Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.5.1.18 (glutathione S-transferase)
22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Anthraquinone dyes are utilized by the military in colored-smoke grenades. During production, workers in munitions plants may be exposed to fugitive emissions of these dyes or mixtures thereof. The effects of a prototype violet dye mixture (VDM) consisting of Disperse Red 11 (DR11), [1,4-diamino-2-methoxy-anthraquinone] and Disperse Blue 3 (DB3) [1-methylamino-4-hydroxyethylamino-anthraquinone] on F344 male and female rats have been investigated. Acute 1-day inhalation exposures (6 hr) to VDM were conducted at 1000, 300, 100, 70, 40, and 10 mg/m3, with an additional exposure to 40 mg/m3 6 hr/day for 5 days; 4.22 +/- 2.1 microns (MMAD +/- delta g). Lung burdens of dye, general histopathology, and/or liver function were evaluated at 0, 3, and 7 days postexposure. Unexpected lethality due to severe liver damage was observed with acute exposures of > or = 300 mg/m3 and in the 5-day 40 mg/m3 exposures. Centrilobular degeneration and necrosis of liver cells was concentration-dependent with inhalation of VDM > or = 40 mg/m3. In addition, nasal olfactory epithelium exhibited degeneration and necrosis with acute exposures > or = 10 mg/m3. Lung instillations at 250, 500, and 1000 micrograms of the VDM revealed no lung or liver toxicity. Because per os exposure due to preening was suspected as a major exposure route, a gavage study with the VDM and its two component dyes DR11 and DB3 (800 mg/kg) was undertaken. One day following gavage with DR11 or DB3, serum enzymes indicative of liver toxicity (LDH, SGPT, SDH, and ICDH) were slightly elevated (1-6x control). However, rats gavaged with VDM had serum enzyme levels 10-100x control by Day 1 after gavage, indicating acute liver toxicity. Activities of liver enzymes involved in xenobiotic and glutathione metabolism were also acutely affected. All of the dyes caused various degrees of induction of glucose-6-phosphate dehydrogenase, glutathione reductase, glutathione peroxidase, and nonprotein sulfhydryls. The enzymes involved in xenobiotic metabolism (glutathione S-transferase, NADPH cytochrome-c reductase, and P450) were also elevated by the two component dyes, in contrast to their significant depression with VDM treatment. The similarity between the liver and olfactory epithelium effects of these compounds and the lack of pulmonary tissue effects is not fully understood, but the interaction of the individual dyes as VDM emphasizes the need to assess chemicals such as the anthraquinones as their likely-to-be-encountered mixtures.
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PMID:Toxicity of an anthraquinone violet dye mixture following inhalation exposure, intratracheal instillation, or gavage. 812 3

Ingestion of aflatoxin B1 (AFB1) represents a major risk factor in the aetiology of human hepatocellular carcinoma. In the rat, the harmful effects of AFB1 can be prevented by the administration of certain drugs which induce hepatic detoxification enzymes. We have previously shown that treatment of rats with the chemoprotector ethoxyquin (EQ) results in a marked increase in expression of the Alpha-class glutathione S-transferase (GST) Yc2 subunit which has high activity towards AFB1-8,9-epoxide [Hayes, Judah, McLellan, Kerr, Peacock and Neal (1991) Biochem. J. 279, 385-398]. To allow an assessment of whether the increased expression of GST Yc2 represents a general adaptive resistance mechanism to chemical stress, that is invoked by both chemoprotectors and carcinogens, we have examined the effects of EQ, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), phenobarbital (PB), AFB1, 3-methylcholanthrene (3-MC) and clofibrate on the AFB1-glutathione-conjugating activity and the GST subunit levels in rat liver. In addition, the effect of these drugs on the hepatic levels of an aldehyde reductase (AFB1-AR) that metabolizes the cytotoxic dialdehydic form of AFB1 has been studied as this enzyme also appears to be important in chemoprotection. Administration of the antioxidants EQ, BHA or BHT, as well as PB, led to a marked increase in levels of the GST Yc2 subunit in rat liver, and this increase coincided with a substantial rise in the GST activity towards AFB1-8,9-epoxide; neither AFB1, 3-MC nor clofibrate caused induction of Yc2 or any of the GST subunits examined. Among the xenobiotics studied, EQ was found to be the most effective inducing agent for the Yc2 subunit as well as Yc1, Yb1 and Yf. However, PB was equally as effective as EQ in increasing levels of the Ya-type subunits, although it was not found to be as potent an inducer of the other GST subunits, including Yc2. In addition to induction of GST, EQ caused a substantial increase in the hepatic content of AFB1-AR. Both BHA and BHT were also able to induce this enzyme but, by contrast, PB was found to be a poor inducer of AFB1-AR. AFB1, 3-MC and clofibrate were unable to serve as inducers of this reductase. The presence of Alpha-class GST, including the Yc2 subunit, was examined in various rat tissues. Constitutive expression of Yc2 was found in the epididymis at levels comparable with that observed in the liver from EQ-treated rats.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Regulation of aflatoxin B1-metabolizing aldehyde reductase and glutathione S-transferase by chemoprotectors. 819 22

