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)

The activities of several different phase I and phase II drug-metabolizing enzymes were measured in freshly isolated oval cells from rats fed a choline-deficient/DL-ethionine-supplemented diet for 6 weeks and also in vitro in the established oval cell line OC/CDE 6. No cytochrome P450 was spectrophotometrically measurable in both preparations and two cytochrome P450-dependent monoxygenase activities, aminopyrine N-demethylase and ethoxyresorufin O-deethylase, could not be detected in the oval cells of both sources. However, cytosolic glutathione transferase, microsomal epoxide hydrolase and UDP-glucuronosyltransferase activities were clearly measurable in oval cells. Similar enzyme activities were found in freshly isolated and cultured oval cells. The highest activities of these three enzymes were detected during the exponential growth phase of the cultured cells; thereafter the activities decreased until the cells reached confluency. Changes in phenol UDP-glucuronosyltransferase (UGT1A1) mRNA levels paralleled the variations in UDP-glucuronosyltransferase activity, i.e. they were high in exponentially growing oval cells and low in confluent cell cultures. Taking into account that oval cells are able to proliferate in the livers of rats continuously fed a choline-deficient/DL-ethionine-supplemented diet and that none of the analyzed drug metabolizing enzymes are involved in the activation or detoxication of DL-ethionine, the described pattern might be part of a more general, nonspecific, protection mechanism enabling these cells to overcome the cytotoxic effects of a variety of carcinogens and to proliferate even in their presence. Furthermore, the expression of microsomal epoxide hydrolase, cytosolic glutathione transferase and UDP-glucuronosyltransferase appears to depend on the proliferative status of the cells.
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PMID:Drug-metabolizing enzyme activities in freshly isolated oval cells and in an established oval cell line from carcinogen-fed rats. 807 23

In this study, we demonstrate the utility of a broad class of spectrophotometric substrates for the assay of cytosolic epoxide hydrolase purified from murine liver. These substrates, epoxy esters or carbonates, cyclize spontaneously upon or during hydrolysis of the epoxide functionality. The alcohol released by cyclization may then be assayed directly or by coupling to a second reaction. The alcohol produced, or its secondary reaction products, can be selected to give an absorption in the visible or near-uv range of the spectrum. This allows the synthesis of a wide variety of useful spectrophotometric substrates. 4-Nitrophenyl (2S,3S)-2,3-epoxy-3-phenylpropyl carbonate, at pH 6.4 and 25 degrees C, had a Vmax of 22 mumol min-1 mg-1 and a Km of 16 microM when assayed with a conventional spectrophotometer. When assayed under the same conditions with a 96-well plate reader, the measured Vmax was 15 mumol min-1 mg-1 and the Km was 6.6 microM. Some of these compounds were also found to be substrates for glutathione S-transferase, microsomal epoxide hydrolase, and porcine liver carboxylesterase. Indeed, 4-nitrophenyl 3,4-epoxy-3-phenylbutanoate was a 3.4-fold better substrate for porcine liver carboxylesterase than 4-nitrophenyl acetate when initial rates of hydrolysis were measured under the same conditions.
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PMID:Spectrophotometric substrates for cytosolic epoxide hydrolase. 813 50

Chemically reactive epoxide metabolites have been implicated in various forms of drug and chemical toxicity. Naphthalene, which is metabolized to a 1,2-epoxide, has been used as a model compound in this study in order to investigate the effects of perturbation of detoxication mechanisms on the in vitro toxicity of epoxides in the presence of human liver microsomes. Naphthalene (100 microM) was metabolized to cytotoxic, protein-reactive and stable, but not genotoxic, metabolites by human liver microsomes. The metabolism-dependent cytotoxicity and covalent binding to protein of naphthalene were significantly higher in the presence of phenobarbitone-induced mouse liver microsomes than with human liver microsomes. The ratio of trans-1,2-dihydrodiol to 1-naphthol was 8.6 and 0.4 with the human and the induced mouse microsomes, respectively. The metabolism-dependent toxicity of naphthalene toward human peripheral mononuclear leucocytes was not affected by the glutathione transferase mu status of the co-incubated cells. Trichloropropene oxide (TCPO; 30 microM), an epoxide hydrolase inhibitor, increased the human liver microsomal-dependent cytotoxicity (19.6 +/- 0.9% vs 28.7 +/- 1.0%; P = 0.02) and covalent binding to protein (1.4 +/- 0.3% vs 2.8 +/- 0.2%; P = 0.03) of naphthalene (100 microM), and reversed the 1,2-dihydrodiol to 1-naphthol ratio from 6.6 (without TCPO) to 2.6, 0.6 and 0.1 at TCPO concentrations of 30, 100 and 500 microM, respectively. Increasing the human liver microsomal protein concentration reduced the cytotoxicity of naphthalene, while increasing its covalent binding to protein and the formation of the 1,2-dihydrodiol metabolite. Co-incubation with glutathione (5 mM) reduced the cytotoxicity and covalent binding to protein of naphthalene by 68 and 64%, respectively. Covalent binding to protein was also inhibited by gestodene, while stable metabolite formation was reduced by gestodene (250 microM) and enoxacin (250 microM). The study demonstrates that human liver cytochrome P450 enzymes metabolize naphthalene to a cytotoxic and protein-reactive, but not genotoxic, metabolite which is probably an epoxide. This is rapidly detoxified by microsomal epoxide hydrolase, the efficiency of which can be readily determined by measurement of the ratio of the stable metabolites, naphthalene 1,2-dihydrodiol and 1-naphthol.
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PMID:An investigation of the formation of cytotoxic, genotoxic, protein-reactive and stable metabolites from naphthalene by human liver microsomes. 824 Apr 7

In order to investigate the role of the microsomal epoxide hydrolase (mEH) in the detoxification of arene oxides in the presence of a high endogenous glutathione S-transferase (GST) activity-a situation found in several organs--we expressed the rat mEH cDNA in BHK21 Syrian hamster cells. These cells have high GST activities but contain an extremely low endogenous mEH enzyme activity. We obtained several cell clones which expressed the mEH heterologously, as determined by immunoblotting. The cell clone BHK21-mEH/Mz1 had the highest level of mEH protein. Immunofluorescence showed that the level of expression was almost homogeneous throughout the cell population. Total protein isolated from the cell line BHK21-mEH/Mz1 had a specific mEH activity of 123 pmol/min/mg protein, as determined with benzo[a]pyrene 4,5-oxide (B[a]P 4,5-oxide), which was 60 times higher than the activity in the parental cell line and eight times lower than the activity found in rat hepatocytes. However, BHK21-mEH/Mz1 cell homogenates were found to catalyze the conjugation of B[a]P 4,5-oxide to glutathione extremely well. The ratio of the GST enzyme activity to the mEH enzyme activity towards this substrate was 23 in the BHK21-mEH/Mz1 cell line. For hepatocytes this ratio was only six. Despite their already high potential to inactivate B[a]P 4,5-oxide by conjugation to glutathione, BHK21-mEH/Mz1 cells were better protected against the toxic and mutagenic effects of B[a]P 4,5-oxide than the parental cell line due to the expression of the mEH. The mEH, however, failed to protect the cells from the toxic and mutagenic effects of the bay region epoxide anti-7-methylbenz[a]anthracene-3,4-diol 1,2-oxide.
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PMID:Studies on the importance of microsomal epoxide hydrolase in the detoxification of arene oxides using the heterologous expression of the enzyme in mammalian cells. 831 4

We have studied the expression of different xenobiotic metabolizing enzymes in primary operable breast cancer of no special type. The expression of two forms of cytochrome P450, microsomal epoxide hydrolase, and three classes of glutathione S-transferase was investigated using immunohistochemistry. The tumours were characterized by consistent expression of microsomal epoxide hydrolase and by variable expression of the two forms of cytochrome P450 and the three types of glutathione S-transferase. Cytochrome P450 1A and cytochrome P450 3A were identified in 39 and 22 per cent of tumours, respectively. In each case, immunostaining was present only in areas of invasive carcinoma. Epoxide hydrolase was identified in 89 per cent of tumours and glutathione S-transferases pi, mu, and alpha were identified in 56, 65, and 44 per cent of tumours, respectively. Immunoreactivity for epoxide hydrolase and glutathione S-transferases was identified in both tumours and non-neoplastic breast tissue. The presence of different xenobiotic metabolizing enzymes may have a role in determining the intrinsic drug resistance of breast cancer to a variety of anti-cancer drugs, and the expression of these enzymes can readily be assessed using immunohistochemistry.
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PMID:Expression of xenobiotic metabolizing enzymes in breast cancer. 849 28

1. Among nitrogen heterocycles based on the planar phenanthrene structure are three (1,7- and 4,7-phenanthroline and phenanthridine) which selectively increase rat hepatic phase II drug metabolizing enzyme activities without increasing cytochrome P450 concentration. Of six monooxygenase activities investigated, only ethoxyresorufin dealkylase was raised but this was only minor. 2. The detergent-activated UDP-glucuronosyltransferase activities towards morphine, 4-nitrophenol, and 1-naphthol were increased up to five-, three- and two-fold of control respectively. Microsomal epoxide hydrolase activity towards cis-stilbene oxide was increased up to three-fold and cytosolic glutathione S-transferase activity towards 1-chloro-2, 4-dinitrobenzene reached twice the control value. 3. Cytosolic 4-nitrophenol sulphotransferase activity was not increased by any compound and like some monooxygenase reactions, was decreased by 4,7- and 1,7-phenanthrolines. 4. 1,10-Phenanthroline and two compounds which lack a heterocyclic nitrogen atom, (phenanthrene and 9-phenanthrol), failed to elicit any induction of enzyme activities. 5. Changes in microsomal epoxide hydrolase activity showed high correlation (r = 0.97) with changes in UDP-glucuronosyltransferase (4-nitrophenol) activity.
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PMID:Selective induction of rat liver phase II enzymes by N-heterocycle analogues of phenanthrene: a response exhibiting high correlation between UDP-glucuronosyltransferase and microsomal epoxide hydrolase activities. 849 89

