Critical role of reactive oxygen species and mitochondrial permeability transition in microcystin-induced rapid apoptosis in rat hepatocytes.
Microcystin-LR (M-LR) is a specific hepatotoxin. At present, the exact toxic mechanism of its action remains unclear though apoptosis is believed to be involved. This study was designed to investigate the role of reactive oxygen species (ROS) and mitochondrial permeability transition (MPT) in the M-LR-induced apoptotic process. Morphologic changes such as cell shrinkage, externalization of cell membrane phosphatidylserine, DNA fragmentation, and nuclear condensation suggest that M-LR causes rapid apoptosis in hepatocytes. Confocal microscopy revealed that M-LR exposure led to the onset of MPT and mitochondrial depolarization, evidenced by (1) redistribution of calcein fluorescence from cytosol to mitochondria, and (2) loss of mitochondrial tetramethyrhodamine methyl ester (TMRM) fluorescence; both occurred before apoptosis. Moreover, there was a significant and rapid increase of ROS level before the onset of MPT and loss of MMP, indicating a critical role of ROS in M-LR-induced apoptosis. Deferoxamine (DFO), an iron chelator, prevented the increase of ROS production, delayed the onset of MPT, and, subsequently, cell death. In addition, a specific MPT inhibitor, cyclosporin A (CsA), blocked the M-LR-induced ROS formation, onset of MPT, and mitochondrial depolarization as well as cell death. Thus, we conclude that the M-LR-induced ROS formation leads to the onset of MPT and apoptosis.
Ding
W X
WX
Center for Environmental and Occupational Health, Department of Community, Occupational and Family Medicine, Faculty of Medicine, National University of Singapore.
Shen
H M
HM
Ong
C N
CN
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D016764:Cell Polarity / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002614:Chelating Agents / Q000494:pharmacology; D016572:Cyclosporine / Q000494:pharmacology; D003676:Deferoxamine / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000502:physiology*; D008297:Male; D008387:Marine Toxins / Q000494:pharmacology*; D052998:Microcystins; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D010456:Peptides, Cyclic / Q000494:pharmacology*; D010539:Permeability; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism / Q000502:physiology*; D013997:Time Factors
D002614:Chelating Agents; D008387:Marine Toxins; D052998:Microcystins; D010456:Peptides, Cyclic; D017382:Reactive Oxygen Species; D016572:Cyclosporine; D007770:L-Lactate Dehydrogenase; C057862:cyanoginosin LR; D003676:Deferoxamine