Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “GLUTATHIONE”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,405 records · Page 78Linked to original sources
PubMed1992

p-aminophenol nephrotoxicity: biosynthesis of toxic glutathione conjugates.

p-Aminophenol causes necrosis of the pars recta of the proximal tubules in rats, and its nephrotoxicity may be due to glutathione-dependent bioactivation reactions. We have investigated the hepatic metabolism of p-aminophenol in Wistar rats and the cytotoxicity of formed glutathione S-conjugates in rat renal epithelial cells. After ip application of p-aminophenol (100 mg/kg), the following metabolites were identified in rat bile: 4-amino-2-(glutathion-S-yl)phenol, 4-amino-3-(glutathion-S-yl)-phenol, 4-amino-2,5-bis(glutathion-S-yl)phenol, 4-amino-2,3,5(or 6)-tris(glutathion-S-yl)phenol, an aminophenol conjugate (likely a sulfate or glucuronide), acetaminophen glucuronide, and 3-(glutathion-S-yl)acetaminophen. 4-Amino-3-(glutathion-S-yl)phenol, 4-amino-2,5-bis(glutathion-S-yl)phenol, and 4-amino-2,3,5(or 6)-tris(glutathion-S-yl)phenol induced a dose- and time-dependent loss of cell viability in rat kidney cortical cells. Cell killing was significantly reduced by inhibition of gamma-glutamyl transpeptidase with Acivicin. p-Aminophenol was also toxic to renal epithelial cells. Coincubation of p-aminophenol with tetraethylammonium bromide, a competitive inhibitor of the organic cation transporter, and with SKF-525A, an inhibitor of cytochrome P450, protected cells from p-aminophenol-induced toxicity. p-Aminophenol would thus be accumulated in the kidney mainly by organic cation transport systems, which are concentrated in the S-1 segment of the proximal tubule. However, p-aminophenol toxicity in vivo is directed toward the S-2 and S-3 segments, which are rich in gamma-glutamyl transpeptidase. These results and the observation that biliary cannulation and glutathione depletion reduce p-aminophenol nephrotoxicity suggest that the biosynthesis of toxic glutathione conjugates is responsible for p-aminophenol nephrotoxicity in vivo. The aminophenol glutathione S-conjugates formed induce p-aminophenol nephrotoxicity by a pathway dependent on gamma-glutamyl transpeptidase.

Aminophenols↗
PubMed1994

Reduced expression of glutathione S-transferase Yb2 during progression of chemically induced hepatocellular carcinomas in Fischer 344 rats.

We followed the expression of several glutathione S-transferase subunits in altered foci, liver neoplasms and metastases produced in male Fischer 344 rats by a modified Solt-Farber protocol, to determine whether components of the resistant phenotype are lost during neoplastic progression. At 6 mo after initiation, altered foci and persistent nodules displayed increased immunohistochemical expression of glutathione S-transferase subunits Yf (pi-class), Ya (alpha-class) and Yb1 (mu-class) in comparison with normal or surrounding liver tissue. However, although most altered foci exhibited little change in glutathione S-transferase Yb2 (mu-class) subunit expression, 5% of Yf-positive foci and nodules were partially or completely deficient in Yb2 expression. At 12 and 18 mo after initiation, most grossly visible hepatocellular tumors retained induced expression of glutathione S-transferase subunits Yf, Ya and Yb1, but 63% of the carcinomas, 88% of the primary metastatic carcinomas and 94% of the pulmonary metastases were deficient in Yb2 expression. These differences in glutathione S-transferase subunit expression were confirmed by quantitative analysis by reverse-phase HPLC of S-hexylglutathione affinity-purified glutathione S-transferases from advanced tumors. Cytosolic glutathione S-transferase activity for trans-4-phenyl-3-buten-2-one in advanced tumors ranged from 42% to 66% of the activity in matched surrounding liver, whereas glutathione S-transferase activities for 1-chloro-2,4-dinitrobenzene were increased by 140% to 161%. These studies demonstrate that progression of hepatocellular carcinomas in the resistant hepatocyte model of carcinogenesis in which several glutathione S-transferase subunits are induced is associated with the loss of a major constitutive mu-class hepatic glutathione S-transferase. Although the mechanism and role of the reduction or loss of glutathione S-transferase Yb2 during malignant progression are unknown, we propose that loss of glutathione S-transferase Yb2 in some preneoplastic populations of hepatocytes might be conducive to further DNA damage by presently unknown environmental or endogenous compounds that are normally detoxified preferentially by glutathione S-transferase isoenzymes containing this subunit.

