Evaluation of mitochondrial function in isolated rat hepatocytes and mitochondria during oxidative stress

Abstract

The aim of our study was to evaluate mitochondrial changes in isolated rat hepatocytes and isolated liver mitochondria exposed in vitro to a potent non-specific peroxidating agent tert-butylhydroperoxide(tBHP).Decrease in mitochondrial membrane potential(MMP) in tBHP treated hepatocytes in culture was documented by significant reduction of Rhodamine 123 accumulation and by fluorescence microscopy using JC-1 as a fluorescence probe.Dramatic impact of oxidative stress induced by tBHP on mitochondria confirms also substantial decrease of MMP measured using tetraphenylphosphonium-selective electrode.Swelling of mitochondria,which was prevented by Cyclosporine A evidences the role of mitochondrial permeability transition:The high sensitivity of respiratory Complex I could be caused by oxidation on NADH and NADPH by tBHP with following disruption of mitochondrial calcium homeostasis.Increased mitochondrial calcium concentration stimulated formation of reactive oxygen species in mitochondria.