Mitochondrial effects with ceramide-induced cardiac apoptosis are different from those of palmitate.
The objective of this study was to evaluate whether ceramide, palmitate, and inhibitors of mitochondrial electron transport chain shared similar effects on the mitochondria of intact cardiomyocytes in order to determine the likelihood that ceramide and palmitate utilize similar mitochondrial mechanisms or pathways to apoptosis. In embryonic chick cardiomyocytes, ceramide, 100 microM for 24h, induced a 42.9+/-5.8% increase in cell death assessed by the MTT assay, and a significant (P<0.01) 3.9+/-0.6-fold increase in apoptosis assessed by propidium iodide staining of permeabilized cells. Mitochondrial potential (delta psi (m)), as demonstrated microscopically and by flow cytometry of cardiomyocytes stained with a J-aggregate dye, was markedly and significantly reduced by ceramide, palmitate, and two different inhibitors of the mitochondrial electron transport chain-rotenone and antimycin A. In contrast, the effect on mitochondria as assessed by CMX-Ros oxidation was dramatically different, as palmitate, rotenone, and antimycin A each produced a reduction, while ceramide increased CMX-Ros fluorescence. Further ceramide-induced cardiomyocyte apoptosis and loss of delta psi (m) operated through a cyclosporine-insensitive pathway similar to rotenone and antimycin A but distinct from palmitate which induced apoptosis though a cyclosporine-sensitive mechanism in these cells. These data suggest that ceramide acts on the mitochondria of intact cells through a cyclosporine-insensitive mechanism likely from a combination of actions including production of mitochondrial oxidants. The discordant findings between ceramide and palmitate suggest that palmitate-induced cell death is not primarily mediated by de novo ceramide synthesis.
Kong
Jennifer Y
JY
Department of Medicine, University of British Columbia, Room D410, 2733 Heather Street, Vancouver, BC, Canada V5Z 3J5.
Rabkin
Simon W
SW
D000818:Animals; D000968:Antimycin A / Q000031:analogs & derivatives* / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D002478:Cells, Cultured; D002642:Chick Embryo; D016572:Cyclosporine / Q000494:pharmacology; D004579:Electron Transport / Q000187:drug effects; D005456:Fluorescent Dyes; D006321:Heart / Q000187:drug effects; D008564:Membrane Potentials / Q000187:drug effects; D008929:Mitochondria, Heart / Q000187:drug effects* / Q000378:metabolism*; D009206:Myocardium / Q000166:cytology; D009930:Organic Chemicals; D019308:Palmitic Acid / Q000494:pharmacology*; D012402:Rotenone / Q000494:pharmacology; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology*
D005456:Fluorescent Dyes; C064769:N-acetylsphingosine; D009930:Organic Chemicals; C107472:red dye CMXRos; D012402:Rotenone; C032456:antimycin; D019308:Palmitic Acid; D000968:Antimycin A; D016572:Cyclosporine; D013110:Sphingosine