levels and influence 

micronuclei levels in polycyclic aromatic hydrocarbon-exposed individuals. Int J Cancer 125: 1692-1697. 

http://dx.doi.org/10.1002/ijc.24492.

Pavanello, S; Pesatori, AC; Dioni, L; Hoxha, M; Bollati, V; Siwinska, E; Mielzyńska, D; Bolognesi, C; Bertazzi, 

PA; Baccarelli, A. (2010). Shorter telomere length in peripheral blood lymphocytes of workers exposed to 

polycyclic aromatic hydrocarbons. Carcinogenesis 31: 216-221. http://dx.doi.org/10.1093/carcin/bgp278.

Payton, M; Hu, H; Sparrow, D; Weiss, ST. (1994). Low-level lead exposure and renal function in the Normative 

Aging Study. Am J Epidemiol 140: 821-829. 

Payton, M; Riggs, KM; Spiro, A, III; Weiss, ST; Hu, H. (1998). Relations of bone and blood lead to cognitive 

function: The VA Normative Aging Study. Neurotoxicol Teratol 20: 19-27. 

Pekcici, R; Kavlakoğlu, B; Yilmaz, S; Şahin, M; Delibaşi, T. (2010). Effects of lead on thyroid functions in leadexposed workers. Central European Journal of Medicine 5: 215-218. http://dx.doi.org/10.2478/s11536-009-

0092-8.

Peng, S; Hajela, RK; Atchison, WD. (2002). Characteristics of block by Pb2+ of function of human neuronal L-, N-, 

and R-type Ca2+ channels transiently expressed in human embryonic kidney 293 cells. Mol Pharmacol 62: 

1418-1430. http://dx.doi.org/10.1124/mol.62.6.1418.

Pentyala, S; Ruggeri, J; Veerraju, A; Yu, Z; Bhatia, A; Desaiah, D; Vig, P. (2010). Microsomal Ca2+ flux 

modulation as an indicator of heavy metal toxicity. Indian J Exp Biol 48: 737-743. 

Pérez-Bravo, F; Ruz, M; Morán-Jiménez, MJ; Olivares, M; Rebolledo, A; Codoceo, J; Sepúlveda, V; Jenkin, A; 

Santos, JL; Fontanellas, A. (2004). Association between aminolevulinate dehydrase genotypes and blood 

lead levels in children from a lead-contaminated area in Antofagasta, Chile. Arch Environ Contam Toxicol 

47: 276-280. http://dx.doi.org/10.1007/s00244-004-2215-1.

Perlstein, T; Weuve, J; Schwartz, J; Sparrow, D; Wright, R; Litonjua, A; Nie, H; Hu, H. (2007). Cumulative 

community-level lead exposure and pulse pressure: The Normative Aging Study. Environ Health Perspect 

115: 1696-1700. http://dx.doi.org/10.1289/ehp.10350.

Peters, JL; Kubzansky, L; McNeely, E; Schwartz, J; Spiro, A, III; Sparrow, D; Wright, RO; Nie, H; Hu, H. (2007). 

Stress as a potential modifier of the impact of lead levels on blood pressure: The Normative Aging Study. 

Environ Health Perspect 115: 1154-1159. http://dx.doi.org/10.1289/ehp.10002.
February 2012 5-602 Draft – Do Not Cite or Quote

Petit, TL; LeBoutillier, JC. (1979). Effects of lead exposure during development on neocortical dendritic and 

synaptic structure. Exp Neurol 64: 482-492. http://dx.doi.org/10.1016/0014-4886(79)90226-7.

Petri, S; Calingasan, NY; Alsaied, OA; Wille, E; Kiaei, M; Friedman, JE; Baranova, O; Chavez, JC; Beal, MF.

(2007). The lipophilic metal chelators DP-109 and DP-460 are neuroprotective in a transgenic mouse 

model of amyotrophic lateral sclerosis. J Neurochem 102: 991-1000. http://dx.doi.org/10.1111/j.1471-

4159.2007.04604.x.

Piao, F; Cheng, F; Chen, H; Li, G; Sun, X; Liu, S; Yamauchi, T; Yokoyama, K. (2007). Effects of zinc 

coadministration on lead toxicities in rats. Ind Health 45: 546-551. 

Piccinini, F; Favalli, L; Chiari, MC. (1977). Experimental investigations on the conctraction induced by lead in 

arterial smooth muscle. Toxicology 8: 43-51. http://dx.doi.org/10.1016/0300-483X(77)90022-1.

Pillai, P; Pandya, C; Bhatt, N; Gupta, SS. (In Press) Biochemical and reproductive effects of gestational/lactational 

exposure to lead and cadmium with respect to testicular steroidogenesis, antioxidant system, endogenous 

sex steroid and cauda-epididymal functions. Andrologia. http://dx.doi.org/10.1111/j.1439-

0272.2010.01109.x.

Pillai, P; Patel, R; Pandya, C; Gupta, S. (2009). Sex-specific effects of gestational and lactational coexposure to lead 

and cadmium on hepatic phase I and phase II xenobiotic/steroid-metabolizing enzymes and antioxidant 

status. J Biochem Mol Toxicol 23: 419-431. http://dx.doi.org/10.1002/jbt.20305.

Pillai, P; Pandya, C; Gupta, S; Gupta, S. (2010). Biochemical and molecular effects of gestational and lactational 

coexposure to lead and cadmium on ovarian steroidogenesis are associated with oxidative stress in F1 

generation rats. J Biochem Mol Toxicol 24: 384-394. http://dx.doi.org/10.1002/jbt.20351.

Pilsner, JR; Hu, H; Ettinger, A; Sánchez, BN; Wright, RO; Cantonwine, D; Lazarus, A; Lamadrid-Figueroa, H; 

Mercado-García, A; Téllez-Rojo, MM; Hernández-Avila, M. (2009). Influence of prenatal lead exposure on 

genomic methylation of cord blood DNA. Environ Health Perspect 117: 1466-1471. 

http://dx.doi.org/10.1289/ehp.0800497.

Pilsner, JR; Hu, H; Wright, RO; Kordas, K; Ettinger, AS; Sánchez, BN; Cantonwine, D; Lazarus, AL; Cantoral, A; 

Schnaas, L; Téllez-Rojo, MM; Hernández-Avila, M. (2010). Maternal MTHFR genotype and haplotype 

predict deficits in early cognitive development in a lead-exposed birth cohort in Mexico City. Am J Clin 

Nutr 92: 226-234. http://dx.doi.org/10.3945/ajcn.2009.28839.

Pine, MD; Hiney, JK; Dearth, RK; Bratton, GR; Dees, WL. (2006). IGF-1 administration to prepubertal female rats 

can overcome delayed puberty caused by maternal Pb exposure. Reprod Toxicol 21: 104-109. 

http://dx.doi.org/10.1016/j.reprotox.2005.07.003.

Pineda-Zavaleta, AP; García-Vargas, G; Borja-Aburto, VH; Acosta-Saavedra, LC; Aguilar, EV; Gómez-Muñoz, A; 

Cebrián, ME; Calderón-Aranda, ES. (2004). Nitric oxide and superoxide anion production in monocytes 

from children exposed to arsenic and lead in region Lagunera, Mexico. Toxicol Appl Pharmacol 198: 283-

290. http://dx.doi.org/10.1016/j.taap.2003.10.034.

Pinkerton, LE; Biagini, RE; Ward, EM; Hull, RD; Deddens, JA; Boeniger, MF; Schnorr, TM; MacKenzie, BA; 

Luster, MI. (1998). Immunologic findings among lead-exposed workers. Am J Ind Med 33: 400-408. 

http://dx.doi.org/10.1002/(SICI)1097-0274(199804)33:4<400::AID-AJIM11>3.0.CO;2-2.

Pinto, D; Ceballos, JM; Garcia, G; Guzman, P; Del Razo, LM; Vera, E; Gomez, H; Garcia, A; Gonsebatt, ME.

(2000). Increased cytogenetic damage in outdoor painters. Mutat Res Genet Toxicol Environ Mutagen 467: 

105-111. http://dx.doi.org/10.1016/S1383-5718(00)00024-3.

Pizent, A; Macan, J; Jurasovic, J; Varnai, VM; Milkovic-Kraus, S; Kanceljak-Macan, B. (2008). Association of 

toxic and essential metals with atopy markers and ventilatory lung function in women and men. Sci Total 

Environ 390: 369-376. http://dx.doi.org/10.1016/j.scitotenv.2007.10.049.

Pizent, A; Colak, B; Kljakovic Gaspic, Z; Telisman, S. (2009). Prostate-specific antigen (PSA) in serum in relation 

to blood lead concentration and alcohol consumption in men. Arh Hig Rada Toksikol 60: 69-78. 

http://dx.doi.org/10.2478/10004-1254-60-2009-1901.
February 2012 5-603 Draft – Do Not Cite or Quote

Plusquellec, P; Muckle, G; Dewailly, E; Ayotte, P; Jacobson, SW; Jacobson, JL. (2007). The relation of low-level 

prenatal lead exposure to behavioral indicators of attention in Inuit infants in Arctic Quebec. Neurotoxicol 

Teratol 29: 527-537. http://dx.doi.org/10.1016/j.ntt.2007.07.002.

Plusquellec, P; Muckle, G; Dewailly, E; Ayotte, P; Bégin, G; Desrosiers, C; Després, C; Saint-Amour, D; Poitras, 

K. (2010). The relation of environmental contaminants exposure to behavioral indicators in Inuit 

preschoolers in Arctic Quebec. Neurotoxicology 31: 17-25. http://dx.doi.org/10.1016/j.neuro.2009.10.008.

Pocock, SJ; Smith, M; Baghurst, P. (1994). Environmental lead and children's intelligence: A systematic review of 

the epidemiological evidence [Review]. Br Med J 309: 1189-1197. 

http://dx.doi.org/10.1136/bmj.309.6963.1189.

Pollack, AZ; Schisterman, EF; Goldman, LR; Mumford, SL; Albert, PS; Jones, RL; Wactawski-Wende, J. (2011). 

Cadmium, lead, and mercury in relation to reproductive hormones and anovulation in premenopausal 

women. Environ Health Perspect 119: 1156-1161. http://dx.doi.org/10.1289/ehp.1003284.

Ponce-Canchihuamán, JC; Pérez-Méndez, O; Hernández-Muñoz, R; Torres-Durán, PV; Juárez-Oropeza, MA.

(2010). Protective effects of Spirulina maxima on hyperlipidemia and oxidative-stress induced by lead 

acetate in the liver and kidney. Lipids Health Dis 9: 35. http://dx.doi.org/10.1186/1476-511X-9-35.

Poreba, R; Gac, P; Poreba, M; Derkacz, A; Pilecki, W; Antonowicz-Juchniewicz, J; Andrzejak, R. (2010). 

[Relationship between chronic exposure to lead, cadmium and manganese, blood pressure values and 

incidence of arterial hypertension]. Med Pr 61: 5-14. 

Poreba, R; Poreba, M; Gać, P; Andrzejak, R. (2011). Ambulatory blood pressure monitoring and structural changes 

in carotid arteries in normotensive workers occupationally exposed to lead. Hum Exp Toxicol 30: 1174-

1180. http://dx.doi.org/10.1177/0960327110391383.

Poręba, R; Poręba, M; Gać, P; Pilecki, W; Andrzejak, R. (2010). Impaired endothelial function assessed by flowmediated vasodilatation in workers occupationally exposed to lead. Environ Toxicol Pharmacol 29: 266-

270. http://dx.doi.org/10.1016/j.etap.2010.03.001.

Poręba, R; Poręba, M; Gać, P; Steinmetz-Beck, A; Beck, B; Pilecki, W; Andrzejak, R; Sobieszczańska, M. (2011a). 

Electrocardiographic changes in workers occupationally exposed to lead. Ann Noninvasive Electrocardiol 

16: 33-40. http://dx.doi.org/10.1111/j.1542-474X.2010.00406.x.

Poręba, R; Gać, P; Poręba, M; Antonowicz-Juchniewicz, J; Andrzejak, R. (2011b). Relationship between 

occupational exposure to lead and local arterial stiffness and left ventricular diastolic function in 

individuals with arterial hypertension. Toxicol Appl Pharmacol 254: 342-348. 

http://dx.doi.org/10.1016/j.taap.2011.05.009.

Prentice, RC; Kopp, SJ. (1985). Cardiotoxicity of lead at various perfusate calcium concentrations: Functional and 

metabolic responses of the perfused rat heart. Toxicol Appl Pharmacol 81: 491-501. 

http://dx.doi.org/10.1016/0041-008X(85)90420-X.

Prins, JM; Park, S; Lurie, DI. (2010). Decreased expression of the voltage-dependent anion channel in differentiated 

PC-12 and SH-SY5Y cells following low-level Pb exposure. Toxicol Sci 113: 169-176. 

http://dx.doi.org/10.1093/toxsci/kfp249.

Prospective Studies Collaboration. (2002). Age-specific relevance of usual blood pressure to vascular mortality: A 

meta-analysis of individual data for one million adults in 61 prospective studies. Lancet 360: 1903-1913. 

http://dx.doi.org/10.1016/S0140-6736(02)11911-8.

Prozialeck, WC; Grunwald, GB; Dey, PM; Reuhl, KR; Parrish, AR. (2002). Cadherins and NCAM as potential 

targets in metal toxicity [Review]. Toxicol Appl Pharmacol 182: 255-265. 

http://dx.doi.org/10.1006/taap.2002.9422.

Pugh Smith, P; Nriagu, JO. (2011). Lead poisoning and asthma among low-income and African American children 

in Saginaw, Michigan. Environ Res 111: 81-86. http://dx.doi.org/10.1016/j.envres.2010.11.007.

Purdy, RE; Smith, JR; Ding, Y; Oveisi, F; Vaziri, ND; Gonick, HC. (1997). Lead-induced hypertension is not 

associated with altered vascular reactivity in vitro. Am J Hypertens 10: 997-1003. 

http://dx.doi.org/10.1016/S0895-7061(97)00108-8.
February 2012 5-604 Draft – Do Not Cite or Quote

Putnam, CD; Arvai, AS; Bourne, Y; Tainer, JA. (2000). Active and inhibited human catalase structures: Ligand and 

NADPH binding and catalytic mechanism. J Mol Biol 296: 295-309. 

http://dx.doi.org/10.1006/jmbi.1999.3458.

Pyatt, DW; Zheng, JH; Stillman, WS; Irons, RD. (1996). Inorganic lead activates NF-kB in primary human CD4+ T 

lymphocytes. Biochem Biophys Res Commun 227: 380-385. http://dx.doi.org/10.1006/bbrc.1996.1516.

Qian, L; Flood, PM. (2008). Microglial cells and Parkinson's disease. Immunol Res 41: 155-164. 

http://dx.doi.org/10.1007/s12026-008-8018-0.

Qian, Y; Harris, ED; Zheng, Y; Tiffany-Castiglioni, E. (2000). Lead targets GRP78, a molecular chaperone, in C6 

rat glioma cells. Toxicol Appl Pharmacol 163: 260-266. http://dx.doi.org/10.1006/taap.1999.8878.

Qian, Y; Zheng, Y; Ramos, KS; Tiffany-Castiglioni, E. (2005). GRP78 compartmentalized redistribution in Pbtreated glia: Role of GRP78 in lead-induced oxidative stress. Neurotoxicology 26: 267-275. 

http://dx.doi.org/10.1016/j.neuro.2004.09.002.

Qian, ZM; Morgan, EH. (1990). Effect of lead on the transport of transferrin-free and transferrin-bound iron into 

rabbit reticulocytes. Biochem Pharmacol 40: 1049-1054. http://dx.doi.org/10.1016/0006-2952(90)90492-4.

Qiao, YF; Jiang, YS; Pang, DZ. (2006). [Expression of renal nuclear factor-kappaB, transforming growth factor-beta 

and fibronectin of rats exposed to lead]. Zhonghua Lao Dong Wei Sheng Zhi Ye Bing Za Zhi 24: 139-142. 

Qu, W; Diwan, BA; Liu, J; Goyer, RA; Dawson, T; Horton, JL; Cherian, MG; Waalkes, MP. (2002). The 

metallothionein-null phenotype is associated with heightened sensitivity to lead toxicity and an inability to 

form inclusion bodies. Am J Pathol 160: 1047-1056. http://dx.doi.org/10.1016/S0002-9440(10)64925-5.

Queirolo, EI; Ettinger, AS; Stoltzfus, RJ; Kordas, K. (2010). Association of anemia, child and family characteristics 

with elevated blood lead concentrations in preschool children from Montevideo, Uruguay. Arch Environ 

Occup Health 65: 94-100. http://dx.doi.org/10.1080/19338240903390313.

Queiroz, ML; da Rocha, MC; Torello, CO; de Souza Queiroz, J; Bincoletto, C; Morgano, MA; Romano, MR; 

Paredes-Gamero, EJ; Barbosa, CM; Calgarotto, AK. (2011). Chlorella vulgaris restores bone marrow 

cellularity and cytokine production in lead-exposed mice. Food Chem Toxicol 49: 2934-2941. 

http://dx.doi.org/10.1016/j.fct.2011.06.056.

Queiroz, MLS; Almeida, M; Gallão, MI; Höehr, NF. (1993). Defective neutrophil function in workers 

occupationally exposed to lead. Basic Clin Pharmacol Toxicol 72: 73-77. http://dx.doi.org/10.1111/j.1600-

0773.1993.tb00293.x.

Queiroz, MLS; Costa, FF; Bincoletto, C; Perlingeiro, RCR; Dantas, DCM; Cardoso, MP; Almeida, M. (1994a). 

Engulfment and killing capabilities of neutrophils and phagocytic splenic function in persons 

occupationally exposed to lead. International Journal of Immunopharmacology 16: 239-244. 

http://dx.doi.org/10.1016/0192-0561(94)90018-3.

Queiroz, MLS; Perlingeiro, RCR; Bincoletto, C; Almeida, M; Cardoso, MP; Dantas, DCM. (1994b). 

Immunoglobulin levels and cellular immune function in lead exposed workers. Immunopharmacol 

Immunotoxicol 16: 115-128. http://dx.doi.org/10.3109/08923979409029904.

Quinlan, GJ; Halliwell, B; Moorhouse, CP; Gutteridge, JMC. (1988). Action of lead(II) and aluminium(III) ions on 

iron-stimulated lipid peroxidation in liposomes, erythrocytes and rat liver microsomal fractions. Biochim 

Biophys Acta 962: 196-200. http://dx.doi.org/10.1016/0005-2760(88)90159-2.

Quintanar-Escorza, MA; González-Martínez, MT; Navarro, L; Maldonado, M; Arévalo, B; Calderón-Salinas, JV.

(2007). Intracellular free calcium concentration and calcium transport in human erythrocytes of leadexposed workers. Toxicol Appl Pharmacol 220: 1-8. http://dx.doi.org/10.1016/j.taap.2006.10.016.

Quintanar-Escorza, MA; González-Martínez, MT; del Pilar, IOM; Calderón-Salinas, JV. (2010). Oxidative damage 

increases intracellular free calcium [Ca2+]i concentration in human erythrocytes incubated with lead. 

