Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C1260386 (GSH)
 38,102 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Rat hepatocytes prepared by collagenase digestion or EDTA dissociation were examined in culture for comparison of culture stability and morphology, and retention of selected adult rat liver characteristics. Cells prepared by EDTA perfusion followed by Percoll centrifugation were deemed to form confluent monolayer cultures more rapidly and monolayers remained intact for up to 21 days without signs of nonparenchymal cell growth or loss of primary hepatocyte appearance. The spectrally determined cytochrome P-450 content remained constant through eight days in culture. Collagenase-prepared cells contained an identical amount of P-450 but within 72 hr lost greater than 80% of the spectrally detectable P-450. Glutathione (GSH) content was higher in the EDTA-prepared hepatocytes and remained constant with only a modest effect of transferrin and selenium (T/S) supplementation, while GSH levels in collagenase-prepared cells increased, thereafter decreased with time in culture and was dependent on T/S supplementation. Cells prepared with EDTA also displayed an increase in GSH efflux rate in response to chronic GSH depletion by ethacrynic acid. gamma-Cystathionase (CNase) activity was retained at initial levels in EDTA-prepared hepatocytes supplemented with T/S and declined only about 25% in unsupplemented cells. Collagenase-prepared cells lost 75% of CNase activity by 72 hr. The established marker of hepatocyte neoplastic transformation, gamma-glutamyl transpeptidase (GGT), increased rapidly in collagenase-prepared cells. The accumulation of GGT was slowed by T/S supplementation. GGT activity did not increase in EDTA-prepared hepatocytes. Evaluation of morphological and biochemical criteria suggest that hepatocytes prepared without collagenase present superior model systems for the study of biochemical events through more extended culture times.
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PMID:Rat hepatocytes prepared without collagenase: prolonged retention of differentiated characteristics in culture. 285 31

Previous methods to deplete in vivo concentrations of reduced glutathione (GSH) have not been able to lower tissue GSH levels for extended periods, have been toxic, and can alter the metabolism of xenobiotics. A possible alternative to lower in vivo concentrations of GSH may be the use of buthionine-S,R-sulfoximine (BSO) in the drinking water of laboratory animals to inhibit the biosynthesis of GSH. It has been previously reported that 20 mM BSO in the drinking water given to mice was able to lower GSH levels in a variety of tissues after 15 days. In order to more fully characterize the in vivo depletion of GSH in tissues by ingestion of BSO and determine if this method would be suitable in studies requiring depressed levels of GSH for extended periods, we added different amounts of this agent to the drinking water given to mice for various times up to 28 days. We found that ingested BSO at the highest concentration used in drinking water (30 mM) was able to maximally lower GSH concentrations in mouse lungs, lung lavage fluid, liver, kidneys, and blood to 59.0 +/- 3.6%, 35.0 +/- 5.1%, 44.3 +/- 1.5%, 69.5 +/- 3.9%, and 70.0 +/- 6.0% of control mice, respectively, for up to 28 days. These lowered concentrations of tissue GSH returned to control levels after mice were returned to untreated drinking water for 7 days. The potential toxicity of such treatments was also evaluated. Levels of alkaline phosphatase, lactate dehydrogenase, glucose-6-phosphate dehydrogenase, glutathione peroxidase, and glutathione reductase in lungs and lung lavage fluid, and total and differential cell counts from lung lavage fluid were not different between control and BSO-treated mice. This showed that BSO treatment did not produce indications of lung injury as measured by these biochemical parameters. Serum aspartyl transferase and gamma-glutamyl transpeptidase activities were unaffected by the BSO treatments, indicating normal liver functions. Lung and liver cytochrome P-450 concentrations were also not different between controls and BSO-treated animals. Thus, BSO in the drinking water of mice was able to effectively lower in vivo levels of GSH without eliciting acute toxic responses.
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PMID:Effects of the long-term depletion of reduced glutathione in mice administered L-buthionine-S,R-sulfoximine. 286 40

Hemin-thiolate complexes, as chemical models for cytochrome P-450 monooxygenases, have been shown to cause strand scission of DNA. Circular super-coiled DNA was degraded to open-circular and linear forms by these complexes in 30 min at pH 7.8 under aerobic conditions, the degradation depending on the structure of the thiol ligand and the ratio of thiol ligand to hemin concentration. The relationship between the structure of the thiol ligand and DNA strand cleaving activity was examined. Complete cleavage of DNA was observed by complexes containing TGE and ME at 400-600 moles excess of thiol ligand to hemin, those containing Cys, CysMe, and CysEt at 50-200 moles excess, and those containing MPG, GSH, penta- and nona-peptides at 5-20 moles excess. Complexes containing NACys and MEA caused no cleavage of DNA. Inhibition experiments suggested the involvement of active oxygen species in the cleavage.
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PMID:DNA strand scission by hemin-thiolate complexes as models of cytochrome P-450. 301 Sep 88

