bluuebunny/crossref_metadata_2025 huggingface dataset:
Potentiation of carbon tetrachloride hepatotoxicity by chlordecone: Dose‐response relationships and increased covalent binding in vivo
Authors: Britton Robert S. , Dolak James A. , Glende Eric A. , Recknagel Richard O.
Published: October 2006

Chlordecone greatly potentiates carbon tetrachloride (CC14) hepatotoxicity. In order to quantitate the degree of this potentiation, the effects of a range of doses of CC14 on two microsomal enzymatic functions and liver enzyme release were examined in chlordecone‐treated and control rats. Male Sprague‐Dawley rats were pretreated with 15 mg chlordecone per kilogram body weight (BW) intragastrically or with vehicle. After 48 hours, 0 to 250 μ1 CC14 per 100 g body weight were given intraperitoneally (IP), and the rats were killed 24 hours later. Chlordecone treatment produced approximately a 17‐fold potentiation of the CC14 dependent loss of cytochrome P‐450 and glucose‐6‐phosphatase activity, so that a dose of 6 μ1 CC14 per 100 g body weight in the chlordecone‐treated animals resulted in a similar amount of damage as observed with 100 μ1 CC14 per 100 g body weight in controls. A similar potentiation by chlordecone was seen with CC14‐ induced increases in serum glutamic‐oxaloacetic transaminase (SGOT) levels. Chlordecone treatment also increased hepatic cytochrome P‐450 levels by 67% and resulted in an increase in the covalent binding of [14‐C]‐CC14‐derived metabolites to microsomal protein and lipid in vivo.
Chlordecone does not interfere with hepatic repair after carbon tetrachloride or partial hepatectomy
Authors: Dolak James A. , Britton Robert S. , Glende Jr. Eric A. , Recknagel Richard O.
Published: October 2006

The effect of chlordecone (CD) on hepatic repair, measured either as recovery of microsomal enzymatic functions or as the induction of cytosolic thymidine kinase (TK) activity, was evaluated in rats given carbon tetrachloride (CC14). Carbon tetrachloride was administered to CD‐potentiated and control animals using doses of this hepatotoxin which produce similar degrees of damage at 24 hours in both groups of animals (6 and 100 μ1 CC14 per 100 g body weight, respectively). Chlordecone had no significant effect on the time course of recovery of microsomal cytochrome P‐450 content or glucose‐6‐phosphatase activity following CC14 administration. Hepatic TK activity was measured 48 hours after CC14 administration as a biochemical index of the hepatic regenerative response. Thymidine kinase activity was increased eightfold in CD‐treated rats receiving 6 μ1 CC14 per 100 g body weight, whereas in controls a similar induction of TK activity was produced by 100 μ1 CC14 per 100 g body weight. Therefore, the TK response in CD‐treated rats receiving CC14 is appropriate for the amount of damage produced, suggesting that CD does not inhibit the hepatic regenerative response to CC14‐induced injury. The effect of CD on hepatic repair was also examined in rats receiving a two‐thirds partial hepatectomy. Pretreatment of animals with CD had no significant effect on the increase in TK activity produced 24 hours after partial hepatectomy. These results offer no support for the idea that CD impairs hepatic repair after either partial hepatectomy or CC14 administration.
Changes in lung ATP concentration in the rat after low‐level phosgene exposure
Authors: Currie William D. , Hatch Gary E. , Frosolono Michael F.
Published: October 2006

Inhibition of mitochondrial respiratory activity and decreased lung adenosine triphosphate (ATP) concentration occur following exposure to 240 ppm·min phosgene. To determine the relationship between energy stores and the onset of phosgene‐induced pulmonary edema, we measured the ATP concentration in rapidly frozen rat lung tissue before and during pulmonary edema. Male Sprague‐Dawley rats were exposed to phosgene for four hours at concentrations of 0.05 to 1.0 ppm (12, 30, 60,120, and 240 ppm·min). Lung wet and dry weight and ATP concentration were measured immediately after exposure and for three days postexposure. The accumulation of lavage fluid protein (LFP) was also measured as an index of damage or edema due to phosgene. Lung dry weight was significantly elevated one day postexposure to 0.5 ppm phosgene, while the LFP was elevated by 0.2 ppm phosgene. Time course studies at these doses of phosgene showed that decreased ATP levels preceded the onset of edema or increase in lung weight. The ATP values expressed on a per‐lung basis showed that ATP levels were significantly lowered immediately following phosgene exposure, suggesting that the ATP changes were not the result of edema. This study is the first demonstration of a biochemical change that occurs following exposure to phosgene at a level significantly below the threshold limit value for this gas.
Effects of tricyclohexylhydroxytin on the kinetics of adenosine triphosphatase system and protection by thiol reagents
Authors: Prasada Rao Kodavanti S. , Chetty Sreeramulu C. , Desaiah Durisala
Published: October 2006

Tricyclohexylhydroxytin, commonly known as Plictran® inhibited Na+, K+ ‐ATPase activity of rat brain synaptosomes in a concentration‐dependent manner with median inhibitory concentration (IC‐50) of 2 μM. Both K+ ‐stimulated para‐nitrophenylphosphatase and [3‐H]‐ouabain binding to synaptosomes were also inhibited by Plictran with IC‐50 values of 11 and 30 μM, respectively. Altered pH and Na+, K+ ‐ATPase activity curves demonstrated comparable inhibition in buffered neutral and alkaline pH ranges, and no inhibition was observed in acidic pH. The inhibition of Na+, K+ ‐ATPase was independent of temperature. Kinetic studies of substrate (ATP) activation of Na+, K+ ‐ATPase indicated uncompetitive inhibition. Results also showed noncompetitive inhibition for p‐nitrophenylphosphate and uncompetitive inhibition for K+ activations of p‐nitrophenylphosphatase. Preincubation of synaptosomes with dithiothreitol, a sulfhydryl (SH) agent, resulted in the complete protection of Plictran inhibition of Na+, K+ ‐ATPase, K+ ‐para‐nitrophenylphosphatase, and [3‐H]‐ouabain binding. The protection was specific and concentration dependent since cysteine and glutathione did not afford protection. These results indicate that Plictran inhibited Na+, K+ ‐ATPase by interacting with dephosphorylation of the enzyme‐phosphoryl complex and exerted a similar effect to that of SH‐blocking agents.
Induction of single‐strand breaks and interstrand cross‐links in liver DNA after the administration of 2‐acetylaminofluorene and trans‐4‐acetylaminostilbene to rats
Authors: Ruthsatz M. , Neumann H.‐G.
Published: October 2006

