Cytosolic activation of 2-aminoanthracene: implications in its use as diagnostic mutagen in the Ames test.

The metabolic activation of 2-aminoanthracene to mutagens in the Ames test was investigated using hepatic S9, microsomal and cytosolic fractions from control and Aroclor 1254-treated rats as activation systems. Microsomal and S9 preparations from control animals could activate 2-aminoanthracene, but the efficiency of activation was suppressed by pretreatment of animals with Aroclor 1254. Cytosolic fractions from Aroclor 1254-treated rats could readily activate the promutagen more readily than microsomes. The cytosolic activation of 2-aminoanthracene required NADPH and could not be accounted for by possible microsomal contamination. The molybdenum oxygenases appear not to contribute to the cytosolic activation of this promutagen. It is concluded that (a) the microsomal activation of 2-aminoanthracene is catalysed more effectively by enzyme systems other than the P450 I family and (b) an enzyme system capable of activating this carcinogen in vitro is present in the hepatic cytosol. The implications of these findings in the use of 2-aminoanthracene as a positive control in the Ames test are discussed.     

Arylamine activation following chronic ethanol ingestion by rats: studies on the liver S9, microsomal and cytosolic fractions and comparison with Aroclor 1254 pretreatment

That enzyme fractions derived from animals chronically fed alcohol can alter the metabolism of carcinogenic xenobiotic compounds has been documented. To further understand this relationship the mutagenicity of 3 aromatic amines was determined in the Ames test, employing activation systems derived from rats maintained on an alcohol-containing liquid diet, an isocaloric control liquid diet or Aroclor 1254-pretreated animals fed standard laboratory chow. Depending upon protein and substrate concentrations, S9 from ethanol-fed rats was 30-50% less efficient than S9 from pair-fed rats in activating arylamines (2-aminofluorene, 2-aminoanthracene and 2-acetylaminofluorene) to mutagens in Salmonella typhimurium TA98 and TA100. Cytosolic fractions from ethanol-fed animals always resulted in greater arylamine activation than that of controls whereas the opposite was true of the microsomal compartment in which the ethanol-treated group was consistently less active than the controls. The cytosolic N-acetyltransferase activities with respect to 2 different substrates, isoniazid and 2-aminofluorene, were unaffected by ethanol consumption, indicating that this activity probably does not account for the different activation profiles exhibited by the ethanol and pair-fed cytosolic systems. Both the cytosolic and microsomal compartments are required for maximal expression of the mutagenicity of each arylamine however, each compartment can activate arylamines independently of the other. Reconstituting cytosol with microsomes from ethanol- and pair-fed rats, but not Aroclor-pretreated rats, resulted in a synergistic activation of the aromatic amines and displayed an effect similar to that of S9. Compared to Aroclor pretreatment and pair-fed controls, microsomes from ethanol-fed rats displayed the least capacity for activating any of the arylamines to mutagens. Microsomes from Aroclor-pretreated rats accounted for at least 80% of the S9-mediated activation of each of the arylamines to mutagenic metabolites which was in marked contrast to the contribution of the microsomal fractions to the S9 activity in the ethanol- (5-20% of S9 activity) and pair-fed systems (22-30% of S9 activity). The data indicate that 2 opposing reactions occur in S9, a cytosolic activity that augments and a microsomal activity that attenuates the mutagenicity of arylamines. Both activities are modified by ethanol consumption and Aroclor pretreatment.     

The Salmonella/mammalian microsome mutagenicity test: comparison of human and rat livers as activating systems

The mutagenicity of several test compounds was verified by the Salmonella/microsome mutagenicity test (Ames test), using both human liver and rat liver (untreated or pretreated with Aroclor 1254) S9 under identical experimental conditions. Aflatoxin B1, 3-methylcholanthrene, and cigarette-smoke condensate were less mutagenic in the presence of human-liver S9 than in the presence of rat-liver S9 (particularly after treatment with Aroclor 1254). The opposite was observed with 2-aminonanthracene and to a lesser degree with 2-aminofluorene; correlation studies indicate that the two compounds were activated by the same or by very similar enzymes, probably cytochrome P-450s. These results clearly indicate that human-liver S9, as an activating system, behaves differently than rat-liver S9; therefore, it may constitute a useful, additional tool for the study of mutagenicity and probably, carcinogenicity in man.     