Animals↗
PubMed2004

Acivicin-induced alterations in renal and hepatic glutathione concentrations and in gamma-glutamyltransferase activities.

gamma-Glutamyltransferase (gamma-GT) catalyzes the hydrolysis of glutathione, glutathione S-conjugates, and gamma-substituted l-glutamate derivatives. Acivicin is an irreversible inhibitor of gamma-GT that has been used to study the role of gamma-GT in glutathione homeostasis and glutathione-dependent bioactivation reactions. The present studies were undertaken because of reported conflicting effects of acivicin on the nephrotoxicity of some haloalkenes that undergo glutathione-dependent bioactivation. The objective of this study was to test the hypothesis that acivicin may alter renal glutathione concentrations; acivicin-induced changes in renal glutathione concentrations may alter the susceptibility of the kidney to the nephrotoxic effects of haloalkenes. Hence, diurnal and acivicin-induced changes in renal and hepatic glutathione concentrations along with renal and hepatic gamma-GT activities were investigated. The previously observed diurnal variations in hepatic glutathione concentrations in fed rats were confirmed, but no diurnal variations were observed in renal glutathione concentrations or in renal or hepatic gamma-GT activities. Renal and hepatic glutathione concentrations and gamma-GT activities were measured in tissue homogenates from rats given 0, 0.1, or 0.2 mmol acivicin/kg (i.p.) and killed 0, 2, 4, 8, 12, or 24 hr later. Renal glutathione concentrations were increased above control values in acivicin-treated rats, whereas acivicin had no effect on hepatic glutathione concentrations. Renal gamma-GT activities decreased within 2 hr after giving acivicin and remained decreased for 24 hr. Acivicin had no effect on hepatic gamma-GT activities, except at 24 hr after treatment when values in acivicin-treated rats were elevated compared with controls. Although the present studies do not afford an explanation of the mechanism whereby acivicin increases the nephrotoxicity of some haloalkenes, they do indicate that acivicin is not a reliable probe to investigate the role of gamma-GT in haloalkene-induced nephrotoxicity.

Animals↗
PubMed1992

Glutathione S-transferases in human testicular germ cell tumors: changes of expression and activity.

Glutathione S-transferases are involved in the detoxification of carcinogens and xenobiotics and are potentially associated with the development of drug-resistance. Forty-six testicular germ cell tumors and 33 adjacent normal testicular tissue specimens were analyzed at the RNA level for the expression of glutathione S-transferase alpha and pi. Glutathione S-transferase alpha was expressed in 31 of the 33 normal testicular tissues (94%) but in only three of the 46 germ cell tumors (7%). Glutathione S-transferase pi mRNA was detected in all normal and malignant testicular tissue samples. Thirteen testicular germ cell tumors and eight normal testicular tissue samples were analyzed at the protein level. The mean specific activity of total cytosolic glutathione S-transferase in tumor tissue was decreased by about 80% as compared to normal testicular tissue. Protein analysis of the glutathione S-transferase subunits of normal testicular tissue demonstrated the presence of the glutathione S-transferase classes alpha, mu and pi, with a predominance of the mu class. In testicular germ cell tumors the glutathione S-transferase subunit pattern showed a predominance of glutathione S-transferase pi representing 88% +/- 3% of total glutathione S-transferase. Since all three glutathione S-transferase isoenzyme classes contribute to the resistance to antineoplastic drugs, the altered glutathione S-transferase isoenzyme pattern and the decrease of glutathione S-transferase activity may play a role in the high inherent drug sensitivity of human testicular germ cell tumors.

Glutathione Transferase↗
PubMed2000

Glutathione-related factors are not correlated with sensitivity of human tumour cells to actinomycin D.