2‐Acetylaminofluorene (AAF) or trans‐4‐acetylaminostilbene (AAS) was orally or intraperitoneally administered to female Wistar rats. DNA from liver cells was analyzed for single‐strand breaks by the alkaline elution assay. Only borderline effects were observed with doses (100 μMol/kg) used in animal carcinogenesis experiments. Even high doses of AAF (1,000 μMol/kg) were not effective. Methyl methanesulfonate (MMS) in vivo and gamma irradiation in vitro were shown to produce dose‐dependent DNA single‐strand breaks (positive control). Only a marginal effect was obtained with 100 μMollkg MMS. The elution rate of DNA was increased by a factor of 34 in liver cells in vitro with 400 rad of gamma irradiation. Only a fraction of this rate could be demonstrated immediately after irradiation in vivo, and no lesions were found two hours later. This strongly indicates the rapid repair of single‐strand breaks. Additional experiments showed that AAS, a nonhepatocarcinogen, produced more interstrand cross‐links in the rat liver DNA than did AAF.
Effects of cadmium‐metallothionein on renal organic ion transport and lipid peroxidation in rats
Authors: Suzuki Catherine A. M. , Cherian M. George
Published: October 2006

The effects of cadmium‐metallothionein (Cd‐MT) on organic ion uptake in renal cortical slices and lipid peroxidation in the kidney were studied in rats. For in vitro studies, slices were prepared from kidneys of control animals and incubated in buffer containing either cadmium chloride (CdCl2) or Cd‐MT in equimolar Cd concentrations ranging from 5 × 10−6 to 2 × 10−4 M. Uptake into the slices of the organic anion p‐aminohippuric acid (PAH) was found to be inhibited by both forms of Cd in a dose‐dependent manner. Although this inhibition was slightly greater in the presence of Cd‐MT, accumulation of Cd into the slices was approximately 12 times greater with CdCl2 than Cd‐MT. Tetraethylammonium (TEA) uptake was less sensitive to the inhibitory effects of both CdCl2 and Cd‐MT, although a dose‐dependent inhibition did occur with higher Cd concentrations.

To study the in vivo effects of Cd‐MT on transport function and lipid peroxidation in the kidney, rats were injected with Cd‐MT (0.3 mg Cd per kilogram body weight [bw]) and sacrificed at specific time intervals. Similar to the in vitro studies, PAH uptake into the renal cortical slices was markedly inhibited within 12 hours after Cd‐MT injection whereas inhibition of TEA uptake was less and not observed until 48 hours after injection. Only a small increase (1.4‐fold) in lipid peroxidation, as measured by generation of malondialdehyde (MDA), in the kidney was detected at four hours postinjection, and no further increase was observed at later time periods.

The results suggest that Cd‐MT affects the transport of organic anions and cations during its renal uptake but that lipid peroxidation may play only a minor role in Cd‐MT‐induced renal toxicity.
Morphological and biochemical effects of gentamicin and cyclosporin‐a on urinary cell phospholipids and phospholipases in man
Authors: Chatterjee Subroto , Bose Sharmistha
Published: October 2006

The morphology, lipid composition, and activity of sphingomyelinase (E.C. 3.1.4.12) and phospholipases A (E.C. 3.1.1.32) and C (E.C. 3.1.4.3) were studied in the urinary cells from four normal subjects, four patients receiving gentamicin (G), and four patients receiving cyclosporin‐A (CsA). We report that abnormal urinary excretion of proximal tubular cells occurred in patients receiving G and CsA. Membrane‐enclosed sudanophilic material and numerous vacuoles were found in the cytoplasm of the proximal tubular cells from both patients receiving G and those receiving CsA. Patients receiving G shed higher levels of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and sphingomyelin (SM) in the order of 78%, 38%, and 30% relative to normal. In contrast, the excretions of phosphatidylinositol (PI) and PC were 50% and 30% lower, respectively, in patients receiving CsA as compared to control. Sphingomyelin levels, however, were moderately elevated in these patients' urinary renal tubular cells.

The activity of acid sphingomyelinase was one half the normal level in the cells of patients receiving G and CsA. The most striking result was a tenfold decrease in the activity of neutral sphingomyelinase in patients receiving G. In contrast, the activity of neutral sphingomyelinase in patients receiving CsA was similar to control. Phospholipase A activity was decreased and increased 35% and 15%, respectively, in urinary proximal tubular cells from patients receiving G and CsA.