Toxicol In Vitro 24: 1338-1346. http://dx.doi.org/10.1016/j.tiv.2010.05.002.

Quintanilla-Vega, B; Smith, DR; Kahng, MW; Hernandez, JM; Albores, A; Fowler, BA. (1995). Lead-binding 

proteins in brain tissue of environmentally lead-exposed humans. Chem Biol Interact 98: 193-209. 

http://dx.doi.org/10.1016/0009-2797(95)03646-6.
February 2012 5-605 Draft – Do Not Cite or Quote

Rabbani-Chadegani, A; Abdosamadi, S; Fani, N; Mohammadian, S. (2009). A comparison of the effect of lead 

nitrate on rat liver chromatin, DNA and histone proteins in solution. Arch Toxicol 83: 565-570. 

http://dx.doi.org/10.1007/s00204-008-0362-2.

Rabbani-Chadegani, A, III; Fani, N; Abdossamadi, S; Shahmir, N. (2011). Toxic effects of lead and nickel nitrate on 

rat liver chromatin components. J Biochem Mol Toxicol 25: 127-134. http://dx.doi.org/10.1002/jbt.20368.

Rabinowitz, MB. (1988). Lead and pregnancy. Birth 15: 236-241. http://dx.doi.org/10.1111/j.1523-

536X.1988.tb01117.x.

Rabinowitz, MB; Allred, EN; Bellinger, DC; Leviton, A; Needleman, HL. (1990). Lead and childhood propensity to 

infectious and allergic disorders: Is there an association? Bull Environ Contam Toxicol 44: 657-660. 

http://dx.doi.org/10.1007/BF01701784.

Rafalowska, U; Struzyńska, L; Dabrowska-Bouta, B; Lenkiewicz, A. (1996). Is lead toxicosis a reflection of altered 

energy metabolism in brain synaptosomes? Acta Neurobiol Exp (Wars) 56: 611-617. 

Rafique, M; Khan, N; Perveen, K; Naqvi, A. (2009). The effects of lead and zinc on the quality of semen of albino 

rats. J Coll Physicians Surg Pak 19: 510-513. 

Raghavan, SRV; Gonick, HC. (1977). Isolation of low-molecular-weight lead-binding protein from human 

erythrocytes. Proc Soc Exp Biol Med 155: 164-167. 

Raghavan, SRV; Culver, BD; Gonick, HC. (1980). Erythrocyte lead-binding protein after occupational exposure: I 

Relationship to lead toxicity. Environ Res 22: 264-270. http://dx.doi.org/10.1016/0013-9351(80)90138-3.

Raghavan, SRV; Culver, BD; Gonick, HC. (1981). Erythrocyte lead-binding protein after occupational exposure: II. 

Influence on lead inhibition of membrane Na+, K+ - adenosinetriphosphatase. J Toxicol Environ Health 7: 

561-568. http://dx.doi.org/10.1080/15287398109530001.

Raine, A; Lencz, T; Bihrle, S; LaCasse, L; Colletti, P. (2000). Reduced prefrontal gray matter volume and reduced 

autonomic activity in antisocial personality disorder. Arch Gen Psychiatry 57: 119-127. 

Raitakari, OT; Juonala, M; Kähönen, M; Taittonen, L; Laitinen, T; Mäki-Torkko, N; Järvisalo, MJ; Uhari, M; 

Jokinen, E; Rönnemaa, T; Akerblom, HK; Viikari, JS. (2003). Cardiovascular risk factors in childhood and 

carotid artery intima-media thickness in adulthood: The Cardiovascular Risk in Young Finns Study. JAMA 

290: 2277-2283. http://dx.doi.org/10.1001/jama.290.17.2277.

Rajah, T; Ahuja, YR. (1995). In vivo genotoxic effects of smoking and occupational lead exposure in printing press 

workers. Toxicol Lett 76: 71-75. http://dx.doi.org/10.1016/0378-4274(94)03200-9.

Rajah, TT; Ahuja, YR. (1996). In vivo genotoxicity of alcohol consumption and lead exposure in printing press 

workers. Alcohol 13: 65-68. http://dx.doi.org/10.1016/0741-8329(95)02014-4.

Rajan, P; Kelsey, KT; Schwartz, JD; Bellinger, DC; Weuve, J; Sparrow, D; Spiro, A, III; Smith, TJ; Nie, H; Hu, H; 

Wright, RO. (2007). Lead burden and psychiatric symptoms and the modifying influence of the deltaaminolevulinic acid dehydratase (ALAD) polymorphism: The VA Normative Aging Study. Am J 

Epidemiol 166: 1400-1408. http://dx.doi.org/10.1093/aje/kwm220.

Rajan, P; Kelsey, KT; Schwartz, JD; Bellinger, DC; Weuve, J; Spiro, A, III; Sparrow, D; Smith, TJ; Nie, H; 

Weisskopf, MG; Hu, H; Wright, RO. (2008). Interaction of the delta-aminolevulinic acid dehydratase 

polymorphism and lead burden on cognitive function: The VA Normative Aging Study. J Occup Environ 

Med 50: 1053-1061. http://dx.doi.org/10.1097/JOM.0b013e3181792463.

Rajaraman, P; Stewart, PA; Samet, JM; Schwartz, BS; Linet, MS; Zahm, SH; Rothman, N; Yeager, M; Fine, HA; 

Black, PM; Loeffler, J; Shapiro, WR; Selker, RG; Inskip, PD. (2006). Lead, genetic susceptibility, and risk 

of adult brain tumors. Cancer Epidemiol Biomarkers Prev 15: 2514-2520. http://dx.doi.org/10.1158/1055-

9965.EPI-06-0482.

Ramesh, GT; Manna, SK; Aggarwal, BB; Jadhav, AL. (1999). Lead activates nuclear transcription factor -kB, 

activator protein-1, and amino-terminal c-Jun kinase in pheochromocytoma cells. Toxicol Appl Pharmacol 

155: 280-286. http://dx.doi.org/10.1006/taap.1999.8624.
February 2012 5-606 Draft – Do Not Cite or Quote

Ramesh, GT; Manna, SK; Aggarwal, BB; Jadhav, AL. (2001). Lead exposure activates nuclear factor kappa B, 

activator protein-1, c-Jun N-terminal kinase and caspases in the rat brain. Toxicol Lett 123: 195-207. 

http://dx.doi.org/10.1016/S0378-4274(01)00395-2.

Rana, SVS. (2008). Metals and apoptosis: Recent developments. J Trace Elem Med Biol 22: 262-284. 

http://dx.doi.org/10.1016/j.jtemb.2008.08.002.

Razmiafshari, M; Zawia, NH. (2000). Utilization of a synthetic peptide as a tool to study the interaction of heavy 

metals with the zinc finger domain of proteins critical for gene expression in the developing brain. Toxicol 

Appl Pharmacol 166: 1-12. http://dx.doi.org/10.1006/taap.2000.8950.

Razmiafshari, M; Kao, J; d'Avignon, A; Zawia, NH. (2001). NMR identification of heavy metal-binding sites in a 

synthetic zinc finger peptide: Toxicological implications for the interactions of xenobiotic metals with zinc 

finger proteins. Toxicol Appl Pharmacol 172: 1-10. http://dx.doi.org/10.1006/taap.2001.9132.

Reddy, GR; Zawia, NH. (2000). Lead exposure alters Egr-1 DNA-binding in the neonatal rat brain. Int J Dev 

Neurosci 18: 791-795. http://dx.doi.org/10.1016/S0736-5748(00)00048-4.

Reddy, SY; Pullakhandam, R; Dinesh Kumar, B. (2010). Thiamine reduces tissue lead levels in rats: Mechanism of 

interaction. Biometals 23: 247-253. http://dx.doi.org/10.1007/s10534-009-9282-8.

Regunathan, S; Sundaresan, R. (1984). Pyruvate metabolism in the brain of young rats intoxicated with organic and 

inorganic lead. J Neurochem 43: 1346-1351. http://dx.doi.org/10.1111/j.1471-4159.1984.tb05393.x.

Rehman, S; Rehman, S; Chandra, O; Abdulla, M. (1995). Evaluation of malondialdehyde as an index of lead 

damage in rat brain homogenates. Biometals 8: 275-279. http://dx.doi.org/10.1007/BF00141599.

Reif, A; Fritzen, S; Finger, M; Strobel, A; Lauer, M; Schmitt, A; Lesch, K. (2006). Neural stem cell proliferation is 

decreased in schizophrenia, but not in depression. Mol Psychiatry 11: 514-522. 

http://dx.doi.org/10.1038/sj.mp.4001791.

Rendón-Ramirez, A; Cerbón-Solórzano, J; Maldonado-Vega, M; Quintanar-Escorza, MA; Calderón-Salinas, JV.

(2007). Vitamin-E reduces the oxidative damage on δ-aminolevulinic dehydratase induced by lead 

intoxication in rat erythrocytes. Toxicol In Vitro 21: 1121-1126. 

http://dx.doi.org/10.1016/j.tiv.2007.04.019.

Reshma Anjum, M; Sainath, SB; Suneetha, Y; Sreenivasula Reddy, P. (2010). Lead acetate induced reproductive 

and paternal mediated developmental toxicity in rats. Ecotoxicol Environ Saf 74: 793–799. 

http://dx.doi.org/10.1016/j.ecoenv.2010.10.044.

Reuhl, KR; Rice, DC; Gilbert, SG; Mallett, J. (1989). Effects of chronic developmental lead exposure on monkey 

neuroanatomy: Visual system. Toxicol Appl Pharmacol 99: 501-509. http://dx.doi.org/10.1016/0041-

008X(89)90157-9.

Revis, NW; Zinsmeister, AR; Bull, R. (1981). Atherosclerosis and hypertension induction by lead and cadmium 

ions: An effect prevented by calcium ion. PNAS 78: 6494-6498. 

Reza, B; Ali, N; Azhdar, H; Alireza, A; Ali, K. (2008). Effects of low-level lead exposure on blood pressure and 

function of the rat isolated heart. Indian J Pharmacol 40: 69-72. http://dx.doi.org/10.4103/0253-

7613.41041.

Rhodes, D; Spiro, A, III; Aro, A; Hu, H. (2003). Relationship of bone and blood lead levels to psychiatric 

symptoms: The Normative Aging Study. J Occup Environ Med 45: 1144-1151. 

http://dx.doi.org/10.1097/01.jom.0000094995.23808.7b.

Rice, C; Ghorai, JK; Zalewski, K; Weber, DN. (2011). Developmental lead exposure causes startle response deficits 

in zebrafish. Aquat Toxicol 105: 600-608. http://dx.doi.org/10.1016/j.aquatox.2011.08.014.

Rice, D; Barone, S, Jr. (2000). Critical periods of vulnerability for the developing nervous system: Evidence from 

humans and animal models. Environ Health Perspect 108: 511-533. 

http://dx.doi.org/10.1289/ehp.00108s3511.

Rice, DC; Karpinski, KF. (1988). Lifetime low-level lead exposure produces deficits in delayed alternation in adult 

monkeys. Neurotoxicol Teratol 10: 207-214. http://dx.doi.org/10.1016/0892-0362(88)90019-0.
February 2012 5-607 Draft – Do Not Cite or Quote

Rice, DC. (1990). Lead-induced behavioral impairment on a spatial discrimination reversal task in monkeys exposed 

during different periods of development. Toxicol Appl Pharmacol 106: 327-333. 

http://dx.doi.org/10.1016/0041-008X(90)90251-O.

Rice, DC; Gilbert, SG. (1990). Sensitive periods for lead-induced behavioral impairment (nonspatial discrimination 

reversal) in monkeys. Toxicol Appl Pharmacol 102: 101-109. http://dx.doi.org/10.1016/0041-

008X(90)90087-B.

Rice, DC. (1992). Effect of lead during different developmental periods in the monkey on concurrent discrimination 

performance. Neurotoxicology 13: 583-592. 

Rice, DC. (1996). Behavioral effects of lead: Commonalities between experimental and epidemiologic data. Environ 

Health Perspect 104: 337-351. http://dx.doi.org/10.1289/ehp.96104s2337.

Rice, DC. (1997). Effects of lifetime lead exposure in monkeys on detection of pure tones. Fundam Appl Toxicol 

36: 112-118. http://dx.doi.org/10.1006/faat.1996.2268.

Rice, DC. (1998). Effects of lifetime lead exposure on spatial and temporal visual function in monkeys. 

Neurotoxicology 19: 893-902. 

Richardt, G; Federolf, G; Habermann, E. (1986). Affinity of heavy metal ions to intracellular Ca2+-binding proteins. 

Biochem Pharmacol 35: 1331-1335. http://dx.doi.org/10.1016/0006-2952(86)90278-9.

Richetti, S; Rosemberg, D; Ventura-Lima, J; Monserrat, J; Bogo, M; Bonan, C. (2010). Acetylcholinesterase activity 

and antioxidant capacity of zebrafish brain is altered by heavy metal exposure. Neurotoxicology 32: 116-

122. http://dx.doi.org/10.1016/j.neuro.2010.11.001.

Riddell, TJ; Solon, O; Quimbo, SA; Tan, CM; Butrick, E; Peabody, JW. (2007). Elevated blood-lead levels among 

children living in the rural Philippines. Bull World Health Organ 85: 674-680. 

Ris, MD; Dietrich, KN; Succop, PA; Berger, OG; Bornschein, RL. (2004). Early exposure to lead and 

neuropsychological outcome in adolescence. J Int Neuropsychol Soc 10: 261-270. 

http://dx.doi.org/10.1017/S1355617704102154.

Rizzi, E; Castro, MM; Fernandes, K; Barbosa, F; Arisi, GM; Garcia-Cairasco, N; Bendhack, LM; Tanus-Santos, JE; 

Gerlach, RF. (2009). Evidence of early involvement of matrix metalloproteinase-2 in lead-induced 

hypertension. Arch Toxicol 83: 439-449. http://dx.doi.org/10.1007/s00204-008-0363-1.

Robins, JM; Cullen, MR; Connors, BB; Kayne, RD. (1983). Depressed thyroid indexes associated with occupational 

exposure to inorganic lead. Arch Intern Med 143: 220-224. http://dx.doi.org/10.1001/archinte.143.2.220.

Rodriguez-Iturbe, B; Vaziri, ND; Herrera-Acosta, J; Johnson, RJ. (2004). Oxidative stress, renal infiltration of 

immune cells, and salt-sensitive hypertension: All for one and one for all. Am J Physiol 286: F606-F616. 

http://dx.doi.org/10.1152/ajprenal.00269.2003.

Rodriguez-Iturbe, B; Sindhu, RK; Quiroz, Y; Vaziri, ND. (2005). Chronic exposure to low doses of lead results in 

renal infiltration of immune cells, NF-"kappa"B activation, and overexpression of tubulointerstitial 

angiotensin II. Antioxid Redox Signal 7: 1269-1274. http://dx.doi.org/10.1089/ars.2005.7.1269.

Roels, H; Lauwerys, R; Konings, J; Buchet, JP; Bernard, A; Green, S; Bradley, D; Morgan, W; Chettle, D. (1994). 

Renal function and hyperfiltration capacity in lead smelter workers with high bone lead. Occup Environ 

Med 51: 505-512. http://dx.doi.org/10.1136/oem.51.8.505.

Rojo-Tellez, M. (2008). Email from Martha Tellez Rojo, Direction of Evaluation of Programs and Quantitative 

Methods, Center of Investigation in Evaluation and Polls, National institute of Public Health to Jee-Young 

Kim, USEPA, Subject - Questions on the Tellez-Rojo et al. (2006) paper. (EPA-HQ-OAR-2006-0735-

5123). Rojo-Tellez, M. http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OAR-2006-0735-5123.

Roncal, C; Mu, W; Reungjui, S; Kim, KM; Henderson, GN; Ouyang, X; Nakagawa, T; Johnson, RJ. (2007). Lead, 

at low levels, accelerates arteriolopathy and tubulointerstitial injury in chronic kidney disease. Am J 

Physiol Renal Physiol 293: F1391-F1396. http://dx.doi.org/10.1152/ajprenal.00216.2007.
February 2012 5-608 Draft – Do Not Cite or Quote

Rondo, PH; Carvalho, MDF; Souza, MC; Moraes, F. (2006). Lead, hemoglobin, zinc protoporphyrin and ferritin 

concentrations in children. Rev Saude Publica 40: 71-76. http://dx.doi.org/10.1590/S0034-

89102006000100012.

Ronis, MJ; Badger, TM; Shema, SJ; Roberson, PK; Shaikh, F. (1998a). Effects on pubertal growth and reproduction 

in rats exposed to lead perinatally or continuously throughout development. J Toxicol Environ Health A 53: 

327-341. http://dx.doi.org/10.1080/009841098159312.

Ronis, MJJ; Badger, TM; Shema, SJ; Roberson, PK; Shaikh, F. (1996). Reproductive toxicity and growth effects in 

rats exposed to lead at different periods during development. Toxicol Appl Pharmacol 136: 361-371. 

http://dx.doi.org/10.1006/taap.1996.0044.

Ronis, MJJ; Gandy, J; Badger, T. (1998b). Endocrine mechanisms underlying reproductive toxicity in the 

developing rat chronically exposed to dietary lead. J Toxicol Environ Health A 54: 77-99. 

http://dx.doi.org/10.1080/009841098158935.

Ronis, MJJ; Aronson, J; Gao, GG; Hogue, W; Skinner, RA; Badger, TM; Lumpkin, CK, Jr. (2001). Skeletal effects 

of developmental lead exposure in rats. Toxicol Sci 62: 321-329. http://dx.doi.org/10.1093/toxsci/62.2.321.

Roshan, VD; Assali, M; Moghaddam, AH; Hosseinzadeh, M; Myers, J. (2011). Exercise training and antioxidants: 

effects on rat heart tissue exposed to lead acetate. Int J Toxicol 30: 190-196. 

http://dx.doi.org/10.1177/1091581810392809.

Rossi-George, A; Virgolini, MB; Weston, D; Cory-Slechta, DA. (2009). Alterations in glucocorticoid negative 

feedback following maternal Pb, prenatal stress and the combination: A potential biological unifying 

mechanism for their corresponding disease profiles. Toxicol Appl Pharmacol 234: 117-127. 

http://dx.doi.org/10.1016/j.taap.2008.10.003.

Rossi-George, A; Virgolini, M; Weston, D; Thiruchelvam, M; Cory-Slechta, D. (2011). Interactions of lifetime lead 

exposure and stress: Behavioral, neurochemical and HPA axis effects. Neurotoxicology 32: 83-99. 

http://dx.doi.org/10.1016/j.neuro.2010.09.004.

Rothenberg, SJ; Schnaas, L; Cansino-Ortiz, S; Perroni-Hernandez, E; De La Torre, P; Neri-Mendez, C; Ortega, P; 

Hidalgo-Loperena, H; Svendsgaard, D. (1989). Neurobehavioral deficits after low level lead exposure in 

neonates: The Mexico City pilot study. Neurotoxicol Teratol 11: 85-93. http://dx.doi.org/10.1016/0892-

0362(89)90046-9.