Lipid peroxidation in the rat-liver S9 fraction: influence of membrane lipid composition

These studies describe the influence of membrane fatty acid composition on peroxidation processes in rat-liver S9 fractions. Lipid peroxidation may be expected to affect enzyme activity and cofactors of importance for the performance of the Salmonella Mutagenicity Test, as well as to contribute to the formation of chemically reactive degradation products that are mutagenic. Lipid peroxidation products were measured as derivatives of 2-thiobarbituric acid (TBA). The amount of TBA-reactive compounds (TBA-C), formed during incubation of S9 fractions from rats fed a diet containing sunflower-seed oil, was 8 times higher than that produced in S9 fractions prepared from rats fed diets containing coconut oil or hydrogenated lard as their only sources of fat. S9 fractions from livers of Aroclor 1254 treated rats showed a marked increase in peroxidation yields for all 3 dietary groups investigated as compared to S9 fractions from non-induced animals. The coconut oil and hydrogenated lard dietary groups showed a 13-fold increase in the yield of TBA-reactive material, while a 2-fold increase was found for the sunflower-seed oil group. The variations in the glutathione (GSH) levels and the degradation of unsaturated fatty acids were also studied in response to Aroclor 1254 treatment, fatty acid composition of the diets and incubation at 37 degrees C. Pronounced variations in the GSH levels were observed in response to Aroclor 1254 treatment and incubation conditions. A positive correlation between production of TBA-reactive material and degradation of unsaturated fatty acids was verified for S9 fractions from the coconut oil and hydrogenated lard dietary groups. Furthermore, the effect of Fe2+ on lipid peroxidation was studied in all 3 dietary groups. The rate of lipid peroxidation was increased in all groups but only the coconut oil and hydrogenated lard dietary groups showed increased total yields of TBA-C upon administration of Aroclor 1254 to rats. Lipid peroxidation processes cause chemical alterations in liver homogenates. Therefore, these effects ought to be considered both in the preparation and in the use of the S9 fraction in different test systems.     

Diethylstilbestrol and 11 derivatives: a mutagenicity study with Salmonella typhimurium.

Diethylstilbestrol was tested for mutagenicity with his- S. typhimurium strains under 10 different matabolic situations (no exogenous metabolizing system; S9 mix from liver homogenate of rats induced with Aroclor 1254, with or without inhibition of epoxide hydratase; liver and/or kidney S9 mix from control or hamsters treated with Aroclor 1254; horse-radish peroxidase + H2O2). Under none of these conditions did diethylstilbestrol give any indication of a mutagenic effect. Furthermore, 11 metabolites and other derivatives of diethylstilbestrol, 2 of them potent inducers of sister-chromatid exchange in cultured fibroblasts, were not mutagenic with any of the 4 tester strains (S. typhimurium TA100, TA98, TA1537, TA1535) in the presence or absence of S9 mix from liver homogenate of rats induced with Aroclor 1254. Thus, one of the few known human carcinogens is very resistant to detection by the mammalian enzyme-mediated Salmonella typhimurium mutagenicity test (Ames test). This is especially remarkable since the metabolizing systems used included: (1) some of very high metabolic activity (S9 mix from liver homogenate of rats and hamsters induced with Aroclor 1254); (2) metabolizing systems from organs susceptible to the carcinogenic activity of diethylstilbestrol (hamster kidney); as well as (3) a mixture of (1) and (2) in case both activities are required for the carcinogenic effect in the whole animal.     