We conclude that deficient neutral sphingomyelinase activity precedes phospholipid (PL) overloading and gross pathological changes in patients receiving gentamicin but not in patients receiving cyclosporin‐A.
Modulations in the biotransformation of tobacco extract and N′‐nitrosonornicotine under differential dietary protein status
Authors: Ammigan N. , Nair U. J. , Amonkar A. J. , Bhide S. V.
Published: October 2006

The modulation of the phase I and phase II biotransformation enzymes upon treatment with tobacco extract (TE) and N'‐nitrosonornicotine (NNN) was investigated using male Sprague‐Dawley rats fed differential protein diets. It was observed that the animals fed a low protein diet showed an overall decrease in the basal levels of hepatic and pulmonary phase I and II enzymes. TE and NNN significantly decreased the detoxifying system in the low‐proteinfed animals. Animals fed 20% protein, however, showed significant increases in glutathione and glutathione S‐transferase upon treatment. Furthermore, TE and NNN treatment brought about a significant depletion in the hepatic pool of vitamin A with a concomitant increase in the vitamin C levels.
Hepatic polyamines and related enzymes following chlordecone‐potentiated carbon tetrachloride toxicity in rats
Authors: Rao Shrinivas B. , Young Robert A. , Mehendale Harihara M.
Published: October 2006

Chlordecone potentiation of the hepatotoxic and lethal effects of CCl4 has been well established. Recent studies have shown that the suppression of hepatocellular regeneration results in an accelerated progression of liver injury leading to complete hepatic failure. Since polyamines are involved in cell division, these studies were designed to investigate the polyamine levels and associated enzymes in the livers of rats treated with a low‐dose combination of CD and CCl4. For comparison, a large toxic dose of CCl4 was also employed. The extent of liver toxicity in rats fed 10 parts per million chlordecone (CD) for 15 days and subsequently injected with a single dose of CCl4 (100 μL/kg body weight) or a high dose of CCl4 alone (2.5 mL/kg body weight) was similar 6 and 24 hr later as assessed by plasma transaminase levels. There was significant elevation in liver ornithine decarboxylase, S‐adeno‐sylmethionine decarboxylase, and putrescine at 24 hr and spermidine N1‐acetyltransferase, N1‐acetylputrescine, putreanine, putrescine, and N1‐acetylspermidine at 6 hr in rats treated with the high dose of CCl4 alone compared to the combination treatment. Spermidine levels decreased up to 6 hr and then increased up to 24 hr for both treatments. Spermine continuously decreased up to 24 hr for the CD and CCl4 low‐dose combination treatment compared to rats treated with a high dose of CCl4 alone. Spermidine levels were lower than in controls and rose towards control value between 6 and 24 hr after the combination treatment and the high dose of CCl4. Results indicate that the CD and CCl4 low‐dose combination treatment increased liver toxicity, resulting in compromised polyamine metabolism that is coincidental with suppressed hepatocellular regeneration, which leads to an accelerated progressive phase of liver injury and culminates in complete hepatic failure.
Comparison of the formation of benzo[a]pyrene diolepoxide‐dna adducts in vitro by rat and human microsomes: Evidence for the involvement of p‐450IA1 and p‐450IA2
Authors: Vähäkangas Kirsi , Raunio Hannu , Pasamen Markku , Sivonen Päivi , Park Sang‐Shin , Gelboin Harry V. , Pelkonen Olavi
Published: October 2006

The involvement of cytochrome P‐450 isozymes in the activation of benzo[a]pyrene (BP) by human placental and liver microsomes was studied in vitro using monoclonal antibodies (Mab) toward the major 3‐methylcholanthrene (MC)‐inducible and phenobarbital‐inductible rat liver P‐450 isozymes (Mab 1–7–1 and Mab 2–66‐3, respectively). Microsomes from human placenta and liver and rat liver were incubated with BP and DNA, and BP‐diolepoxide‐DNA (BPDE‐DNA) adducts were measured by synchronous fluorescence spectrophotometry (SFS). The only BP metabolite giving the same fluorescence peak as chemically modified BPDE‐DNA was BP‐7,8‐dihydrodiol. Five (smokers) out of 29 human placentas (smokers and nonsmokers), and five out of nine human livers were able to metabolically activate BP to BPDE‐DNA adducts in this system. The Mab 1–7–1 totally inhibited the formation of BPDE‐DNA adducts in placental microsomal incubations. Inhibition using rat or human liver microsomes was 50–60% and about 90%, respectively. The Mab 2–66‐3 had no effect in any of the microsome types. Adduct formation was inhibited more strongly and at lower concentrations of Mab 1–7–1 compared with the inhibition of AHH activity. This study is a clear indication of the major role of P‐450IA1 (P‐450c) in human placenta and probably P‐450IA2 (P‐450d) in human liver in BP activation, while other isozymes also take part in the activation in rat liver. Furthermore, this clearly indicates that AHH activity and BP activation are not necessarily associated.
Covalent binding to proteins of reactive intermediates resulting from prostaglandin h synthase‐catalyzed oxidation of stilbene and steroid estrogens
Authors: Freyberger Alexius , Degen Gisela H.
Published: October 2006

Prostaglandin H synthase (PSH) is known to metabolically activate a variety of xeno‐biotics in vitro by means of its peroxidase activity. Recently, stilbene and steroid estrogens have been found to be cooxidized by ram seminal vesical microsomes, a rich source of PHS, to nonextractable metabolites bound to microsomal protein. To investigate further the nature of this protein binding, different radiolabeled estrogens were incubated with purified PHS, holoenzyme in the presence of various amounts of albumin (BSA), and radioactivity bound to protein was determined after gel electrophoretic separation. Diethylstilbestrol (DES), its analog hexestrol, and the steroid estrogens estrone and 2‐hydroxy‐estrone were cooxidized by PHS in vitro to metabolites that bound covalently to PHS and to BSA. Although a preferential binding of DES to PHS was found in the presence of excess BSA, reactive intermediates derived from DES, or from the other estrogens, were sufficiently stable to react with the competing nucleophile BSA as well.

With respect to the metabolic reactions catalyzed by PHS, in addition to one‐electron oxidation of phenolic functions, PHS catalyzed the aromatic hydroxylation of synthetic and steroid estrogens as shown by 3H2O release from regiospecifically labeled compounds and confirmed by product identification. Although DES was extensively metabolized by PHS, its aromatic hydroxylation was minor by comparison to estradiol, a difference possibly related to the compounds' redox potentials.

