bluuebunny/crossref_metadata_2025 huggingface dataset:
Subchronic Toxicity of Pentostatin in Wistar Rats
Authors: Courtney Cynthia L. , Hawkins Kathleen L. , Rothwell Charles E.
Published: April 2009

Pentostatin, an adenosine deaminase inhibitor, has been approved for the treatment of refractory hairy cell leukemia. In a preclinical toxicity study, Wistar rats were administered 0, 1, 10. 25, and 50 mg/kg (0, 6, 60, 150, and 300 mg/m2, respectively) pentostatin intravenously once a week for 26 wk (1.5-75-fold above the therapeutic dose in humans). Lymphoplasmacytic thyroiditis was present in 20% of females given 25 mg/kg and in 20 and 47% of males and females given 50 mg/kg, respectively. Thyroiditis was still present 4 wk following drug withdrawal. Thyroiditis was characterized by glandular enlargement, follicular epithelial hyperplasia and degeneration, colloid depletion, and interstitial infiltrates of lymphocytes and plasma cells. Drug-related changes in other tissues included lymphoid depletion of T-cell regions of thymus, spleen, and lymph nodes; bronchiolization of alveolar ducts with accumulation of mucus and foamy macrophages; testicular atrophy with sperm granulomas; dermoepidermal lymphocytic infiltrates with ulceration and alopecia; and hepatocytomegaly.
Characterization of a Renal Epithelial Cell Model of Apoptosis Using Okadaic Acid and the NRK-52E Cell Line
Authors: Davis Myrtle A. , Smith Mary W. , Chang Seung H. , Trump Benjamin F.
Published: April 2009

Apoptotic cell death plays an important role in the pathogenesis of renal tubular epithelial damage and repair following tubular injury. Presently, the cellular factors involved in regulating apoptotic pathways in the kidney are unknown. To address the possibility that protein phosphorylation may regulate apoptosis in kidney cells, okadaic acid (OKA), a specific inhibitor of protein phosphatases 1A and 2A, was tested for its morphologic and biochemical effects on normal rat kidney epithelial cells (NRK-52E) in culture. As revealed by the DNA-specific stain DAPI, nuclei of cells treated with 1.0 μM okadaic acid contained irregular clumps of dense chromatin. Additional morphologic alterations typical of apoptosis were apparent within 2 hr after treatment with 1.0 μM OKA, including marked cellular rounding, cytoplasmic condensation, and cytoplasmic blebs. Ultrastructurally, 1.0 μM OKA caused cytoplasmic bleb formation, cellular fragmentation, condensation of heterochromatin into clumps, and segregation of nucleoli. At this stage, the cytoplasmic fragments and blebs contained many normal mitochondria. The attached, rounded cells also effectively excluded propidium iodide, demonstrating maintenance of membrane integrity despite pronounced morphologic alterations. A 2-fold increase in intracellular free Ca2+ was apparent 90 min after treatment with 1.0 μM okadaic acid. Transverse alternating field electrophoresis revealed the appearance of large DNA fragments of approximately 300-kbp. The appearance of these 300-kbp fragments correlated temporally with the observed elevation in intracellular calcium and the onset of morphologic alterations. However, preloading cells with EGTA-AM, an intracellular calcium chelator, obliterated the calcium elevation and had no effect on OKA-induced morphology, DNA fragmentation, or cell death. Detectable internucleosomal fragmentation occurred much later than the onset of morphologic changes (24-hr treatment time) and did not correlate with elevations in cellular calcium. These studies support the hypothesis that during apoptosis, chromatin condensation reflects chromatin cleavage at nuclease-sensitive sites between hexameric rosettes. These results also suggest that morphologic and nuclear alterations in the pathway of OKA-induced apoptosis occur independent of observed increases in intracellular calcium.
Proliferating Cell Nuclear Antigen Staining of Fischer-344/N Rat Spleens Affected by Large Granular Lymphocyte Leukemia
Authors: Stefanski Steven A. , Greenwell Arnold , Merrick B. Alex , Brown Talmage T. , Reynolds Steven H.
Published: April 2009

Large granular lymphocyte (LGL) leukemia commonly occurs in the Fischer-344/N rat. The high spontaneous incidence complicates the interpretation of results from chronic carcinogenicity studies that use this rat strain. As a result, a comprehensive characterization of LGL leukemia is necessary to help understand the leukemogenic process and the applicability of staging for assessing the progression of this disease. In the current study, the proliferation rate of LGL leukemia cells from untreated control Fischer-344 (F-344) rats in 3 stages of leukemia compared to nonleukemic age-matched rats was determined by immunohistochemical staining for proliferating cell nuclear antigen (PCNA). In histologic sections of spleen from aged F-344/N rats affected by LGL leukemia, there was a significant increase of both PCNA labeling and mitotic indices that was most advanced in the spleen of rats with more severe LGL leukemia. These results support biological significance for the morphologic staging system currently in use.
Suppressive Effect of β-Carotene on the Development of Pulmonary Foam Cells in Rats with Hyper β-Lipoproteinemia
Authors: Shibuya Kazumoto , Tajima Masanori , Saitoh Toshiki , Yamate Jyoji , Nunoya Tetsuo
Published: April 2009