Glutathione (GSH) contents and activities of glutathione S-transferases (GST), glutathione reductase (GSH-RD), glutathione peroxidase (GSHpx) and glutathione conjugate export pump (GS-X pump) were determined in eight human tumour cell lines with different sensitivities to melphalan, a substrate of glutathione conjugation, and actinomycin D which has not been shown to be detoxified by glutathione-related mechanisms. Chang liver cells with highest GSH content and highest activities of GST, GSH-RD, GSHpx and GS-X pump were found to be most resistant to melphalan. Statistical analysis showed significant correlations between sensitivities of the human tumour cells to melphalan and the glutathione-related factors (r = 0.72-0.79; except for GST, r = 0.65, P = 0.08), while there were no significant correlations observed between sensitivities of the human tumour cells to actinomycin D and all the glutathione-related factors tested (r = -0.25-0.14). Significant correlations of the glutathione-related factors to resistance of human tumour cells to melphalan, a substrate of glutathione conjugation, but not to resistance of the human tumour cells to actinomycin D which has not been shown to be detoxified by glutathione-related mechanisms suggested that glutathione-related mechanisms contribute to drug resistance by increased detoxification of the drugs involved.

Antibiotics, Antineoplastic↗
PubMed1998

Glutathione homeostasis is disturbed in CD4-positive lymphocytes of HIV-seropositive individuals.

BACKGROUND: Numerous in vitro tests have suggested that a disturbed cellular glutathione homeostasis plays an important role in the pathogenesis of human immunodeficiency virus (HIV) infection. METHODS: Using validated high-performance liquid chromatography (HPLC) methods, glutathione concentrations were determined in plasma and in cytosol of CD4+ lymphocytes and CD14+ cells of HIV-seropositive individuals and healthy control subjects. We measured concentrations of total glutathione, which is the sum of reduced (GSH) and oxidised (GSSG) glutathione and mixed disulphides of which there are two fractions: soluble mixed disulphides (GSSR) and protein bound glutathione (ProSSG). Also, non-protein-bound glutathione was measured, which is the sum of GSH, GSSG and GSSR. Thirty-five healthy control subjects and 35 HIV-infected individuals participated in the study. RESULTS: We found that in CD4+ lymphocytes from HIV-seropositive individuals, total glutathione levels were significantly higher than in healthy control subjects, whereas the fraction of non-protein-bound glutathione was not different. This can only be explained by an increase in the protein-bound fraction of glutathione indicating the presence of oxidative stress in CD4+ lymphocytes of HIV-seropositive individuals. Glutathione measurements of cytosol of CD14+ cells and plasma were, however, not compatible with significant increased oxidation. Glutathione precursors (cysteine, cysteinylglycine, glutamylcysteine and homocysteine) and products of lipid peroxidation (thiobarbituric acid-reactive substances) were also measured in plasma and did not differ between healthy control subjects and HIV-seropositive individuals. CONCLUSION: We conclude that the glutathione homeostasis is disturbed in CD4+ lymphocytes of HIV-seropositive individuals. The glutathione redox dysbalance in CD4+ lymphocytes could be important in the pathogenesis of HIV infection and have implications for therapy.

Adult↗
PubMed1996

gamma-Glutamyl transpeptidase mediation of tumor glutathione utilization in vivo.