In this report we describe the heterologous expression of glutathione S-transferase (GST) and cytochrome P450 reductase (Red) in E. coli and Salmonella typhimurium. The same expression vectors could be applied to both systems and high levels of catalytically active GST and Red were obtained. Interestingly the level of expression was invariably higher in S. typhimurium. The level of the alpha class GST being up to 20% of the total bacterial protein. A further advantage of the salmonella system is that strains were used which can be applied to mutagenicity tests. This system was validated by demonstrating increasing mutation frequency of halogenated hydrocarbons in strains expressing the GST and increased cytotoxicity of mitomycin C in cells expressing P450 reductase.
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PMID:Heterologous expression of drug-metabolizing enzymes in cellular and whole animal models. 823 79

The alkylating agent BCNU [1,3-bis(2-chloroethyl)-1-nitrosourea] can be inactivated through denitrosation reactions catalyzed by both cytosolic and microsomal enzymes. While previous studies have identified a class mu glutathione S-transferase [rat transferase 4-4 (Yb2)] as a major catalyst of the cytosolic denitrosation reaction, the enzymatic catalysts of BCNU denitrosation in microsomal membranes have not been identified. In the present study, both NADPH and glutathione (GSH) were found to support BCNU denitrosation catalyzed by isolated rat liver microsomes. Treatment of rats with the microsomal enzyme inducers phenobarbital and dexamethasone increased NADPH-dependent liver microsomal BCNU denitrosation up to fivefold without major effect on the GSH-dependent denitrosation activity. Although the NADPH-dependent activity was fully inhibited by antibody to NADPH-P450 reductase, purified NADPH-P450 reductase catalyzed BCNU denitrosation at rates that could only account for approximately 2-3% of the microsomal activity. Other experiments, including selective inhibition of NADPH-dependent microsomal BCNU denitrosation by chemical and antibody inhibitors of cytochrome P450, competitive inhibition of P450-catalyzed cyclophosphamide and ifosfamide activation by BCNU, and reconstitution of the denitrosation reaction by purified P450 enzyme 2B1 (major phenobarbital-inducible P450 form), established an important role for cytochrome P450 in BCNU denitrosation. By contrast, GSH-dependent microsomal BCNU denitrosation was unaffected by cytochrome P450 inhibitors, but was inhibited, with varying degrees of selectivity, by the microsomal glutathione S-transferase inhibitors ethacrynic acid, bromosulfophthalein, and indomethacin. These studies establish that BCNU inactivation can be catalyzed by two independent microsomal enzyme systems and suggest that therapeutically useful improvements in BCNU antitumor activity might be achieved through differential inhibition of these enzyme systems in tumor as compared to extratumoral sites.
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PMID:Denitrosation of the anti-cancer drug 1,3-bis(2-chloroethyl)-1-nitrosourea catalyzed by microsomal glutathione S-transferase and cytochrome P450 monooxygenases. 827 24