The effect of β-carotene (BC) on the development of pulmonary foam cells (PFCs) was studied in rats with diet-induced hyper β-lipoproteinemia. Rats were fed a standard diet; a hyper-β-lipoproteinemic diet (HB) consisting of the standard diet, 4% cholesterol, and 1% cholic acid; or the standard diet plus 0.1% BC; or the HB diet plus 0.1% BC diet (HBC). Rats in the HB and HBC groups developed hyper-β-lipoproteinemia, but no significant differences were observed in serum levels of total cholesterol, phospholipid, and β- lipoprotein (B-LP) between both groups. The percentages of foamy, lipid-ingested monocytes (FMs) to the number of blood monocytes (BMs), the number and size of lipid droplets in FMs, the percentages of PFCs to the number of alveolar macrophages from bronchopulmonary lavage fluid, and the score of PFC development in the lungs of rats in the HBC group were reduced compared to those of rats in the HB group. There were no differences in latex-phagocytotic activity of BMs among rats in the control, HB, BC, and HBC groups. BC suppressed the foamy transformation of BMs and development of PFCs deriving from the influx of FMs into the alveoli of hyper-β-lipoproteinemic rats. Based on the present results, it is presumed that the antioxidative property of BC may prevent an oxidative modification of B-LP under the hyper-β-lipoproteinemic condition, leading to a decrease in the uptake of oxidatively modified B-LP by BMs.
An Introduction to Drug Disposition: The Basic Principles of Absorption, Distribution, Metabolism, and Excretion
Authors: Caldwell John , Gardner Iain , Swales Nicola
Published: April 2009

A knowledge of the fate of a drug, its disposition (absorption, distribution, metabolism, and excretion, known by the acronym ADME) and pharmacokinetics (the mathematical description of the rates of these processes and of concentration-time relationships), plays a central role throughout pharmaceutical research and development. These studies aid in the discovery and selection of new chemical entities, support safety assessment, and are critical in defining conditions for safe and effective use in patients. ADME studies provide the only basis for critical judgments from situations where the behavior of the drug is understood to those where it is unknown: this is most important in bridging from animal studies to the human situation. This presentation is intended to provide an introductory overview of the life cycle of a drug in the animal body and indicates the significance of such information for a full understanding of mechanisms of action and toxicity.
Ancillary Approaches to Toxicokinetic Evaluations
Authors: Takacs Adrienne R.
Published: April 2009

The principal objective of toxicokinetic studies is to assess systemic exposure of the toxicity species to test compound during nonclinical toxicity studies by generation of pharmacokinetic data on the rate, extent, and duration of exposure. Toxicokinetics are normally integrated within the toxicity studies (concomitant); however, there are a number of circumstances when prospective, retrospective, or other ancillary types of toxicokinetic studies are performed in support of toxicity studies. The need for retrospective toxicokinetic studies arises when the original toxicity study was conducted with insufficient plasma concentration determinations, especially when pilot toxicity studies show lack of toxicity or unusual organ toxicities. Examples of ancillary toxicokinetic studies from the Drug Metabolism and Drug Safety departments at The R. W. Johnson Pharmaceutical Research Institute and from the scientific literature are provided that highlight the challenges associated with these alternate approaches. Each example includes a discussion of the rationale for the supportive toxicokinetic study, the specific experimental designs, the data interpretation, and the usefulness of the toxicokinetic data in decision-making. The ancillary toxicokinetic studies cited provided important information on establishing systemic exposure during long-term toxicity studies, allowing comparisons to human exposures, elucidating compound-related toxicities, and explaining lack of toxicity due to autoinduction and resulting low levels of parent compound.
Diet, Overfeeding, and Moderate Dietary Restriction in Control Sprague-Dawley Rats: I. Effects on Spontaneous Neoplasms
Authors: Keenan Kevin P. , Soper Keith A. , Smith Peter F. , Ballam Gordon C. , Clark Robert L.
Published: April 2009

This study was designed to compare the effects of ad libitum (AL) overfeeding and moderate dietary restriction (DR) of two different diets on Sprague-Dawley (SD) rat 2-yr survival and the development of spontaneous neoplasms. SD rats were fed Purina Rodent Chow 5002 or a modified Rodent Chow 5002-9 containing lower protein, fat, metabolizable energy and increased fiber by AL or by DR at 65% of the AL amount by measurement or time (6.5 hr). At 106 wk, rats fed the 5002-9 diet AL did not have significantly improved survival over rats fed the 5002 diet AL. The 5002 diet fed DR by time (6.5 hr) improved survival for males but not females. Only DR by measurement of both diets resulted in lower mortality for both sexes. The most common cause of death in rats of both sexes fed either diet AL was pituitary tumors followed by mammary gland tumors in females and renal and cardiovascular disease in males. The overall tumor incidence by 106 wk was remarkably similar between AL and DR groups. However, compared to the 5002 AL group, a decrease in the age-adjusted (Peto analysis) incidence of pituitary adenoma was observed in all other male groups. This effect was noted in the female DR by measurement groups only. For males, compared to the 5002 AL group, a decrease in the age-adjusted incidence of pancreatic islet carcinoma was observed in the DR by measurement groups only. In females, compared to the 5002 AL group, the only other difference in tumor incidence was the mammary gland tumors, which showed a significant decrease in the age-adjusted tumor incidence or multiplicity in the 5002-9 AL, 5002-9 DR, and 5002 DR groups. Additional analyses of mammary gland tumors showed growth time (time from initial palpation until death), tumor doubling time, and tumor volume were generally not statistically significantly different between AL and DR groups, although AL females could sustain larger tumor volumes. Compared to the 5002 AL group, there were no other significant differences in the age-adjusted incidence of any other tumor site in animals fed a modified diet or subjected to moderate DR of either diet. The conclusion from this study is that moderate DR delays death due to fatal cardiovascular or renal degenerative disease and spontaneous tumors, particularly those of the pituitary and mammary gland. However, moderate DR appears only to delay the time of onset, but not the progression, of these spontaneous tumors whether measured by age-adjusted incidence, growth time, tumor doubling time, or the time between initial detection and death.
Indirect Immunohistochemistry and Immunoelectron Microscopy Distribution of Eight Epidermal-Dermal Junction Epitopes in the Pig and in Isolated Perfused Skin Treated with Bis (2-Chloroethyl) Sulfide
Authors: Monteiro-Riviere Nancy A. , Inman Alfred O.
Published: April 2009