BACKGROUND: Glutathione is a tripeptide used by cells to protect against oxidative and free radical damage. It may also be involved in biochemical mechanisms that cause some tumors to become resistant to anticancer drugs. gamma-Glutamyl transpeptidase (GGTP) is a membrane-bound enzyme that cleaves extracellular glutathione, providing cells with amino acids necessary for intracellular synthesis of this compound. Increased expression of GGTP has been found in a number of human tumors; however, few studies have examined the contribution of GGTP to tumor glutathione metabolism in vivo. PURPOSE: Our goals were to study the utilization of host glutathione by 3-methylcholanthrene (MCA)-induced sarcomas grown in rats and to evaluate the involvement of tumor GGTP in this process. METHODS: The left ovaries of 21 female Fischer 344 rats were isolated by laparotomy and placed in subcutaneous positions through stab wounds in the abdominal wall. A 3-mm cube of MCA sarcoma was then sutured to each of the isolated ovaries. The MCA implants obliterated the ovarian tissue, yielding isolated tumors with one arterial supply (the ovarian artery) and one draining vein (the ovarian vein, referred to as the tumor vein). After 2 weeks of tumor growth, blood was drawn from the tumor vein, the inferior vena cava (IVC), and the aorta of 16 animals. Glutathione and cysteine concentrations in plasma samples from this blood were determined by high-performance liquid chromatography and used to calculate glutathione and cysteine utilization ratios for the tumor and the systemic circulations ([(concentration aorta-concentration tumor vein)/concentration aorta] x 100 and [(concentration aorta-concentration IVC)/concentration aorta ] x 100, respectively). The utilization ratios from these control animals were compared with those from acivicin (AT-125; an irreversible GGTP inhibitor)-treated rats (the remaining five animals). Data are presented as mean +/- standard deviation; reported P values are from two-tailed tests of statistical significance. RESULTS: In the control animals, glutathione and cysteine concentrations were significantly lower in the tumor vein (3.55 +/- 1.9 and 5.69 +/- 2.8 microM, respectively) and in the IVC (5.65 +/- 2.3 and 7.17 +/- 2.4 microM, respectively) than in the artery (12.48 +/- 5.7 and 12.33 +/- 5.9 microM, respectively; all P values < .05). In addition, the glutathione utilization ratio was significantly higher for the tumor circulation than for the systemic circulation (69% +/- 14% versus 52% +/- 14%; P < .003). The combined glutathione and cysteine utilization ratio was also significantly higher for the tumor circulation than for the systemic circulation (116% +/- 35% versus 88% +/- 28%; P < .02). Treatment with AT-125 lowered the tumor glutathione utilization ratio significantly (45% +/- 12% for treated animals versus 69% +/- 14% for control animals; P < .005). CONCLUSIONS: Our results show that glutathione and cysteine in the host circulation are used by MCA sarcomas. The significant reduction in tumor utilization of serum glutathione after treatment with AT-125, a GGTP inhibitor, indicates that GGTP is important in tumor glutathione metabolism.

Animals↗
PubMed2004

The glutathione system as a stress marker in plant ecophysiology: is a stress-response concept valid?

Environmental stress impacts cause an increased formation of reactive oxygen species (ROS) in the chloroplasts (photo-oxidative stress). The role of glutathione in the antioxidative defence system provides a rationale for its use as a stress marker. However, responses of glutathione concentrations and redox states are not consistent among the large number of available publications. In the present review the hypothesis that stress responses of the glutathione system follow a general ecophysiological stress-response concept is investigated. In this view, an initial response phase would be followed by an acclimation phase where a new steady-state is established. Alternatively, if successful acclimation is not achieved, degradation of the system will follow. Recent publications dealing with responses to photochilling, salinity, and drought are analysed as to whether the results fit the concept. In general, an initial stress response was related to changes in the glutathione redox state, whereas acclimation was marked by increased glutathione concentrations, increased related enzyme activities, and/or a more reduced redox state of glutathione. The latter was interpreted as overcompensation leading to enhanced regeneration of glutathione. Deterioration effects upon strong stress impacts were related to progressive degradation and oxidation of the glutathione pool. A time-course analysis, which has rarely been done in the published literature, showed this sequence of events. When apple trees were subjected to progressing drought, the initial response was a slight oxidation of the glutathione pool, followed by increased glutathione concentrations. When the stress increased, glutathione concentrations dropped and redox state became more oxidized, which marked the degradation of the system. In spite of the general congruency of these results with the suggested stress-response concept, several limitations have to be highlighted: The importance of the glutathione system relative to other components of the photoprotective and antioxidative defence system, as well as relative to stress avoidance strategies, has to be established. It is suggested that a variety of parameters taking into account alternative protection pathways (e.g. photorespiration, light dissipation) and other components of the antioxidative systems should be measured. Within such response patterns the glutathione system is a valuable stress marker in ecophysiological studies.

Acclimatization↗
PubMed1993

Effect of glutathione on endothelial prostacyclin synthesis after anoxia.

We previously observed decreased prostacyclin (PGI2) formation after reoxygenation of anoxic endothelium. In the present study, the effects of glutathione on endothelial prostaglandin (PG) H synthase activity after reoxygenation were explored. Intracellular glutathione content decreased 70% after 24 h of anoxia; reoxygenation did not produce any additional decrease in glutathione content. Intracellular glutathione was maintained in the reduced state by the endothelium even during the oxidant stress caused by reoxygenation or the addition of peroxide. Glutathione depletion produced by DL-buthionine-(S,R)-sulfoximine (BSO), 1,3-bis(chloroethyl)1-nitrosourea (BCNU), or incubation in a sulfhydryl-free medium resulted in increased sensitivity of PGH synthase to the effects of added H2O2. However, glutathione depletion resulting from BSO or culture in sulfhydryl-free medium during anoxia did not increase the sensitivity of PGH synthase to reoxygenation. In addition, anoxia did not make the endothelium more sensitive to H2O2. Glutathione peroxidase and glutathione reductase activities were preserved after anoxia-reoxygenation. When glutathione reductase was inhibited with BCNU during reoxygenation, PGI2 release was decreased further. These findings demonstrate that, although anoxia decreases endothelial glutathione content, the endothelium is able to utilize its remaining glutathione to protect against additional oxidant stress because glutathione peroxidase and glutathione reductase retain their activity.

