Mechanism of clofibrate hepatotoxicity: mitochondrial damage and oxidative stress in hepatocytes.
Peroxisome proliferators have been found to induce hepatocarcinogenesis in rodents, and may cause mitochondrial damage. Consistent with this, clofibrate increased hepatic mitochondrial oxidative DNA and protein damage in mice. The present investigation aimed to study the mechanism by which this might occur by examining the effect of clofibrate on freshly isolated mouse liver mitochondria and a cultured hepatocyte cell line, AML-12. Mitochondrial membrane potential (Delta Psi(m)) was determined by using the fluorescent dye 5,5',6,6'-tetrachloro-1,1', 3,3'-tetraethyl-benzimidazolylcarbocyanine iodide (JC-1) and tetramethylrhodamine methyl ester (TMRM). Application of clofibrate at concentrations greater than 0.3 mM rapidly collapsed the Delta Psi(m) both in liver cells and in isolated mitochondria. The loss of Delta Psi(m) occurred prior to cell death and appeared to involve the mitochondrial permeability transition (MPT), as revealed by calcein fluorescence studies and the protective effect of cyclosporin A (CsA) on the decrease in Delta Psi(m). Levels of reactive oxygen species (ROS) were measured with the fluorescent probes 5-(and-6)-carboxy-2',7'-dichlorofluorescein diacetate (DCFDA) and dihydrorhodamine 123 (DHR123). Treatment of the hepatocytes with clofibrate caused a significant increase in intracellular and mitochondrial ROS. Antioxidants such as vitamin C, deferoxamine, and catalase were able to protect the cells against the clofibrate-induced loss of viability, as was CsA, but to a lesser extent. These results suggest that one action of clofibrate might be to impair mitochondrial function, so stimulating formation of ROS, which eventually contribute to cell death.
Qu
B
B
Department of Biochemistry, Faculty of Medicine, National University of Singapore, Singapore, Singapore.
Li
Q T
QT
Wong
K P
KP
Tan
T M
TM
Halliwell
B
B
D000818:Animals; D000924:Anticholesteremic Agents / Q000633:toxicity*; D002470:Cell Survival; D002478:Cells, Cultured; D002994:Clofibrate / Q000633:toxicity*; D016572:Cyclosporine / Q000494:pharmacology; D005609:Free Radicals; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D007473:Ion Channels; D008099:Liver / Q000502:physiology*; D008564:Membrane Potentials / Q000502:physiology; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008930:Mitochondria, Liver / Q000187:drug effects* / Q000378:metabolism; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D018384:Oxidative Stress / Q000187:drug effects*; D017382:Reactive Oxygen Species / Q000378:metabolism
D000924:Anticholesteremic Agents; D005609:Free Radicals; D007473:Ion Channels; D008565:Membrane Proteins; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D017382:Reactive Oxygen Species; D016572:Cyclosporine; D002994:Clofibrate