Sulfur mustard (bis [2-chloroethyl] sulfide, HD) is a potent cutaneous vesicant that causes gross blisters by separation of the epidermal-dermal junction (EDJ). The EDJ of the skin is a highly specialized and complex structure composed of several components and plays a major role in the integrity of the skin. The isolated perfused porcine skin flap (IPPSF) was dosed with 0.2 mg/ml (n = 4), 5.0 mg/ml (n = 4), and 10.0 mg/ml (n = 5) HD or ethanol (n = 4) for 8 hr (dose-response study) and 10.0 mg/ml HD or ethanol for 1, 3, 5, and 8 hr (n = 4/treatment) (time-response study). Successful EDJ mapping was carried out in normal pig skin (NPS), ethanol-treated IPPSFs, and HD-treated IPPSFs using the following antibodies: laminin, type IV collagen, fibronectin, GB3 (Nicein), bullous pemphigoid (BP), and epidermolysis bullosa acquisita (EBA). Two mouse anti-human monoclonal antibodies, L3d and 19-DEJ-l (Uncein), did not cross-react with the EDJ of the pig. Antibody staining in NPS, ranging from very intense for laminin and type IV collagen to weak for fibronectin, was generally more discrete than in the IPPSF. No differences in staining were noted between the ethanol and nonblistered areas of the HD-treated IPPSFs. In HD-blistered areas, BP stained only the epidermal hemidesmosomes, and laminin, fibronectin, and GB3 stained primarily the dermis with fragments attached to the basal pole of the stratum basale cells, while type IV collagen and EBA stained only the dermis. Mapping of these epitopes determined that the precise plane of EDJ separation in the HD-treated skin occurred beneath the hemidesmosomes within the upper portion of the lamina lucida. The conservation of human epitopes in the EDJ of the pig further emphasizes the similarities between human skin and pig skin. Therefore, pig skin and the IPPSF may be used to study HD-induced vesication and blistering diseases.
Incidence of Nonneoplastic Lesions in Historical Control Male and Female Fischer-344 Rats from 90-Day Toxicity Studies
Authors: Dixon Darlene , Heider Katharina , Elwell Michael R.
Published: April 2009

The incidence of all spontaneously occurring histologic lesions was determined for control Fischer-344 (F-344) rats from 90-day (13-wk) prechronic National Toxicology Program (NTP) toxicity studies. A total of 319 rats, represented by control groups of 10 males and 10 females each from dosed feed ( n = 8), inhalation ( n = 4), and gavage ( n = 4) studies were included in the review. All protocol required tissues routinely collected for evaluation were reexamined for potential nonneoplastic and neoplastic lesions. Histopathologic findings in tissues included a spectrum of degenerative and inflammatory lesions. The most common lesions in male rats were nephropathy [145/160 (90.6%)] and cardiomyopathy [125/158 (79.1%)]. These changes were also present in the female rats, however, at much lower incidence rates [nephropathy = 30/157 (19.1%); cardiomyopathy = 36/158 (22.8%)]. Other less frequently occurring lesions included inflammation of the preputial [36/152 (23.7%)] and clitoral [34/155 (21.9%)] glands and inflammation of the liver consisting of either foci of mononuclear inflammatory cells [19/159 (11.9%) in males and 33/159 (20.8%) in females] or focal granulomatous inflammation [1/159 (0.6%) in males and 14/159 (8.8%) in females]. Pancreatic acinar cell atrophy occurred in both males [11/160 (6.9%)] and females [8/159 (5.0%)]. A variety of other less common nonneoplastic lesions were identified in both sexes of rats. Also recorded in this review are histologic changes generally considered to be components of the normal morphology of a particular tissue or organ for the F-344 rat (i.e., extramedullary hematopoiesis and hemosiderin deposition in the spleen, renal mineralization, uterine dilation, etc.). These findings were included and discussed due to potential treatment effects that may result in an increase or decrease in these changes compared to controls. Neoplasms were not observed in rats from the prechronic studies evaluated.
Apoptosis in the Central Nervous System of Developing Mouse Fetuses from 5-Azacytidine-Administered Dams
Authors: Hossain Mokbul Md. , Nakayama Hiroyuki , Goto Naoaki
Published: April 2009

To determine the effect of 5-azacytidine (5Az), a cytidine analogue, on the developing fetus, different strains of mice were injected with various doses of the agent, ip, at 11 days of gestation. Light microscopically, severely pyknotic cell death profiles were observed in different layers of the developing brain and spinal cord (central nervous system) on the day after the injection. Electron microscopy of the pyknotic cells revealed the shrinkage of the cell body, to various degrees margination of the nuclear chromatin, condensation of nuclear materials, fragmentation of the affected cells, and, finally, neat "bite-size" bodies engulfed by surrounding normal neuroepithelial cells and macrophage-like cells. DNA fragmentation was detected in the nuclei of the pyknotic cells by an in situ detection method for fragmented DNA ends. These changes were consistent with the criteria of apoptosis.
Histomorphogenesis of Soman-Induced Encephalocardiomyopathy in Sprague-Dawley Rats
Authors: Tryphonas Leander , Clement John G.
Published: April 2009