Rothenberg, SJ; Karchmer, S; Schnaas, L; Perroni, E; Zea, F; Alba, JF. (1994a). Changes in serial blood lead levels 

during pregnancy. Environ Health Perspect 102: 876-880. http://dx.doi.org/10.1289/ehp.94102876.

Rothenberg, SJ; Poblano, A; Garza-Morales, S. (1994b). Prenatal and perinatal low level lead exposure alters 

brainstem auditory evoked responses in infants. Neurotoxicology 15: 695-699. 

Rothenberg, SJ; Poblano, A; Schnaas, L. (2000). Brainstem auditory evoked response at five years and prenatal and 

postnatal blood lead. Neurotoxicol Teratol 22: 503-510. http://dx.doi.org/10.1016/S0892-0362(00)00079-9.

Rothenberg, SJ; Schnaas, L; Salgado-Valladares, M; Casanueva, E; Geller, AM; Hudnell, HK; Fox, DA. (2002a). 

Increased ERG a- and b-wave amplitudes in 7- to 10-year-old children resulting from prenatal lead 

exposure. Invest Ophthalmol Vis Sci 43: 2036-2044. 

Rothenberg, SJ; Rothenberg, JC. (2005). Testing the dose-response specification in epidemiology: Public health and 

policy consequences for lead. Environ Health Perspect 113: 1190-1195. 

http://dx.doi.org/10.1289/ehp.7691.

Rotruck, JT; Pope, AL; Ganther, HE; Swanson, AB; Hafeman, DG; Hoekstra, WG. (1973). Selenium: Biochemical 

role as a component of glutathione peroxidase. Science 179: 588-590. 

http://dx.doi.org/10.1126/science.179.4073.588.

Rousseau, MC; Straif, K; Siemiatycki, J. (2005). IARC carcinogen update. Environ Health Perspect 113: A580-

A581. http://dx.doi.org/10.1289/ehp.113-a580.

Rousseau, MC; Parent, ME; Nadon, L; Latrellie, B; Siemiatycki, J. (2007). Occupational exposure to lead 

compounds and risk of cancer among men: A population-based case-control study. Am J Epidemiol 166: 

1005-1014. http://dx.doi.org/10.1093/aje/kwm183.
February 2012 5-609 Draft – Do Not Cite or Quote

Roy, A; Bellinger, D; Hu, H; Schwartz, J; Ettinger, AS; Wright, RO; Bouchard, M; Palaniappan, K; Balakrishnan, 

K. (2009a). Lead exposure and behavior among young children in Chennai, India. Environ Health Perspect 

117: 1607-1611. http://dx.doi.org/10.1289/ehp.0900625.

Roy, A; Hu, H; Bellinger, DC; Mukherjee, B; Modali, R; Nasaruddin, K; Schwartz, J; Wright, RO; Ettinger, AS; 

Palaniapan, K; Balakrishnan, K. (2011). Hemoglobin, lead exposure, and intelligence quotient: Effect 

modification by the DRD2 Taq IA polymorphism. Environ Health Perspect 119: 144-149. 

http://dx.doi.org/10.1289/ehp.0901878.

Roy, AA; Baxla, SP; Gupta, T; Bandyopadhyaya, R; Tripathi, SN. (2009b). Particles emitted from indoor 

combustion sources: size distribution measurement and chemical analysis. Inhal Toxicol 21: 837-848. 

http://dx.doi.org/10.1080/08958370802538050.

Roy, AK; Dhir, H; Sharma, A. (1992). Modification of metal-induced micronuclei formation in mouse bone marrow 

erythrocytes by Phyllanthus fruit extract and ascorbic acid. Toxicol Lett 62: 9-17. 

http://dx.doi.org/10.1016/0378-4274(92)90072-R.

Roy, NK; Rossman, TG. (1992). Mutagenesis and comutagenesis by lead compounds. Mutat Res 298: 97-103. 

http://dx.doi.org/10.1016/0165-1218(92)90034-W.

Rubio, J; Riqueros, MI; Gasco, M; Yucra, S; Miranda, S; Gonzales, GF. (2006). Lepidium meyenii (Maca) reversed 

the lead acetate induced - Damage on reproductive function in male rats. Food Chem Toxicol 44: 1114-

1122. http://dx.doi.org/10.1016/j.fct.2006.01.007.

Rybicki, BA; Johnson, CC; Uman, J; Gorell, JM. (1993). Parkinson's disease mortality and the industrial use of 

heavy metals in Michigan. Mov Disord 8: 87-92. http://dx.doi.org/10.1002/mds.870080116.

Sá, I; da Costa, MJ; Cunha, EM. (In Press) Lead hepatotoxicology: A study in an animal model. Toxicol Ind Health. 

http://dx.doi.org/10.1177/0748233711407240.

Sabbioni, E; Marafante, E. (1976). Identification of lead-binding components in rat liver: In vivo study. Chem Biol 

Interact 15: 1-20. http://dx.doi.org/10.1016/0009-2797(76)90124-1.

Saimi, Y; Kung, C. (2002). Calmodulin as an ion channel subunit. Annu Rev Physiol 64: 289-311. 

http://dx.doi.org/10.1146/annurev.physiol.64.100301.111649.

Sainath, SB; Meena, R; Supriya, C, h; Reddy, KP; Reddy, PS. (2011). Protective role of Centella asiatica on leadinduced oxidative stress and suppressed reproductive health in male rats. Environ Toxicol Pharmacol 32: 

146-154. http://dx.doi.org/10.1016/j.etap.2011.04.005.

Sakai, T; Morita, Y. (1996). delta-Aminolevulinic acid in plasma or whole blood as a sensitive indicator of lead 

effects, and its relation to the other heme-related parameters. Int Arch Occup Environ Health 68: 126-132. 

http://dx.doi.org/10.1007/BF00381245.

Sakata, S; Shimizu, S; Ogoshi, K; Hirai, K; Ohno, Y; Kishi, T; Sherchand, JB; Utsumi, M; Shibata, M; Takaki, M; 

Ueda, M; Mori, I. (2007). Inverse relationship between serum erythropoietin and blood lead concentrations 

in Kathmandu tricycle taxi drivers. Int Arch Occup Environ Health 80: 342-345. 

http://dx.doi.org/10.1007/s00420-006-0125-4.

Sakuma, H; Kusama, M; Yamaguchi, K; Matsuki, T; Sugawara, S. (1984). The distribution of cigarette smoke 

components between mainstream and sidestream smoke: II. Bases. Beitraege zur Tabakforschung 

International 12: 199-209. 

Salawu, EO; Adeeyo, OA; Falokun, OP; Yusuf, UA; Oyerinde, A; Adeleke, AA. (2009). Tomato (Lycopersicon 

esculentum) prevents lead-induced testicular toxicity. Journal of Human Reproductive Sciences 2: 30-34. 

http://dx.doi.org/10.4103/0974-1208.51346.

Saldiva, PHN; Clarke, RW; Coull, BA; Stearns, RC; Lawrence, J; Krishna-Murthy, GG; Diaz, E; Koutrakis, P; Suh, 

H; Tsuda, A; Godleski, JJ. (2002). Lung inflammation induced by concentrated ambient air particles is 

related to particle composition. Am J Respir Crit Care Med 165: 1610-1617. 

http://dx.doi.org/10.1164/rccm.2106102.
February 2012 5-610 Draft – Do Not Cite or Quote

Saleh, M; Mathison, J; Wolinski, M; Bensinger, S; Fitzgerald, P; Droin, N; Ulevitch, R; Green, D; Nicholson, D.

(2006). Enhanced bacterial clearance and sepsis resistance in caspase-12-deficient mice. Nature 440: 1064-

1068. http://dx.doi.org/10.1038/nature04656.

Sanchez-Fructuoso, AI; Blanco, J; Cano, M; Ortega, L; Arroyo, M; Fernandez, C; Prats, D; Barrientos, A. (2002a). 

Experimental lead nephropathy: Treatment with calcium disodium ethylenediaminetetraacetate. Am J 

Kidney Dis 40: 59-67. http://dx.doi.org/10.1053/ajkd.2002.33936.

Sanchez-Fructuoso, AI; Cano, M; Arroyo, M; Fernandez, C; Prats, D; Barrientos, A. (2002b). Lead mobilization 

during calcium disodium ethylenediaminetetraacetate chelation therapy in treatment of chronic lead 

poisoning. Am J Kidney Dis 40: 51-58. http://dx.doi.org/10.1053/ajkd.2002.33913.

Sandhir, R; Julka, D; Gill, KD. (1994). Lipoperoxidative damage on lead exposure in rat brain and its implications 

on membrane bound enzymes. Pharmacol Toxicol 74: 66-71. http://dx.doi.org/10.1111/j.1600-

0773.1994.tb01077.x.

Sandhir, R; Gill, KD. (1995). Effect of lead on lipid peroxidation in liver of rats. Biol Trace Elem Res 48: 91-97. 

http://dx.doi.org/10.1007/BF02789081.

Sanna, E; Vallascas, E. (2011). Hair lead levels to evaluate the subclinical impact of lead on growth in Sardinian 

children (Italy). Am J Hum Biol 23: 740-746. http://dx.doi.org/10.1002/ajhb.21203.

Santibanez, M; Vioque, J; Alguacil, J; Barber, X; de la Hera, G; Kauppinen, T. (2008). Occupational exposures and 

risk of oesophageal cancer by histological type: a case-control study in eastern Spain. Occup Environ Med 

65: 774-781. http://dx.doi.org/10.1136/oem.2007.037929.

Santos, FW; Rocha, JBT; Nogueira, CW. (2006a). 2,3-dimercaptopropanol, 2,3-dimercaptopropane-1-sulfonic acid 

and meso-2,3-dimercaptosuccinic acid increase lead-induced inhibition of delta-aminolevulinate 

dehydratase in vitro and ex vivo. Toxicol In Vitro 20: 317-323. http://dx.doi.org/10.1016/j.tiv.2005.08.006.

Santos, MRV; Marchioro, M; Antoniolli, AR. (2006b). Lead effects on non-adrenergic non-cholinergic relaxations 

in the rat gastric fundus. Toxicol In Vitro 20: 38-42. http://dx.doi.org/10.1016/j.tiv.2005.05.005.

Saraiva, MCP; Taichman, RS; Braun, T; Nriagu, J; Eklund, SA; Burt, BA. (2007). Lead exposure and periodontitis 

in US adults. J Periodontal Res 42: 45-52. http://dx.doi.org/10.1111/j.1600-0765.2006.00913.x.

Sarasua, SM; Vogt, RF; Henderson, LO; Jones, PA; Lybarger, JA. (2000). Serum immunoglobulins and lymphocyte 

subset distributions in children and adults living in communities assessed for lead and cadmium exposure. J 

Toxicol Environ Health A 60: 1-15. http://dx.doi.org/10.1080/009841000156556.

Sarasua, SM; Mueller, P; Kathman, S; Campagna, D; Uddin, MS; White, MC. (2003). Confirming the utility of four 

kidney biomarker tests in a longitudinal follow-up study. Ren Fail 25: 797-817. 

Sarnat, JA; Marmur, A; Klein, M; Kim, E; Russell, AG; Sarnat, SE; Mulholland, JA; Hopke, PK; Tolbert, PE.

(2008). Fine particle sources and cardiorespiratory morbidity: An application of chemical mass balance and 

factor analytical source-apportionment methods. Environ Health Perspect 116: 459-466. 

http://dx.doi.org/10.1289/ehp.10873.

Savery, LC; Grlickova-Duzevik, E; Wise, SS; Thompson, WD; Hinz, JA; Thompson, LH; Wise, JP, Sr. (2007). Role 

of the Fancg gene in protecting cells from particulate chromate-induced chromosome instability. Mutat Res 

Genet Toxicol Environ Mutagen 626: 120-127. http://dx.doi.org/10.1016/j.mrgentox.2006.09.005.

Saxena, G; Flora, SJS. (2004). Lead-induced oxidative stress and hematological alterations and their response to 

combined administration of calcium disodium EDTA with a thiol chelator in rats. J Biochem Mol Toxicol 

18: 221-233. http://dx.doi.org/10.1002/jbt.20027.

Saxena, G; Pathak, U; Flora, SJS. (2005). Beneficial role of monoesters of meso-2,3-dimercaptosuccinic acid in the 

mobilization of lead and recovery of tissue oxidative injury in rats. Toxicology 214: 39-56. 

http://dx.doi.org/10.1016/j.tox.2005.05.026.

Schanne, FAX; Dowd, TL; Gupta, RK; Rosen, JF. (1989). Lead increases free Ca2+ concentration in cultured 

osteoblastic bone cells: simultaneous detection of intracellular free Pb2+ by 19F NMR. PNAS 86: 5133-

5135. 
February 2012 5-611 Draft – Do Not Cite or Quote

Schanne, FAX; Long, GJ; Rosen, JF. (1997). Lead induced rise in intracellular free calcium is mediated through 

activation of protein kinase C in osteoblastic bone cells. Biochim Biophys Acta 136: 247-254. 

http://dx.doi.org/10.1016/S0925-4439(97)00006-9.

Schell, LM; Denham, M; Stark, AD; Gomez, M; Ravenscroft, J; Parsons, PJ; Aydermir, A; Samelson, R. (2003). 

Maternal blood lead concentration, diet during pregnancy, and anthropometry predict neonatal blood lead 

in a socioeconomically disadvantaged population. Environ Health Perspect 111: 195-200. 

http://dx.doi.org/10.1289/ehp.5592.

Schell, LM; Denham, M; Stark, AD; Parsons, PJ; Schulte, EE. (2009). Growth of infants' length, weight, head and 

arm circumferences in relation to low levels of blood lead measured serially. Am J Hum Biol 21: 180-187. 

http://dx.doi.org/10.1002/ajhb.20842.

Schirrmacher, K; Wiemann, M; Bingmann, D; Busselberg, D. (1998). Effects of lead, mercury, and methyl mercury 

on gap junctions and [CA2+]i in bone cells. Calcif Tissue Int 63: 134-139. 

http://dx.doi.org/10.1007/s002239900503.

Schnaas, L; Rothenberg, SJ; Flores, MF; Martinez, S; Hernandez, C; Osorio, E; Velasco, SR; Perroni, E. (2006). 

Reduced intellectual development in children with prenatal lead exposure. Environ Health Perspect 114: 

791-797. http://dx.doi.org/10.1289/ehp.8552.

Schneider, JS; Anderson, DW; Wade, TV; Smith, MG; Leibrandt, P; Zuck, L; Lidsky, TI. (2005). Inhibition of 

progenitor cell proliferation in the dentate gyrus of rats following post-weaning lead exposure. 

Neurotoxicology 26: 141-145. http://dx.doi.org/10.1016/j.neuro.2004.06.006.

Schneider, ML; Moore, CF; Gajewski, LL; Larson, JA; Roberts, AD; Converse, AK; DeJesus, OT. (2008). Sensory 

processing disorder in a primate model: Evidence from a longitudinal study of prenatal alcohol and prenatal 

stress effects. Child Dev 79: 100-113. http://dx.doi.org/10.1111/j.1467-8624.2007.01113.x.

Schober, SE; Mirel, LB; Graubard, BI; Brody, DJ; Flegal, KM. (2006). Blood lead levels and death from all causes, 

cardiovascular disease, and cancer: Results from the NHANES III Mortality Study. Environ Health 

Perspect 114: 1538-1541. http://dx.doi.org/10.1289/ehp.9123.

Schrauzer, GN. (1987). Effects of selenium antagonists on cancer susceptibility: New aspects of chronic heavy 

metal toxicity. J UOEH 9: 208-215. 

Schrauzer, GN. (2008). Effects of selenium and low levels of lead on mammary tumor development and growth in 

MMTV-infected female mice. Biol Trace Elem Res 125: 268-275. http://dx.doi.org/10.1007/s12011-008-

8172-1.

Schwark, WS; Haluska, M; Blackshear, P; Magana, T. (1985). Lifetime lead intoxication: influence on the 

amygdaloid kindling model of epileptogenesis. Toxicology 36: 49-60. http://dx.doi.org/10.1016/0300-

483X(85)90006-X.

Schwartz, BS; Lee, BK; Stewart, W; Ahn, KD; Kelsey, K; Bresssler, J. (1997a). Associations of subtypes of 

hemoglobin with delta-aminolevulinic acid dehydratase genotype and dimercaptosuccinic acid-chelatable 

lead levels. Arch Environ Occup Health 52: 97-103. http://dx.doi.org/10.1080/00039899709602871.

Schwartz, BS; Lee, BK; Stewart, W; Sithisarankul, P; Strickland, PT; Ahn, KD; Kelsey, K. (1997b). deltaAminolevulinic acid dehydratase genotype modifies four hour urinary lead excretion after oral 

administration of dimercaptosuccinic acid. Occup Environ Med 54: 241-246. 

Schwartz, BS; Lee, BK; Lee, GS; Stewart, WF; Simon, D; Kelsey, K; Todd, AC. (2000a). Associations of blood 

lead, dimercaptosuccinic acid-chelatable lead, and tibia lead with polymorphisms in the vitamin D receptor 

and delta-aminolevulinic acid dehydratase genes. Environ Health Perspect 108: 949-954. 

http://dx.doi.org/10.2307/3435053.

Schwartz, BS; Lee, BK; Lee, GS; Stewart, WF; Lee, SS; Hwang, KY; Ahn, KD; Kim, YB; Bolla, KI; Simon, D; 

Parsons, PJ; Todd, AC. (2001). Associations of blood lead, dimercaptosuccinic acid-chelatable lead, and 

tibia lead with neurobehavioral test scores in South Korean lead workers. Am J Epidemiol 153: 453-464. 

http://dx.doi.org/10.1093/aje/153.5.453.
February 2012 5-612 Draft – Do Not Cite or Quote

Schwartz, BS; Lee, BK; Bandeen-Roche, K; Stewart, W; Bolla, K; Links, J; Weaver, V; Todd, A. (2005). 

Occupational lead exposure and longitudinal decline in neurobehavioral test scores. Epidemiology 16: 106-

113. http://dx.doi.org/10.1097/01.ede.0000147109.62324.51.

Schwartz, BS; Chen, S; Caffo, B; Stewart, WF; Bolla, KI; Yousem, D; Davatzikos, C. (2007). Relations of brain 

volumes with cognitive function in males 45 years and older with past lead exposure. Neuroimage 37: 633-

641. http://dx.doi.org/10.1016/j.neuroimage.2007.05.035.

Schwartz, J; Otto, D. (1987). Blood lead, hearing thresholds, and neurobehavioral development in children and 

youth. Arch Environ Occup Health 42: 153-160. http://dx.doi.org/10.1080/00039896.1987.9935814.

Schwartz, J; Otto, D. (1991). Lead and minor hearing impairment. Arch Environ Occup Health 46: 300-305. 

http://dx.doi.org/10.1080/00039896.1991.9934391.

Schwartz, J. (1991). Lead, blood pressure, and cardiovascular disease in men and women. Environ Health Perspect 

91: 71-75. http://dx.doi.org/10.1289/ehp.919171.

