Preconditioning blocks cardiocyte apoptosis:: role of KATP channels and PKC-ε

The aims of this study were to determine whether preconditioning blocks cardiocyte apoptosis and to determine the role of mitochondrial ATP-sensitive K+ (K-ATP) channels and the protein kinase C epsilon-isoform (PKC-epsilon) in this effect. Ventricular myocytes from 10-day-old chick embryos were used. In the control series, 10 h of simulated ischemia followed by 12 h of reoxygenation resulted in 42 +/- 3% apoptosis (n = 8). These results were consistent with DNA laddering and TdT-mediated dUTP nick-end labeling (TUNEL) assay. Preconditioning, elicited with three cycles of 1 min of ischemia separated by 5 min of reoxygenation before subjection to prolonged simulated ischemia, markedly attenuated the apoptotic process (28 +/- 4%, n = 8). The selective mitochondrial K-ATP channel opener diazoxide (400 mumol/l), given before ischemia, mimicked preconditioning effects to prevent apoptosis (22 +/- 4%, n = 6). Pretreatment with 5-hydroxydecanoate (100 mumol/l), a selective mitochondrial K-ATP channel blocker, abolished preconditioning (42 +/- 2%, n = 6). In addition, the effects of preconditioning and diazoxide were blocked with the specific PKC inhibitors Go-6976 (0.1 mumol/l) or chelerythrine (4 mumol/l), given at simulated ischemia and reoxygenation. Furthermore, preconditioning and diazoxide selectively activated PKC-epsilon in the particulate fraction before simulated ischemia without effect on the total fraction, cytosolic fraction, and PKC delta-isoform. The specific PKC activator phorbol 12-myristate 13-acetate (0.2 mumol/l), added during simulated ischemia and reoxygenation, mimicked preconditioning to block apoptosis. Opening mitochondrial K-ATP channels blocks cardiocyte apoptosis via activating PKC-epsilon in cultured ventricular myocytes. Through this signal transduction, preconditioning blocks apoptosis and preserves cardiac function in ischemia-reperfusion.