Aflatoxin B1 metabolism by 3-methylcholanthrene-induced hamster hepatic cytochrome P-450s

We have studied the activation of aflatoxin B1 by hamster liver microsomes and purified hamster cytochrome P-450 isozymes using a umu mutagen test. The hamster liver microsomes or S-9 fractions were much more active than rat liver microsomes or S-9 fractions in the activation of umu gene expression by aflatoxin B1 metabolites. 3-Methyl-cholanthrene treatment increased aflatoxin B1 activation by hamster liver microsomes. Two major 3-methylcholanthrene-inducible cytochrome P-450 isozymes, P-450 MC1 (IIA) and P-450 MC4 (IA2), were purified from 3-methylcholanthrene-treated hamster liver microsomes, and the metabolism of aflatoxin B1 by these two cytochromes was studied. In the reconstituted enzyme system, both P-450 MC1 and P-450 MC4 were highly active in the activation of aflatoxin B1, and antibodies against these P-450s specifically inhibited these activities. Antibody against P-450 MC1 inhibited the activation of aflatoxin B1 by 20% in the presence of 3-methyl-cholanthrene-treated hamster liver microsomes. In contrast, antibody against P-450 MC4 stimulated the activity by 175%. These results indicated that hamster P-450 MC1 might convert aflatoxin B1 to more toxic metabolite(s), whereas P-450 MC4 might convert aflatoxin B1 to less toxic metabolite(s), than aflatoxin B1 in liver microsomes. The metabolite(s) produced by both hamster cytochrome P-450 MC1 and MC4 were genotoxic in the umu mutagen test.     

Effect of hepatic cytochrome P-450 inducing agents on mutagen activity in the host-mediated assay.

Intraperitoneal treatment of female BALB/c mice with either phenobarbitone or beta-naphthoflavone led to the induction of various hepatic enzymes associated with xenobiotic metabolism and to increased abilities of hepatic S9 fractions to convert the dietary carcinogen 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) to an active bacterial mutagen. In the case of another carcinogen, aflatoxin B1 an increase in in vitro hepatic activation was seen only in mice treated with phenobarbitone. In contrast, pretreatment with either phenobarbitone or beta-naphthoflavone reduced the in vivo activity of both aflatoxin B1 and MeIQx in the host mediated bacterial mutation assay. These data indicate that, for some carcinogens at least, the host-mediated assay may be used to predict the carcinogenic consequences of hepatic enzyme induction.     

Effects of harman and norharman on the metabolism and genotoxicity of 2-acetylaminofluorene in cultured rat hepatocytes

Monolayers of rat hepatocytes metabolize 0.25 m M 2-acetylaminofluorene (AAF) to various ether-extractable, water-soluble as well as covalently bound products. The major ether-extractable metabolite formed is 2-aminofuorene (AF), followed by 7-OH-AAF and 9-OH-AAF. Pretreatment of rats with the inducer Aroclor 1254 (PCB) increased the metabolism of AAF and caused an increased DNA repair synthesis in hepatocytes exposed to AAF or AF. With N-OH-AAF, a decreased genotoxic response in PCB-treated cells compared to control cells was seen. The addition of harman and norharman decreased the metabolism of AAF to ether-extractable metabolites, water-soluble metabolites and metabolites covalently bound to macromolecules. In contrast, the DNA-repair synthesis caused by the same concentrations of AAF was increased by harman. One explanation for this apparent discrepancy could be that the aromatic amines changed the metabolism of harman and norharman in such a way that these compounds were converted into genotoxic metabolites.Abbreviations AAF 2-acetylaminofluorene - AF 2-aminofluorene - DMSO dimethylsulfoxide - HPLC high performance liquid chromatography - N-OH-AAF N-ydroxy-2-acetylaminofluorene - PCB polychlorinated biphenyls, Aroclor 1254 - TCDD 2,3,7,8-tetrachlorodibenzo-p-dioxin - TdR thymidine - Trp-P-1 3-amino-1,4dimethyl-5H-pyrido(4,3b)indole - Trp-P-2 3-amino-l-methyl-5H-pyrido(4,3b)indole - UDS unscheduled DNA synthesis     