Schwartz, J. (1994). Low-level lead exposure and children's IQ: A meta-analysis and search for a threshold. Environ 

Res 65: 42-55. http://dx.doi.org/10.1006/enrs.1994.1020.

Schweitzer, JB; Faber, TL; Grafton, ST; Tune, LE; Hoffman, JM; Kilts, CD. (2000). Alterations in the functional 

anatomy of working memory in adult attention deficit hyperactivity disorder. Am J Psychiatry 157: 278-

280. 

Sciarillo, WG; Alexander, G; Farrell, KP. (1992). Lead exposure and child behavior. Am J Public Health 82: 1356-

1360. 

Scinicariello, F; Murray, HE; Moffett, DB; Abadin, HG; Sexton, MJ; Fowler, BA. (2007). Lead and deltaaminolevulinic acid dehydratase polymorphism: Where does it lead? A meta-analysis. Environ Health 

Perspect 115: 35-41. http://dx.doi.org/10.1289/ehp.9448.

Scinicariello, F; Yesupriya, A; Chang, MH; Fowler, BA. (2010). Modification by ALAD of the association between 

blood lead and blood pressure in the U.S. population: Results from the Third National Health and Nutrition 

Examination Survey. Environ Health Perspect 118: 259-264. http://dx.doi.org/10.1289/ehp.0900866.

Seki, T; Arai, Y. (1993). Highly polysialylated NCAM expression in the developing and adult rat spinal cord 

[Letter]. Dev Brain Res 73: 141-145. http://dx.doi.org/10.1016/0165-3806(93)90056-G.

Selevan, SG; Rice, DC; Hogan, KA; Euling, SY; Pfahles-Hutchens, A; Bethel, J. (2003). Blood lead concentration 

and delayed puberty in girls. N Engl J Med 348: 1527-1536. http://dx.doi.org/10.1056/NEJMoa020880.

Selvin-Testa, A; Lopez-Costa, JJ; Nessi-de Avinon, AC; Saavedra, JP. (1991). Astroglial alterations in rat 

hippocampus during chronic lead exposure. Glia 4: 384-392. http://dx.doi.org/10.1002/glia.440040406.

Senger, MR; Rico, EP; Arizi, MDB; Frazzon, APG; Dias, RD; Bogo, MR; Bonan, CD. (2006). Exposure to Hg2+ 

and Pb2+ changes NTPDase and ecto-5 '-nucleotidase activities in central nervous system of zebrafish 

(Danio rerio). Toxicology 226: 229-237. http://dx.doi.org/10.1016/j.tox.2006.07.012.

Shabani, A; Rabbani, A. (2000). Lead nitrate induced apoptosis in alveolar macrophages from rat lung. Toxicology 

149: 109-114. http://dx.doi.org/10.1016/S0300-483X(00)00232-8.

Shadick, NA; Kim, R; Weiss, S; Liang, MH; Sparrow, D; Hu, H. (2000). Effect of low level lead exposure on 

hyperuricemia and gout among middle aged and elderly men: the normative aging study. J Rheumatol 27: 

1708-1712. 

Shah, F; Kazi, TG; Afridi, HI; Baig, JA; Khan, S; Kolachi, NF; Wadhwa, SK; Shah, AQ. (2010). Environmental 

exposure of lead and iron deficit anemia in children age ranged 1-5 years: A cross sectional study. Sci Total 

Environ 408: 5325-5330. http://dx.doi.org/10.1016/j.scitotenv.2010.07.091.

Shaik, AP; Jamil, K. (2009). Individual susceptibility and genotoxicity in workers exposed to hazardous materials 

like lead. J Hazard Mater 168: 918-924. http://dx.doi.org/10.1016/j.jhazmat.2009.02.129.
February 2012 5-613 Draft – Do Not Cite or Quote

Shakoor, A; Gupta, PK; Singh, YP; Kataria, M. (2000). Beneficial effects of aluminum on the progression of leadinduced nephropathy in rats. Basic Clin Pharmacol Toxicol 87: 258-260. http://dx.doi.org/10.1034/j.1600-

0773.2000.pto870603.x.

Shan, G; Tang, T; Zhang, X. (2009). The protective effect of ascorbic acid and thiamine supplementation against 

damage caused by lead in the testes of mice. J Huazhong Univ Sci Technolog Med Sci 29: 68-72. 

http://dx.doi.org/10.1007/s11596-009-0114-4.

Sharifi, AM; Darabi, R; Akbarloo, N; Larijani, B; Khoshbaten, A. (2004). Investigation of circulatory and tissue 

ACE activity during development of lead-induced hypertension. Toxicol Lett 153: 233-238. 

http://dx.doi.org/10.1016/j.toxlet.2004.04.013.

Sharifi, AM; Mousavi, SH. (2008). Studying the effects of lead on DNA fragmentation and proapoptotic bax and 

antiapoptotic Bcl-2 protein expression in PC12 cells. Toxicol Mech Meth 18: 75-79. 

http://dx.doi.org/10.1080/15376510701665814.

Sharifi, AM; Mousavi, SH; Jorjani, M. (2010). Effect of chronic lead exposure on pro-apoptotic Bax and antiapoptotic Bcl-2 protein expression in rat hippocampus in vivo. Cell Mol Neurobiol 30: 769-774. 

http://dx.doi.org/10.1007/s10571-010-9504-1.

Sharma, A; Sharma, V; Kansal, L. (2010a). Amelioration of lead-induced hepatotoxicity by Allium sativum extracts 

in Swiss albino mice. 5: 4621-4630. http://dx.doi.org/10.4176/091107.

Sharma, V; Sharma, A; Kansal, L. (2010b). The effect of oral administration of Allium sativum extracts on lead 

nitrate induced toxicity in male mice. Food Chem Toxicol 48: 928-936. 

http://dx.doi.org/10.1016/j.fct.2010.01.002.

Shelkovnikov, SA; Gonick, HC. (2001). Influence of lead on rat thoracic aorta contraction and relaxation. Am J 

Hypertens 14: 873-878. http://dx.doi.org/10.1016/S0895-7061(01)02149-5.

Shelton, KR; Egle, PM. (1982). The proteins of lead-induced intranuclear inclusion bodies. J Biol Chem 257: 

11802-11807. 

Shelton, KR; Cunningham, JG; Klann, E; Merchant, RE; Egle, PM; Bigbee, JW. (1990). Low-abundance 32-

kilodalton nuclear protein specifically enriched in the central nervous system. J Neurosci Res 25: 287-294. 

http://dx.doi.org/10.1002/jnr.490250304.

Shen, XM; Yan, CH; Guo, D; Wu, SM; Li, RQ; Huang, H; Ao, LM; Zhou, JD; Hong, ZY; Xu, JD; Jin, XM; Tang, 

JM. (1998). Low-level prenatal lead exposure and neurobehavioral development of children in the first year 

of life: A prospective study in Shanghai. Environ Res 79: 1-8. http://dx.doi.org/10.1006/enrs.1998.3851.

Shih, RA; Glass, TA; Bandeen-Roche, K; Carlson, MC; Bolla, KI; Todd, AC; Schwartz, BS. (2006). Environmental 

lead exposure and cognitive function in community-dwelling older adults. Neurology 67: 1556-1562. 

http://dx.doi.org/10.1212/01.wnl.0000239836.26142.c5.

Shin, JH; Lim, KM; Noh, JY; Bae, ON; Chung, SM; Lee, MY; Chung, JH. (2007). Lead-induced procoagulant 

activation of erythrocytes through phosphatidylserine exposure may lead to thrombotic diseases. Chem Res 

Toxicol 20: 38-43. http://dx.doi.org/10.1021/tx060114+.

Shinkai, Y; Yamamoto, C; Kaji, T. (2010). Lead induces the expression of ER chaperones GRP78 and GRP94 in 

vascular endothelial cells via the JNK-AP-1 pathway. Toxicol Sci 114: 378-386. 

http://dx.doi.org/10.1093/toxsci/kfq008.

Shinozuka, H; Ohmura, T; Katyal, SL; Zedda, AI; Ledda-Columbano, GM; Columbano, A. (1996). Possible roles of 

nonparenchymal cells in hepatocyte proliferation induced by lead nitrate and by tumor necrosis factor 

alpha. Hepatology 23: 1572-1577. http://dx.doi.org/10.1002/hep.510230638.

Siddiqui, MKJ; Jyoti; Singh, S; Mehrotra, PK; Singh, K; Sarangi, R. (2006). Comparison of some trace elements 

concentration in blood, tumor free breast and tumor tissues of women with benign and malignant breast 

lesions: An Indian study. Environ Int 32: 630-637. http://dx.doi.org/10.1016/j.envint.2006.02.002.

Silberstein, T; Saphier, O; Paz-Tal, O; Trimarchi, JR; Gonzalez, L; Keefe, DL. (2006). Lead concentrates in ovarian 

follicle compromises pregnancy. J Trace Elem Med Biol 20: 205-207. 

http://dx.doi.org/10.1016/j.jtemb.2006.05.001.
February 2012 5-614 Draft – Do Not Cite or Quote

Silkin, YA; Silkina, EN; Sherstobitov, AO; Gusev, GP. (2001). Activation of potassium channels in erythrocytes of 

marine teleost Scorpaena porcus. Membr Cell Biol 14: 773-782. 

Silveira, EA; Lizardo, JH; Souza, LP; Stefanon, I; Vassallo, DV. (2010). Acute lead-induced vasoconstriction in the 

vascular beds of isolated perfused rat tails is endothelium-dependent. Braz J Med Biol Res 43: 492-499. 

http://dx.doi.org/10.1590/S0100-879X2010007500027.

Simões, MR; Ribeiro Júnior, RF; Vescovi, MV; de Jesus, HC; Padilha, AS; Stefanon, I; Vassallo, DV; Salaices, M; 

Fioresi, M. (2011). Acute lead exposure increases arterial pressure: role of the Renin-Angiotensin system. 

PLoS ONE 6: e18730. http://dx.doi.org/10.1371/journal.pone.0018730.

Simons, TJB. (1986). Passive transport and binding of lead by human red blood cells. J Physiol 378: 267-286. 

Simons, TJB. (1993a). Lead-calcium interactions in cellular lead toxicity. Neurotoxicology 14: 77-85. 

Simons, TJB. (1993b). Lead transport and binding by human erythrocytes in vitro. Pflugers Arch 423: 307-313. 

http://dx.doi.org/10.1007/BF00374410.

Simons, TJB. (1995). The affinity of human erythrocyte porphobilinogen synthase for Zn2+ and Pb2+. FEBS J 234: 

178-183. http://dx.doi.org/10.1111/j.1432-1033.1995.178_c.x.

Simsek, N; Karadeniz, A; Kalkan, Y; Keles, ON; Unal, B. (2009). Spirulina platensis feeding inhibited the anemiaand leucopenia-induced lead and cadmium in rats. J Hazard Mater 164: 1304-1309. 

http://dx.doi.org/10.1016/j.jhazmat.2008.09.041.

Singh, B; Chandran, V; Bandhu, HK; Mittal, BR; Bhattacharya, A; Jindal, SK; Varma, S. (2000). Impact of lead 

exposure on pituitary-thyroid axis in humans. Biometals 13: 187-192. 

http://dx.doi.org/10.1023/A:1009201426184.

Singh, J; Pritchard, DE; Carlisle, DL; Mclean, JA; Montaser, A; Orenstein, JM; Patierno, SR. (1999). Internalization 

of carcinogenic lead chromate particles by cultured normal human lung epithelial cells: Formation of 

intracellular lead-inclusion bodies and induction of apoptosis. Toxicol Appl Pharmacol 161: 240-248. 

http://dx.doi.org/10.1006/taap.1999.8816.

Singh, US; Saxena, DK; Singh, C; Murthy, RC; Chandra, SV. (1991). Lead-induced fetal nephrotoxicity in irondeficient rats. Reprod Toxicol 5: 211-217. http://dx.doi.org/10.1016/0890-6238(91)90053-I.

Sivaprasad, R; Nagaraj, M; Varalakshmi, P. (2003). Combined efficacies of lipoic acid and meso-2,3-

dimercaptosuccinic acid on lead-induced erythrocyte membrane lipid peroxidation and antioxidant status in 

rats. Hum Exp Toxicol 22: 183-192. http://dx.doi.org/10.1191/0960327103ht335oa.

Skoczynska, A; Stojek, E. (2005). The impact of subchronic lead poisoning on the vascular effect of nitric oxide in 

rats. Environ Toxicol Pharmacol 19: 99-106. http://dx.doi.org/10.1016/j.etap.2004.05.004.

Slivkova, J; Popelkova, M; Massanyi, P; Toporcerova, S; Stawarz, R; Formicki, G; Lukac, N; Putaa, A; Guzik, M.

(2009). Concentration of trace elements in human semen and relation to spermatozoa quality. J Environ Sci 

Health A Tox Hazard Subst Environ Eng 44: 370-375. http://dx.doi.org/10.1080/10934520802659729.

Smejkalova, J. (1990). The chromosomal aberrations investigation in children permanently living in the lead 

polluted area. Sb Ved Pr Fak Karlovy Univ Hradci Kralovea 33: 539-564. 

Smith, CM; Hu, H; Wang, X; Kelsey, KT. (1995a). ALA-D genotype is not associated with HT or HB levels among 

workers exposed to low levels of lead. Med Lav 86: 229-235. 

Smith, CM; Wang, X; Hu, H; Kelsey, KT. (1995b). A polymorphism in the delta-aminolevulinic acid dehydratase 

gene may modify the pharmacokinetics and toxicity of lead. Environ Health Perspect 103: 248-253. 

http://dx.doi.org/10.1289/ehp.95103248.

Smith, DR; Kahng, MW; Quintanilla-Vega, B; Fowler, BA. (1998). High-affinity renal lead-binding proteins in 

environmentally-exposed humans. Chem Biol Interact 115: 39-52. http://dx.doi.org/10.1016/S0009-

2797(98)00060-X.
February 2012 5-615 Draft – Do Not Cite or Quote

Snyder, JE; Filipov, NM; Parsons, PJ; Lawrence, DA. (2000). The efficiency of maternal transfer of lead and its 

influence on plasma IgE and splenic cellularity of mice. Toxicol Sci 57: 87-94. 

http://dx.doi.org/10.1093/toxsci/57.1.87.

Snyder, RD; Lachmann, PJ. (1989). Thiol involvement in the inhibition of DNA repair by metals in mammalian 

cells. Mol Toxicol 2: 117-128. 

Sobekova, A; Holovska, K; Lenartova, V; Legath, J; Javorsky, P. (2009). The alteration of glutathione peroxidase 

activity in rat organs after lead exposure. Acta Physiol Hung 96: 37-44. 

http://dx.doi.org/10.1556/APhysiol.96.2009.1.4.

Sokol, RZ; Madding, CE; Swerdloff, RS. (1985). Lead toxicity and the hypothalamic-pituitary-testicular axis. Biol 

Reprod 33: 722-728. http://dx.doi.org/10.1095/biolreprod33.3.722.

Sokol, RZ. (1987). Hormonal effects of lead acetate in the male rat: mechanism of action. Biol Reprod 37: 1135-

1138. http://dx.doi.org/10.1095/biolreprod37.5.1135.

Sokol, RZ; Berman, N. (1991). The effect of age of exposure on lead-induced testicular toxicity. Toxicology 69: 

269-278. http://dx.doi.org/10.1016/0300-483X(91)90186-5.

Solon, O; Riddell, TJ; Quimbo, SA; Butrick, E; Aylward, GP; Lou Bacate, M; Peabody, JW. (2008). Associations 

between cognitive function, blood lead concentration, and nutrition among children in the central 

Philippines. J Pediatr 152: 237-243. http://dx.doi.org/10.1016/j.jpeds.2007.09.008.

Songdej, N; Winters, PC; McCabe, MJ, Jr; Wijngaarden, EV. (2010). A population-based assessment of blood lead 

levels in relation to inflammation. Environ Res 110: 272-277. 

http://dx.doi.org/10.1016/j.envres.2009.12.008.

Spector, JT; Navas-Acien, A; Fadrowski, J; Guallar, E; Jaar, B; Weaver, VM. (2011). Associations of blood lead 

with estimated glomerular filtration rate using MDRD, CKD-EPI and serum cystatin C-based equations. 

Nephrol Dial Transplant 26: 2786-2792. http://dx.doi.org/10.1093/ndt/gfq773.

Srivastava, D; Hurwitz, RL; Fox, DA. (1995). Lead- and calcium-mediated inhibition of bovine rod cGMP 

phosphodiesterase: Interactions with magnesium. Toxicol Appl Pharmacol 134: 43-52. 

http://dx.doi.org/10.1006/taap.1995.1167.

Srivastava, V; Dearth, RK; Hiney, JK; Ramirez, LM; Bratton, GR; Dees, W. (2004). The effects of low-level Pb on 

steroidogenic acute regulatory protein (StAR) in the prepubertal rat ovary. Toxicol Sci 77: 35-40. 

http://dx.doi.org/10.1093/toxsci/kfg249.

Stacchiotti, A; Morandini, F; Bettoni, F; Schena, I; Lavazza, A; Grigolato, PG; Apostoli, P; Rezzani, R; Aleo, MF.

(2009). Stress proteins and oxidative, damage in a renal derived cell line exposed to inorganic mercury and 

lead. Toxicology 264: 215-224. http://dx.doi.org/10.1016/j.tox.2009.08.014.

Stackpole, MM; Wise, SS; Goodale, BC; Duzevik, EG; Munroe, RC; Thompson, WD; Thacker, J; Thompson, LH; 

Hinz, JM; Wise, JP. (2007). Homologous recombination repair protects against particulate chromateinduced chromosome instability in Chinese hamster cells. Mutat Res-Fundam Mol Mech Mutagen 625: 

145-154. http://dx.doi.org/10.1016/j.mrfmmm.2007.06.003.

Staessen, JA; Lauwerys, RR; Buchet, JP; Bulpitt, CJ; Rondia, D; Van Renterghem, Y; Amery, A. (1992). 

Impairment of renal function with increasing blood lead concentrations in the general population. N Engl J 

Med 327: 151-156. http://dx.doi.org/10.1056/NEJM199207163270303.

Staessen, JA; Nawrot, T; Den Hond, E; Thijs, L; Fagard, R; Hoppenbrouwers, K; Koppen, G; Nelen, V; Schoeters, 

G; Vanderschueren, D; Van Hecke, E; Verschaeve, L; Vlietinck, R; Roels, HA. (2001). Renal function, 

cytogenetic measurements, and sexual developments in adolescents in relation to environmental pollutants: 

A feasibility study of biomarkers. Lancet 357: 1660-1669. http://dx.doi.org/10.1016/S0140-

6736(00)04822-4.

Stangle, DE; Smith, DR; Beaudin, SA; Strawderman, MS; Levitsky, DA; Strupp, BJ. (2007). Succimer chelation 

improves learning, attention, and arousal regulation in lead-exposed rats but produces lasting cognitive 

impairment in the absence of lead exposure. Environ Health Perspect 115: 201-209. 

http://dx.doi.org/10.1289/ehp.9263.
February 2012 5-616 Draft – Do Not Cite or Quote

Stayner, L; Steenland, K; Dosemeci, M; Hertz-Picciotto, I. (2003). Attenuation of exposure-response curves in 

occupational cohort studies at high exposure levels. Scand J Work Environ Health 29: 317-324. 

