CHEMICAL HYPOXIA TRIGGERS APOPTOSIS OF CULTURED NEONATAL RAT CARDIAC MYOCYTES - MODULATION BY CALCIUM-REGULATED PROTEASES AND PROTEIN-KINASES

Myocardial infarctions and stroke arise primarily as a result of hypox ia/ischemia-induced cell injury. However, the molecular mechanism of c ardiac cell death due to hypoxia has not been elucidated. We showed he re that chemical hypoxia induced by 1 mM azide triggered apoptosis of isolated neonatal rat ventricular cardiac myocytes but had no effect o n cardiac fibroblasts. The azide-induced cardiomyocyte apoptosis could be characterized by a reversible initiation phase (0-6 h after azide exposure) during which cytosolic ATP levels remained little affected. This was followed by an irreversible execution phase (12-18 h) exhibit ing prominent internucleosomal DNA fragmentation, cell membrane leakag e, mitochondrial dysfunction, and increased calpain messenger RNA. Blo cking extracellular calcium influx or intracellular calcium release wa s each effective in suppressing myocyte apoptosis. Cell death was also found to be mediated by calcium sensitive signal transduction events based on the use of specific antagonists. Consistent with the inductio n of calpain expression during apoptosis, blocking de novo protein syn thesis and calpain activity inhibited cell death. These regulatory fea tures coupled with the ease of the cell system suggest that the myocyt e apoptosis model described here should be useful in the study of even ts leading to the demise of the myocardium.