Arachidonic Acid↗
PubMed1990

Evidence for carrier-mediated transport of glutathione across the blood-brain barrier in the rat.

Information on the origin of brain glutathione and the possibility of its transport from blood to brain is limited. We found a substantial uptake of 35S-labeled glutathione by the rat brain using the carotid artery injection technique. The brain uptake index of glutathione with and without an irreversible gamma-glutamyl transpeptidase inhibitor, acivicin, was similar. No significant differences in the regional uptake of labeled glutathione were found in rats pretreated with acivicin. The brain uptake index of tracer glutathione was similar to that of cysteine tracer and was lower than that of phenylalanine. The transport of oxidized glutathione (glutathione disfulfide) across the blood-brain barrier was not significantly different from that of sucrose, an impermeable marker. Brain radioactivity 15 s after carotid artery injection of labeled glutathione to rats pretreated with acivicin was predominantly in the form of glutathione. The in vivo glutathione uptake was saturable with an apparent Km of 5.84 mM. Amino acids, amino acid analogues, and other compounds [cysteine, phenylalanine, glutathione disulfide, gamma-glutamylglutamate, gamma-glutamyl p-nitroanilide, 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid (BCH)] did not affect glutathione transport. Our data suggest that glutathione is transported across the blood-brain barrier by a saturable and specific mechanism.

Animals↗
PubMed1984

Characterization of glutathione peroxidase in frog retina.

The properties and kinetic parameters of glutathione peroxidase were measured in retinal homogenates from frogs (R. pipiens) using a spectrophotometric assay in which the oxidation of glutathione is coupled to the oxidation of NADPH+ by exogenous glutathione reductase. The standard assay utilized 5.0 mM glutathione and 0.6 mM cumene hydroperoxide in 50 mM sodium phosphate buffer at pH 7.0. All solutions were bubbled with argon and all reactions were carried out under an atmosphere of argon. The enzyme activity was linear with protein concentration at different concentrations of both substrates. Determination of the pH optimum was complicated by a large increase in non-enzymatic oxidation of glutathione at alkaline pH. The highest ratio of enzymatic to non-enzymatic activity was at pH 7.0. Increasing glutathione concentration showed less effect on the spontaneous reaction than increasing the cumene hydroperoxide concentration. Glutathione peroxidase Km value for glutathione was 3.86 mM and for cumene hydroperoxide was 0.55 mM. Vmax for glutathione at 0.6 mM cumene hydroperoxide was 138 nmoles glutathione oxidized/min/mg protein, while at 5.0 mM glutathione the value for cumene hydroperoxide was 146 nmoles glutathione oxidized/min/mg protein. These studies demonstrate that glutathione peroxidase is active in the retina and establish the optimal experimental conditions for determination of the enzymatic activity.

Animals↗
PubMed1991

Inhibition of glutathione synthesis in the liver leads to S-adenosyl-L-methionine synthetase reduction.

The hepatic levels of glutathione in rats treated with buthionine sulfoximine (4 mmol/kg), an inhibitor of glutathione synthesis, were 72.5% +/- 4.9% of those determined in control animals. This decrease in glutathione concentration was prevented by the administration of glutathione monoethyl ester (7.5 mmol/kg). S-Adenosyl-L-methionine-synthetase activity in the liver of rats treated with buthionine sulfoximine was 39.4% +/- 6.5% of that determined in control animals. Again, glutathione monoethyl ester prevented the effect of buthionine sulfoximine on S-adenosyl-L-methionine-synthetase activity. There was a close correlation (r = 0.936) between the hepatic levels of glutathione and S-adenosyl-L-methionine-synthetase activity. The hepatic concentration of S-adenosyl-L-methionine in buthionine sulfoximine-treated animals was 59.7% +/- 3.7% of that measured in control rats. Contrasting with the protective effects mentioned above, glutathione monoester had no preventive action on buthionine sulfoximine-induced S-adenosyl-L-methionine depletion. Electron microscopic examination of liver samples of rats after buthionine sulfoximine administration showed evidence of liver degeneration, which was attenuated by glutathione monoethyl ester treatment. Glutathione (7.5 mmol/kg) treatment was less effective than glutathione monoethyl ester in attenuating buthionine sulfoximine effects on hepatic S-adenosyl-L-methionine metabolism and morphology. The reduction of S-adenosyl-L-methionine-synthetase activity observed after treatment with buthionine sulfoximine and its prevention by glutathione monoethyl ester, as well as the correlation between the activity of this enzyme and glutathione levels, indicate that glutathione plays an important role in maintaining S-adenosyl-L-methionine-synthetase activity in the liver.