Metabolic conversion of IQ and MeIQ to bacterial mutagens

The metabolic conversion of 2-amino-3-methyl- and 2-amino-3,4-dimethyl-imidazo[4,5-f]quinoline (IQ and MeIQ respectively) to bacterial mutagens was studied using a bacterial mutation assay. Studies were performed using S9 fractions derived from either corn oil (uninduced) or Aroclor-1254-treated Sprague-Dawley rats. Aroclor 1254 treatment lowered the S9 protein concentration required for optimum levels of mutagenesis, enhanced the numbers of mutants observed and altered the effects of metabolic inhibitors and cofactors added to the assay. Studies with uninduced preparations revealed that IQ and MeIQ exhibited similar responses to the effects of metabolic inhibitors and cofactors involved in detoxication reactions. Both IQ and MeIQ activation appeared to be inhibited by the biogenic amines tryptamine and tyramine and inactivated by conjugation with either acetyl coenzyme A or glutathione.     

Differential time course of induction of rat liver microsomal cytochrome P-450 isozymes and epoxide hydrolase by Aroclor 1254

The time course of induction of rat liver microsomal cytochromes P-450a, P-450b + P-450e, P-450c, and P-450d and epoxide hydrolase has been determined in immature male rats administered a single large dose [1500 mumol (500 mg)/kg body wt] of the polychlorinated biphenyl mixture Aroclor 1254. Differential regulation of these xenobiotic-metabolizing enzymes was indicated by their characteristic patterns of induction. The rate of induction of cytochrome P-450a and epoxide hydrolase was relatively slow, and steady-state levels of these enzymes were maintained from approximately Days 9 to 15 after Aroclor 1254 treatment. In contrast, cytochrome P-450c was maximally induced 2 days after Aroclor 1254 treatment and remained at a constant level through Day 15. Steady-state levels of cytochrome P-450d, beginning 1 week after Aroclor 1254 treatment, were preceded by a fairly rapid rate of induction and possibly by a small decline from maximal levels observed around Days 4 to 5. Like those of the other cytochrome P-450 isozymes and epoxide hydrolase, the levels of cytochromes P-450b + P-450e were constant from Day 9 to 15 after Aroclor 1254 treatment. However, an unexpected but reproducible decline (approximately 25%) in total cytochrome P-450 content observed between Days 4 and 9 after Aroclor 1254 treatment principally reflected a dramatic and totally unanticipated decrease (approximately 45%) in the level of cytochromes P-450b + P-450e. This transient decline in the level of cytochromes P-450b + P-450e was not due to an unusual effect of a mixture of polychlorinated biphenyls, since identical results were obtained with two individual congeners, namely 2,3,4,5,4'-penta- and 2,3,4,5,3',4'-hexachlorobiphenyl, that induced the same isozymes as Aroclor 1254. In contrast, when rats were treated with 2,4,5,2',4',5'-hexachlorobiphenyl, which induces cytochromes P-450a and P-450b + P-450e and epoxide hydrolase but not cytochromes P-450c or P-450d, maximal levels of cytochromes P-450b + P-450e were attained on Day 4 and no decrease was observed over the next 11 days. These results suggest that there may be an interaction in the regulation of induction of certain individual cytochrome P-450 isozymes.     

Aflatoxin B1-4-hydroxylase is associated with cytochrome P3-450 in C57BL/6 mouse liver

Messenger RNA from the livers of Aroclor 1254 treated mice was used to produce a cDNA library. cDNA clones corresponding to cytochromes P1-450 and P3-450 were isolated from this library by screening with a probe for the rat cytochrome P-450c gene. Specific non-cross hybridizing probes for P1-450 and P3-450 were prepared from unique restriction fragments. The radiolabeled probes were hybridized to RNA from mice treated with a low (15 mg/kg) and high (150 mg/kg, 400 mg/kg) doses of beta-naphthoflavone. The low dose of beta-naphthoflavone was found to induce only P3-450 mRNA, whereas higher doses induced both P1-450 and P3-450 mRNA. Similarly, a low dose of beta-naphthoflavone induced aflatoxin B1-4-hydroxylase, whereas higher doses induced both aflatoxin B1-4-hydroxylase and aryl hydrocarbon hydroxylase activities. These results suggest that P3-450 mRNA codes for the cytochrome that is associated with aflatoxin B1-4-hydroxylase activity.     