The effect of phenobarbital pretreatment on acinar distribution of microsomal drug metabolizing enzymes was investigated by analysis of periportal (pp) and perivenous (pv) enriched rat hepatocytes isolated by collagenase gradient perfusion. In untreated animals the activities of cytochrome P-450, NADPH-cytochrome c reductase and microsomal ethanol oxidation were significantly higher in pv cells. Phenobarbital produced a 45% increase of the yielded microsomes related to the hepatocytic protein but did not change their relative distribution. The content of reduced glutathione (GSH) was lower in hepatocytes from the pv area. The GSH content was more than 20% increased after phenobarbital treatment in both subclasses of cells, but the distribution pattern remained unchanged. The higher activity of drug metabolizing enzymes in the pv area of untreated animals may account for the higher cytotoxicity of numerous drugs to the perivenous hepatocytes. A 3-day treatment with phenobarbital equalized the pp-pv difference by producing more induction of the periportal cytochrome P-450-mediated drug and ethanol oxidation capacities in microsomes derived from periportally enriched hepatocytes.
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PMID:The distribution of cytochrome P-450-mediated drug oxidation and glutathione in periportal and perivenous rat hepatocytes after phenobarbital treatment. 308 68

Both 4,4'-dichlorobiphenyl (4,4'-DCB) and 3-methylcholanthrene (3-MC) caused a substantial increase of phenacetin-induced hepatic glutathione (GSH) depletion, whereas phenobarbital (PB) had no effect, suggesting that 4,4'-DCB possesses cytochrome P-448 inducing activity. The O-deethylation of phenacetin by liver microsomes from control and PB- and 4,4'-DCB-treated rats showed biphasic Michaelis-Menten kinetics, in contrast to the monophasic course after pretreatment with 3-MC. Hepatic phenacetin levels indicated that in vivo interaction with only a high affinity site is involved in the O-deethylation of phenacetin. 4,4'-DCB and 3-MC caused marked increases in intrinsic clearance and extraction ratio of phenacetin, whereas control values were obtained after PB-treatment. Because of an absence of a spectral change at low phenacetin concentrations, it could not be demonstrated whether the observed differences in metabolism should be ascribed to a change in binding of phenacetin to cytochrome P-450. The results of this study indicate that after pretreatment with various enzyme inducers the phenacetin-induced hepatic GSH depletion strongly correlates with microsomal phenacetin O-deethylation. Further, these findings suggest a discrepancy between 4,4'-DCB and PB in cytochrome P-450 inducing activity, as 4,4'-DCB mimics 3-MC in the induction of phenacetin O-deethylase. The difference between 4,4'-DCB and PB is discussed in relation to the multiplicity and induction of cytochrome P-450 isoenzymes.
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PMID:The 3-methylcholanthrene-mimetic effect of 4,4'-dichlorobiphenyl-treatment on phenacetin-induced hepatic glutathione depletion and liver microsomal phenacetin O-deethylation in rats. 309 30

The ability of hepatic microsomes from senescent rats to metabolize the two potent hepatocarcinogens dimethylnitrosamine (DMN) and aflatoxin B1 (AFB1) was investigated. Seven and 24-month-old male Sprague-Dawley rats were used. Liver weights, and microsomal protein per gram tissue weight were higher, whereas cytochrome P-450 and cytochrome b5 were significantly lower in older rats. Glutathione S-transferases and NADPH cytochrome c reductase activities were dramatically reduced in senescent rats. There was no difference in the formation of formaldehyde from DMN in vitro (31 vs. 34 pmol/nmol P-450) between the young and old rats. In contrast, increased microsome mediated binding of AFB1 to DNA was observed in older rats (116 vs. 228 pmol/nmol P-450) suggesting the possibility of either quantitative or qualitative changes in P-450 species. Additionally the cytoplasmic GSH S-transferases from older rats affected lower inhibition of binding of AFB1 to DNA. These results indicated differential abilities in the hepatic microsomal metabolism of these two carcinogens which may cause differential effects of these carcinogens in senescent rats.
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PMID:Differential effects on the metabolism of dimethylnitrosamine and aflatoxin B1 by hepatic microsomes from senescent rats. 310 18