Acetaminophen↗
PubMed1990

Cholestasis, altered junctional permeability, and inverse changes in sinusoidal and biliary glutathione release by vasopressin and epinephrine.

The mechanism for the vasopressin- and epinephrine-induced decrease in bile formation and increase in sinusoidal efflux of glutathione was investigated in rat livers perfused with recirculating fluorocarbon emulsion. Vasopressin and epinephrine transiently decreased bile flow and excretion of endogenous bile acids and glutathione and increased the bile/perfusate ratio of [14C]sucrose, suggesting an increase in junctional permeability, but had no effect on the bile/perfusate ratio of [3H]polyethylene glycol-900. The decreased biliary glutathione was balanced by an increase in sinusoidal efflux, such that total hepatic release remained unchanged. The adrenergic antagonist dihydroergotamine blocked the effects of epinephrine. To examine whether an increase in junctional permeability per se could account for the changes in glutathione efflux, biliary permeability was increased by either bile duct ligation, lowering of perfusate Ca2+ concentration with ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), or addition of taurolithocholate, a cholestatic bile acid. All three maneuvers produced a decrease in biliary glutathione excretion and a concomitant increase in sinusoidal glutathione efflux, whereas total glutathione release was largely unaffected. The effects of EGTA were partially reversed if CaCl2 was reintroduced into the perfusate. Because the GSH/GSSG ratio in perfusate could not be measured in this experimental system due to the spontaneous oxidation of GSH to GSSG, additional experiments in the nonrecirculating mode examined the effects of vasopressin and bile duct ligation on sinusoidal release of GSH and GSSG. In control livers there was no detectable GSSG in perfusate (less than 0.5 nmol.min-1.g-1). After vasopressin administration, the additional sinusoidal glutathione was mainly as GSH, although there was also a significant amount of GSSG (1-2 nmol.min-1.g-1). The additional glutathione released into perfusate after bile duct ligation was 47% as GSSG. When vasopressin was administered to livers whose bile duct had been ligated, its ability to enhance sinusoidal glutathione release was diminished, suggesting that the effects of vasopressin and bile duct ligation are not additive. These observations support previous findings that vasopressin and epinephrine can modulate hepatocyte tight junctional permeability and demonstrate that these hormones produce cholestasis and inverse changes in sinusoidal and biliary glutathione efflux. Other maneuvers that increased biliary permeability to [14C]sucrose also produced cholestasis and a redistribution of glutathione efflux from bile to perfusate, suggesting that an increase in junctional permeability may allow biliary glutathione to reflux from bile to plasma.

Animals↗
PubMed1986

Differential sensitization of human ovarian carcinoma and mouse L1210 cells to cisplatin and melphalan by glutathione depletion.