Mutagenic activation of 2-amino-3-methylimidazo[4,5-f]-quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline (MeIQ) by subcellular fractions and cells isolated from small intestine,kidney and liver of the rat

The mutagenic activity of the pyrolysis products 2-amino-3-methylimidazo[4,5-f]-quinoline and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline in Salmonella typhimurium TA98 using rat intestinal and renal subcellular fractions as activation systems was approximately 1 and 5 revertants per nmol, respectively. This was 1,000 times less than the activity with a subcellular fraction from rat liver. The mutagenic activity of both compounds was considerably increased using intestinal, renal and hepatic preparations isolated from PCB (Aroclor 1254)-pretreated rats, compared to preparations from control animals. In addition, both compounds displayed a moderate direct-acting mutagenic activity at concentrations above 10^-5 M. Isolated cells from small intestine, kidney and liver incubated in nucleopore chambers were able to convert both compounds into products which mutated bacteria outside the chambers. The concentrations of chemicals required to yield responses of a similar magnitude were approximately 3 orders of magnitude higher in the intestinal and renal systems compared to the hepatic system. The formation of metabolites mutagenic for Salmonella typhimurium by hepatic subcellular and cellular systems was shown to be superior to the respective intestinal and renal systems.Abbreviations AHH arylhydrocarbon hydroxylase - IQ 2-amino-3-methylimidazo[4,5-f]-quinoline - MC 3-methylcholanthrene - MeIQ 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline - PCB polychlorinated biphenyls (Aroclor 1254) - S9 the 9,000 g supernatant tissue fraction - TCDD 2,3,7,8-tetrachlorodibenzo-p-dioxin     

1-Hydroxypyrene as an indicator of the mutagenicity of coal tar after activation with human liver preparations

Liver S9 fractions were prepared from male Wistar rats, either non-induced or induced with Aroclor 1254 and from 5 human kidney transplant donors. The preparations were compared for their ability to metabolize the premutagens present in coal tar to mutagenic metabolites in the Salmonella mutagenicity assay towards strain TA98. Low levels of mutagenicity of coal tar were seen with human S9 preparations. The differences between the S9 mix of the 5 donors in capacity to activate premutagens were approximately 6-fold. The activation of coal tar by rat liver S9 preparations was higher than by the human S9 preparations. The metabolic conversion of pyrene in coal tar to 1-hydroxypyrene by the same human S9 preparations was determined in a parallel assay. 3 human preparations showed a high correlation between the formation of 1-hydroxypyrene and bioactivation of coal tar to mutagenic metabolites. The slope values of the individual regression lines were equal, suggesting that 1-hydroxypyrene is a good indicator for the activation of premutagens present in coal tar.     

Metabolic activation of promutagens in the yeast-microsome test. II. Activation of cyclophosphamide and 2-aminofluorene

The effect of recombinogenic and mutagenic activation of promutagens, cyclophosphamide and 2-aminofluorene was studied in S9 mouse liver preparations. Cyclophosphamide was activated to induce reversions of an ochre mutation and heteroallelic reversion in yeast tester strains r2089-15V-P4 and P3288, respectively. Metabolic activation of this chemical was greatly enhanced by pretreatment of mice with AROGLOR--1254. 2-aminofluorene was a potent recombinogen after metabolic activation, but proved a poor inducer of reversions of the ochre mutation. For the stationary yeast cells, activated 2-aminofluorene was shown to be not recombinogenic, while for logarithmic cells grown in galactose medium it was moderately recombinogenic, being highly active in this respect for logarithmic cells grown in glucose medium. We recommend to use these compounds as positive controls for exogenous activation in the yeast/microsome test.     