The stabilities of several drug oxidation and conjugation pathways in human adult hepatocytes have been investigated during 72 hr culture. Cytochrome P-450-dependent mixed function oxidase was measured by the O-dealkylations of ethoxyresorufin (EROD), pentoxyresorufin (PROD) and benzyloxyresorufin (BROD), which are probes for different isozymes of cytochrome P-450 in the rat. EROD declined to 64% of initial fresh cell values after 72 hr in culture, whereas PROD increased to 162% and BROD remained relatively constant. Addition of phenobarbitone to the culture medium selectively increased PROD to a greater extent than EROD and did not affect BROD. NADPH-cytochrome c reductase and NADH-cytochrome b5 reductase were markedly labile during culture, declining to 32% and 22% of fresh cell values respectively. Epoxide hydrolase (EH) showed a large transient increase (2-5-fold) in enzyme activity 24 hr after culture, declining to fresh cell values by 48 hr. UDP-glucuronyltransferase (GT) activity towards phenolphthalein and 1-naphthol also increased (2-3-fold) during the 72 hr of culture, the greater and more rapid increase being observed with phenolphthalein glucuronidation. Sulphotransferase activity declined rapidly within 24 hr of culture, whereas reduced glutathione (GSH) levels and GSH conjugation were maintained at fresh cell values for 72 hr.
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PMID:Human adult hepatocytes in primary monolayer culture. Maintenance of mixed function oxidase and conjugation pathways of drug metabolism. 311 81

The peroxidase-H2O2 catalyzed oxidation of certain drugs in the presence of GSH resulted in extensive oxidation to thiyl radicals and GSSG. NADH or arachidonate in place of GSH was also readily oxidized. Extensive oxygen uptake ensued resulting in the formation of superoxide radicals and H2O2. Only catalytic amounts of drugs and low peroxide levels were required, indicating a radox cycling mechanism. Active drugs included morphine, phenothiazines, aminopyrine, p-phenetidine, acetaminophen and 4-N,N-(CH3)2-aminophenol. Other drugs, including dopamine and methyl-alpha-dopa, did not catalyze oxygen uptake, nor was GSH oxidized to GSSG. Instead, GSH was depleted by GSH conjugate formation. Drugs of the former group, e.g. acetaminophen, aminopyrine or N,N-(CH3)2-aniline, have also been found by other investigators to form GSSG and hydrogen peroxide when added to hepatocytes or when perfused through an isolated liver. Although cytochrome P-450 normally catalyzes a two-electron oxidation of drugs, serious consideration should be given to some one-electron oxidation occurring as well and resulting in radical formation, oxygen activation and GSSG formation.
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PMID:Oxygen activation during drug metabolism. 311 75

Hepatic 1,2-dibromoethane (DBE) metabolism proceeds via two pathways: oxidation by cytochrome P-450 and direct conjugation with the ubiquitous tripeptide glutathione (GSH) via the GSH S-transferases. The toxicity of DBE in monolayers of hepatocytes was assessed to establish whether the toxicity of this compound is increased under conditions of reductive metabolism at low oxygen concentrations. Our previous studies with t-butyl hydroperoxide and the calcium ionophore A23187 suggested that hypoxia would exacerbate toxicity that was mediated through lipid peroxidation or loss of calcium homeostasis. Monolayers of hepatocytes were exposed for 2 hr to 0, 14, 140, 1400, or 14,000 ppm of DBE in an atmosphere of either 1, 2, or 20% oxygen. Toxicity was measured by leakage of aspartate aminotransferase (AST) and trypan blue exclusion. The time course of the development of cytotoxicity was examined by assaying cell death both immediately following a 2-hr exposure and 24 hr later. The LC50 of DBE vapor was found to be approximately 14,000 ppm when assayed immediately after exposure but only 140 ppm when assayed 24 hr after exposure. The similarity of the percentages of DBE-induced cell death after incubations at 1, 2, and 20% oxygen demonstrates that the toxicity of DBE is oxygen-independent. We conclude that while DBE is highly toxic to rat hepatocytes, hypoxia does not appear to contribute to the toxicity of DBE, even under conditions of low oxygen concentrations. This result is in direct contrast to a previous report where we showed that the toxicity of halothane is potentiated under hypoxic conditions.
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PMID:Toxicity of 1,2-dibromoethane in primary hepatocyte monolayer cultures: lack of dependence on oxygen concentration. 313 87

Objectives of this study were to compare the effects of sex, nutritional status and L-2-oxothiazolidine carboxylate (OTC) treatment on tissue constituents frequently involved in responses to chemical toxins. Four groups of adult CD-1 mice were studied: fed females, fed males, fasted males, and fasted males three hours after treatment with OTC (10 mmoles/kg, sc). Female fed mice were found to differ from male fed mice as follows: lower tissue GSH in liver and kidney but not lung; lower hepatic microsomal cytochrome P-450 content and cytosolic GSH transferase activities, particularly using CDNB as substrate; and higher hepatic GSH peroxidase but similar GSSG reductase activities. Overnight fasting was associated with a decrease in hepatic and renal GSH and hepatic cytochrome P-450. OTC treatment was only found to increase hepatic GSH and decrease renal GSH. Thus in fasted CD-1 male mice, the intracellular cysteine precursor, OTC, has an apparently selective effect on tissue GSH contents without confounding effects on hepatic GSH utilizing or restoring activities.
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PMID:Effects of a cysteine precursor, L-2-oxothiazolidine-carboxylate, nutritional status, and sex on tissue glutathione and hepatic GSH-utilizing enzymes of CD-1 mice. 317 43


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