Thus, cooxidation of estrogens in vitro resulted in phenoxy radicals, semiquinones and quinones, reactive intermediates capable of protein binding that may contribute to the adverse effects of stilbene and steroid estrogen observed in vivo and in short‐term assays.
Plasma concentrations of pituitary hormones in 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin‐treated male rats
Authors: Moore Robert W. , Parsons Jonathan A. , Bookstaff Robert C. , Peterson Richard E.
Published: October 2006

Experiments were conducted to test the hypothesis that acute TCDD toxicity is associated with pituitary hypofunction. Sexually mature male Sprague‐Dawley rats were given graded doses of TCDD (0–100 μg/kg) and evaluated 7 days later. Despite pronounced hypophagia and body weight loss, plasma concentrations of growth hormone (GH), follicle‐stimulating hormone (FSH), and luteinizing hormone (LH) were not significantly affected by any dose of TCDD. Only prolactin (PRL) concentrations were reduced, while, as previously reported, thyroid‐stimulating hormone concentrations were elevated. Also, plasma LH, PRL, and adrenocorticotropic hormone (ACTH) concentrations were not significantly affected 1, 2, 3, 4, 5, or 7 days after a single dose of TCDD (50 μg/kg). We conclude that (1) pituitary hypofunction is not a major cause of the initial stages of acute TCDD toxicity, (2) growth retardation in TCDD‐treated rats is not the result of a deficiency of GH, (3) alterations in plasma corticosterone concentrations are due to altered responsiveness of the adrenal to ACTH stimulation rather than to changes in plasma ACTH concentrations, and (4) that impaired spermatogenesis is not associated with a decrease in plasma FSH concentrations. In addition, the lack of a consistent effect on plasma PRL concentrations suggests that alterations in plasma PRL concentrations do not play a critical role in the toxicity of TCDD. Finally, because TCDD treatment causes a serious androgenic deficiency without increasing the rates at which androgens are catabolized or excreted, the fact that plasma LH concentrations were unaffected indicates that TCDD treatment must reduce the responsiveness of the testis to LH stimulation. In addition, because the expected compensatory response to low plasma androgen concentrations (a pronounced elevation in plasma LH concentrations) was absent, these observations also indicate that TCDD treatment must interfere with the regulation of plasma LH concentrations.
Covalent interaction of chloroacetic and acetic acids with cholesterol
Authors: Bhat Hari K. , Ansari G. A. S.
Published: October 2006

The covalent interaction of chloroacetic acid with rat liver lipids was studied in vivo. Rats were given a single oral dose (8.75 mg/kg, 50 μCi) of 1‐[14C]chloroacetic acid and sacrificed after 24 hours. Lipids extracted from the livers were separated into neutral lipids and phospholipids by solid‐phase extraction using sep‐pak silica cartridges. The neutral lipid fraction was further fractionated by preparative thin‐layer chromatography followed by reverse‐phase high‐performance liquid chromatography. The fraction corresponding to the retention time of standard cholesteryl chloroacetate gave a pseudomolecular ion peak at m/z 480/482 ratio: (3:1) on ammonia chemical ionization mass spectrometry, and the fragmentation pattern was found to be similar to that of the standard sample. Under similar conditions, acetic acid resulted in the formation of cholesteryl acetate. The effect of such conjugation reactions on the cell membrane and their contribution to toxicity is presently unknown.
Increased susceptibility to pentobarbital following mouse cytomegalovirus infection: Relative roles of viral‐induced interferon and viral infection of the liver
Authors: Catignani Joan C. , Ménache M. G. , Selgrade Maryjane K.
Published: October 2006

The purpose of this study was to determine the relative roles of viral‐induced interferon (IFN) and viral infection of the liver in mouse cytomegalovirus (MCMV)‐induced depression of cytochrome P‐450 (cyt P‐450) levels and enhancement of pentobarbital‐induced sleeping time (PEN‐ST). This was done by establishing the temporal relationship among the IFN response, viral infection of the liver, suppression of cyt P‐450 levels, and enhancement of PEN‐ST, by determining the effect of anti‐IFN antibody treatment on all of these responses, and by manipulating factors known to influence viral pathogenesis and host response to virus such as animal age, virulence of the virus, and dose of virus. In general, manipulation of these factors toward increased stimulation of host immune responses resulted in greater depression of cyt P‐450. The data are consistent with the hypothesis that some IFN‐dependent mechanism may have contributed to the effects of MCMV infection on both cyt P‐450 levels and PEN‐ST; however, the temporal relationship among the various responses measured following viral infection suggested that the effect of the IFN response may be indirect and due to modulation of other host defense mechanisms. Use of anti‐IFN antisera to definitively establish a role for IFN in the effects observed here proved unsuccessful. Effects on PEN‐ST and cyt P‐450 levels did not appear to be related to the magnitude of infection in the liver. Because suppression of cyt P‐450 levels and enhancement of PEN‐ST were not always parallel, it is possible that the enhanced sensitivity to pentobarbital observed in infected mice was the cumulative result of effects on cyt P‐450 as well as other components involved in the distribution and metabolism of the drug.
Appendix to the sulfur‐cyanolysis sites of serum albumin: Metabolite competition studies. Tight‐binding inhibitions that yield atypical henderson plots
Authors: Westley John , Jarabak Rebecca
Published: October 2006

Ordinary tight‐binding inhibition in steady‐state enzyme systems is conveniently evaluated by means of the Henderson plot. This is a linear plotting form that has an ordinate intercept equal to the total enzyme concentration. However, there are two experimental situations that yield deviations from the common Henderson plot form. These are inhibitor binding in a separate, noninhibitory mode that depletes the concentration of free inhibitor, and partial inhibition, i.e., the retention of partial activity by the enzyme‐inhibitor complex. Noninhibitory depletion results in Henderson plots with elevated ordinate intercepts. Competitive partial inhibition yields a characteristic pattern of parabolic Henderson plots.
Differences in induction of hepatic cytochrome p450 isozymes by mice in eight methylenedioxyphenyl compounds
Authors: Lewandowski Margaret , Chui Y. C. , Levi Patricia E. , Hodgson Ernest
Published: October 2006

