Prevention of staurosporine-induced cell death in embryonic chick cardiomyocyte is more dependent on caspase-2 than caspase-3 inhibition and is independent of sphingomyelinase activation and ceramide generation.
Apoptosis was induced in embryonic chick cardiomyocytes by staurosporine. Treatment of cardiomyocytes with the preferential caspase-2 inhibitor, z-VDVAD-fmk (100 microM), produced a significant (P < 0.05) although small reduction in the amount of cell death. Ac-DVED-cmk (100 microM), which preferentially inhibits caspase-3 but inhibits to a lesser extent caspase-6, -7, -8, and -10, produced a minimal decrease in cell death. The combination of the caspase-3 and -2 inhibitors produced an additive reduction in cell death after staurosporine (1 microM for 6 h) from 80.4 +/- 0.7 to 54.6 +/- 1.3%. The ability of staurosporine to activate caspase-3 was confirmed in these cardiomyocytes by measurement of caspase-3 activity. A role for ceramide formation, from sphingomyelin to induce caspase activation was unlikely, as there were no changes in cellular ceramide or sphingomyelin after staurosporine treatment of cardiomyocytes when sphingomyelin was labeled by [(3)H]palmitate for 24 h. Neither were there any changes in sphingomyelinase activity. While staurosporine effectively suppressed PKC activity, phorbol 12-myristate 13 acetate did not alter staurosporine-induced cell death or DNA fragmentation. These results demonstrate that, in this model of cardiac cell death, caspase-2 inhibition is of considerable importance, caspase-3 inhibition is of lesser significance but may produce additional effects in the combination with caspase-2 inhibition, and ceramide production from sphingomyelin is not operative in the pathway leading to caspase activation and cell death.
Rabkin
S W
SW
University of British Columbia, Vancouver, British Columbia, Canada. rabkin@interchange.ubc.ca
D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology*; D053143:Caspase 2; D053148:Caspase 3; D061945:Caspase Inhibitors; D002518:Ceramides / Q000096:biosynthesis; D002642:Chick Embryo; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D006321:Heart / Q000187:drug effects; D009206:Myocardium / Q000166:cytology; D009842:Oligopeptides / Q000494:pharmacology; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism; D013109:Sphingomyelins / Q000378:metabolism; D019311:Staurosporine / Q000494:pharmacology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D061945:Caspase Inhibitors; D002518:Ceramides; D015853:Cysteine Proteinase Inhibitors; D009842:Oligopeptides; D013109:Sphingomyelins; C427308:benzoylcarbonyl-valyl-aspartyl-valyl-alanyl-aspartyl-fluoromethyl ketone; D013108:Sphingomyelin Phosphodiesterase; D053143:Caspase 2; D053148:Caspase 3; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate