Oxidative damage of rat liver mitochondria during exposure to t-butyl hydroperoxide. Role of Ca2 + ions in oxidative processes

Abstract

AimsThe present study was designed for further evaluation of the biochemical mechanism of hepatic mitochondrial dysfunction under oxidative damages induced by organic hydroperoxide, tert-butyl hydroperoxide (tBHP), for estimation of the molecular targets impaired during oxidative stress, and for investigation of the role of Ca2+ ions in mitochondrial oxidative reactions and of the protective effect of melatonin during mitochondrial peroxidative damage. Main methodsMitochondria were isolated by differential centrifugation from the rat liver. The effects of tBHP exposure, EDTA, Ca2+ ions and melatonin on mitochondrial respiratory activity, mitochondrial enzyme activities and redox status were measured. Key findingsThe present study provides evidence that tBHP (at low concentrations of 0.02–0.065mM, in EDTA-free medium) induced uncoupling of the oxidation and phosphorylation processes and decreased the efficiency of the phosphorylation reaction. This effect depended on the respiratory substrate used. The presence of EDTA prevented oxidative impairment of mitochondrial respiration, but Ca2+ ions in the medium enhanced oxidant-induced mitochondrial damage considerably. In the presence of 0.5mM EDTA, tBHP (at high concentrations, 0.5–2mM) considerably oxidized mitochondrial reduced glutathione, enhanced accumulation of membrane lipid peroxidation products and mixed protein–glutathione disulfides and led to an inhibition of oxoglutarate dehydrogenase and succinate dehydrogenase. SignificanceDirect oxidative modification of enzymatic complexes of the respiratory chain and mitochondrial matrix, mitochondrial reduced glutathione depletion, protein glutathionylation, membrane lipid peroxidation and Ca2+ overload are the main events of mitochondrial peroxidative damages. Experiments in vitro demonstrated that melatonin inhibited the mitochondrial peroxidative damage, preventing redox-balance changes and succinate dehydrogenase inactivation.