Steenland, NK; Thun, MJ; Ferguson, CW; Port, FK. (1990). Occupational and other exposures associated with male 

end-stage renal disease: A case/control study. Am J Public Health 80: 153-157. 

Steinthorsdottir, SD; Eliasdottir, SB; Indridason, OS; Agustsdottir, IM; Palsson, R; Edvardsson, VO. (2011). 

Prevalence of hypertension in 9- to 10-year-old icelandic school children. J Clin Hypertens (Greenwich) 

13: 774-779. http://dx.doi.org/10.1111/j.1751-7176.2011.00496.x.

Sterling, G; O'Neill, K; McCafferty, M; O'Neill, J. (1982). Effect of chronic lead ingestion by rats on glucose 

metabolism and acetylcholine synthesis in cerebral cortex slices. J Neurochem 39: 592-596. 

http://dx.doi.org/10.1111/j.1471-4159.1982.tb03989.x.

Stevens, LA; Coresh, J; Schmid, CH; Feldman, HI; Froissart, M; Kusek, J; Rossert, J; Van Lente, F; Bruce, RD; 

Zhang, YP; Greene, T; Levey, AS. (2008). Estimating GFR using serum cystatin C alone and in 

combination with serum creatinine: A pooled analysis of 3,418 individuals with CKD. Am J Kidney Dis 

51: 395-406. http://dx.doi.org/10.1053/j.ajkd.2007.11.018.

Stewart, PW; Blaine, C; Cohen, M; Burright, RG; Donovick, PJ. (1996). Acute and longer term effects of meso-2,3 

dimercaptosuccinic acid (DMSA) on the behavior of lead-exposed and control mice. Physiol Behav 59: 

849-855. http://dx.doi.org/10.1016/0031-9384(95)02185-X.

Stewart, WF; Schwartz, BS; Simon, D; Kelsey, K; Todd, AC. (2002). ApoE genotype, past adult lead exposure, and 

neurobehavioral function. Environ Health Perspect 110: 501-505. http://dx.doi.org/10.1289/ehp.02110501.

Stewart, WF; Schwartz, BS; Davatzikos, C; Shen, D; Liu, D; Wu, X; Todd, AC; Shi, W; Bassett, S; Youssem, D.

(2006). Past adult lead exposure is linked to neurodegeneration measured by brain MRI. Neurology 66: 

1476-1484. http://dx.doi.org/10.1212/01.wnl.0000216138.69777.15.

Stiles, KM; Bellinger, DC. (1993). Neuropsychological correlates of low-level lead exposure in school-age children: 

a prospective study. Neurotoxicol Teratol 15: 27-35. 

Stoleski, S; Karadzinska-Bislimovska, J; Stikova, E; Risteska-Kuc, S; Mijakoski, D; Minov, J. (2008). Adverse 

effects in workers exposed to inorganic lead. Arh Hig Rada Toksikol 59: 19-29. 

http://dx.doi.org/10.2478/10004-1254-59-2008-1866.

Strömberg, U; Schütz, A; Skerfving, S. (1995). Substantial decrease of blood lead in Swedish children, 1978-94, 

associated with petrol lead. Occup Environ Med 52: 764-769. http://dx.doi.org/10.1136/oem.52.11.764.

Struzynska, L; Dabrowska-Bouta, B; Rafałowska, U. (1997a). Acute lead toxicity and energy metabolism in rat 

brain synaptosomes. Acta Neurobiol Exp (Wars) 57: 275-281. 

Struzynska, L; Walski, M; Gadamski, R; Dabrowska-Bouta, B; Rafalowska, U. (1997b). Lead-induced 

abnormalities in blood-brain barrier permeability in experimental chronic toxicity. Mol Chem Neuropathol 

31: 207-224. http://dx.doi.org/10.1007/BF02815125.

Struzynska, L; Dabrowska-Bouta, B; Koza, K; Sulkowski, G. (2007). Inflammation-like glial response in leadexposed immature rat brain. Toxicol Sci 95: 156-162. http://dx.doi.org/10.1093/toxsci/kfl134.

Su, Y; Ding, Y; Jiang, M; Hu, X; Zhang, Z. (2007). Protein 4.2 Komatsu (D175Y) associated with the lack of 

interaction with ankyrin in human red blood cells. Blood Cells Mol Dis 38: 221-228. 

http://dx.doi.org/10.1016/j.bcmd.2006.11.004.

Sun, H; Wang, HL; Wang, S. (2007). D-serine relieves chronic lead exposure-impaired long-term potentiation in the 

CA1 region of the rat hippocampus in vitro. Neurosci Lett 417: 118-122. 

http://dx.doi.org/10.1016/j.neulet.2007.01.085.

Sun, L; Hu, J; Zhao, Z; Li, L; Cheng, H. (2003). Influence of exposure to environmental lead on serum 

immunoglobulin in preschool children. Environ Res 92: 124-128. http://dx.doi.org/10.1016/S0013-

9351(02)00090-7.

Sun, LR; Suszkiw, JB. (1995). Extracellular inhibition and intracellular enhancement of Ca2+ currents by Pb2+ in 

bovine adrenal chromaffin cells. J Neurophysiol 74: 574-581. 
February 2012 5-617 Draft – Do Not Cite or Quote

Sun, Y; Sun, DH; Zhou, ZJ; Zhu, GY; Lei, LJ; Zhang, HY; Chang, XL; Jin, TY. (2008a). Estimation of benchmark 

dose for bone damage and renal dysfunction in a Chinese male population occupationally exposed to lead. 

Ann Occup Hyg 52: 527-533. http://dx.doi.org/10.1093/annhyg/men031.

Sun, Y; Sun, DH; Zhou, ZJ; Zhu, GY; Zhang, HY; Chang, XL; Lei, LJ; Jin, TY. (2008b). Osteoporosis in a Chinese 

population due to occupational exposure to lead. Am J Ind Med 51: 436-442. 

http://dx.doi.org/10.1002/ajim.20567.

Sundstrom, R; Muntzing, K; Kalimo, H; Sourander, P. (1985). Changes in the integrity of the blood-brain barrier in 

suckling rats with low dose lead encephalopathy. Acta Neuropathol 68: 1-9. 

http://dx.doi.org/10.1007/BF00688948.

Surkan, PJ; Zhang, A; Trachtenberg, F; Daniel, DB; McKinlay, S; Bellinger, DC. (2007). Neuropsychological 

function in children with blood lead levels <10 μg/dL. Neurotoxicology 28: 1170-1177. 

http://dx.doi.org/10.1016/j.neuro.2007.07.007.

Surkan, PJ; Schnaas, L; Wright, RJ; Téllez-Rojo, MM; Lamadrid-Figueroa, H; Hu, H; Hernández-Avila, M; 

Bellinger, DC; Schwartz, J; Perroni, E; Wright, RO. (2008). Maternal self-esteem, exposure to lead, and 

child neurodevelopment. Neurotoxicology 29: 278-285. http://dx.doi.org/10.1016/j.neuro.2007.11.006.

Suszkiw, J; Toth, G; Murawsky, M; Cooper, GP. (1984). Effects of Pb2+ and Cd2+ on acetylcholine release and 

Ca2+ movements in synaptosomes and subcellular fractions from rat brain and Torpedo electric organ. 

Brain Res 323: 31-46. http://dx.doi.org/10.1016/0006-8993(84)90262-2.

Suszkiw, JB. (2004). Presynaptic disruption of transmitter release by lead. Neurotoxicology 25: 599-604. 

http://dx.doi.org/10.1016/j.neuro.2003.09.009.

Suzuki, T; Morimura, S; Diccianni, MB; Yamada, R; Hochi, SI; Hirabayashi, M; Yuki, A; NomuraK; Kitagawa, T; 

Imagawa, M; Muramatsu, M. (1996). Activation of glutathione transferase P gene by lead requires 

glutathione transferase P enhancer I. J Biol Chem 271: 1626-1632. 

http://dx.doi.org/10.1074/jbc.271.3.1626.

Swarup, D; Naresh, R; Varshney, VP; Balagangatharathilagar, M; Kumar, P; Nandi, D; Patra, RC. (2007). Changes 

in plasma hormones profile and liver function in cows naturally exposed to lead and cadmium around 

different industrial areas. Res Vet Sci 82: 16-21. http://dx.doi.org/10.1016/j.rvsc.2006.05.002.

Taketani, S; Tanaka, A; Tokunaga, R. (1985). Reconstitution of heme-synthesizing activity from ferric ion and 

porphyrins, and the effect of lead on the activity. Arch Biochem Biophys 242: 291-296. 

http://dx.doi.org/10.1016/0003-9861(85)90504-1.

Tanner, CM; Chen, B; Wang, W; Peng, M; Liu, Z; Liang, X; Kao, LC; Gilley, DW; Goetz, CG; Schoenberg, BS.

(1989). Environmental factors and Parkinson's disease: A case-control study in China. Neurology 39: 660-

664. 

Tapisso, JT; Marques, CC; Mathias, ML; Ramalhinho, MG. (2009). Induction of micronuclei and sister chromatid 

exchange in bone-marrow cells and abnormalities in sperm of Algerian mice (Mus spretus) exposed to 

cadmium, lead and zinc. Mutat Res Genet Toxicol Environ Mutagen 678: 59-64. 

http://dx.doi.org/10.1016/j.mrgentox.2009.07.001.

Tarantini, L; Bonzini, M; Apostoli, P; Pegoraro, V; Bollati, V; Marinelli, B; Cantone, L; Rizzo, G; Hou, L; 

Schwartz, J; Bertazzi, PA; Baccarelli, A. (2009). Effects of particulate matter on genomic DNA 

methylation content and iNOS promoter methylation. Environ Health Perspect 117: 217-222. 

http://dx.doi.org/10.1289/ehp.11898.

Tavakoli-Nezhad, M; Barron, AJ; Pitts, DK. (2001). Postnatal inorganic lead exposure decreases the number of 

spontaneously active midbrain dopamine neurons in the rat. Neurotoxicology 22: 259-269. 

http://dx.doi.org/10.1016/S0161-813X(01)00010-9.

te Velde, SJ; Ferreira, I; Twisk, JW; Stehouwer, CD; van Mechelen, W; Kemper, HC; Study, AGaHL. (2004). 

Birthweight and arterial stiffness and blood pressure in adulthood--results from the Amsterdam Growth and 

Health Longitudinal Study. Int J Epidemiol 33: 154-161. http://dx.doi.org/10.1093/ije/dyh011.
February 2012 5-618 Draft – Do Not Cite or Quote

Technische Universität Braunschweig. (2011). BRENDA: Comprehensive enzyme information system. Available 

online at http://www.brenda-enzymes.org (accessed January 10, 2011).

Teijon, C; Olmo, R; Blanco, D; Romero, A; Teijon, JM. (2006). Low doses of lead: Effects on reproduction and 

development in rats. Biol Trace Elem Res 111: 151-165. http://dx.doi.org/10.1385/BTER:111:1:151.

Teijon, C; Blanco, MD; Romero, CS; Beneit, JV; Villarino, AL; Guerrero, S; Olmo, R. (2010). Study of response of 

thymic and submaxillary lymph node lymphocytes to administration of lead by different routes. Biol Trace 

Elem Res 135: 74-85. http://dx.doi.org/10.1007/s12011-009-8495-6.

Telisman, S; Colak, B; Pizent, A; Jurasovic, J; Cvitkovic, P. (2007). Reproductive toxicity of low-level lead 

exposure in men. Environ Res 105: 256-266. http://dx.doi.org/10.1016/j.envres.2007.05.011.

Tellez-Rojo, MM; Bellinger, DC; Arroyo-Quiroz, C; Lamadrid-Figueroa, H; Mercado-Garcia, A; Schnaas-Arrieta, 

L; Wright, RO; Hernandez-Avila, M; Hu, H. (2006). Longitudinal associations between blood lead 

concentrations lower than 10 microg/dL and neurobehavioral development in environmentally exposed 

children in Mexico City. Pediatrics 118: e323-e330. http://dx.doi.org/10.1542/peds.2005-3123.

Terziivanov, D; Filipova, M; Janků, I; Balík, J; Filip, V; Stika, L. (1983). Changes in electroretinogram and serum 

potassium during L-DOPA treatment in parkinsonism. Arch Psychiatr Z Neurol 232: 507-513. 

http://dx.doi.org/10.1007/BF00344065.

Tessitore, L; Sesca, E; Pani, P; Dianzani, MU. (1995). Sexual dimorphism in the regulation of cell turnover during 

liver hyperplasia. Chem Biol Interact 97: 1-10. http://dx.doi.org/10.1016/0009-2797(94)03602-1.

Tian, L; Lawrence, DA. (1995). Lead inhibits nitric oxide production in vitro by murine splenic macrophages. 

Toxicol Appl Pharmacol 132: 156-163. http://dx.doi.org/10.1006/taap.1995.1096.

Tian, L; Lawrence, DA. (1996). Metal-induced modulation of nitric oxide production in vitro by murine 

macrophages: Lead, nickel, and cobalt utilize different mechanisms. Toxicol Appl Pharmacol 141: 540-

547. http://dx.doi.org/10.1006/taap.1996.0320.

Tian, Y; Green, P; Stamova, B; Hertz-Picciotto, I; Pessah, IN; Hansen, R; Yang, X; Gregg, JP; Ashwood, P; 

Jickling, G; Van de Water, J; Sharp, F. (2011). Correlations of gene expression with blood lead levels in 

children with autism compared to typically developing controls. Neurotox Res 19: 1-13. 

http://dx.doi.org/10.1007/s12640-009-9126-x.

Tiffany-Castiglioni, E; Sierra, EM; Wu, JN; Rowles, TK. (1989). Lead toxicity in neuroglia. Neurotoxicology 10: 

417-443. 

Timchalk, C; Lin, Y; Weitz, KK; Wu, H; Gies, RA; Moore, DA; Yantasee, W. (2006). Disposition of lead (Pb) in 

saliva and blood of Sprague-Dawley rats following a single or repeated oral exposure to Pb-acetate. 

Toxicology 222: 86-94. http://dx.doi.org/10.1016/j.tox.2006.01.030.

Tokar, EJ; Diwan, BA; Waalkes, MP. (2010). Early life inorganic lead exposure induces testicular teratoma and 

renal and urinary bladder preneoplasia in adult metallothionein-knockout mice but not in wild type mice. 

Toxicology 276: 5-10. http://dx.doi.org/10.1016/j.tox.2010.06.006.

Tomokuni, K; Ichiba, M. (1990). Effect of lead on the activity of erythrocyte porphobilinogen deaminase in vivo 

and in vitro. Toxicol Lett 50: 137-142. http://dx.doi.org/10.1016/0378-4274(90)90003-5.

Tomoum, HY; Mostafa, GA; Ismail, NA; Ahmed, SM. (2010). Lead exposure and its association with pubertal 

development in school-age Egyptian children: Pilot study. Pediatrics International 52: 89-93. 

http://dx.doi.org/10.1111/j.1442-200X.2009.02893.x.

Tomsig, JL; Suszkiw, JB. (1993). Intracellular mechanism of Pb(2+)-induced norepinephrine release from bovine 

chromaffin cells. Am J Physiol Cell Physiol 265: C1630-C1636. 

Tomsig, JL; Suszkiw, JB. (1995). Multisite interactions between Pb2+ and protein kinase C and its role in 

norepinephrine release from bovine adrenal chromaffin cells. J Neurochem 64: 2667-2673. 

http://dx.doi.org/10.1046/j.1471-4159.1995.64062667.x.
February 2012 5-619 Draft – Do Not Cite or Quote

Tonelli, M; Wiebe, N; Hemmelgarn, B; Klarenbach, S; Field, C; Manns, B; Thadhani, R; Gill, J. (2009). Trace 

elements in hemodialysis patients: A systematic review and meta-analysis [Review]. BMC Med 7: 25. 

http://dx.doi.org/10.1186/1741-7015-7-25.

Tong, IS; Lu, Y. (2001). Identification of confounders in the assessment of the relationship between lead exposure 

and child development. Ann Epidemiol 11: 38-45. 

Tong, S; Baghurst, P; McMichael, A; Sawyer, M; Mudge, J. (1996). Lifetime exposure to environmental lead and 

children's intelligence at 11-13 years: The Port Pirie cohort study. Br Med J 312: 1569-1575. 

http://dx.doi.org/10.1136/bmj.312.7046.1569.

Tong, S; Baghurst, PA; Sawyer, MG; Burns, J; McMichael, AJ. (1998). Declining blood lead levels and changes in 

cognitive function during childhood: the Port Pirie cohort study. JAMA 280: 1915-1919. 

http://dx.doi.org/10.1001/jama.280.22.1915.

Tong, S; McMichael, AJ; Baghurst, PA. (2000). Interactions between environmental lead exposure and 

sociodemographic factors on cognitive development. Arch Environ Occup Health 55: 330-335. 

Trope, I; Lopez-Villegas, D; Cecil, KM; Lenkinski, RE. (2001). Exposure to lead appears to selectively alter 

metabolism of cortical gray matter. Pediatrics 107: 1437-1443. http://dx.doi.org/10.1542/peds.107.6.1437.

Tsaih, SW; Korrick, S; Schwartz, J; Amarasiriwardena, C; Aro, A; Sparrow, D; Hu, H. (2004). Lead, diabetes, 

hypertension, and renal function: The normative aging study. Environ Health Perspect 112: 1178-1182. 

http://dx.doi.org/10.1289/ehp.7024.

Tsao, DA; Yu, HS; Cheng, JT; Ho, CK; Chang, HR. (2000). The change of beta-adrenergic system in lead-induced 

hypertension. Toxicol Appl Pharmacol 164: 127-133. http://dx.doi.org/10.1006/taap.1999.8871.

Tsuang, M. (2000). Schizophrenia: Genes and environment. Biol Psychiatry 47: 210-220. 

http://dx.doi.org/10.1016/S0006-3223(99)00289-9.

Tuppurainen, M; Wagar, G; Kurppa, K; Sakari, W; Wambugu, A; Froseth, B; Alho, J; Nykyri, E. (1988). Thyroid 

function as assessed by routine laboratory tests of workers with long-term lead exposure. Scand J Work 

Environ Health 14: 175-180. 

Turgut, S; Polat, A; Inan, M; Turgut, G; Emmungil, G; Bican, M; Karakus, TY; Genc, O. (2007). Interaction 

between anemia and blood levels of iron, zinc, copper, cadmium and lead in children. Indian J Pediatr 74: 

827-830. http://dx.doi.org/10.1007/s12098-007-0147-2.

Turkez, H; Geyikoglu, F; Tatar, A; Keles, MS; Kaplan, I. (In Press) The effects of some boron compounds against 

heavy metal toxicity in human blood. Exp Toxicol Pathol. http://dx.doi.org/10.1016/j.etp.2010.06.011.

U.S. EPA (U.S. Environmental Protection Agency). (1986a). Air quality criteria for lead [EPA Report]. (EPA/600/8-

83/028aF-dF). Research Triangle Park, NC. http://cfpub.epa.gov/ncea/cfm/recordisplay.cfm?deid=32647.

