Cyanide-induced death of dopaminergic cells is mediated by uncoupling protein-2 up-regulation and reduced Bcl-2 expression.
Cyanide is a potent inhibitor of mitochondrial oxidative metabolism and produces mitochondria-mediated death of dopaminergic neurons and sublethal intoxications that are associated with a Parkinson-like syndrome. Cyanide toxicity is enhanced when mitochondrial uncoupling is stimulated following up-regulation of uncoupling protein-2 (UCP-2). In this study, the role of a pro-survival protein, Bcl-2, in cyanide-mediated cell death was determined in a rat dopaminergic immortalized mesencephalic cell line (N27 cells). Following pharmacological up-regulation of UCP-2 by treatment with Wy14,643, cyanide reduced cellular Bcl-2 expression by increasing proteasomal degradation of the protein. The increased turnover of Bcl-2 was mediated by an increase of oxidative stress following UCP-2 up-regulation. The oxidative stress involved depletion of mitochondrial glutathione (mtGSH) and increased H2O2 generation. Repletion of mtGSH by loading cells with glutathione ethyl ester reduced H2O2 generation and in turn blocked the cyanide-induced decrease of Bcl-2. To determine if UCP-2 mediated the response, RNAi knock down was conducted. The RNAi decreased cyanide-induced depletion of mtGSH, reduced H2O2 accumulation, and inhibited down-regulation of Bcl-2, thus blocking cell death. To confirm the role of Bcl-2 down-regulation in the cell death, it was shown that over-expression of Bcl-2 by cDNA transfection attenuated the enhancement of cyanide toxicity after UCP-2 up-regulation. It was concluded that UCP-2 up-regulation sensitizes cells to cyanide by increasing cellular oxidative stress, leading to an increase of Bcl-2 degradation. Then the reduced Bcl-2 levels sensitize the cells to cyanide-mediated cell death.
Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, IN 47907-1333, USA.
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Borowitz
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Isom
D000818:Animals; D016923:Cell Death / Q000187:drug effects*; D002461:Cell Line, Transformed; D003486:Cyanides / Q000633:toxicity*; D018076:DNA, Complementary; D004298:Dopamine / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects; D005978:Glutathione / Q000187:drug effects / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D007473:Ion Channels / Q000187:drug effects* / Q000235:genetics; D008636:Mesencephalon / Q000166:cytology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000187:drug effects* / Q000235:genetics; D018384:Oxidative Stress / Q000187:drug effects; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000187:drug effects* / Q000235:genetics; D011743:Pyrimidines / Q000494:pharmacology; D034622:RNA Interference; D051381:Rats; D014162:Transfection; D000071246:Uncoupling Protein 2; D015854:Up-Regulation / Q000187:drug effects
D003486:Cyanides; D018076:DNA, Complementary; D007473:Ion Channels; D024101:Mitochondrial Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011743:Pyrimidines; C000604391:Ucp2 protein, rat; D000071246:Uncoupling Protein 2; C006253:pirinixic acid; D006861:Hydrogen Peroxide; D005978:Glutathione; D004298:Dopamine