Effects of Aroclor 1254 on In Vivo Oocyte Maturation in the Mouse

Polychlorinated biphenyls (PCBs) are stable, lipophilic compounds that accumulate in the environment and in the food chain. Though some studies provided evidence that PCBs had adverse effects on reproductive function, most of these results were from in vitro models. Therefore we investigated the effect of Aroclor 1254 (a commercial PCBs mixture) treatments on in vivo maturation and developmental potential of mouse oocytes. In the present study, female ICR mice were treated with different doses (12.5, 25 and 50 mg/kg) of Aroclor 1254 (a commercial PCB mixture) once every 72 hours by intraperitoneal injection for 9 days. After three treatments of Aroclor 1254, the mice were superovulated to collect oocytes one day after the last exposure. The effects of Aroclor 1254 on oocyte maturation, fertilization, and preimplantation embryonic development were investigated. Immunofluorescence-stained oocytes were observed under a confocal microscope to assess the effects of Aroclor 1254 on spindle morphology. Parthenogenic activation and the incidence of cumulus apoptosis in cumulus-oocyte complexes were observed as well. Oocytes exposed to different doses of Aroclor 1254 in vivo were associated with a significant decrease in outgrowth potential, abnormal spindle configurations, and the inhibition of parthenogenetic activation of ovulated oocytes. Furthermore, the incidence of apoptosis in cumulus cells was increased after exposed to Aroclor 1254. These results may provide reference for the treatment of reproductive diseases such as infertility or miscarriage caused by environmental contaminants.     

EDTA affects cytochrome P450-dependent biotransformation reactions during incubations for the liver microsomal assay

In order to optimize the condition of the liver microsomal assay (LMA), studies were carried out to determine the effects of EDTA on mixed-function oxidase activity and its stability under the exact incubation conditions for the LMA. Aminopyrine N-demethylase (APD) and p-nitroanisole O-demethylase (p-NAD) activities as well as lipid peroxidation development (LP) in S9 liver fractions from beta-naphthoflavone and sodium phenobarbital (beta-NF + PB)- or Aroclor 1254 (AC)-treated mice were examined during a period of preincubation with EDTA ranging from 1 to 40 mM. At 5 mM EDTA, we obtained a strong inhibition of the microsomal LP as well as the greatest value of the mean specific activity (Asp) for both APD and pNAD activities. In agreement with the biochemical data, the presence of 5 mM EDTA in the incubation mixtures for the LMA significantly increased the mitotic gene conversion, mitotic crossing-over and point-reverse mutation of the well-known premutagen cyclophosphamide (30 mM) on the diploid D7 strain of Saccharomyces cerevisiae as the outcome of a greater metabolic activity. We concluded that the systematic use of 5 mM EDTA in LMA mixtures could improve the reliability and sensitivity of such a test.     

Mutagenicity of some derivatives of dipyrido[1,2-a:3',2'-d]imidazoles in Salmonella typhimurium with metabolic activation by rat liver and small intestine subcellular fractions

The mutagenic effect of 2-amino-dipyrido[1,2-a:3',2'-d]imidazole (Glu-P-2) was compared with that of the 3-amino, 3-nitro, or 3-N-hydroxylated derivatives of the same base ring with methyl groups at positions 4 and 6 of the molecule. The compounds were tested in Salmonella typhimurium strain TA98 without metabolic activation and in the presence of different concentrations of subcellular fractions from livers or small intestines of rats pretreated with different P448/P450 inducers. The 4,6-dimethyl compounds are always more mutagenic than Glu-P-2. Pretreatment with Aroclor 1254 (ARO) is the most effective inducer in the activation of the 2- and 3-amino compounds by liver S9, whereas the same fraction decreases the mutagenicity of the 3-nitro derivative. S9 from small intestine increased the mutagenic effect of the 3-nitro and 3-N-hydroxylated compounds, but it was unable to activate the amino compounds.     