Although myocardial damage caused by soman has been previously reported, its relation to brain damage is unclear. In order to clarify this relationship, we examined the histomorphogenesis of central nervous system (CNS) and myocardial lesions in Sprague-Dawley rats, given atropine methylnitrate (20 mg/kg) and HI-6 (125 mg/kg) ip 10 min before a single injection of 0 or 130 μg soman/kg (sc) and sacrificed 45 min and 1.5 hr, 3 hr, 24 hr, and 72 hr later. Bilaterally symmetrical CNS damage began with vacuolation of the neuropil and was followed by astrocytic degeneration and neuronal necrosis culminating in liquefaction necrosis and focal hemorrhage. The cerebral cortex, limbic system, thalamus, and substantia nigra were common target sites. Repair in affected sites was characterized by capillary endothelial cell proliferation, microgliosis, and reversal of microvacuolation. Myocardial damage began with myocytolysis and contraction bands and evolved into coagulative myocytolysis and replacement fibrosis with a transient recruitment of acute inflammatory cells. The left ventricle, especially its free wall and papillary muscles, was consistently affected. There was good correlation among seizures, CNS damage, and myocardial lesions at all times following treatment. The results support the view that CNS lesions are associated with protracted seizure activity and provide evidence that myocardial damage is neurogenic.
An Overview of Current Cytochrome P450 Technology for Assessing the Safety and Efficacy of New Materials
Authors: Parkinson Andrew
Published: April 2009

Studies of xenobiotic biotransformation by in vitro techniques are destined to play an increasingly important role in assessing the safety and efficacy of drugs and other new chemical entities. The first part of this article summarizes some of the in vitro techniques that have been developed to evaluate xenobiotics as inducers of liver microsomal cytochrome P450. The second part provides an overview of reaction phenotyping, an in vitro technology for determining which human P450 enzyme or enzymes are involved in the biotransformation of xenobiotics.
Use of Precision-Cut Liver Slices as an In Vitro Tool for Evaluating Liver Function
Authors: Gandolfi A. Jay , Wijeweera Jayanthika , Brendel Klaus
Published: April 2009

Precision-cut liver slices have been developed as an in vitro tool for assessing liver viability and function and for examining hepatotoxicants. Liver slices from a variety of species (including human) are prepared using mechanical slicers that produce reproducible slices of a uniform thickness, which allows optimum exchange of nutrients, waste, and gases. Slices are incubated in dynamic systems that allow the slices to be maintained viable in culture for 1-10 days. The viability of slices can be assessed by ion content (K+, Na+ATPase status), intermediary metabolism, energy status (ATP), respiration, biosynthetic ability, and biotransformation activity. In addition, liver tissue slices allow the opportunity for extensive microscopic evaluation (light and electron) as well as newer technologies such as confocal microscopy. Assessment of the toxic potential of a chemical can be performed after a short-term or constant exposure by evaluating the viability parameters. Liver slices have been used extensively for rank-ordering the toxicity of chemicals as well as for examining the mechanisms of liver injury. Liver slices in culture also can be used for an examination of the induction of new enzymes such as cytochrome P-450 and the expression of stress proteins or peroxisomal enzymes. Finally, liver slices offer a system for evaluating whole or cryopreserved liver as well as regeneration of liver tissue after toxic insult. Liver slices have been shown to be a valid in vitro system for examining liver function and offer a bridge between in vivo and cell culture systems.
Simultaneous Measurement of Hundreds of Liver Proteins: Application in Assessment of Liver Function
Authors: Anderson N. Leigh , Taylor John , Hofmann Jean-Paul , Esquer-Blasco Ricardo , Swift Susan , Anderson Norman G.
Published: April 2009

Proteins implement most biological functions at the molecular level. As one might expect based on this fact, it appears that the altered functional states associated with toxic effects involve changes in the abundance or structure of proteins. Although numerous specific assays exist to measure changes in the abundance of individual proteins, practical limitations have prevented widespread use of multiple protein assays for the global characterization of toxicity. Recent developments in protein analytical technology are rapidly changing this picture. Two-dimensional gel electrophoresis, a technique capable of resolving and quantitating hundreds of proteins simultaneously, is becoming an automated, high-throughput tool. In parallel, techniques have been developed that allow the resulting deluge of protein measurements to be organized into a prototype Molecular Effects Database™ describing xenobiotic effects in rodent liver. This database can detect, classify, and characterize a broad range of liver toxicity mechanisms. It currently contains approximately 10 million protein measurements, including data on the liver effects of 43 compounds, with a further 50 compounds to be added in 1995. Observed effects range from very broad (sex steroids alter levels of 45% of all liver proteins) to very specific (e.g., hepatic hydroxymethyl glutaryl coenzyme A reductase inhibitors). Companion 2-dimensional databases describing rodent brain and kidney have been initiated, as have linkages to the genomic sequence databases. Assimilation of this approach into research and regulatory toxicology poses an interesting challenge—one that is likely to lead to a radically more sophisticated understanding of toxicity and its biological basis.
Biochemical Basis of Hepatocellular Injury
Authors: Kedderis Gregory L.
Published: April 2009