1. The effects of garlic oil (GO) on the expression of P4502E1, glutathione S-transferase (GST) and microsomal epoxide hydrolase (mEH) were assessed by metabolic activities, immunoblot and RNA blot analyses in the rat. 2. p-Nitrophenol (PNP) hydroxylase activity decreased in the hepatic microsomes isolated from rats treated with GO at 200 mg/kg b.w. by 10-30% as compared with control. Pyrazine-inducible P4502E1 expression was decreased by approximately 40% following concomitant treatment of animals with GO at the dose of 200 mg/kg from day 1 to 3 post-treatment, as evidenced by PNP hydroxylase activity. The rates of aniline hydroxylase and NDMA demethylase activities in GO-treated animals were consistent with those of PNP hydroxylase activity. Treatment of animals with 500 mg/kg GO resulted in suppression of P4502E1-mediated catalytic activities, as monitored by both PNP and aniline hydroxylase activities, whereas the effects at the dose of 1000 mg/kg were identical with those at 500 mg/kg b.w. 3. Immunoblot analyses of hepatic microsomes, using an anti-P4502E1 antibody, showed that GO minimally suppressed constitutive P4502E1 expression at 24, 48 and 72 h post-treatment at the daily doses from 200 to 1000 mg/kg b.w., as compared with vehicle-treated animals. Time-dependent pyrazine induction of P4502E1, however, was substantially blocked by concomitant treatment of animals with 200 mg/kg GO to the levels of control. Treatment at the dose of 1000 mg/kg failed to further suppress P4502E1 levels. GO treatment caused no changes in the levels of P4502E1 mRNA, as assessed by slot blot analyses. 4. Cytosol produced from the GO-treated rat showed approximately 40% increases in GST conjugating activity toward 1-chloro-2,4-dinitrobenzene, whereas mEH protein levels were 1.5-2.0-fold greater than control with similar increases in the mRNA levels noted. 5. These results demonstrate that GO suppresses inducible P4502E1 expression more significantly than constitutive expression, and that GO induces GST and mEH expression to a certain extent.
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PMID:Effects of garlic oil on rat hepatic P4502E1 expression. 857 58

Within the selective induction of phase II enzymes following treatment with dipyridyls or N-heterocyclic analogs of phenanthrene, strong correlations (r > or = 0.70) are observed between the increase of microsomal epoxide hydrolase (mEH) activity and UDP-glucuronosyltransferase (UGT) activities towards 4-nitrophenol, 1-naphthol and morphine. The present study investigates whether this correlation is maintained with inducing agents known to also increase phase I enzyme activities. Rats were treated with beta-naphthoflavone, isosafrole, phenobarbital, ethanol, dexamethasone and clofibric acid regimens in which P450 isozyme induction could be confirmed. Comparisons between the responses of mEH, UGT and glutathione S-transferase (GST) activities were made. mEH activity was increased by beta-naphthoflavone, isosafrole, phenobarbital and clofibric acid. The elevation in mEH activity by these agents showed modest but significant correlations with GST activities toward all the substrates monitored (r values range between 0.49 and 0.65) and a strong correlation with UGT activity towards only one substrate, morphine (r = 0.70). This study suggests that induction of mEH activity correlates with the increases in select phase II enzyme activities whether it is accompanied by P450 induction or not.
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PMID:Correlations of the induction of microsomal epoxide hydrolase activity with phase II drug conjugating enzyme activities in rat liver. 861 56

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

Human bronchial epithelial cells (BEC), a primary defense against inhaled materials, are the progenitor cells for bronchogenic carcinomas and have important metabolic capabilities. We used reverse transcriptase-polymerase chain reaction (RT-PCR) to identify xenobiotic metabolism enzymes expressed in primary BEC and alveolar macrophages (AM) of non-smoking volunteers. Cytochromes P450 (CYP) 1A1, 1B1, 2B7, 2E1, and 4B1 and microsomal epoxide hydrolase (mEH) were expressed in BEC but not AM. CYP2F1 was expressed in BEC, but it was expressed at barely detectable levels or not at all in AM. NADPH oxidoreductase (NADPH OR), microsomal glutathione transferase (GST 12), glutathione transferase mu, phenol sulfotransferase (PST), thermolabile phenol sulfotransferase (TL PST), and the clara cell-specific gene, CC10 were expressed in both BEC and AM. CYP3A4 and glucuronosyl transferases-1 and 2 were not expressed in either BEC or AM. In contrast to primary BEC, of the genes evaluated, the immortalized human bronchial epithelial cell line BEP2D constitutively expressed only CYP1A1, CYP2E1, NADPH OR, glucuronosyl transferase 1, GST 12, GST mu, PST, TL PST, and CC10. The loss of xenobiotic metabolism enzyme gene expression in the BEP2D cell line may result from either reduced exposure to inducing agents, or loss of differentiative characteristics in culture. It is clear from the data comparing BEC and AM that there are important intertissue differences in expression of xenobiotic metabolism enzymes.
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PMID:Xenobiotic metabolism enzyme gene expression in human bronchial epithelial and alveolar macrophage cells. 884 77


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