Metabolic activation of promutagens, detectable in Ames' Salmonella assay, by 5000 X g supernatant of rat ventral prostate

Induction of aryl hydrocarbon hydroxylase (AHH) and 7-ethoxyresorufin-O-deethylase (7-EOD) activities as well as of benzo[a]pyrene (BP) metabolite formation in rat prostatic microsomes has been demonstrated after treatment with beta-naphthoflavone (BNF). The capacity to convert promutagenic compounds to ultimate mutagenic metabolites in the Ames' Salmonella assay by 5000 X g supernatant of rat ventral prostate was investigated. Male rats were treated with BNF, polychlorinated biphenyls (PCB; Arochlor 1254), phenobarbital (PB) and the vehicle, corn oil. PCB or BNF pretreatment increased the AHH- and 7-EOD activities 100-200-fold in the rat prostate 5000 X g supernatant (S-5 fraction). Epoxide hydrolase (EH) and glutathione-S-transferase (GST) activities were not affected while UDP-glucuronosyltransferase (UDP-GT) was increased 2.2- and 2.5-fold by PCB and BNF, respectively. PB did not significantly affect any of the enzyme activities measured. A dose-dependent increase in mutagenic response versus amount of 5000 X g supernatant and promutagen (aflatoxin B1 (AFB), 2-aminofluorene (2-AF), BP) was observed. The most pronounced activation was obtained with S-5 fraction from BNF- or PCB-treated rats. The great sensitivity of prostatic AHH to certain inducers and the capacity of the prostate to produce mutagenic metabolites might be of importance for initiation of prostatic cancer by environmental factors.     

Protective Role of Quercetin on Polychlorinated Biphenyls (Aroclor‐1254) Induced Oxidative Stress and Apoptosis in Liver of Adult Male Rats

The present study aims to investigate the protective effect of quercetin against Aroclor‐1254–induced hepatotoxicity in rats. Male Wistar rats were grouped into Group I control received vehicle (corn oil; 1 mL/kg bwt); Group II quercetin alone (50 mg/kg bwt/day orally); Group III Aroclor‐1254 (2 mg/kg bwt/day intraperitoneally); Group IV Aroclor‐1254 + quercetin treated for 30 days. The Aroclor‐1254 treatment caused significant alteration in the biochemical parameters (hydrogen peroxide, lipid peroxidation, reduced glutathione levels, and alkaline phosphatase activity). The expressions of apoptotic and antiapoptotic proteins and the liver histology of Aroclor‐1254–exposed rats showed cytoplasmic degeneration along with infiltration of polymorphonuclear cells. Whereas simultaneous treatment with quercetin normalized all the biochemical parameters, consequently it inhibited apoptosis mediated by Aroclor‐1254 by downregulating aryl hydrocarbon receptor, p53 and apoptotic protein (Bax, caspase‐9, caspase‐3) and upregulating the antiapoptotic protein (Bcl‐2) expression patterns; thereby, quercetin reduces alteration in hepatocellular morphology. Thus quercetin exhibited hepatoprotective effect. © 2012 Wiley Periodicals, Inc. J BiochemMol Toxicol 26:522‐532, 2012; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.21466     

Non-mutagenicity for Salmonella of the chlorinated hydrocarbons aroclor 1254, 1,2,4-trichlorobenzene, mirex and kepone.

A polychlorinated biphenyl mixture, Aroclor 1254, two commercial grade insecticides, mirex and kepone, and a pesticide breakdown product, 1,2,4-trichlorobenzene were evaluated for mutagenicity and hepatic enzyme induction potential in the Salmonella/microsomal assay. None was found to revert strains TA1535, TA1537, TA98 or TA100 when tested with or without metabolic activation. Liver microsomal extracts (S9) from rats induced with 1,2,4-trichlorobenzene were shown to differ from S9 of either control or Aroclor 1254-induced rats in the capacity to activate 2-aminoanthracene mutagenesis.