The hepatotoxic response elicited by a chemical agent depends on the concentration of the toxicant (parent compound or metaholite) delivered to the hepatocytes across the liver acinus via blood flow. Hepatotoxicants produce characteristic patterns of cytolethality in specific zones of the acinus due to the differential expression of enzymes and the concentration gradients of cofactors and toxicant in blood across the acinus. Most hepatotoxic chemicals produce necrosis, characterized by swelling in contiguous tracts of cells and inflammation. This process has been contrasted with apoptosis, where cells and organelles condense in an orderly manner under genetic control. Biotransformation can activate a chemical to a toxic metabolite or decrease toxicity. Quantitative or qualitative species differences in biotransformation pathways can lead to significant species differences in hepatotoxicity. Fasted rodents are more susceptible to the hepatotoxic effects of many chemicals due to glutathione depletion and cytochrome P-450 induction. Freshly isolated hepatocytes are the most widely used in vitro system to study mechanisms of cell death. Hepatotoxicants can interact directly with cell macromolecules or via a reactive metabolite. The reactive metabolite can alkylate critical cellular macromolecules or induce oxidative stress. These interactions generally lead to a loss of calcium homeostasis prior to plasma membrane lysis. Mitochondria have been shown to be important cellular targets for many hepatotoxicants. Decreasing hepatocellular adenosine triphosphate concentrations compromise the plasma membrane calcium pump, leading to increased cellular calcium concentrations. Calcium-dependent endonucleases produce double-strand breaks in DNA before cell lysis. These biochemical pathways induced by necrosis-causing toxicants are similar to the biochemical pathways involved in apoptosis, suggesting that apoptosis and necrosis differ in intracellular and extracellular control points rather than in the biochemistry involved in cell death.
Quantitation of Multistage Carcinogenesis in Rat Liver
Authors: Pitot Henry C. , Dragan Yvonne P , Teeguarden Justin , Hsia Steven , Campbell Harold
Published: April 2009

A well characterized model of multistage carcinogenesis is that of hepatocarcinogenesis in the rat. The histopathology as well as the cell and molecular biology of the stages of initiation, promotion, and progression have been elucidated to varying degrees in this system. Putatively single initiated hepatocytes are identified by their expression of the ubiquitous marker of hepatocarcinogenesis, glutathione-S-transferase π (GSTP). 0.5-1.0 X 106 GSTP-positive "initiated" hepatocytes developed within 14 days after initiation with a subcarcinogenic dose of diethylnitrosamine (DEN). Approximately 1% of these cells develop clonally into altered hepatic foci (AHF) in animals administered promoting agents, such as phenobarbital, chronically for 4-8 mo. Hepatocytes within AHF during the stage of promotion exhibit normal diploid karyotypes but various phenotypes depending on the chemical nature of the promoting agent. Continued administration of the promoting agent results in the infrequent development of hepatocellular carcinomas; however, administration of a complete carcinogen or a progressor agent during the stage of promotion results in substantial numbers of hepatic neoplasms. In order to quantitate the development of the stage of progression more accurately, markers selective for this stage have been sought. Transforming growth factor-α (TGF-α) appears to be such a marker of progression. About 500 TGF-α-positive lesions develop spontaneously following initiation and continued promotion, usually within GSTP-positive AHF, but administration of a single dose of a progressor agent such as ethylnitrosourea may increase this number 3-fold or more. Some agents such as γ radiation and hydroxyurea, when administered as single or a few closely spaced multiple doses, result in no increased number in TGF-α-positive lesions but a markedly enhanced increase in their growth rate. By monitoring gene expression using quantitative stereology, the stages of hepatocarcinogenesis can be analyzed and quantified in sufficient detail so that the animal data can be utilized in biomathematical modeling to develop more accurate models for estimation of human cancer risks.
Enhanced Arsenite-Induced Hepatic Morphological and Biochemical Changes in Phenobarbital-Pretreated Rats
Authors: Albores Arnulfo , Cebrián Mariano E. , García-Vargas Gonzalo G. , Connelly John C. , Price Shirley C. , Hinton Richard H. , Bach Peter H. , Bridges James W.
Published: April 2009

Changes in liver morphology and biochemistry have been assessed 16 hr after a sc injection of sodium arsenite [As(III), 75 μmol/ kg] to control and phenobarbital (PB)-pretreated (80 mg/kg, ip daily for 3 days) adult male Wistar rats. As(III) administration to PBpretreated rats [PB + As(III)] caused hydropic degeneration, total loss of glycogen, necrosis in some centrilobular zones, and an increase in lipid vacuoles around the periportal area. Electron microscopy showed an increased number of vacuoles and autophagosomes containing organelle-like material. There was a 30% decrease in total hepatic cytochrome P-450 (CYP450). O -dealkylation of ethoxy- and pentoxyresorufin and N-demethylation of benzphetamine decreased to 42, 32, and 30% of control values, respectively. 5-Aminolevulinic acid dehydrase decreased 25% from controls, and metal chelatase activities decreased to 25% of the PB-treated group. Injection of As(III) alone resulted in a mild increase in lipid-containing vacuoles around the periportal zone, a moderate loss of glycogen in midzonal areas, and, by electron microscopy, a dilatation of the bile canaliculi and an increase of the number of myelin-like structures. CYP450 content and the O-dealkylation of ethoxy- and pentoxyresorufin and N-demethylation of benzphetamine all decreased between 30 and 50%. These results demonstrate the greater susceptibility of the liver to injury following PB with compounds not requiring metabolic activation. The metabolic basis of these changes are unclear but may result from an increased demand for metabolic energy due to PB induction and decreased adenosine triphosphate synthesis caused by arsenic.
Histopathology of Acetaminophen-Induced Liver Changes: Role of Interleukin 1α and Tumor Necrosis Factor α
Authors: Blazka Mark E. , Elwell Michael R. , Holladay Steve D. , Wilson Ralph E. , Luster Michael I.
Published: April 2009

