Involvement of caspase 3-and 8-like proteases in ceramide-induced apoptosis of cardiomyocytes

Ceramides are the metabolic products of sphingolipids of the eukaryotic cel l membranes and are believed to function as signaling molecules in a variet y of biological processes. Ceramide induces apoptosis in cultured cardiomyo cytes. However, the molecular pathway underlying ceramide-induced apoptosis is not clear. In this study, we investigated the role of the cysteinyl asp artate-specific proteases (caspases) in cardiomyocyte apoptosis induced by ceramide. Treatment of in vitro cultured rat neonatal cardiomyocytes with c eramide results in robust cell death, of which the majority is apoptotic, a s shown by positive staining for terminal deoxyribonuclease transferase-med iated deoxyuridine triphosphate nick end-labeling and the appearance of pyk notic nuclei with Hoechst staining. Caspase 3- and 8-like protease activiti es are induced in cardiomyocytes by ceramide treatment. Addition of the tet rapeptide inhibitors for caspases attenuated ceramide-induced apoptosis. Th e nonselective caspase inhibitor (B-D-FMK) and the caspase 3 (Z-DEVD-FMK) a nd caspase 8 (Z-IETD-FMK) inhibitors reduced ceramide-induced cardiomyocyte death and significantly inhibited the activation of caspase 3. However, th e inhibitors specific for caspases 1, 2, 4, 6, and 9 have no significant ef fects on cardiomyocyte survival under the same conditions. These data sugge st that caspases 3- and 8-related proteases are involved in ceramide-induce d cardiomyocyte apoptosis.