We have investigated the role of glutathione in determining the macromolecular binding and cytotoxicity of cisplatin (DDP) and melphalan (LPAM) in human ovarian carcinoma cells and DDP-resistant L1210 mouse leukemia cells. Glutathione reacted avidly with DDP in normal saline with a bimolecular rate constant of 16.2 M-1 hr-1. Glutathione had no effect on the rate of hydrolysis of LPAM, consistent with the SN1-like reaction mechanism of LPAM. Glutathione protected calf thymus DNA and bovine serum albumin from DDP platination and LPAM alkylation. Glutathione also protected nuclei isolated from human ovarian carcinoma cells from DDP platination. The importance of intracellular glutathione in determining the cytotoxicity of DDP and LPAM was assessed by depletion of glutathione with buthionine sulfoximine in three cell types. Exposure to 0.5 mM buthionine sulfoximine for 20-28 hr depleted glutathione to levels that were 10-20% of control levels. COLO 316 and 2008 human ovarian carcinoma cells, and ZCR9 mouse leukemia cells were all sensitized to LPAM cytotoxicity by this level of glutathione depletion. The dose modification factors, defined as the IC50 control cells/IC50 depleted cells, were: 2.6 +/- 0.5 for COLO 316 cells, 1.6 +/- 0.1 for 2008 cells, and 2.1 +/- 1.1 for ZCR9 cells. In contrast, glutathione depletion had a minimal effect on DDP cytotoxicity in these cells with dose modification factors of: 1.2 +/- 0.2 for COLO 316 cells, 0.8 +/- 0.3 for 2008 cells, and 1.1 +/- 0.1 for ZCR9 cells. The differential potentiation of DDP and LPAM cytotoxicity by glutathione depletion in these cells, despite the similar protection that glutathione affords macromolecules from drug binding, suggests that there are fundamental differences in the intracellular interaction of these electrophilic drugs with glutathione.

Animals↗
PubMed1980

Effect of salicylates and phenobarbital on hepatic glutathione in the rat.

Phenobarbital and salicylates were shown to have opposite effects on hepatic glutathione. Phenobarbital increased glutathione concentration by approximately 20 to 30%. This increase occurred within 48 hr and could be attributed almost exclusively to an increase in bound glutathione. No changes in ATP, substrate amino acids for glutathione synthesis or the level of gamma-glutamylcysteine synthetase, the rate limiting enzymatic step in glutathione synthesis, were found with phenobarbital. Phenobarbital, which induces hepatic proteins that bind glutathione, increased bound glutathione but did not affect unbound glutathione. Therefore, the concentration of the latter probably regulates glutathione synthesis. Salicylates (aspirin and sodium salicylate) were found to deplete hepatic glutathione in both saline- and phenobarbital-treated rats. Maximum depletion (approximately 40%) was seen 4 to 6 hr after salicylate administration and returned toward the control level by 12 hr. The salicylate effect was not related to a change in gamma-glutamylcysteine synthetase, gamma-glutamyl transpeptidase or the concentrations of free hepatic glycine, glutamate, cysteine and methionine. An increase in concentration of glutathione both in vivo in plasma from salicylate-treated rats and in vitro in buffer from the incubation of liver slices with salicylate suggests that glutathione leakage from hepatocytes is an important factor in salicylate-induced hepatic glutathione depletion.

Amino Acids↗
PubMed1985

The responses of glutathione and antioxidant enzymes to hyperoxia in developing lung.

Total glutathione levels and the activity of enzymes associated with antioxidant protection in neonatal lung are increased in response to hyperoxia. Glutathione levels in developing rat lung decreased from 24 nmol/mg protein on day 19 of gestation to approximately 12 nmol/mg protein at birth. The initial decrease in glutathione may be due to emergence of other antioxidant systems. Newborn rats placed in 100% oxygen showed a rapid and sustained increase in total glutathione levels which was primarily due to an increase in reduced glutathione. Explants obtained from 16-wk gestation human fetal lung or from 17- to 18-day fetal rat lung also showed increased total and reduced glutathione when cultured in 95% oxygen, 5% CO2 as compared with explants cultured in room air. Type II cells isolated from neonatal rats maintained in oxygen for 6 days also showed glutathione levels twice those found in cells isolated from animals in room air. The activity of antioxidant enzymes (glucose-6-phosphate dehydrogenase, glutathione peroxidase, glutathione reductase) was increased in lungs of newborn rats exposed to 100% oxygen either at birth or 2 days of age. Antioxidant enzyme activity of lung explants cultured in 95% oxygen, 5% CO2 was also higher than in explants maintained in room air. These results suggest that the increases in glutathione and of antioxidant enzymes in vivo and in vitro are a direct effect of oxygen exposure in lung and that the increase of both glutathione and antioxidant enzyme activity is intrinsic to the lung cell itself. It is likely that increases in glutathione in lung represent an important protective mechanism against oxidant injury.

Animals↗
PubMed2001

Proposed reductive metabolism of artemisinin by glutathione transferases in vitro.