Administration of 500 mg/kg acetaminophen (APAP) to female B6C3F1 mice resulted in well-documented pathophysiological changes in the liver manifested as increased serum concentration of liver enzymes (aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and serum sorbitol dehydrogenase), centrilobular congestion, and hepatocellular degeneration and necrosis. The role of proinflammatory cytokines, including tumor necrosis factor a (TNF-α) and interleukin 1α (IL-1α), on the hepatotoxicity of APAP was examined at 4,8, 12, and 24 hr following APAP administration. Neutralization of TNF-α or IL-1α with specific antibodies partially prevented the hepatotoxic effects of APAP at the 4- and 8-hr time points. In addition, prior administration of anti-TNF-α antibodies shortened the recovery time following APAP treatment. While IL-1 receptor antagonist (IL-1ra) had only a modest protective effect against APAP-induced liver damage, as determined by serum enzyme release, IL-1ra had no effect on the degree of hepatic congestion or necrosis at any of the time points examined. On the other hand, administration of antibodies against IL-1ra exacerbated APAP-induced liver toxicity. These results suggest that TNF-α and IL-1α play an important role in the degree of damage and recovery that the liver undergoes following APAP intoxication.
Chemicals Associated with Decreases in the Incidence of Mononuclear Cell Leukemia in the Fischer Rat
Authors: Elwell Michael R. , Dunnick June K. , Hailey James R. , Haseman Joseph K.
Published: April 2009

A significant treatment-related decrease in the incidence of mononuclear cell leukemia (MCL) was identified in Fischer-344/N rats for 20 chemicals tested in the National Toxicology Program's 2-yr carcinogenicity bioassay. Fourteen of the 20 chemicals caused decreases of MCL in both male and female rats; 6 of the 20 caused a significant decrease only in males and a marginal or no decrease in female rats. Seventeen of the chemicals associated with a decrease in MCL had a free aromatic amine or nitro functional groups that could be metabolized to free amines. With 1 exception, all 14 chemicals causing a decrease in MCL in both sexes produced spleen toxicity in the 13-wk studies. Reduced body weight and decreased survival, related either to toxicity or to an increase in other types of lethal neoplasms, did not contribute to the decreases in MCL observed in chemical exposure groups. Thirteen of the 20 chemicals were positive in Salmonella tests, and 15 were associated with increases in neoplasms at other sites in rats and/or mice, suggesting that different metabolites could be responsible for these varied biological effects.
Extensive Handling of Rats Leads to Mild Urinary Bladder Hyperplasia
Authors: Cohen Samuel M. , Cano Martin , Anderson Traci , Garland Emily M.
Published: April 2009

The urinary and urothelial effects of the frequent handling necessary for obtaining fresh-voided urine specimens were evaluated in 5-wk-old male F-344 rats fed control diet or diet containing 7.5% sodium saccharin. Frequent handling consisted of holding rats by the back of the neck in a position to obtain fresh-voided urine directly into centrifuge tubes 3 times per week for 10 weeks, whereas seldomly handled control rats received this treatment only twice during the entire 10 weeks. The urothelium of frequently handled rats fed control diet showed superficial necrosis and regenerative hyperplasia as observed by light and scanning electron microscopy. These changes were not observed in rats fed control diet that were seldomly handled. The necrosis and hyperplasia were not as pronounced in frequently handled rats fed control diet as in seldomly handled, sodium-saccharin-treated rats, but handling also potentiated the severity of the changes produced by sodium saccharin feeding. The urothelial exfoliation and consequent regenerative hyperplasia are likely secondary effects of stress.
Hepatic Neoplasms from Internally Deposited 144CeCl3
Authors: Hahn Fletcher F. , Muggenburg Bruce A. , Boecker Bruce B.
Published: April 2009

Fifty-five dogs were exposed by inhalation to graded activity levels of 144CeCl3, a relatively soluble form of the β-emitting radionuclide. A large portion of the 144Ce translocated from lung to liver and skeleton. Significant radiation doses were delivered to the respiratory tract, liver, and skeleton; however, the liver received the greatest cumulative absorbed dose. Liver tumors were the most frequently observed neoplasms in these exposed dogs and included 7 primary hepatic hemangiosarcomas, 1 cholangiocarcinoma, 1 hepatocellular carcinoma, 1 fibrosarcoma, 4 biliary cystadenomas, and 1 fibroma. The dose to the liver in these dogs ranged from 11 to 250 Gy with a median of 57 Gy. Autoradiographs showed a relative uniform distribution of β dose to the liver. All the malignant tumors and 1 cystadenoma were primary causes of death. The morphologic features of the hemangiosarcomas and associated hepatic lesions were similar to those described for hemangiosarcomas induced in people exposed to Thorotrast. Biliary cystadenomas were associated with degenerative lesions in the liver but not with other neoplasms in the liver. These results indicate that the liver is an important target organ for effects from internally deposited 144Ce.
Review Article
Authors: Kuijpers Marga H.M. , Van De Kooij Aart J. , Slootweg Pieter J.
Published: April 2009

Microscopic examination of the incisors of rats and mice may reveal toxicologically significant changes. First, the incisor morphology reflects the nutritional status of the animal: fluctuations of mineral metabolism and vitamin availability are disclosed by the rodent incisors, because the incisors continue to grow during life. Similarly, direct or indirect changes of mineral metabolism by a test substance are reflected in the morphological appearance of the incisor dentin. In addition, hormonal disturbances may give rise to typical structural alterations of the incisor in the test animal. Certain chemicals may have deleterious effects upon the odontogenic tissues, resulting in tooth malformation and malocclusion and eventually in odontomas. Apparent nasal tumors may turn out to be of dental origin. Nasal luminal masses that are discussed within this scope are dental malformation, dental callus, and true odontogenic tumors. According to our experience, odontogenic tumors might possibly develop within the scope of a reaction to mechanical tooth trauma as well. In carcinogenicity studies, this consideration deserves attention when evaluating treatment-related putative odontogenic tumors.
Application ofIn VivoConfocal Microscopy to the Understanding of Surfactant-Induced Ocular Irritation*Oryzias latipes
Authors: Jester James V. , Maurer James K. , Petroll W. Matthew , Wilkie David A. , Parker Ron D. , Cavanagh H. Dwight
Published: April 2009

