Effect of extracellular Ca++ omission on isolated hepatocytes. II. Loss of mitochondrial membrane potential and protection by inhibitors of uniport Ca++ transduction.
Incubation of isolated rat hepatocytes in Ca++-free medium generates an oxidative stress which causes significant cell injury. Ruthenium red and La , which block Ca++ uptake through the mitochondrial uniport, totally prevented malondialdehyde formation, glutathione and protein thiol oxidation and vitamin E loss induced by Ca++ omission. Accordingly, these agents also prevented leakage of intracellular K+ and lactate dehydrogenase. Similar protective effects were provided by the Ca++ chelator ethylene glycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid. The absence of extracellular Ca++ resulted in a marked decline of the mitochondrial transmembrane potential which could be prevented by ruthenium red, ethylene glycol bis(beta-aminoethyl ether)-N,N'tetraacetic acid, the antioxidant vitamin E and the iron chelator, desferrioxamine. In contrast, oxidative stress induced by treatment with the redox active agent paraquat and 1,3-bis(2-chloroethyl)-1-nitrosourea had little effect on mitochondrial transmembrane potential and malondialdehyde formation and lactate dehydrogenase leakage were not affected by ruthenium red or La . These results indicate that the incubation of rat hepatocytes in the absence of extracellular Ca++ creates an unusual oxidative stress which markedly affects mitochondrial function. The ability of vitamin E and desferrioxamine to inhibit the loss of mitochondrial transmembrane potential indicates that oxidative damage is involved in producing mitochondrial dysfunction. Furthermore, the potent inhibitory effects of ruthenium red and La suggest that Ca++ movement through the uniport, perhaps indicative of mitochondrial Ca++ cycling, plays a major role in generating this oxidative stress and promoting cell injury.
D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002478:Cells, Cultured; D003676:Deferoxamine / Q000494:pharmacology*; D005978:Glutathione / Q000378:metabolism; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D007811:Lanthanum / Q000494:pharmacology; D008099:Liver / Q000187:drug effects / Q000502:physiology*; D008297:Male; D008315:Malondialdehyde / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D008930:Mitochondria, Liver / Q000187:drug effects / Q000502:physiology*; D051381:Rats; D011919:Rats, Inbred Strains; D012428:Ruthenium / Q000494:pharmacology*; D012430:Ruthenium Red / Q000494:pharmacology*; D013367:Submitochondrial Particles / Q000187:drug effects / Q000502:physiology*; D014810:Vitamin E / Q000494:pharmacology*
D012430:Ruthenium Red; D014810:Vitamin E; D008315:Malondialdehyde; D007811:Lanthanum; D012428:Ruthenium; D007770:L-Lactate Dehydrogenase; D005978:Glutathione; D003676:Deferoxamine; D002118:Calcium