Eight methylenedioxyphenyl (MDP) compounds were examined for their ability to induce cytochrome P450 (P450) in mouse liver. Induction by safrole, isosafrole, and dihydrosafrole was studied in both C57BL/6N (Ah‐responsive) and DBA/2N (Ahnonresponsive) male mice after IP administration of 200 mg/kg/day MDP compound for 3 days. Hepatic P450 content, ethylmorphine N‐demethylase, ethoxy‐resorufin O‐deethylase, and acetanilide hydroxylase activities were induced to the same extent in both strains of mice. Benzo(a)pyrene hydroxylase activity, however, was not induced in either C57 or DBA mice. The similarity of results in both strains of mice indicated induction of these P450 isozymes by these three MDP compounds is not mediated by the Ah receptor. Induction of P450 by butylbenzodioxole (n‐butyl‐BD), tertiarybutylbenzodioxole (t‐butyl‐BD), methylbenzodioxole (methyl‐BD), nitrobenzodioxole (nitro‐BD), and bromobenzodioxole (bromo‐BD) was examined only in C57BL/6N mice. Methyl‐BD, nitro‐BD, and bromo‐BD did not induce hepatic microsomal proteins or selected P450 monooxygenase activities. In contrast, n‐butyl‐BD, and t‐butyl‐BD induced P450 content, ethylmorphine N‐demethylase, acetanilide hydroxylase, and ethoxyresorufin O‐deethylase activities. Benzo(a)pyrene hydroxylase was not induced by any of the treatments. Induction of these P450 activities is consistent with induction of P450 IIB1 and P450 IA2, but not induction of P450 IA1. Western blot analysis with antibodies to P450 isozymes induced with either phenobarbital (Pb) or 3‐methylcholanthrene (3‐MC) confirmed that both IIB1 and IA2 were induced, but that IA1 was not induced.
The effects of soman in vitro on catechol‐o‐methyltransferase and monoamine oxidase activities in rabbit tissues
Authors: Hsu Chia‐Hsuh , Hu Chao‐Yu , Robinson Casey P.
Published: October 2006

Soman (pinacolyl methylphosphonofluoridate) not only increases acetylcholine levels by inhibiting cholinesterases, it also alters the levels of some other neurotransmitters including norepinephrine, dopamine, and serotonin. Soman also causes an alteration in the activities of the enzymes metabolizing norepinephrine when it is administered to animals. Because these alterations may result from indirect effects on the enzymes, the effects of in vitro application of soman on catechol‐O‐methyl‐transferase (COMT) and monoamine oxidase (MAO) activities in rabbit tissues were investigated. Enzyme activities were determined in rabbit lung, liver, cerebellum, cerebrum, brain stem, mesenteric artery, pulmonary artery, renal artery, central ear artery, thoracic aorta, and diaphragm. MAO and COMT activities were not affected by soman in any tissues tested, except the lung and liver, where the activity of COMT was increased (p < 0.05). Thus, reported effects of soman in vivo on norepinephrine, dopamine, or serotonin concentrations, and MAO and COMT activities do not seem to result from direct effects on the activities of these aminemetabolizing enzymes.
Role of inducer binding in cytochrome P‐450 IA2‐mediated uroporphyrinogen oxidation
Authors: Jacobs Judith M. , Sinclair Peter R. , Lambrecht Richard W. , Sinclair Jacqueline F. , Jacobs Nicholas J.
Published: October 2006

The oxidation of uroporphyrinogen, an intermediate of the heme biosynthetic pathway, by methylcholanthrene‐inducible isozyme(s) of cytochrome P‐450 has been proposed to play a role in the development of chemically induced uroporphyria. Prior work from this laboratory [10] indicated that although addition of 3,4,3′,4′‐tetrachlorobiphenyl is required for uroporphyrinogen oxidation by methylcholanthrene‐induced chick embryo liver microsomes, this biphenyl is not required for the oxidation catalyzed by hepatic microsomes from methylcholan‐threne‐induced rodents. Here we investigated whether rodent microsomes catalyze uroporphyrinogen oxidation without addition of 3,4,3′,4′‐tetrachlorobiphenyl because the chemical used as an inducer remains bound to cytochrome P‐450. Hepatic microsomes containing almost no residual inducer were isolated from rats treated with a low dose of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD). These microsomes oxidized uroporphyrinogen at high rates without addition of 3,4,3′,4′‐tetrachlorobiphenyl. Inducerfree microsomal cytochrome P‐450 was also obtained by inducing cytochrome P‐450 in rats and mice with isosafrole, which was then removed from the isolated microsomes by butanol treatment. This procedure resulted in microsomes with high activity for uroporphyrinogen oxidation. Furthermore, addition of chlorobiphenyl to these inducer‐free microsomes was inhibitory. Hepatic microsomes from isosafroleinduced C57BL / 6 and DBA mice, rendered inducerfree by butanol treatment, oxidized uroporphyrinogen at the same rate even though these two strains differ markedly in their susceptibility to chemically induced uroporphyria. We conclude that uroporphyrinogen oxidation is catalyzed by cytochrome P‐450 that is free of inducer.
Tissue distribution, metabolism, and elimination of perfluorooctanoic acid in male and female rats
Authors: Heuvel John P. Vanden , Kuslikis Benedict I. , Van Rafelghem Marc J. , Peterson Richard E.
Published: October 2006