Complementary DNA sequences encoding the mature form of pea ferredoxin-NADP+ reductase were cloned in-frame at the 3' end of the Schistosoma japonicum glutathione S-transferase gene in the expression vector pGEX-3X (Smith and Johnson, Gene 67, 31-40, 1988). A spacer sequence linking the two genes was modified to provide a proteolytic site just before the first amino acid residue of mature pea reductase. When introduced into competent Escherichia coli cells and induced, the resulting plasmid (pGF205) directed the expression of a 60-kDa immunoreactive peptide that results from the fusion between glutathione S-transferase and ferredoxin-NADP+ reductase sequences. The fused protein could be purified in a single step by selective absorption onto glutathione-agarose beads, followed by elution with free glutathione. It showed both transferase and reductase activities. Removal of the transferase portion by cleavage with the restriction protease Xa rendered ferredoxin-NADP+ reductase electrophoretically homogeneous. The purified transgenic enzyme showed kinetic and spectroscopic properties that were similar to those reported for the plant flavoprotein, indicating that, even when fused to the 27-kDa transferase portion, the reductase was still able to assemble FAD and to acquire an active conformation in the bacterial host. The expression-purification protocol employed here allows the isolation of up to 1 mg of active ferredoxin-NADP+ reductase/g of transformed cells. The system is potentially useful for the purification of activity-impaired forms of the flavoprotein.
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PMID:One-step purification of plant ferredoxin-NADP+ oxidoreductase expressed in Escherichia coli as fusion with glutathione S-transferase. 828 51

Previous studies reported that, in the absence of drug exposure, multidrug resistance, including resistance to Adriamycin (ADR), could develop in primary rat hepatocyte cultures (B. Carr, Proc. Am. Assoc. Cancer Res., 29:1158, 1988). However, the hepatocytes in that report were cultured on plastic without the benefit of an extracellular matrix (ECM). Because the ECM regulates hepatic gene expression, we have critically evaluated in primary cultures of rat hepatocytes how the ECM affects hepatic ADR resistance, the level of the drug efflux transporter associated with MDR, P-glycoprotein (pgp), and transport of a prototypical pgp substrate, vincristine. Hepatocytes cultured on type I collagen (Vitrogen) had greater resistance to ADR toxicity accompanied by parallel increases in the level of pgp mRNA, decreased drug accumulation, and enhanced drug efflux when compared with the hepatocytes maintained on the basement membrane matrix Matrigel. The development of ADR resistance coincided with the time course of increased pgp mRNA but was not coincident with the time course of expression of either the placental isozyme of glutathione S-transferase or P-450 reductase, proteins associated with MDR in some resistance models. Southern blot analysis revealed neither gross changes in pgp gene structure or gene copy number to account for the increase in pgp RNA levels for hepatocytes cultured on Vitrogen. ECM also regulated xenobiotic-inducible expression of hepatic pgp, since chemotherapeutic agents, including vincristine and colchicine, induced pgp mRNA exclusively in hepatocytes cultured on Vitrogen. The critical matrix proteins in Matrigel responsible for regulation of pgp were determined by the selective addition of its components to the culture environment. The presentation of the individual matrix elements as a rigid substratum to the hepatocyte did not decrease pgp mRNA. In contrast, the presentation to the same hepatocytes of either laminin or type IV collagen in a nonrigid state (solubly in the medium) selectively decreased hepatocellular pgp mRNA. We conclude that primary rat hepatocytes develop ADR resistance with time in culture due to increased expression of pgp and that ECM proteins represent endogenous physiological modulators of both basal and chemotherapeutically inducible expression of hepatic P-glycoprotein.
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PMID:Extracellular matrix regulation of multidrug resistance in primary monolayer cultures of adult rat hepatocytes. 842 4