U.S. EPA (U.S. Environmental Protection Agency). (1986b). Air quality criteria for lead: Volume I of IV [EPA 

Report]. (EPA-600/8-83/028aF). Research Triangle Park, NC. 

http://cfpub.epa.gov/ncea/cfm/recordisplay.cfm?deid=32647.

U.S. EPA (U.S. Environmental Protection Agency). (2006b). Air quality criteria for lead: Volume I of II. 

(EPA/600/R-05/144aF). Research Triangle Park, NC: US EPA. 

http://cfpub.epa.gov/ncea/CFM/recordisplay.cfm?deid=158823.

U.S. EPA (U.S. Environmental Protection Agency). (2006g). Table 5-7, studies reporting lead-induced shifts in Th1 

versus Th2 cytokines [EPA Report]. In Air quality criteria for lead: Volume I of II (pp. 5-225-225-226). 

(EPA/600/R-5/144aF).

U.S. EPA (U.S. Environmental Protection Agency). (2006h). Table AX5.9.6., pattern of lead-induced macrophage 

immunotoxicity [EPA Report]. In Air quality criteria for lead: Volume II of II (pp. AX5-155). (EPA/600/R05/144bF).

U.S. Renal Data System. (2009). USRDS 2009 annual data report: Atlas of chronic kidney disease and end-stage 

renal disease in the United States. Bethesda, MD. http://www.usrds.org/adr_2009.htm.
February 2012 5-620 Draft – Do Not Cite or Quote

Ueda, D; Kishimoto, T; Dekio, S; Tada, M. (1997). Inhibitory effect of lead on tube formation by cultured human 

vascular endothelial cells. Hum Cell 10: 283-291. 

Ujihara, H; Sasa, M; Ban, T. (1995). Selective blockade of P-type calcium channels by lead in cultured hippocampal 

neurons. Jpn J Pharmacol 67: 267-269. http://dx.doi.org/10.1254/jjp.67.267.

Ukaejiofo, EO; Thomas, N; Ike, SO. (2009). Haematological assessment of occupational exposure to lead handlers 

in Enugu urban, Enugu State, Nigeria. Niger J Clin Pract 12: 58-64. 

Undeger, U; Basaran, N; Canpinar, H; Kansu, E. (1996). Immune alterations in lead-exposed workers. Toxicology 

109: 167-172. http://dx.doi.org/10.1016/0300-483X(96)03333-1.

Upadhyay, AK; Mathur, R; Bhadauria, M; Nirala, SK. (2009). Therapeutic influence of zinc and ascorbic acid 

against lead induced biochemical alterations. Therapie 64: 383-388. 

http://dx.doi.org/10.2515/therapie/2009055.

Urch, B; Brook, JR; Wasserstein, D; Brook, RD; Rajagopalan, S; Corey, P; Silverman, F. (2004). Relative 

contributions of PM2.5 chemical constituents to acute arterial vasoconstriction in humans. Inhal Toxicol 

16: 345-352. 

Ustundag, A; Duydu, Y. (2007). The influence of melatonin and N-acetylcysteine in delta-aminolevulinic acid and 

lead induced genotoxicity in lymphocytes in vitro. Biol Trace Elem Res 117: 53-64. 

http://dx.doi.org/10.1007/BF02698083.

Uzbekov, MG; Bubnova, NI; Kulikova, GV. (2007). Effect of prenatal lead exposure on superoxide dismutase 

activity in the brain and liver of rat fetuses. Bull Exp Biol Med 144: 783-785. 

http://dx.doi.org/10.1007/s10517-007-0431-1.

Uzu, G; Sauvain, JJ; Baeza-Squiban, A; Riediker, M; Sánchez Sandoval Hohl, M; Val, S; Tack, K; Denys, S; 

Pradère, P; Dumat, C. (2011). In vitro assessment of the pulmonary toxicity and gastric availability of leadrich particles from a lead recycling plant. Environ Sci Technol 45: 7888-7895. 

http://dx.doi.org/10.1021/es200374c.

Vaglenov, A; Carbonell, E; Marcos, R. (1998). Biomonitoring of workers exposed to lead genotoxic effects, its 

modulation by polyvitamin treatment and evaluation of the induced radioresistance. Mutat Res Genet 

Toxicol Environ Mutagen 418: 79-92. http://dx.doi.org/10.1016/S1383-5718(98)00111-9.

Vaglenov, A; Creus, A; Laltchev, S; Petkova, V; Pavlova, S; Marcos, R. (2001). Occupational exposure to lead and 

induction of genetic damage. Environ Health Perspect 109: 295-298. 

Valencia, I; Castillo, EE; Chamorro, G; Bobadilla, RA; Castillo, C. (2001). Lead induces endothelium- and Ca2+-

independent contraction in rat aortic rings. Pharmacol Toxicol 89: 177-182. 

http://dx.doi.org/10.1111/j.0901-9928.2001.890406.x.

Valentino, M; Governa, M; Marchiseppe, I; Visona, I. (1991). Effects of lead on polymorphonuclear leukocyte 

(PMN) functions in occupationally exposed workers. Arch Toxicol 65: 685-688. 

http://dx.doi.org/10.1007/BF02098038.

Valentino, M; Rapisarda, V; Santarelli, L; Bracci, M; Scorcelletti, M; Di Lorenzo, L; Cassano, F; Soleo, L. (2007). 

Effect of lead on the levels of some immunoregulatory cytokines in occupationally exposed workers. Hum 

Exp Toxicol 26: 551-556. http://dx.doi.org/10.1177/0960327107073817.

Valverde, M; Fortoul, TI; Diaz-Barriga, F; Majia, J; Del Castillo, ER. (2002). Genotoxicity induced in CD-1 mice 

by inhaled lead: Differential organ response. Mutagenesis 17: 55-61. 

http://dx.doi.org/10.1093/mutage/17.1.55.

Van De Vyver, FL; D'Haese, PC; Visser, WJ; Elseviers, MM; Knippenberg, LJ; Lamberts, LV; Wedeen, RP; De 

Broe, ME. (1988). Bone lead in dialysis patients. Kidney Int 33: 601-607. 

Van Esch, GJ; Kroes, R. (1969). The induction of renal tumours by feeding basic lead acetate to mice and hamsters. 

Br J Cancer 23: 765-771. http://dx.doi.org/10.1038/bjc.1969.95.
February 2012 5-621 Draft – Do Not Cite or Quote

Van Larebeke, N; Koppen, G; Nelen, V; Schoeters, G; Van Loon H Albering H Riga L Vlietinck, R; Kleinjans, J.

(2004). Differences in HPRT mutant frequency among middle-aged Flemish women in association with 

area of residence and blood lead levels. Biomarkers 9: 71-84. 

http://dx.doi.org/10.1080/13547500310001652160.

Van Wijngaarden, E; Dosemeci, M. (2006). Brain cancer mortality and potential occupational exposure to lead: 

Findings from the National Longitudinal Mortality Study, 1979-1989. Int J Cancer 119: 1136-1144. 

http://dx.doi.org/10.1002/ijc.21947.

Van Wijngaarden, E; Campbell, JR; Cory-Slechta, DA. (2009). Bone lead levels are associated with measures of 

memory impairment in older adults. Neurotoxicology 30: 572-580. 

http://dx.doi.org/10.1016/j.neuro.2009.05.007.

Vander, AJ. (1988). Chronic effects of lead on the renin-angiotensin system. Environ Health Perspect 78: 77-83. 

http://dx.doi.org/10.1289/ehp.887877.

Vargas-Robles, H; Romo, E; Sanchez-Mendoza, A; Rios, AP; Soto, V; Avila-Casado, C; Medina, A; Escalante, B.

(2007). Lead exposure effect on angiotensin II renal vasoconstriction. Hum Exp Toxicol 26: 499-507. 

http://dx.doi.org/10.1177/0960327106077597.

Vassallo, DV; Lebarch, EC; Moreira, CM; Wiggers, GA; Stefanon, I. (2008). Lead reduces tension development and 

the myosin ATPase activity of the rat right ventricular myocardium. Braz J Med Biol Res 41: 789-795. 

http://dx.doi.org/10.1590/S0100-879X2008000900008.

Vaziri, ND; Ding, Y; Ni, Z; Gonick, HC. (1997). Altered nitric oxide metabolism and increased oxygen free radical 

activity in lead-induced hypertension: effect of lazaroid therapy. Kidney Int 52: 1042-1046. 

http://dx.doi.org/10.1038/ki.1997.426.

Vaziri, ND; Wang, XQ. (1999). cGMP-mediated negative-feedback regulation of endothelial nitric oxide synthase 

expression by nitric oxide. Hypertension 34: 1237-1241. 

Vaziri, ND; Liang, K; Ding, Y. (1999a). Increased nitric oxide inactivation by reactive oxygen species in leadinduced hypertension. Kidney Int 56: 1492-1498. http://dx.doi.org/10.1046/j.1523-1755.1999.00670.x.

Vaziri, ND; Ding, Y; Ni, Z. (1999b). Nitric oxide synthase expression in the course of lead-induced hypertension. 

Hypertension 34: 558-562. 

Vaziri, ND; Wang, XQ; Oveisi, F; Rad, B. (2000). Induction of oxidative stress by glutathione depletion causes 

severe hypertension in normal rats. Hypertension 36: 142-146. http://dx.doi.org/10.1161/01.HYP.36.1.142.

Vaziri, ND; Ding, Y. (2001). Effect of lead on nitric oxide synthase expression in coronary endothelial cells: Role of 

superoxide. Hypertension 37: 223-226. http://dx.doi.org/10.1161/01.HYP.37.2.223.

Vaziri, ND; Lin, CY; Farmand, F; Sindhu, RK. (2003). Superoxide dismutase, catalase, glutathione peroxidase and 

NADPH oxidase in lead-induced hypertension. Kidney Int 63: 186-194. http://dx.doi.org/10.1046/j.1523-

1755.2003.00711.x.

Vaziri, ND; Ding, Y; Ni, Z; Barton, CH. (2005). Bradykinin down-regulates, whereas arginine analogs up-regulates, 

endothelial nitric-oxide synthase expression in coronary endothelial cells. J Pharmacol Exp Ther 313: 121-

126. http://dx.doi.org/10.1124/jpet.104.076497.

Vaziri, ND; Khan, M. (2007). Interplay of reactive oxygen species and nitric oxide in the pathogenesis of 

experimental lead-induced hypertension. Clin Exp Pharmacol Physiol 34: 920-925. 

http://dx.doi.org/10.1111/j.1440-1681.2007.04644.x.

Vaziri, ND. (2008a). Causal link between oxidative stress, inflammation, and hypertension. Iran J Kidney Dis 2: 1-

10. 

Vaziri, ND. (2008b). Mechanisms of lead-induced hypertension and cardiovascular disease. Am J Physiol Heart 

Circ Physiol 295: H454-H465. http://dx.doi.org/10.1152/ajpheart.00158.2008.

Vega-Dienstmaier, JM; Salinas-Pielago, JE; Gutierrez-Campos, MDR; Mandamiento-Ayquipa, RD; Yara-Hokama, 

MDC; Ponce-Canchihuaman, J; Castro-Morales, J. (2006). Lead levels and cognitive abilities in Peruvian 

children. Rev Bras Psiquiatr 28: 33-39. http://dx.doi.org/10.1590/S1516-44462006000100008.
February 2012 5-622 Draft – Do Not Cite or Quote

Veranth, JM; Moss, TA; Chow, JC; Labban, R; Nichols, WK; Walton, JC; Walton, JG; Yost, GS. (2006). 

Correlation of in vitro cytokine responses with the chemical composition of soil-derived particulate matter. 

Environ Health Perspect 114: 341-349. http://dx.doi.org/10.1289/ehp.8360.

Verina, T; Rohde, CA; Guilarte, TR. (2007). Environmental lead exposure during early life alters granule cell 

neurogenesis and morphology in the hippocampus of young adult rats. Neuroscience 145: 1037-1047. 

http://dx.doi.org/10.1016/j.neuroscience.2006.12.040.

Verma, SK; Dua, R; Gill, KD. (2005). Impaired energy metabolism after co-exposure to lead and ethanol. Basic Clin 

Pharmacol Toxicol 96: 475-479. http://dx.doi.org/10.1111/j.1742-7843.2005.pto_96611.x.

Vetter, SW; Leclerc, E. (2003). Novel aspects of calmodulin target recognition and activation. Eur J Biochem 270: 

404-414. http://dx.doi.org/10.1046/j.1432-1033.2003.03414.x.

Vigeh, M; Yokoyama, K; Kitamura, F; Afshinrokh, M; Beygi, A; Niroomanesh, S. (2010). Early pregnancy blood 

lead and spontaneous abortion. Women Health 50: 756-766. 

http://dx.doi.org/10.1080/03630242.2010.532760.

Vigeh, M; Yokoyama, K; Seyedaghamiri, Z; Shinohara, A; Matsukawa, T; Chiba, M; Yunesian, M. (2011). Blood 

lead at currently acceptable levels may cause preterm labour. Occup Environ Med 68: 231-234. 

http://dx.doi.org/10.1136/oem.2009.050419.

Vimpani, GV; Wigg, NR; Robertson, EF; McMichael, AJ; Baghurst, PA; Roberts, RJ. (1985). The Port Pirie cohort 

study: blood lead concentration and childhood developmental assessment. In LJ Goldwater; LM Wysocki; 

RA Volpe (Eds.), Edited proceedings: Lead environmental health - the current issues (pp. 139-146). 

Durham, NC: Duke University. 

Vinceti, M; Guidetti, D; Bergomi, M; Caselgrandi, E; Vivoli, R; Olmi, M; Rinaldi, L; Rovesti, S; Solime, F. (1997). 

Lead, cadmium, and selenium in the blood of patients with sporadic amyotrophic lateral sclerosis. Ital J 

Neurol Sci 18: 87-92. http://dx.doi.org/10.1007/BF01999568.

Virgolini, MB; Chen, K; Weston, DD; Bauter, MR; Cory-Slechta, DA. (2005). Interactions of chronic lead exposure 

and intermittent stress: Consequences for brain catecholamine systems and associated behaviors and HPA 

axis function. Toxicol Sci 87: 469-482. http://dx.doi.org/10.1093/toxsci/kfi269.

Virgolini, MB; Rossi-George, A; Lisek, R; Weston, DD; Thiruchelvam, M; Cory-Slechta, DA. (2008a). CNS effects 

of developmental Pb exposure are enhanced by combined maternal and offspring stress. Neurotoxicology 

29: 812-827. http://dx.doi.org/10.1016/j.neuro.2008.03.003.

Virgolini, MB; Rossi-George, A; Weston, D; Cory-Slechta, DA. (2008b). Influence of low level maternal Pb 

exposure and prenatal stress on offspring stress challenge responsivity. Neurotoxicology 29: 928-939. 

http://dx.doi.org/10.1016/j.neuro.2008.09.010.

Vogetseder, A; Picard, N; Gaspert, A; Walch, M; Kaissling, B; Le Hir, M. (2008). Proliferation capacity of the renal 

proximal tubule involves the bulk of differentiated epithelial cells. Am J Physiol Cell Physiol 294: C22-

C28. http://dx.doi.org/10.1152/ajpcell.00227.2007.

Waalkes, MP; Diwan, BA; Ward, JM; Devor, DE; Goyer, RA. (1995). Renal tubular tumors and atypical 

hyperplasias in B6C3F1 mice exposed to lead acetate during gestation and lactation occur with minimal 

chronic nephropathy. Cancer Res 55: 5265-5271. 

Waalkes, MP; Liu, J; Goyer, RA; Diwan, BA. (2004). Metallothionein-I/II double knockout mice are hypersensitive 

to lead-induced kidney carcinogenesis: Role of inclusion body formation. Cancer Res 64: 7766-7772. 

http://dx.doi.org/10.1158/0008-5472.CAN-04-2220.

Wananukul, W; Sura, T; Salaitanawatwong, P. (2006). Polymorphism of delta-aminolevulinic acid dehydratase and 

its effect on blood lead levels in Thai workers. Arch Environ Occup Health 61: 67-72. 

http://dx.doi.org/10.3200/AEOH.61.2.67-72.

Wang, A; Wang, Q; Song, Q; Xu, J. (2009a). [Study of ALAD and VDR gene polymorphisms associated with lead 

nephrotoxicity susceptibility]. Wei Sheng Yan Jiu 38: 326-329. 
February 2012 5-623 Draft – Do Not Cite or Quote

Wang, CH; Zhang, Y; Liang, JC; Shan, G; Wang, Y; Shi, Q. (2006a). Impacts of ascorbic acid and thiamine 

supplementation at different concentrations on lead toxicity in testis. Clin Chim Acta 370: 82-88. 

http://dx.doi.org/10.1016/j.cca.2006.01.023.

Wang, CL; Chuang, HY; Ho, CK; Yang, CY; Tsai, JL; Wu, TS; Wu, TN. (2002a). Relationship between blood lead 

concentrations and learning achievement among primary school children in Taiwan. Environ Res 89: 12-

18. http://dx.doi.org/10.1006/enrs.2002.4342.

Wang, CY; Lin, YW; Yang, JL. (2008c). Activation of protein kinase C alpha signaling prevents cytotoxicity and 

mutagenicity following lead acetate in CL3 human lung cancer cells. Toxicology 250: 55-61. 

http://dx.doi.org/10.1016/j.tox.2008.06.001.

Wang, FT; Hu, H; Schwartz, J; Weuve, J; Spiro, AS, III; Sparrow, D; Nie, HL; Silverman, EK; Weiss, ST; Wright, 

RO. (2007a). Modifying effects of the HFE polymorphisms on the association between lead burden and 

cognitive decline. Environ Health Perspect 115: 1210-1215. http://dx.doi.org/10.1289/ehp.9855.

Wang, G; Chen, XQ; Lipsky, MM; Whittaker, MH; Fowler, BA. (2005). Effect of combined lead, cadmium and 

arsenic exposure on the expression of cellular protective factors in rat kidneys at LOEL dose levels 

[Abstract]. Toxicologist 85: 237. 

Wang, GS; Fowler, BA. (2008). Roles of biomarkers in evaluating interactions among mixtures of lead, cadmium 

and arsenic. Toxicol Appl Pharmacol 233: 92-99. http://dx.doi.org/10.1016/j.taap.2008.01.017.

Wang, L; Chen, DW; Wang, H; Liu, ZP. (2009b). Effects of lead and/or cadmium on the expression of 

metallothionein in the kidney of rats. Biol Trace Elem Res 129: 190-199. http://dx.doi.org/10.1007/s12011-

008-8288-3.

Wang, L; Wang, H; Hu, MZ; Cao, J; Chen, DW; Liu, Z. (2009c). Oxidative stress and apoptotic changes in primary 

cultures of rat proximal tubular cells exposed to lead. Arch Toxicol 83: 417-427. 

http://dx.doi.org/10.1007/s00204-009-0425-z.