The elimination, tissue distribution, and metabolism of [1‐14C]perfluorooctanoic acid (PFOA) was examined in male and female rats for 28 days after a single ip dose (9.4 μmol/kg, 4 mg/kg). A sex difference in urinary elimination of PFOA‐derived 14C was observed. Female rats eliminated PFOA‐derived radioactivity rapidly in the urine with 91% of the dose being excreted in the first 24 hr. In the same period, male rats eliminated only 6% of the administered 14C in the urine. The sex‐related difference in urinary elimination resulted in the observed difference in the whole‐body elimination half‐life (t1/2) of PFOA in males (t1/2 = 15 days) and females (t1/2 < 1 day). Analysis of PFOA‐derived 14C in tissues showed that the liver and plasma of male rats and the liver, plasma, and kidney of female rats were the primary tissues of distribution. The relatively high concentration of PFOA in the male liver was further examined using an in situ nonrecirculating liver perfusion technique. It was shown that 11% of the PFOA infused was extracted by the liver in a single pass. The ability of the liver to eliminate PFOA into bile was examined in rats whose renal pedicles were ligated to alleviate sex differences in the urinary excretion of PFOA. In a 6‐hr period following IP administration of PFOA, there was no apparent difference in biliary excretion, where both males and females eliminated less than 1% of the PFOA dose via this route. We hypothesized that the sex difference in the persistence of PFOA was due to a more rapid formation of a PFOA‐containing lipid (i.e., a PFOA‐containing mono‐, di‐, or triacylglycerol, cholesteryl ester, methyl ester, or phospholipid) in the male rat. Also, the increased urinary elimination of PFOA in females may have been due to increased metabolism to a PFOA‐glucuronide or sulfate ester. However, no evidence that PFOA is conjugated to form a persistent hybrid lipid was obtained, nor were polar metabolites of PFOA in urine or bile detected. In addition, daily urinary excretion of fluoride in male and female rats before or after PFOA treatment were similar, suggesting that the parent compound is not defluorinated. Thus, the more rapid elimination of PFOA from female rats is not due to formation of a PFOA metabolite.
Effects of hydroxylated polychlorinated biphenyls on mouse liver mitochondrial oxidative phosphorylation
Authors: Narasimhan T. R. , Kim H. L. , Safe S. H.
Published: October 2006

The effects of three tetrachlorobiphenylols [2′,3′,4′,5′‐tetrachloro‐2‐biphenylol (1); 2′,3′,4′,5′‐tetrachloro‐4‐biphenylol (2); and 2′,3′,4′,5′‐tetrachloro‐3‐biphenylol (3)]; three monochlorobiphenylols [5‐chloro‐2‐biphenylol (5), 3‐chloro‐2‐biphenylol (6); and 2‐chloro‐4‐biphenylol (7)] and a tetrachlorobiphenyldiol [3,3′,5,5′‐tetrachloro‐4,4′‐biphenyldiol (4) on respiration, adenosine triphosphatase (ATPase)] activity, and swelling in isolated mouse liver mitochondria have been investigated. Tetrachlorobiphenylols (1–3) and the tetrachlorobiphenyldiol (4) inhibited state‐3 respiration in a concentration‐dependent manner with succinate as substrate (flavin adenine dinucleotide [FAD]‐linked) and the tetrachlorobiphenyldiol (4) caused a more pronounced inhibitory effect on state‐3 respiration than the other congeners. The monochlorobiphenylols 5–7 were less active as inhibitors of state‐3 mitochondrial respiration and significant effects were observed only at higher concentration (≥0.4 μM). However, in the presence of the nicotinamide adenine dinucleotide (NAD)‐linked substrates (glutamate plus malate), hydroxylated PCBs (1–7) significantly inhibited mitochondrial state‐3 respiration in a concentration‐dependent manner. Compounds 5, 6, and 7 uncoupled mitochondrial oxidative phosphorylation only in the presence of FAD‐linked substrate as evidenced by increased oxygen consumption during state‐4 respiratory transition, stimulating ATPase activity, releasing oligomycin‐inhibited respiration, and inducing mitochondrial swelling (5, 6, and 7). Tetrachlorobiphenylols 1, 2, and 3 had no effect on mitochondrial ATPase activity while the tetrachlorobiphenyldiol, 4, decreased the enzyme activity. The possible inhibitory site of electron transport by these compounds and their toxicologic significance is discussed.
Selective erythrocyte potassium efflux following pulse treatment with tellurite
Authors: Klug Robert K. , Kurantsin ‐Mills Joseph
Published: October 2006

Human erythrocytes exposed to 0.1 mM tellurite (K2TeO3) in an isotonic buffered choline chloride medium for 15 min at 37°C, washed, and incubated further in the absence of the chemical in the buffer, exhibited selective leakiness for potassium within minutes. The potassium efflux curve was sigmoidal, with an initially slow leakage followed by a sharp rise (first‐order kinetics) and a plateau by 60 min. After 15 min, 30–50% of the total potassium concentration had leaked from the cells, although less than 1% lysis had occurred. The control cells incubated in buffer with no K2TeO3 exhibited no potassium leakage. The mean volume of the K2TeO3‐treated erythrocytes increased and their median density decreased, indicating changes in the colloid osmotic state and physical characteristics of the cells. However, cells pretreated with K2TeO3 exhibited no significant change in glutathione (GSH) concentration and no membrane lipid peroxidation, unlike cells pretreated with t‐butylhydroperoxide (Deuticke et al., Biochim. Bio phys. Acta, 899, 125–128, 1987). The enhanced potassium permeability of the K2TeO3‐treated erythrocytes preceded the increase in cell volume, intracellular hydration, and a decrease in median density. We suggest that perturbation of the lipid‐protein interaction in the membrane by the oxidant alters the potassium permeability and results in the selective leakage with eventual hemolysis.
TCDD causes stimulation of c‐ras expression in the hepatic plasma membranes in vivo and in vitro
Authors: Tullis Kathryn , Olsen Hugh , Bombick David W. , Matsumura F. , Jankun Jerzy
Published: October 2006