The effects of aging on the activities of drug-metabolizing enzymes and antioxidant enzymes were studied in male and female White-Footed mice (Peromyscus leucopus) at ages of 6, 8, 12, 18, 24, 30, 36, and 48 months. Male mice had significantly higher liver microsomal cytochrome P450 (P450) content and NADPH:cytochrome P450 oxidoreductase (P450 reductase) activities than females at all age groups. Many of the P450-dependent enzyme activities were also generally higher in males. Female mice showed age-dependent decreases in P450 content and the activities of P450 reductase, pentoxyresorufin O-dealkylase (PROD) and N-nitrosodimethylamine demethylase (NDMAd) in the liver from 6 to 24 months; while, the males showed an age-dependent decrease only for the liver PROD activity from 6 to 24 months. The old males (30-month old) appeared to have significantly higher activities for 6 beta-, 2 beta-, 16 alpha- and 16 beta-testosterone and androstenedione formation than the middle-aged (6- to 18-month old) and very old (48-month old) males. Females showed age-dependent decreases for the formation of 6 beta-, 2 beta-, 16 alpha- and 16 beta-testosterone in liver microsomes from 6 to 24 months. Lung microsomes from 6- and 8-month old males had much higher activities of ethoxyresorufin O-deethylase (EROD) and PROD than older males. The total NNK alpha-hydroxylation activities changed in the same pattern as lung microsomal EROD and PROD activities in both male and female mice. The activities of several phase II drug-metabolizing enzymes: glutathione S-transferase (GST), DT-diaphorase, sulfotransferase and UDP-glucuronosyl-transferase (UDPGT) did not show any significant age-dependent changes, with the possible exception that the GST activity in males decreased from 18 to 36 months. Males had about 3-fold higher UDPGT activities than females among all age groups. Glutathione peroxidase activities were drastically lower in old and very old males, and 6 to 24 months old males had significantly higher activities than the corresponding females. In females, superoxide dismutase activities decreased linearly to extremely low levels as mice aged. Catalase activities showed a tendency for increase with age in males. In conclusion, some P450-dependent activities and antioxidant enzymes, but not phase II drug-metabolizing enzymes, showed age-dependent changes; and most of these changes occur from 6 to 24 months. The demographic attributes of the White-Footed mouse are well-suited for physiological and biochemical studies of aging and can complement the more standard laboratory mouse model with its typical two year life span.
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PMID:Age- and gender-related variations in the activities of drug-metabolizing and antioxidant enzymes in the white-footed mouse (Peromyscus leucopus). 849 97

Chronic pancreatitis and pancreatic cancer have both been linked with occupational exposure to organic chemicals. These chemicals are known to be metabolized within the liver by the cytochrome P-450 family of enzymes, and indeed are able to induce levels of these enzymes as evidence of their interaction. The purpose of this study was therefore to see if these enzyme systems were altered in chronic pancreatitis and pancreatic cancer. Immunocytochemistry of four phase I drug-metabolizing enzymes (cytochromes P-450 IIIA1, P-450 IIE, P-450 IA2, and NADPH cytochrome P-450 oxido-reductase) and one phase II enzyme [glutathione S-transferase (GST) 5-5] was therefore performed on pancreas and/or liver biopsy samples from organ donors and compared with patients with chronic pancreatitis or pancreatic cancer. In samples from donor subjects, the types and levels of drug-metabolizing enzymes in hepatocytes were similar to those seen in pancreatic acinar cells. In material from patients with chronic pancreatitis or pancreatic cancer, cytochrome P-450 enzyme levels were greater in both the liver and the pancreas than those seen in the donor group, while GST levels were unchanged. Islets of Langerhans showed high levels of P-450 IA2 in the donor group, with clear induction of P-450 IIIA1 and NADPH cytochrome P-450 oxidoreductase in patients with chronic pancreatitis but not in the pancreatic cancer group. Levels of GST 5-5 were also induced in the islets. The present findings raise the possibility of an aetiological relationship between elevated levels of drug-metabolizing enzymes and the subsequent development of disease.
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PMID:Induction of drug-metabolizing enzymes in human pancreatic cancer and chronic pancreatitis. 850 44

