Heat Shock Pretreatment Inhibits Ischemia - Reperfusion Induced Cardiomyocyte Apoptosis Mechanism

Aim: Myocardial apoptosis has been recognized as one of patterns of cell death during myocardial ischemia-reperfusion injury. Heat shock pretreatment may protect myocardium against various damages by the induction of heat shock proteins. This study was designed to explore the mechanisms of myocardial apoptosis during myocardial ischemia-reperfusion injury from mitochondria and death receptor signaling pathways and to further clarify the molecular mechanisms by which heat shock pretreatment inhibits myocardial apoptosis induced by ischemia-reperfusion injury.Methods: Myocardial ischemia-reperfusion injury was induced by occlusion of left anterior descending branch of coronary artery for 30 min then release for 4, 8, 12, 24 hours in mice. Oxidative damage of neonatal rat cardiomyocytes was induced by exposure to hydrogen peroxide (H2O2, 0.5mmol/L) for different durations. Apoptosis was evaluated by DNA laddering assay and staining with Hoechst 33258. Activities of Caspase-3, 8, 9 were measured with Caspase Colorimetric Assay Kit and Western blot. Expression of heat shock proteins was detected by Western blot analysis. The release of Smac from mitochondria into cytoplasm was observed by Western blot and immunofluorescence. To explore the effect of heat shock pretreatment on myocardium against apoptosis, mice were pretreated with whole body hyperthermia (rectal 42 ℃ for 15 min and recovery for 24 hours) before myocardial ischemia-reperfusion injury, and neonatal rat cardiomyocytes were pretreated with hyperthermia (42℃ for 1 hour and recovery for 12 hours )beforexposure to H2O2.Results: Ischemia-reperfusion injury induced activation of Caspase-3, 8, 9 in myocardium. Heat shock pretreatment induced the expression of several family members of heat shock proteins and inhibited myocardial apoptosis and activation of Caspase-3, 8, 9. Heat shock pretreatment induced expression of HSP70, HSP90 and aB-crystallin, inhibited H2O2-mediatedactivation of caspase-3, 9 and release of Smac from mitochondria and inhibited apoptosis in neonatal rat cardiomyocytes.Conclusion: Ischemia-reperfusion and H2O2 could induce myocardial apoptosis. Mitochondria and death receptor signaling pathways played important roles in myocardial apoptosis induced by ischemia-reperfusion injury. Heat shock pretreatment increased the expression of several heat shock proteins, inhibited the activation of both mitochondria and death receptor signaling pathways, decreased the release of Smac from mitochondria and inhibited apoptosis in cardiomyocytes.