A series of in vivo and in vitro experiments were conducted to determine the effects of 2, 3, 7, 8‐tetrachlorodibenzo‐p‐dioxin (TCDD) administered on the expression of c‐ras. Differences in c‐ras expression between control and TCDD treated groups were determined by immunoassay of p21ras protein, or indirectly measured by the specific binding of 3H‐GTP to hepatic plasma membrane preparations. Intraperitoneal injection of sublethal doses of TCDD significantly elevated (P < 0.05, Student t test) levels of hepatic p21ras protein in Sprague‐Dawley rats and TCDD sensitive C57BL/6J mice. Such an increase occurred at an early stage of poisoning in the C57BL/6J mice. The earliest increase was detectable 6 hr after dosing, and the differencebecame statistically significant by 12 and 24 hr after dosing. In contrast, TCDD tolerant DBA/2J mice had only a marginal increase in hepatic p21ras protein which did not become statistically significant even at 24 hr post‐dosing. TCDD evoked increases in hepatic p21ras protein of C57BL/6J mice were accompanied by the increase in the specific binding of GTP to hepatic plasma membranes. Column chromatography of solubilized rat hepatic membrane proteins on sephadex G‐50 showed TCDD administration increased levels of a 3H‐GTP binding protein with MW of approximately 21 Kd. 3H‐GTP binding in total hepatic membranes was also elevated (P<0.05, Fisher PLSD multiple comparison test) 6 hr and 24 hr after dosing of C57BL/6J mice, but as expected the effect of TCDD was not as conspicuous as that found in the plasma membrane. TCDD treatment increased levels of a 21 Kd protein found in the in vitro translation products of RNA purified from guinea pig liver. This protein was identified as a c‐ras protein based upon its ability to bind GTP, precipitation by a polyclonal antibody against the rasHa and Ki proteins and subsequent SDS‐PAGE which showed a single protein band of ≈ 21 Kd.
Autoionization reaction of phosgene (OCC12) studied by electron paramagnetic resonance/spin trapping techniques
Authors: Arroyo Carmen M. , Feliciano Felix , Kolb Dana L. , Keeler Jill R. , Millette Susan R. , Stotts Richard R.
Published: October 2006

The reaction of phosgene with ni‐trone spin traps was investigated using electron paramagnetic resonance (EPR)/spin trapping techniques. Evidence for the intermediacy of a carba‐moyl monochloride intermediate was obtained. Isotopic substitution of 13C‐phosgene was employed to verify the hyperfine coupling constant assignments. The implications of these observations on pulmonary damage caused by inhalation of phosgene are mentioned.
Characterization of cytochromes P450 in liver and kidney of rats treated with di‐(2‐ethylhexyl)phthalate
Authors: Okita Janice Rice , Castle Pamela J. , Okita Richard T.
Published: October 2006

A polyclonal antibody was made to a liver cytochrome P450 purified from di‐(2‐ethyl‐hexy)Dphthalate (DEHP)‐treated Sprague‐Dawley rats and was used to identify the CYP4A forms in liver and kidney cortex microsomes of control rats and rats treated with this peroxisome proliferator. Three clearly separated major protein bands were recognized on western blots in liver microsomes of control male rats or male rats treated with a single dose of DEHP, which, based on the description of relative mobility, tissue specificity, and sex dependent expression of CYP4A forms (Sundseth and Waxman (1992). J. Biol. Chem., 267, 801–810), correspond to the migration pattern of forms 4A1, 4A2, and 4A3 in clofibrate‐treated rats. The administration of DEHP for 2 or 3 days caused a loss of resolution of two of the protein bands. The protein band corresponding to 4A2 was absent in liver or kidney cortex microsomes of DEHP‐treated or control female rats and was not always visible in the livers of control male rats. The purified P450DEHPsupported the hydroxylation of arachidonic acid at both the 19‐ and 20‐carbon atoms with turnover rates of 1.4×0.2 and 22.7×2.5 nmoles per minute per nmol P450, respectively. No measurable amounts of hydroxylated products were obtained when prostaglandin E1, leukotriene B4, or testosterone were used as substrates. Another member of the CYP4 family, 4B1 from rabbit lung microsomes, was also recognized by this antibody on western blot analysis; however, rabbit lung form 4A4 showed only minimal cross‐reactivity with this antibody.
Acute lindane intoxication: A study on lindane tissue concentration and oxidative stress‐related parameters in liver and erythrocytes
Authors: Junqueira V. B. C. , Bainy A. C. D. , Arisi A. C. M. , Azzalis L. A. , Simizu K. , Pimentel R. , Barros S. B. M. , Videla L. A.
Published: October 2006

Treatment of rats with daily dosis of 20 mg of lindane/kg for 3 consecutive days led to the accumulation of the insecticide in several tissues, including erythrocytes and liver. Lindane did not alter the hematocrit and hemoglobin concentration but reduced methemogiobin levels by 17%. Red blood cells from controls and lindane‐treated rats, exposed to t‐butyl hydroperoxide, exhibited comparable rates of oxygen uptake and visible chemiluminescence, whereas the induction period that precedes oxygen uptake was significantly enhanced in the latter group. Lindane treatment did not modify the activity of erythrocyte glutathione peroxidase, glucose‐6‐phosphate dehydrogenase, catalase, and methemoglobin reductase, being the total content of glutathione and superoxide dismutase activity significantly increased. The liver from lindane‐treated rats showed an enhanced microsomal pro‐oxidant activity, evidenced by higher cytochrome P450 content and NADPH‐cytochrome c reductase and NADPH oxidase activities. The higher enzyme activities led to an increased superoxide anion generation (adrenochrome formation) and lipid peroxidation (measured either by the production of thiobarbituric acid reactants and spontaneous visible chemiluminescence). Concomitantly, liver glutathione content and the activity of glutathione peroxidase‐glutathione reductase couple were augmented by lindane treatment, without any change in superoxide dismutase activity, together with a reduction in that of catalase. Results suggest that lindane does not alter the prooxidant/antioxidant status of the erythrocyte in conditions of a significant cellular accumulation of the insecticide, which might exert direct action on enzymatic systems leading to enhanced superoxide dismutase activity and glutathione content. In the liver, lindane‐induced pro‐oxidant condition was not accompanied by cell injury, probably due to the adaptative increase in some antioxidant mechanisms of the hepatocyte.
Modulation of the rat hepatic cytochrome P450 composition by long‐term streptozotocin‐induced insulin‐dependent diabetes
Authors: Barnett Christopher R. , Flatt Peter R. , Ioannides Costas
Published: October 2006