Ellagic acid (EA), a naturally occurring plant polyphenol possesses broad chemoprotective properties. Dietary EA has been shown to reduce the incidence of N-2-fluorenylacetamide-induced hepatocarcinogenesis in rats and N-nitrosomethylbenzylamine (NMBA)-induced rat esophageal tumors. In this study changes in the expression and activities of specific rat hepatic and esophageal mucosal cytochromes P450 (P450) and phase II enzymes following dietary EA treatment were investigated. Liver and esophageal mucosal microsomes and cytosol were prepared from three groups of Fisher 344 rats which were fed an AIN-76 diet containing no EA or 0.4 or 4.0 g/kg EA for 23 days. In the liver total P450 content decreased by up to 25% and P450 2E1-catalyzed p-nitrophenol hydroxylation decreased by 15%. No changes were observed in P450 1A1, 2B1 or 3A1/2 expression or activities or cytochrome b5 activity. P450 reductase activity decreased by up to 28%. Microsomal epoxide hydrolase (mEH) expression decreased by up to 85% after EA treatment, but mEH activities did not change. The hepatic phase II enzymes glutathione S-transferase (GST), NAD(P)H:quinone reductase [NAD-(P)H:QR] and UDP glucuronosyltransferase (UDPGT) activities increased by up to 26, 17 and 75% respectively. Assays for specific forms of GST indicated marked increases in the activities of isozymes 2-2 (190%), 4-4 (150%) and 5-5 (82%). In the rat esophageal mucosa only P450 1A1 could be detected by Western blot analysis and androstendione was the only P450 metabolite of testosterone detectable. However, there were no differences in the expression of P450 1A1, the formation of androstendione or NAD(P)H:QR activities between control and EA-fed rats in the esophagus. Although there was no significant decrease in overall GST activity, as measured with 1-chloro-2,4-dinitrobenzene (CDNB), there was a significant decrease in the activity of the 2-2 isozyme (66% of control). In vitro incubations showed that EA at a concentration of 100 microM inhibited P450 2E1, 1A1 and 2B1 activities by 87, 55 and 18% respectively, but did not affect 3A1/2 activity. Using standard steady-state kinetic analyses, EA was shown to be a potent non-competitive inhibitor of both liver microsomal ethoxyresorufin O-deethylase and p-nitrophenol hydroxylase activities, with apparent Ki values of approximately 55 and 14 microM respectively. In conclusion, these results demonstrate that EA causes a decrease in total hepatic P450 with a significant effect on hepatic P450 2E1, increases some hepatic phase II enzyme activities [GST, NAD-(P)H:QR and UDPGT] and decreases hepatic mEH expression. It also inhibits the catalytic activity of some P450 isozymes in vitro. Thus the chemoprotective effect of EA against various chemically induced cancers may involve decreases in the rates of metabolism of these carcinogens by phase I enzymes, due to both direct inhibition of catalytic activity and modulation of gene expression, in addition to effects on the expression of phase II enzymes, thereby enhancing the ability of the target tissues to detoxify the reactive intermediates.
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PMID:The effects of dietary ellagic acid on rat hepatic and esophageal mucosal cytochromes P450 and phase II enzymes. 862 97

Xenobiotic metabolism can be influenced by various nutritional factors, including protein. In the present study, we have examined the effect of dietary protein (casein) levels on the ability of rat liver S9 to activate the promutagens aflatoxin B1 (AFB), 2-aminoanthracene (2AN) and benzo[a]pyrene (BAP) in strain TA98 using the spiral Salmonella mutagenicity assay. S9s were derived from individual male F344 rats fed for 6 weeks on semisynthetic diets containing 8%, 12% or 22% methionine-supplemented casein as the sole source of protein (diets were made isocaloric by adjusting the corn starch content). Rats were housed in large, raised-bed cages by groups of three per diet. S9 activation mixtures were prepared at 5 mg of S9 protein/ml of S9 mix. Slopes from the linear portions of the mutagenicity dose-response curves were analyzed by ANOVA comparisons. Assays used to elucidate the phase I activities of microsomal preparations were cytochrome P450 content, cytochrome-c reductase activity, flavin-containing monooxygenase activity, 7-ethoxyresorufin O-deethylation (EROD) activity, N-demethylation of benzphetamine and para-nitrophenol O-deethylation. Phase II activities in cytosolic preparations were assayed by estimation of glutathione (GSH) content and glutathione S-transferase activity through metabolism of 1-chloro-2,4-dinitrobenzene (CDNB). Increased levels of dietary casein increased liver wet weights and decreased the ability of the S9 to activate 2AN. Dietary casein levels did not influence the S9-mediated activation of BAP; and consistent but nonsignificant increases in activation of AFB were produced by S9 from animals fed the 22% casein diet. The phase I and phase II activities measured here were not altered significantly by dietary casein levels; thus, other, more specific enzymatic activities may account for the mutagenesis data. These results illustrate the complex interaction between dietary levels of casein and promutagen activation mechanisms, which prevents drawing broad generalizations regarding the influence of dietary casein levels on the capacity of hepatic S9s to activate promutagens.
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PMID:Effect of dietary casein levels on activation of promutagens in the spiral Salmonella mutagenicity assay. I. Studies with noninduced rat liver S9. 864 64


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