The purpose of this study was to assess the ability of in vivo confocal microscopy (CM) to provide noninvasively derived histopathologic correlates of surfactant-induced eye irritation from which specific pathologic mechanisms can be identified and later evaluated in alternative in vitro models. Rats and rabbits, divided into groups of 5, received 10 μl of an anionic or cationic surfactant in one eye with the other eye used as a control. At specified times, eyes were examined and scored for ocular irritancy using a penlight and slit-lamp. Subsequently, corneas were evaluated by in vivo CM to evaluate epithelial layer thickness and surface epithelial cell area, corneal thickness, depth of necrosis, inflammation, fibrosis, and endothelial injury. At 3 hr, the anionic surfactant produced slight irritation with peak scores of 12.4 and 8.0 out of a possible 110 in the rats and rabbits, respectively. In vivo CM revealed changes limited to the corneal epithelium that decreased in thickness to 78% in rats and 81% in rabbits at 3 hr. This decrease in the thickness correlated with a significant decrease in surface epithelial cell area from 2,061 ± 395 μm2to 567 ± 330 μm2in the rats and 1,523 ± 185 μm2to 934 ± 71 μm2in the rabbits (p < 0.005 and 0.005, respectively). The cationic surfactant produced severe irritation in both the rats and rabbits with peak scores of 85.4 and 80.2 occurring at day 2, respectively. In vivo CM in the rats showed complete loss of corneal epithelium, lysis of keratocytes, and loss of corneal endothelium. In the rabbits, injury appeared limited to the anterior cornea with complete loss of epithelium and loss of keratocytes extending to 52% of the corneal thickness. These findings establish the application of noninvasive, in vivo CM to qualitatively and quantitatively characterize the pathobiology of ocular irritation in situ. This information will be important in the development and evaluation of mechanistically based in vitro alternatives for ocular irritancy testing.
Subchronic Toxicity of Atorvastatin, a Hydroxymethylglutaryl-Coenzyme A Reductase Inhibitor, in Beagle Dogs
Authors: Walsh Kathleen M. , Albassam Mudher A. , Clarke David E.
Published: April 2009

The toxicity of atorvastatin (AT), an inhibitor of hydroxymethylglutaryl-coenzyme A reductase (HMG), was evaluated in beagle dogs. In 4 studies [2-wk rising dose (daily increasing doses for 1 wk; maintenance for 1 wk), 12-wk rising dose (daily dosing with weekly increases in dose), 2-wk toxicity (daily dosing for 2 wk; 3 dose levels), 13-wk toxicity (daily dosing for 13 wk; 3 dose levels)], dogs received up to 400 mg/kg orally. Doses of 180 mg/kg induced moribundity, necessitating euthanasia. Weight losses up to 26% were seen at doses ≥150 mg/kg. Decreases in cholesterol levels were dose-related. Alanine and/or aspartate aminotransferase were increased at doses ≥80 mg/kg; alkaline phosphatase was increased at doses ≥150 mg/kg. Histopathologic findings were seen at ≥150 mg/kg and included hepatocellular eosinophilia related to increased smooth endoplasmic reticulum and cholangiohepatitis and cholecystitis at 150 mg/kg in the 2-wk toxicity study; hepatocellular degeneration, centrilobular bridging, cholecystitis, hemorrhage in gallbladder and brain, demyelination of optic nerve, and skeletal muscle necrosis at ≥280 mg/kg in the 12-wk rising dose study; and erosion and hemorrhage in large intestine, hepatocellular degeneration and necrosis, and inflammation and necrosis of gallbladder epithelium at 320 mg/kg in the 2-wk rising dose study. Doses up to 80 mg/kg for 13 wk did not induce histopathologic lesions in examined organs. AT effectively lowered serum cholesterol in normal lipidemic dogs. Toxicity of AT in dogs was similar to that with other inhibitors of HMG except that lenticular changes were not seen, significant hepatic, testicular, or neurological toxicity was associated only with high doses of AT, and skeletal muscle changes similar to those described in rats and rabbits were identified.
Review Article: Experimental Chemical Carcinogenesis in Fish
Authors: Bunton Tracie E.
Published: April 2009

Tracie Eileen Bunton received a DVM degree from Michigan State University in 1977. After practicing small animal medicine for a year, she completed a combined Comparative Pathology residency/Ph.D. program in 1982 at the University of California at Davis. Her residency training included 3 yr at the California Regional Primate Research Center and, for her thesis, she studied the effects of glucocorticosteroids on lung development in the fetal rhesus macaque.

Dr. Bunton spent 1982-1984 as an Assistant Professor at Michigan State University, becoming a diplomate of the American College of Veterinary Pathologists in 1983. In 1984 she joined the faculty of the Division of Comparative Medicine at the Johns Hopkins University School of Medicine, where she is now an Associate Professor, with adjunct faculty positions in the Department of Pathology and the Graduate Program in Cellular and Molecular Medicine.