The effects of long‐term insulin‐dependent diabetes on the enzymatic activities of hepatic cytochrome P450 isozymes were determined in rats rendered diabetic by the administration of streptozotocin and killed 4, 8, and 12 weeks following treatment. The O‐dealkylations of ethoxy‐resorufin and pentoxyresorufin were elevated in the diabetic animals throughout the study, the extent of increase being similar at all three time points. p‐Nitrophenol hydroxylase activity was induced in the diabetic animals 4 weeks following treatment with streptozotocin, but the extent of increase became less pronounced with the progress of the disease. A modest increase in ethylmorphine N‐demethylase activity was also observed but only in the diabetic animals killed 4 weeks after the induction of diabetes. Finally, lauric acid hydroxylase activity was elevated in the diabetic animals 4 weeks following streptozotocin administration but then declined rapidly with the duration of the disease. It is concluded that the duration of diabetes modulates the hepatic cytochrome P450 profile, with the effect being isoenzyme specific. Mechanisms that may account for these changes are discussed.
2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD)‐induced changes in glucose transporting activity in guinea pigs, mice, and rats in vivo and in vitro
Authors: Enan Essam , Matsumura Fumio
Published: October 2006

The toxic action of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD) on glucose uptake was studied on different species, sexes, and strains of animals by using a nonmetabolizable glucose analog, 3‐0‐methyl D [1‐3H] glucose (3H‐Me‐glc). We have found a drastic reduction in glucose uptake in different organs from male guinea pigs in vivo as well as in vitro. Female guinea pigs were less responsive to this effect of TCDD compared to males. Highly TCDD‐responsive mice strains responded in the same way as the male guinea pigs, while less TCDD‐responsive mice strains did not. A reduction in glucose uptake was found in explant adipose tissue cultures (in vitro) of male mice (highly TCDD‐responsive) after a 4 hour incubation with 10−8 M TCDD. In less responsive mice strains, stimulation was observed using the same model (in vitro). In the case of male rats, as well as male guinea pigs, glucose uptake was highly reduced by 10−8 M TCDD using an in vitro system of explant adipose tissue. The Ah‐receptor blocker, 4,7‐phenanthroline, abolished the effect of TCDD on inhibition of glucose uptake in adipose tissue of male guinea pigs both in vivo and in vitro. Transcription and translation inhibitors (actinomycin D and cycloheximide, respectively) were tested separately or in combination with TCDD using the guinea pig adipose tissue culture model. It was found that actinomycin D could block the effect of TCDD on glucose uptake throughout the experiment time (360 min) while cycloheximide blocked TCDD action for about 60 minutes. Protein kinase inhibitors (e.g., genistein, neomycin) did not change the effect of TCDD on glucose uptake in male guinea pig adipose tissue. Based on these observations, we conclude that TCDD inhibits Me‐glc uptake in various organs in guinea pigs, mice, and rats, and this response is mediated by the Ah‐receptor.
Inhibition of S‐(1,2‐dichlorovinyl)‐L‐cysteine‐induced lipid peroxidation by antioxidants in rabbit renal cortical slices: Dissociation of lipid peroxidation and toxicity
Authors: Davis Joe W. , Petry Thomas W.
Published: October 2006

Precision‐cut, rabbit renal slices were used to examine the effects of three novel antioxidants (U‐74006, U‐74500, and U‐78517) on S‐(1,2‐dichlorovinyl)‐L‐cysteine (DCVC)‐induced lipid peroxidation and toxicity. Slices exposed to DCVC showed a dose‐ and time‐dependent increase in lipid peroxidation (TBARS) and a decrease in cellular viability, as evidenced by the loss of intracellular potassium, during the course of a 3 hour incubation. Subsequent studies employed DCVC concentrations of 100 μM. Microemulsion formulations of U‐78517, U‐74500, and U‐74006 (100 μM) inhibited DCVC‐induced lipid peroxidation by 100±, 50±, and <5% (not significant), respectively. However, none of these antioxidants had a significant effect on DCVC‐dependent cytotoxicity, as indicated by intracellular potassium release. The effects of U‐78517, the most potent of the three antioxidants, were similar to those observed with two model antioxidants, diphenyl‐p‐phenylenedi‐amine (DPPD) and the iron chelator, deferoxamine. Aminooxyacetic (AOAA), an inhibitor of renal cysteine conjugate β‐lyase, had only a minimal effect on DCVC‐induced lipid peroxidation, and no effect on toxicity.

These data represent the first report of DCVC‐induced lipid peroxidation in rabbit renal cortical slices, a system which has been widely used to investigate mechanisms of nephrotoxicity, including that induced by DCVC. Our results demonstrate that DCVC‐induced lipid peroxidation in renal slices can be inhibited by a variety of antioxidant compounds operating by different mechanisms. Because inhibition of lipid peroxidation had minimal effect on DCVC‐dependent cytotoxicity, the data suggest that DCVC‐induced lipid peroxidation is not a major mechanism in the cytotoxicity induced by this compound.