MODULATING ROLE OF ENDOGENOUS REDUCED GLUTATHIONE IN TERT-BUTYL HYDROPEROXIDE-INDUCED CELL INJURY IN ISOLATED RAT HEPATOCYTES

The role of endogenous reduced glutathione (GSH) in tert-butyl hydrope roxide (TBHP)-induced cell injury was examined in isolated rat hepatoc ytes. When liver cell injury was estimated from release of transaminas es from hepatocytes into the incubation medium, cell Injury in hepatoc ytes (2 x 10(6) cells/ml) incubated in Hanks' balanced salt solution ( pH 7.2) containing 1.0 mM TBHP at 37 degrees C was potentiated with en hanced lipid peroxidation by prior depletion of intracellular GSH whic h was induced by diethylmaleate, a GSH depletor. GSH-depleted hepatocy tes were incubated with gamma-glutamylcysteinylethyl ester (gamma-ECOE t), which is known to be converted to GSH via glutathione synthetase a fter its hydrolysis by esterase, at concentrations of 1.0 to 10 mM in order to replenish intracellular GSH. Although TBHP-induced cell injur y and lipid peroxidation were enhanced in GSH-depleted hepatocytes, th ese enhancements were prevented with the consumption of intracellular GSH in GSH-depleted hepatocytes pretreated with 5.0 mM gamma-ECOEt. Th ese preventive effects were observed at any time point during the TBHP treatment over a 60 min period and depended on the concentration of g amma-ECOEt used. But, no preventive effect was found in GSH-depleted h epatocytes pretreated with 5.0 mM GSH. No prevention of the potentiati on of TBHP-induced cell injury found in GSH-depleted hepatocytes occur red in GSH-depleted hepatocytes pretreated with both 5.0 mM gamma-ECOE t and 250 mu M bis-(p-nitrophenyl) phosphate, a nonspecific esterase i nhibitor. gamma-ECOEt treatment caused an increase in intracellular GS H content in GSH-depleted hepatocytes, while treatments of both gamma- ECOEt and the esterase inhibitor caused no increase in intracellular G SH content in the cells. These results indicate that endogenous GSH mo dulates TBHP-induced cell injury and lipid peroxidation in isolated ra t hepatocytes. The present results suggest that endogenous GSH should play a critical role in TBHP-induced cell injury in isolated rat hepat ocytes and that in rat hepatocytes treated with TBHP, enhanced lipid p eroxidation with the consumption of intracellular GSH could be associa ted with the initiation of cell injury. (C) 1997 Elsevier Science Inc.