Dr. Bunton's primary research interest is on mechanisms of toxicity and carcinogenesis in fish related to environmental contamination and on the development of alternative models.
Potentiation of Inhaled Staphylococcal Enterotoxin B-Induced Toxicity by Lipopolysaccharide in Mice
Authors: Leclaire Ross D. , Hunt Robert E. , Bavari Sina , Estep James E. , Nelson Gene O. , Wilhelmsen Catherine L.
Published: April 2009

Nonhuman primates are the established model for evaluating toxic responses to staphylococcal enterotoxins (SEs), as they react similarly to humans. Rodents are generally considered unresponsive to SEs. Binding affinities and T-cell reactivity suggest that SE binds more efficiently to primate major histocompatability complex class II receptors than to mouse receptors. We investigated the potentiation of staphylococcal enterotoxin B (SEB) inhalation toxicity by lipopolysaccharide (LPS) in BALB/c mice. Lethality occurred only when SEB was potentiated by LPS. Neither SEB nor LPS produced lethal effects alone. Temporal responses of interleukin 1α, tumor necrosis factor α, interleukin 2, and interferon-γ evoked by inhaled SEB were enhanced by LPS. By 24 hr after intoxication, serum cytokines decreased to baseline levels, and consistent pulmonary perivascular leukocytic infiltrates were evident histologically. Histologic lesions induced by inhalation exposure to SEB by mice, with or without potentiation by LPS, were similar to those in the rhesus monkey. Predominant pulmonary lesions included severe, diffuse interstitial and alveolar pulmonary edema, leukocytic infiltrates, mild perivascular edema, and alveolar fibrin deposition. Although the mechanism of aerosolized SEB-induced toxicity has not been completely resolved, similarities in histologic lesions, cytokine responses, and acute dose-response suggest the LPS-potentiated mouse model may be a credible alternative to the nonhuman primate model.
Posterior Lens Rupture in Rats
Authors: Tanaka Kohji , Inagaki Satoru , Matsumoto Hiroyoshi , Ikemoto Fumihiko , Doi Kunio
Published: April 2009

Rupture of the posterior lens capsule with extrusion of lens fibers or entire lens cortex and nucleus was found in Sprague-Dawley rats. The lesion was detected during an ophthalmoscopic examination as round shadows that interfered with the view of the fundus. By slit-lamp biomicroscopic and dissecting microscopic examinations, the shadows were related to opaque plaques located on the posterior capsule. By light microscopic examination, the posterior lens capsule was ruptured and lens fibers extruded into the vitreous. Mononuclear and multinucleated giant cells were present at the site of rupture, however, few other inflammatory responses were present. Although the incidence and pathogenesis of the lesion is unknown, the lesion should be considered along with other spontaneous ocular changes when evaluating safety assessment studies.
Immunohistochemical Demonstration of the Gap Junctional Protein Connexin 32 and Proliferating Cell Nuclear Antigen in Glutathione S-Transferase Placental Form-Negative Lesions of Rat Liver Induced by Diethylnitrosamine and Clofibrate
Authors: Ito Sechi , Tateno Chise , Tuda Mina , Yoshitake Akira
Published: April 2009

The distributions of a gap junctional protein, connexin 32 (cx 32), and proliferating cell nuclear antigen (PCNA) were examined immunohistochemically in glutathione S-transferase placental form (GST-P)-negative foci, induced in rat liver by initiation with diethylnitrosamine (DEN, 200 mg/kg) followed by promotion with clofibrate (1% in diet) in an in vivo medium-term assay system for hepatocarcinogenesis. The results were compared to those in GST-P-positive foci induced by DEN alone. The treatment with clofibrate caused the appearance of GST-P-negative foci, increased in size as compared to GST-P-positive foci in the same liver or induced by the DEN alone. The proportion of PCNA-positive hepatocytes in GST-P-negative foci was significantly higher than in the surrounding parenchyma, indicating increased cell proliferation. The numbers of cx 32-positive spots per hepatocyte in GST-P-negative foci were clearly decreased, reaching 65.4% at week 20 and 51.8% at week 30 of values for surrounding normal hepatocytes. In GST-P-positive foci induced by DEN, only a slight decrease (80%) was observed at week 8. These findings show that a positive association between the sustained inhibition of gap junctional intercellular communication and increased cell proliferation of GST-P-negative foci in Fischer-344 male rats induced with DEN and promoted with clofibrate.
Clinical and Histological Assessment of Collagen-Induced Arthritis Progression in the Diabetes-Resistant BB/Wor Rat
Authors: Knoerzer Debbie Barney , Donovan Matt G. , Schwartz Benjamin D. , Mengle-Gaw Laurel J.
Published: April 2009

Collagen-induced arthritis in the diabetes-resistant BB (DR BB)/Wor rat is a severe, aggressive disease initiated by immunization with heterologous native Type II collagen. Onset of clinical symptoms reproducibly occurs in 100% of animals between days 10 and 12 following collagen immunization. Hypertrophy of the synovial lining is the first histological manifestation of the early inflammatory arthritis. A mild inflammatory infiltrate in the synovium rapidly becomes a fibrovascular pannus eroding articular cartilage and subchondral bone. Beginning at the joint margins, an active synovitis is present. Light microscopy and immunohistochemical staining show the infiltrate to be comprised of mononuclear (lymphocytes, macrophages) and polymorphonuclear inflammatory cells. In addition, there is histological evidence for chronic inflammatory nodules and necrotizing vasculitis in connective tissue from diseased joints, both morphologic features associated with rheumatoid arthritis in humans. Subchondral bone erosion appears to be mediated largely by the resorptive action of activated osteoclasts. These histological parameters of disease progression in the DR BB/Wor rat are similar to human rheumatoid arthritis. The extensive degree of similarity in the pathology of DR BB/Wor rat collagen-induced arthritis and human rheumatoid arthritis supports the role of this model as an in vivo disease model for human rheumatoid arthritis.