Artemisinin is a sesquiterpene lactone containing an endoperoxide bridge. It is a promising new antimalarial and is particularly useful against the drug resistant strains of Plasmodium falciparum. It has unique antimalarial properties since it acts through the generation of free radicals that alkylate parasite proteins. Since the antimalarial action of the drug is antagonised by glutathione and ascorbate and has unusual pharmacokinetic properties in humans, we have investigated if the drug is broken down by a typical reductive reaction in the presence of glutathione transferases. Cytosolic glutathione transferases (GSTs) detoxify electrophilic xenobiotics by catalysing the formation of glutathione (GSH) conjugates and exhibit glutathione peroxidase activity towards hydroperoxides. Artemisinin was incubated with glutathione, NADPH and glutathione reductase and GSTs in a coupled assay system analogous to the standard assay scheme with cumene hydroperoxide as a substrate of GSTs. Artemisinin was shown to stimulate NADPH oxidation in cytosols from rat liver, kidney, intestines and in affinity purified preparations of GSTs from rat liver. Using human recombinant GSTs hetelorogously expressed in Escherichia coli, artemisinin was similarly shown to stimulate NADPH oxidation with the highest activity observed with GST M1-1. Using recombinant GSTs the activity of GSTs with artemisinin was at least two fold higher than the reaction with CDNB. Considering these results, it is possible that GSTs may contribute to the metabolism of artemisinin in the presence of NADPH and GSSG-reductase. We propose a model, based on the known reactions of GSTs and sesquiterpenes, in which (1) artemisinin reacts with GSH resulting in oxidised glutathione; (2) the oxidised glutathione is then converted to reduced glutathione via glutathione reductase; and (3) the latter reaction may then result in the depletion of NADPH via GSSG-reductase. The ability of artemisinin to react with GSH in the presence of GST may be responsible for the NADPH utilisation observed in vitro and suggests that cytosolic GSTs are likely to be contributing to metabolism of artemisinin and related drugs in vivo.

Animals↗
PubMed1996

An HPLC radiotracer method for assessing the ability of L-Cysteine prodrugs to maintain glutathione levels in the cultured rat lens

Purpose. To apply a high performance liquid chromatographic radiotracer method to test a variety of L-cysteine prodrugs and one dipeptide prodrug for their ability to synthesize glutathione in cultured rat lenses. Method. Rat lenses were incubated for 48 h in a medium containing [14C(U)]-glycine and prodrugs. Following homogenization and derivatization, lens extracts were analyzed to determine the extent of biosynthetic incorporation of this labeled amino acid into [14C]-glutathione using high performance liquid chromatography with radioisotope and ultraviolet absorption detection. All of the thiazolidine prodrugs contained masked sulfhydryl groups to stabilize them against air oxidation. L-buthionine-(S,R)-sulfoximine - an inhibitor of the first step in glutathione biosyntesis - was present in media containing the dipeptide prodrug. Results. In all cases, a large [14C]-labeled peak eluted just prior to [14C]-glutathione. This peak had some characteristics of the mixed disulfide of glutathione and L-cysteine, viz., L-cysteine/glutathione disulfide, but requires further investigation in order to be positively identified. Of the eleven L-cysteine prodrugs investigated, the most effective was 2(R,S)-methylthiazolidine-4(R)-carboxylic acid, which increased the rate of [14C]-glutathione biosynthesis 35% over that of the controls. A number of other L-cysteine prodrugs were somewhat effective, increasing glutathione synthesis 5-30% over the controls, while several L-cysteine prodrugs were totally ineffective. The only dipeptide prodrug investigated, viz., gamma-L-glutamyl-L-cysteine ethyl ester, increased the biosynthesis of [14C]-glutathione 18% over control. Biosynthetic rates based on ultraviolet absorption of the derivatized glutathione demonstrated a similar pattern, the compounds most effective in synthesizing [14C]-glutathione generally yielding the highest ultraviolet glutathione concentrations and the ineffective compounds showing the lowest concentrations. Conclusions. 2(R,S)-methylthiazolidine-4(R)-carboxylic acid, 2(R,S)-n-propylthiazolidine-4(R)-carboxylic acid and N-acetyl-L-cysteine were the only compounds that were statistically significant in yielding higher levels of both ultraviolet and radioactive glutathione as compared to their respective controls. Thus, these prodrugs have very promising anti-cataract potential. Keywords: glutathione; HPLC; lens; rat; thiazolidine

Journal Article↗