Wang, L; Wang, Z; Liu, J. (2010d). Protective effect of N-acetylcysteine on experimental chronic lead 

nephrotoxicity in immature female rats. Hum Exp Toxicol 29: 581-591. 

http://dx.doi.org/10.1177/0960327109357270.

Wang, L; Wang, H; Li, J; Chen, D; Liu, Z. (2011b). Simultaneous effects of lead and cadmium on primary cultures 

of rat proximal tubular cells: interaction of apoptosis and oxidative stress. Arch Environ Contam Toxicol 

61: 500-511. http://dx.doi.org/10.1007/s00244-011-9644-4.

Wang, Q; Luo, WJ; Zhang, WB; Dai, ZM; Chen, YM; Chen, JY. (2007b). Iron supplementation protects against 

lead-induced apoptosis through MAPK pathway in weanling rat cortex. Neurotoxicology 28: 850-859. 

http://dx.doi.org/10.1016/j.neuro.2007.04.004.

Wang, Q; Zhao, HH; Chen, JW; Hao, QL; Gu, KD; Zhu, YX; Zhou, YK; Ye, LX. (2010f). delta-Aminolevulinic 

acid dehydratase activity, urinary delta-aminolevulinic acid concentration and zinc protoporphyrin level 

among people with low level of lead exposure. Int J Hyg Environ Health 213: 52-58. 

http://dx.doi.org/10.1016/j.ijheh.2009.08.003.

Wang, TL; Kao, TH; Inbaraj, BS; Su, YT; Chen, BH. (2010g). Inhibition effect of poly(γ-glutamic acid) on leadinduced toxicity in mice. J Agric Food Chem 58: 12562–12567. http://dx.doi.org/10.1021/jf1034509.

Wang, YY; Sui, KX; Hong, LI; Ma, HY. (2009e). The effects of lead exposure on placental NF-kappaB expression 

and the consequences for gestation. Reprod Toxicol 27: 190-195. 

http://dx.doi.org/10.1016/j.reprotox.2008.12.006.

Wasserman, G; Graziano, JH; Factor-Litvak, R; Popovac, D; Morina, N; Musabegovic, A; Vrenezi, N; CapuniParacka, S; Lekic, V; Preteni-Redjepi, E; Hadzialjevic, S; Slavkovich, V; Kline, J; Shrout, P; Stein, Z.

(1992). Independent effects of lead exposure and iron deficiency anemia on developmental outcome at age 

2 years. J Pediatr 121: 695-703. 
February 2012 5-624 Draft – Do Not Cite or Quote

Wasserman, GA; Graziano, JH; Factor-Litvak, P; Popovac, D; Morina, N; Musabegovic, A; Vrenezi, N; CapuniParacka, S; Lekic, V; Preteni-Redjepi, E; Hadzialjevic, S; Slavkovich, V; Kline, J; Shrout, P; Stein, Z.

(1994). Consequences of lead exposure and iron supplementation on childhood development at age 4 years. 

Neurotoxicol Teratol 16: 233-240. http://dx.doi.org/10.1016/0892-0362(94)90044-2.

Wasserman, GA; Liu, X; Lolacono, NJ; Factor-Litvak, P; Kline, JK; Popovac, D; Morina, N; Musabegovic, A; 

Vrenezi, N; Capuni-Paracka, S; Lekic, V; Preteni-Redjepi, E; Hadzialjevic, S; Slavkovich, V; Graziano, 

JH. (1997). Lead exposure and intelligence in 7-year-old children: The Yugoslavia prospective study. 

Environ Health Perspect 105: 956-962. 

Wasserman, GA; Staghezza-Jaramillo, B; Shrout, P; Popovac, D; Graziano, J. (1998). The effect of lead exposure on 

behavior problems in preschool children. Am J Public Health 88: 481-486. 

http://dx.doi.org/10.2105/AJPH.88.3.481.

Wasserman, GA; Musabegovic, A; Liu, X; Kline, J; Factor-Litvak, P; Graziano, JH. (2000). Lead exposure and 

motor functioning in 4 1/2-year-old children: The Yugoslavia prospective study. J Pediatr 137: 555-561. 

Wasserman, GA; Factor-Litvak, P. (2001). Methodology, inference and causation: environmental lead exposure and 

childhood intelligence. Arch Clin Neuropsychol 16: 343-352. 

Wasserman, GA; Factor-Litvak, P; Liu, X; Todd, AC; Kline, JK; Slavkovich, V; Popovac, D; Graziano, JH. (2003). 

The relationship between blood lead, bone lead and child intelligence. Child Neuropsychol 9: 22-34. 

http://dx.doi.org/10.1076/chin.9.1.22.14497.

Waterman, SJ; El-Fawal, HAN; Snyder, CA. (1994). Lead alters the immunogenicity of two neural proteins: A 

potential mechanism for the progression of lead-induced neurotoxicity. Environ Health Perspect 102: 1052-

1056. 

Watts, SW; Chai, S; Webb, RC. (1995). Lead acetate-induced contraction in rabbit mesenteric artery: Interaction 

with calcium and protein kinase C. Toxicology 99: 55-65. http://dx.doi.org/10.1016/0300-483X(94)03003-

K.

Weaver, V; Jaar, B. (2010). UpToDate: Lead nephropathy and lead-related nephrotoxicity. Available online at 

http://www.uptodate.com/contents/lead-nephropathy-and-lead-related-nephrotoxicity (accessed October 25, 

2010).

Weaver, VM; Lee, BK; Ahn, KD; Lee, GS; Todd, AC; Stewart, WF; Wen, J; Simon, DJ; Parsons, PJ; Schwartz, BS.

(2003a). Associations of lead biomarkers with renal function in Korean lead workers. Occup Environ Med 

60: 551-562. http://dx.doi.org/10.1136/oem.60.8.551.

Weaver, VM; Schwartz, BS; Ahn, KD; Stewart, WF; Kelsey, KT; Todd, AC; Wen, J; Simon, DJ; Lustberg, ME; 

Parsons, PJ; Silbergeld, EK; Lee, BK. (2003b). Associations of renal function with polymorphisms in the 

"delta"-aminolevulinic acid dehydratase, vitamin D receptor, and nitric oxide synthase genes in Korean 

lead workers. Environ Health Perspect 111: 1613-1619. 

Weaver, VM; Jarr, BG; Schwartz, BS; Todd, AC; Ahn, KD; Lee, SS; Wen, J; Parsons, PJ; Lee, BK. (2005a). 

Associations among lead dose biomarkers, uric acid, and renal function in Korean lead workers. Environ 

Health Perspect 113: 36-42. http://dx.doi.org/10.1289/ehp.7317.

Weaver, VM; Schwartz, BS; Jaar, BG; Ahn, KD; Todd, AC; Lee, SS; Kelsey, KT; Silbergeld, EK; Lustberg, ME; 

Parsons, PJ; Wen, J; Lee, BK. (2005b). Associations of uric acid with polymorphisms in the "delta"-

aminolevulinic acid dehydratase, vitamin D receptor, and nitric oxide synthase genes in Korean lead 

workers. Environ Health Perspect 113: 1509-1515. 

Weaver, VM; Lee, BK; Todd, AC; Ahn, KD; Shi, W; Jaar, BG; Kelsey, KT; Lustberg, ME; Silbergeld, EK; Parsons, 

PJ; Wen, J; Schwartz, BS. (2006). Effect modification by delta-aminolevulinic acid dehydratase, vitamin D 

receptor, and nitric oxide synthase gene polymorphisms on associations between patella lead and renal 

function in lead workers. Environ Res 102: 61-69. http://dx.doi.org/10.1016/j.envres.2006.01.001.

Weaver, VM; Ellis, LR; Lee, BK; Todd, AC; Shi, W; Ahn, KD; Schwartz, BS. (2008). Associations between patella 

lead and blood pressure in lead workers. Am J Ind Med 51: 336-343. http://dx.doi.org/10.1002/ajim.20573.
February 2012 5-625 Draft – Do Not Cite or Quote

Weaver, VM; Griswold, M; Todd, AC; Jaar, BG; Ahn, KD; Thompson, CB; Lee, BK. (2009). Longitudinal 

associations between lead dose and renal function in lead workers. Environ Res 109: 101-107. 

http://dx.doi.org/10.1016/j.envres.2008.09.005.

Weaver, VM. (2010). Further analysis of Korean lead worker data set with a focus on determining the functional 

form of the exposure-response relationship. Available online at (accessed

Weaver, VM; Kim, NS; Jaar, BG; Schwartz, BS; Parsons, PJ; Steuerwald, AJ; Todd, AC; Simon, D; Lee, BK.

(2011). Associations of low-level urine cadmium with kidney function in lead workers. Occup Environ 

Med 68: 250-256. http://dx.doi.org/10.1136/oem.2010.056077.

Wedrychowski, A; Schmidt, WN; Hnilica, LS. (1986). The in vivo cross-linking of proteins and DNA by heavy 

metals. J Biol Chem 261: 3370-3376. 

Wei, H; Wei, D; Yi, S; Zhang, F; Ding, W. (2011). Oxidative stress induced by urban fine particles in cultured 

EA.hy926 cells. Hum Exp Toxicol 30: 579-590. http://dx.doi.org/10.1177/0960327110374207.

Weiss, B. (1988). Neurobehavioral toxicity as a basis for risk assessment. Trends Pharmacol Sci 9: 59-62. 

Weiss, B. (1990). Risk assessment: The insidious nature of neurotoxicity and the aging brain. Neurotoxicology 11: 

305-314. 

Weisskopf, MG; Wright, RO; Schwartz, J; Spiro, A, III; Sparrow, D; Aro, A; Hu, H. (2004). Cumulative lead 

exposure and prospective change in cognition among elderly men: The VA Normative Aging Study. Am J 

Epidemiol 160: 1184-1193. http://dx.doi.org/10.1093/aje/kwh333.

Weisskopf, MG; Proctor, SP; Wright, RO; Schwartz, J; Spiro, A, III; Sparrow, D; Nie, HL; Hu, H. (2007a). 

Cumulative lead exposure and cognitive performance among elderly men. Epidemiology 18: 59-66. 

http://dx.doi.org/10.1097/01.ede.0000248237.35363.29.

Weisskopf, MG; Jain, N; Nie, HL; Sparrow, D; Vokonas, P; Schwartz, J; Hu, H. (2009). A prospective study of 

bone lead concentration and death from all causes, cardiovascular diseases, and cancer in the department of 

veterans affairs normative aging study. Circulation 120: 1056-1064. 

http://dx.doi.org/10.1161/circulationaha.108.827121.

Weisskopf, MG; Weuve, J; Nie, H; Saint-Hilaire, MH; Sudarsky, L; Simon, DK; Hersh, B; Schwartz, J; Wright, 

RO; Hu, H. (2010). Association of cumulative lead exposure with Parkinson's Disease. Environ Health 

Perspect 118: 1609-1613. http://dx.doi.org/10.1289/ehp.1002339.

Wellenius, GA; Coull, BA; Godleski, JJ; Koutrakis, P; Okabe, K; Savage, ST. (2003). Inhalation of concentrated 

ambient air particles exacerbates myocardial ischemia in conscious dogs. Environ Health Perspect 111: 

402-408. http://dx.doi.org/10.1289/ehp.5775.

Wells, E; Jarrett, J; Hong Lin, Y; Caldwell, K; Hibbeln, J; Apelberg, B; Herbstman, J; Halden, R; Witter, F; 

Goldman, L. (2011a). Body burdens of mercury, lead, selenium and copper among Baltimore newborns. 

Environ Res 111: 411-417. http://dx.doi.org/10.1016/j.envres.2010.12.009.

Wells, EM; Navas-Acien, A; Herbstman, JB; Apelberg, BJ; Silbergeld, EK; Caldwell, KL; Jones, RL; Halden, RU; 

Witter, FR; Goldman, LR. (2011b). Low level lead exposure and elevations in blood pressure during 

pregnancy. Environ Health Perspect 119: 664-669. http://dx.doi.org/10.1289/ehp.1002666.

Westerink, RH; Vijverberg, HP. (2002). Ca2+-independent vesicular catecholamine release in PC12 cells by 

nanomolar concentrations of Pb2+. J Neurochem 80: 861-867. http://dx.doi.org/10.1046/j.0022-

3042.2001.00751.x.

Wetmur, JG; Lehnert, G; Desnick, RJ. (1991a). The delta-aminolevulinate dehydratase polymorphism: Higher blood 

lead levels in lead workers and environmentally exposed children with the 1-2 and 2-2 isozymes. Environ 

Res 56: 109-119. http://dx.doi.org/10.1016/S0013-9351(05)80001-5.

Wetmur, JG. (1994). Influence of the common human delta-aminolevulinate dehydratase polymorphism on lead 

body burden. Environ Health Perspect 102: 215-219. 
February 2012 5-626 Draft – Do Not Cite or Quote

Weuve, J; Kelsey, KT; Schwartz, J; Bellinger, D; Wright, RO; Rajan, P; Spiro, A, III; Sparrow, D; Aro, A; Hu, H.

(2006). Delta-aminolevulinic acid dehydratase polymorphism and the relation between low level lead 

exposure and the Mini-Mental Status Examination in older men: The Normative Aging Study. Occup 

Environ Med 63: 746-753. http://dx.doi.org/10.1136/oem.2006.027417.

Weuve, J; Korrick, SA; Weisskopf, MA; Ryan, LM; Schwartz, J; Nie, HL; Grodstein, F; Hu, H. (2009). Cumulative 

exposure to lead in relation to cognitive function in older women. Environ Health Perspect 117: 574-580. 

http://dx.doi.org/10.1289/ehp.11846.

White, RF; Campbell, R; Echeverria, D; Knox, SS; Janulewicz, P. (2009). Assessment of neuropsychological 

trajectories in longitudinal population-based studies of children. J Epidemiol Community Health 63: I15-

I26. http://dx.doi.org/10.1136/jech.2007.071530.

Whitworth, JA; Mangos, GJ; Kelly, JJ. (2000). Cushing, cortisol, and cardiovascular disease. Hypertension 36: 912-

916. 

WHO (World Health Organization). (1948). Preamble to the Constitution of the World Health Organization as 

adopted by the International Health Conference, New York, 19-22 June, 1946. World Health Organization.

WHO (World Health Organization). (1992). International statistical classification of diseases and related health 

problem, tenth revision. Geneva, Switzerland.

Wiebe, JP; Barr, KJ; Buckingham, KD. (1988). Effect of prenatal and neonatal exposure to lead on gonadotropin 

receptors and steroidogenesis in rat ovaries. J Toxicol Environ Health 24: 461-476. 

Wiebe, JP; Barr, KJ. (1988). Effect of prenatal and neonatal exposure to lead on the affinity and number of estradiol 

receptors in the uterus. J Toxicol Environ Health 24: 451-460. 

http://dx.doi.org/10.1080/15287398809531176.

Wiemann, M; Schirrmacher, K; Busselberg, D. (1999). Interference of lead with the calcium release activated 

calcium flux of osteoblast-like cells. Calcif Tissue Int 65: 479-485. 

http://dx.doi.org/10.1007/s002239900736.

Williams, BL; Pennock-Román, M; Suen, HK; Magsumbol, MS; Ozdenerol, E. (2007). Assessing the impact of the 

local environment on birth outcomes: A case for HLM. J Expo Sci Environ Epidemiol 17: 445-457. 

http://dx.doi.org/10.1038/sj.jes.7500537.

Williams, PL; Sergeyev, O; Lee, MM; Korrick, SA; Burns, JS; Humblet, O; DelPrato, J; Revich, B; Hauser, R.

(2010). Blood lead levels and delayed onset of puberty in a longitudinal study of Russian boys. Pediatrics 

125: 1088-1096. http://dx.doi.org/10.1542/peds.2009-2575.

Wise, JP, Sr; Stearns, DM; Wetterhahn, KE; Patierno, SR. (1994). Cell-enhanced dissolution of carcinogenic lead 

chromate particles: the role of individual dissolution products in clastogenesis. Carcinogenesis 15: 2249-

2254. http://dx.doi.org/10.1093/carcin/15.10.2249.

Wise, JP, Sr; Wise, SS; LaCerte, C; Wise, JP; Aboueissa, AM. (2011). The genotoxicity of particulate and soluble 

chromate in sperm whale (physeter macrocephalus) skin fibroblasts. Environ Mol Mutagen 52: 43-49. 

http://dx.doi.org/10.1002/em.20579.

Wise, SS; Holmes, AL; Ketterer, ME; Hartsock, WJ; Fomchenko, E; Katsifis, S; Thompson, WD; Wise, JP, Sr.

(2004). Chromium is the proximate clastogenic species for lead chromate-induced clastogenicity in human 

bronchial cells. Mutat Res Genet Toxicol Environ Mutagen 560: 79-89. 

http://dx.doi.org/10.1016/j.mrgentox.2004.02.009.

Wise, SS; Holmes, AL; Moreland, JA; Xie, H; Sandwick, SJ; Stackpole, MM; Fomchenko, E; Teufack, S; May, AJ, 

Jr; Katsfis, SP; Wise, JP, Sr. (2005). Human lung cell growth is not stimulated by lead ions after lead 

chromate-induced genotoxicity. Mol Cell Biochem 279: 75-84. http://dx.doi.org/10.1007/s11010-005-

8217-0.

Wise, SS; Holmes, AL; Xie, H; Thompson, WD; Wise, JP, Sr. (2006a). Chronic exposure to particulate chromate 

induces spindle assembly checkpoint bypass in human lung cells. Chem Res Toxicol 19: 1492-1498. 

http://dx.doi.org/10.1021/tx0601410.
February 2012 5-627 Draft – Do Not Cite or Quote

Wise, SS; Holmes, AL; Wise, JP, Sr. (2006b). Particulate and soluble hexavalent chromium ate cytotoxic and 

genotoxic to human lung epithelial cells. Mutat Res Genet Toxicol Environ Mutagen 610: 2-7. 

http://dx.doi.org/10.1016/j.mrgentox.2006.06.005.

Wise, SS; Shaffley, F; LaCerte, C; Goertz, CEC; Dunn, JL; Gulland, FMD; Aboueissa, AE; Zheng, TZ; Wise, JP.

(2009). Particulate and soluble hexavalent chromium are cytotoxic and genotoxic to Steller sea lion lung 

cells. Aquat Toxicol 91: 329-335. http://dx.doi.org/10.1016/j.aquatox.2008.12.004.

Wise, SS; Holmes, AL; Qin, Q; Xie, H; Katsifis, SP; Thompson, WD; Wise, JP, Sr. (2010). Comparative 

genotoxicity and cytotoxicity of four hexavalent chromium compounds in human bronchial cells. Chem 

Res Toxicol 23: 365-372. http://dx.doi.org/10.1021/tx900363j.

Wiwanitkit, V; Suwansaksri, J; Soogarun, S. (2008). White blood cell sister chromatid exchange among a sample of 

Thai subjects exposed to lead: Lead-induced genotoxicity. Toxicol Environ Chem 90: 765-768. 

http://dx.doi.org/10.1080/02772240701712758.

Wolf, J; Daley, AJ. (2007). Microbiological aspects of bacterial lower respiratory tract illness in children: Atypical 

pathogens. Paediatr Respir Rev 8: 212-220. http://dx.doi.org/10.1016/j.prrv.2007.07.004.

Wolff, MS; Britton, JA; Boguski, L; Hochman, S; Maloney, N; Serra, N; Liu, ZS; Berkowitz, G; Larson, S; Forman, 

J. (2008). Environmental exposures and puberty in inner-city girls. Environ Res 107: 393-400. 

http://dx.doi.org/10.1016/j.envres.2008.03.006.

Wozniak, K; Blasiak, J. (2003). In vitro genotoxicity of lead acetate: Induction of single and double DNA strand 

breaks and DNA-protein cross-links. Mutat Res Genet Toxicol Environ Mutagen 535: 127-139. 

http://dx.doi.org/10.1016/S1383-5718(02)00295-4.

Wright, JP; Dietrich, KN; Ris, MD; Hornung, RW; Wessel, SD; Lanphear, BP; Ho, M; Rae, MN. (2008). 

Association of prenatal and childhood blood lead concentrations with criminal arrests in early adulthood. 

PLoS Med 5: 732-740. http://dx.doi.org/10.1371/journal.pmed.0050101.

Wright, RO; Tsaih, SW; Schwartz, J; Spiro, A; McDonald, K; Weiss, ST; Hu, H. (2003b). Lead exposure 

biomarkers and mini-mental status exam scores in older men. Epidemiology 14: 713-718. 

http://dx.doi.org/10.1097/01.EDE.0000081988.85964.db.

Wright, RO; Schwartz, J; Wright, RJ; Bollati, V; Tarantini, L; Park, SK; Hu, H; Sparrow, D; Vokonas, P; Baccarelli, 

A. (2010). Biomarkers of lead exposure and DNA methylation within retrotransposons. Environ Health 

Perspect 118: 790-795. http://dx.doi.org/10.1289/ehp.0901429.

Wu, J; Basha, MR; Brock, B; Cox, DP; Cardozo-Pelaez, F; McPherson, CA; Harry, J; Rice, DC; Maloney, B; Chen, 

D; Lahiri, DK; Zawia, NH. (2008a). Alzheimer's disease (AD)-like pathology in aged monkeys after 

infantile exposure to environmental metal lead (pb): Evidence for a developmental origin and 

environmental link for AD. J Neurosci 28: 3-9. http://dx.doi.org/10.1523/jneurosci.4405-07.2008.

Wu, JF; Basha, R; Zawia, NH. (2008b). The environment, epigenetics and amyloidogenesis. J Mol Neurosci 34: 1-7. 

http://dx.doi.org/10.1007/s12031-007-0009-4.

Wu, MT; Kelsey, K; Schwartz, J; Sparrow, D. (2003a). A "delta"-aminolevulinic acid dehydratase (ALAD) 

polymorphism may modify the relationship of low-level lead exposure to uricemia and renal function: The 

Normative Aging Study. Environ Health Perspect 111: 335-341. http://dx.doi.org/10.1289/ehp.5504.

Wu, T; Buck, GM; Mendola, P. (2003b). Blood lead levels and sexual maturation in U.S. girls: The Third National 

Health and Nutrition Examination Survey, 1988-1994. Environ Health Perspect 111: 737-741. 

http://dx.doi.org/10.1289/ehp.6008.

Wu, TN; Shen, CY; Lai, JS; Goo, CF; Ko, KN; Chi, HY; Chang, PY; Liou, SH. (2000). Effects of lead and noise 

exposures on hearing ability. Arch Environ Health 55: 109-114. 

http://dx.doi.org/10.1080/00039890009603396.

Xie, H; Holmes, AL; Wise, SS; Huang, S; Peng, C; Wise, JP, Sr. (2007). Neoplastic transformation of human 

bronchial cells by lead chromate particles. Am J Respir Cell Mol Biol 37: 544-552. 

http://dx.doi.org/10.1165/rcmb.2007-0058OC.
February 2012 5-628 Draft – Do Not Cite or Quote

Xie, H; Wise, SS; Wise, JP, Sr. (2008). Deficient repair of particulate hexavalent chromium-induced DNA double 

strand breaks leads to neoplastic transformation. Mutat Res Genet Toxicol Environ Mutagen 649: 230-238. 

http://dx.doi.org/10.1016/j.mrgentox.2007.09.008.

Xie, Y; Chiba, M; Shinohara, A; Watanabe, H; Inaba, Y. (1998). Studies on lead-binding protein and interaction 

between lead and selenium in the human erythrocytes. Ind Health 36: 234-239. 

http://dx.doi.org/10.2486/indhealth.36.234.

Xu, J; Ji, LD; Xu, LH. (2006). Lead-induced apoptosis in PC 12 cells: Involvement of p53, Bcl-2 family and 

caspase-3. Toxicol Lett 166: 160-167. http://dx.doi.org/10.1016/j.toxlet.2006.06.643.

Xu, J; Lian, L; W, C; Wang, X; Fu, W; Xu, L. (2008). Lead induces oxidative stress, DNA damage and alteration of 

p53 Bax and Bcl-2 expressions in mice. Food Chem Toxicol 46: 1488-1494. 

http://dx.doi.org/10.1016/j.fct.2007.12.016.

Xu, J; Yan, CH; Yang, B; Xie, HF; Zou, XY; Zhong, L; Gao, Y; Tian, Y; Shen, XM. (2009c). The role of 

metabotropic glutamate receptor 5 in developmental lead neurotoxicity. Toxicol Lett 191: 223-230. 

http://dx.doi.org/10.1016/j.toxlet.2009.09.001.

Xu, SZ; Rajanna, B. (2006). Glutamic acid reverses Pb2+-induced reductions of NMDA receptor subunits in vitro. 

Neurotoxicology 27: 169-175. http://dx.doi.org/10.1016/j.neuro.2005.08.005.

Yamanaka, K; Chun, SJ; Boillee, S; Fujimori-Tonou, N; Yamashita, H; Gutmann, DH; Takahashi, R; Misawa, H; 

Cleveland, DW. (2008). Astrocytes as determinants of disease progression in inherited amyotrophic lateral 

sclerosis. Nat Neurosci 11: 251-253. http://dx.doi.org/10.1038/Nn2047.

Yanez, L; Garcia-Nieto, E; Rojas, E; Carrizales, L; Mejia, J; Calderon, J; Razo, I; Diaz-Barriga, F. (2003). DNA 

damage in blood cells from children exposed to arsenic and lead in a mining area. Environ Res 93: 231-

240. http://dx.doi.org/10.1016/j.envres.2003.07.005.

Yang, Y; Raine, A; Lencz, T; Bihrle, S; LaCasse, L; Colletti, P. (2005). Volume reduction in prefrontal gray matter 

in unsuccessful criminal psychopaths. Biol Psychiatry 57: 1103-1108. 

http://dx.doi.org/10.1016/j.biopsych.2005.01.021.

Yazbeck, C; Thiebaugeorges, O; Moreau, T; Goua, V; Debotte, G; Sahuquillo, J; Forhan, A; Foliguet, B; Magnin, 

G; Slama, R; Charles, MA; Huel, G. (2009). Maternal blood lead levels and the risk of pregnancy-induced 

hypertension: The EDEN cohort study. Environ Health Perspect 117: 1526-1530. 

http://dx.doi.org/10.1289/ehp.0800488.

Ye, SH. (1993). Hypoxanthine phosphoribosyl transferase assay of lead mutagenicity on keratinocytes. Acta 

Pharmacol Sin 14: 145-147. 

Yedjou, CG; Milner, JN; Howard, CB; Tchounwou, PB. (2010). Basic apoptotic mechanisms of lead toxicity in 

human leukemia (HL-60) cells. Int J Environ Res Public Health 7: 2008-2017. 

http://dx.doi.org/10.3390/ijerph7052008.

Yen, TH; Lin, JL; Lin-Tan, DT; Hsu, CW; Chen, KH; Hsu, HH. (2011). Blood cadmium levels association with 18-

month mortality in diabetic patients with maintenance haemodialysis. Nephrol Dial Transplant 26: 998-

1005. http://dx.doi.org/10.1093/ndt/gfq448.

Yetkin-Ay, Z; Cadir, B; Uskun, E; Bozkurt, FY; Delibas, N; Gultepe, FM; Ergurhan-Ilhan, I. (2007). The 

periodontal status of indirectly lead-exposed apprentices working in autorepair workshops. Toxicol Ind 

Health 23: 599-606. http://dx.doi.org/10.1177/0748233708090906.

Yiin, SJ; Lin, TH. (1995). Lead-catalyzed peroxidation of essential unsaturated fatty acid. Biol Trace Elem Res 50: 

167-172. http://dx.doi.org/10.1007/BF02789419.

Yin, Y; Zhang, T; Dai, Y; Bao, Y; Chen, X; Lu, X. (2008). The effect of plasma lead on anembryonic pregnancy. 

Ann N Y Acad Sci 1140: 184-189. http://dx.doi.org/10.1196/annals.1454.042.

Yu, CC; Lin, JL; Lin-Tan, DT. (2004). Environmental exposure to lead and progression of chronic renal diseases: A 

four-year prospective longitudinal study. J Am Soc Nephrol 15: 1016-1022. 

http://dx.doi.org/10.1097/01.ASN.0000118529.01681.4F.
February 2012 5-629 Draft – Do Not Cite or Quote

Yu, DY; Li, WF; Deng, B; Mao, XF. (2008). Effects of lead on hepatic antioxidant status and transcription of 

superoxide dismutase gene in pigs. Biol Trace Elem Res 126: 121-128. http://dx.doi.org/10.1007/s12011-

008-8198-4.

Yu, J; Fujishiro, H; Miyataka, H; Oyama, TM; Hasegawa, T; Seko, Y; Miura, N; Himeno, S. (2009). Dichotomous 

effects of lead acetate on the expression of metallothionein in the liver and kidney of mice. Biol Pharm Bull 

32: 1037-1042. http://dx.doi.org/10.1248/bpb.32.1037.

Yuan, W; Holland, SK; Cecil, KM; Dietrich, KN; Wessel, SD; Altaye, M; Hornung, RW; Ris, MD; Egelhoff, JC; 

Lanphear, BP. (2006). The impact of early childhood lead exposure on brain organization: A functional 

magnetic resonance imaging study of language function. Pediatrics 118: 971-977. 

http://dx.doi.org/10.1542/peds.2006-0467.

Yucesoy, B; Turhan, A; Ure, M; Imir, T; Karakaya, A. (1997a). Effects of occupational lead and cadmium exposure 

on some immunoregulatory cytokine levels in man. Toxicology 123: 143-147. 

http://dx.doi.org/10.1016/S0300-483X(97)00107-8.

Yucesoy, B; Turhan, A; Ure, M; Imir, T; Karakaya, A. (1997b). Simultaneous effects of lead and cadmium on NK 

cell activity and some phenotypic parameters. Immunopharmacol Immunotoxicol 19: 339-348. 

http://dx.doi.org/10.3109/08923979709046980.

Yun, S; Hoyer, S. (2000). Effects of low-level lead on glycolytic enzymes and pyruvate dehydrogenase of rat brain 

in vitro: Relevance to sporadic Alzheimer's disease? Journal of Neural Transmission 107: 355-368. 

http://dx.doi.org/10.1007/s007020050030.

Zailina, H; Junidah, R; Josephine, Y; Jamal, HH. (2008). The influence of low blood lead concentrations on the 

cognitive and physical development of primary school children in Malaysia. Asia Pac J Public Health 20: 

317-326. http://dx.doi.org/10.1177/1010539508322697.

Zawia, NH; Harry, GJ. (1995). Exposure to lead-acetate modulates the developmental expression of myelin genes in 

the rat frontal lobe. Int J Dev Neurosci 13: 639-644. http://dx.doi.org/10.1016/0736-5748(95)00032-C.

Zawia, NH; Sharan, R; Brydie, M; Oyama, T; Crumpton, T. (1998). Sp1 as a target site for metal-induced 

perturbations of transcriptional regulation of developmental brain gene expression. Dev Brain Res 107: 

291-298. http://dx.doi.org/10.1016/S0165-3806(98)00023-6.

Zawia, NH; Basha, MR. (2005). Environmental risk factors and the developmental basis for Alzheimer's disease. 

Rev Neurosci 16: 325-337. 

Zeisel, S; Blusztajn, J. (1994). Choline and human nutrition. Annu Rev Nutr 14: 269-296. 

http://dx.doi.org/10.1146/annurev.nu.14.070194.001413.

Zelikoff, JT; Li, JH; Hartwig, A; Wang, XW; Costa, M; Rossman, TG. (1988). Genetic toxicology of lead 

compounds. Carcinogenesis 9: 1727-1732. http://dx.doi.org/10.1093/carcin/9.10.1727.

Zeller, I; Knoflach, M; Seubert, A; Kreutmayer, S; Stelzmüller, M; Wallnoefer, E; Blunder, S; Frotschnig, S; 

Messner, B; Willeit, J; Debbage, P; Wick, G; Kiechl, S; Laufer, G; Bernhard, D. (2010). Lead contributes 

to arterial intimal hyperplasia through nuclear factor erythroid 2-related factor-mediated endothelial 

interleukin 8 synthesis and subsequent invasion of smooth muscle cells. Arterioscler Thromb Vasc Biol 30: 

1733-1740. http://dx.doi.org/10.1161/ATVBAHA.110.211011.

Zeng, H; Chattarji, S; Barbarosie, M; Rondi-Reig, L; Philpot, B; Miyakawa, T; Bear, M; Tonegawa, S. (2001). 

Forebrain-specific calcineurin knockout selectively impairs bidirectional synaptic plasticity and 

working/episodic-like memory. Cell 107: 617-629. http://dx.doi.org/10.1016/S0092-8674(01)00585-2.

Zentner, LE; Rondó, PH; Mastroeni, SS. (2006). Lead contamination and anthropometry of the newborn baby. J 

Trop Pediatr 52: 369-371. http://dx.doi.org/10.1093/tropej/fml009.

Zentner, LEA; Rondo, PHC; Duran, MC; Oliveira, JM. (2008). Relationships of blood lead to calcium, iron, and 

vitamin C intakes in Brazilian pregnant women. Clin Nutr 27: 100-104. 

http://dx.doi.org/10.1016/j.clnu.2007.10.005.

Zeyrek, D; Soran, M; Cakmak, A; Kocyigit, A; Iscan, A. (2009). Serum copper and ainc levels in mothers and cord 

blood of their newborn infants with neural tube defects: A case-control study. Indian Pediatr 46: 675-680. 
February 2012 5-630 Draft – Do Not Cite or Quote

Zhang, A; Hu, H; Sánchez, BN; Ettinger, AS; Park, SK; Cantonwine, D; Schnaas, L; Wright, RO; LamadridFigueroa, H; Tellez-Rojo, MM. (In Press) Association between Prenatal Lead Exposure and Blood Pressure 

in Female Offspring. Environ Health Perspect. http://dx.doi.org/10.1289/ehp.1103736.

Zhang, A; Park, SK; Wright, RO; Weisskopf, MG; Mukherjee, B; Nie, H; Sparrow, D; Hu, H. (2010a). HFE H63D 

polymorphism as a modifier of the effect of cumulative lead exposure on pulse pressure: The Normative 

Aging Study. Environ Health Perspect 118: 1261-1266. http://dx.doi.org/10.1289/ehp.1002251.

Zhang, D; Hu, X; Qian, L; O'Callaghan, JP; Hong, JS. (2010b). Astrogliosis in CNS pathologies: Is there a role for 

microglia? Mol Neurobiol 41: 232-241. http://dx.doi.org/10.1007/s12035-010-8098-4.

Zhang, LF; Peng, SQ; Wang, S. (2005). Influence of lead (Pb2+) on the reactions of in vitro cultured rat aorta to 5-

hydroxytryptamine. Toxicol Lett 159: 71-82. http://dx.doi.org/10.1016/j.toxlet.2005.04.009.

Zhang, LF; Peng, SQ; Sheng, W; Li, BL; Gang, H; Dong, YS. (2007a). Direct effects of lead (Pb2+) on the 

relaxation of in vitro cultured rat aorta to acetylcholine. Toxicol Lett 170: 104-110. 

http://dx.doi.org/10.1016/j.toxlet.2007.02.004.

Zhang, LF; Peng, SQ; Wang, S. (2009a). Decreased aortic contractile reaction to 5-hydroxytryptamine in rats with 

long-term hypertension induced by lead (Pb2+) exposure. Toxicol Lett 186: 78-83. 

http://dx.doi.org/10.1016/j.toxlet.2009.01.004.

Zhao, Y; Wang, L; Shen, HB; Wang, ZX; Wei, QY; Chen, F. (2007). Association between delta-aminolevulinic acid 

dehydratase (ALAD) polymorphism and blood lead levels: A meta-regression analysis. J Toxicol Environ 

Health 70: 1986-1994. http://dx.doi.org/10.1080/15287390701550946.

Zhao, ZY; Li, R; Sun, L; Li, ZY; Yang, RL. (2004). Effect of lead exposure on the immune function of lymphocytes 

and erythrocytes in preschool children. J Zhejiang Univ Sci 5: 1001-1004. 

http://dx.doi.org/10.1007/BF02947614.

Zheng, C; Sköld, M; Li, J; Nennesmo, I; Fadeel, B; Henter, J. (2007). VEGF reduces astrogliosis and preserves 

neuromuscular junctions in ALS transgenic mice. Biochem Biophys Res Commun 363: 989-993. 

http://dx.doi.org/10.1016/j.bbrc.2007.09.088.

Zhu, M; Fitzgerald, EF; Gelberg, KH; Lin, S; Druschel, CM. (2010). Maternal low-level lead exposure and fetal 

growth. Environ Health Perspect 118: 1471-1475. http://dx.doi.org/10.1289/ehp.0901561.

Zimmermann, MB; Muthayya, S; Moretti, D; Kurpad, A; Hurrell, RF. (2006). Iron fortification reduces blood lead 

levels in children in Bangalore, India. Pediatrics 117: 2014-2021. http://dx.doi.org/10.1542/peds.2005-

2440.

Zoeger, N; Roschger, P; Hofstaetter, JG; Jokubonis, C; Pepponi, G; Falkenberg, G; Fratzl, P; Berzlanovich, A; 

Osterode, W; Streli, C; Wobrauschek, P. (2006). Lead accumulation in tidemark of articular cartilage. 

Osteoarthritis Cartilage 14: 906-913. http://dx.doi.org/10.1016/j.joca.2006.03.001.

Zuo, PJ; Qu, W; Cooper, RN; Goyer, RA; Diwan, BA; Waalkes, MP. (2009). Potential role of alpha-synuclein and 

metallothionein in lead-induced inclusion body formation. Toxicol Sci 111: 100-108. 

http://dx.doi.org/10.1093/toxsci/kfp132.

Zuscik, MJ; Ma, L; Buckley, T; Puzas, JE; Drissi, H; Schwarz, EM; O'Keefe, RJ. (2007). Lead induces 

chondrogenesis and alters transforming growth factor-beta and bone morphogenetic protein signaling in 

mesenchymal cell populations. Environ Health Perspect 115: 1276-1282. 

http://dx.doi.org/10.1289/ehp.10028.