Effects of H2O2 on Cardiac Rate and Apoptosis of Primary Rat Cardiomyoblasts in in Vitro Culture Systems

ABSTRACT

PURPOSE: This work was intended to establish experimental conditions for monitoring the effect of ischemic/reperfusion injury on the beating capability of and apoptotic damage to primary rat cardiomyoblasts. METHODS: In an in vitro system, cardiac rate differed depending on the number of days after birth that the cells were isolated. We maintained a mean rate of 62 times per min until 4 or 5 days in culture. To generate ischemic/reperfusion injury, primary rat cardiomyoblasts were treated with hydrogen peroxide (H2O2). RESULTS: Treatment with H2O2 significantly decreased the cardiac rate of primary rat cardiomyoblasts in a time- and dose-dependent manner. Interestingly, the cardiac rate of primary rat cardiomyoblasts abruptly dropped prior to the decrease in cell viability. H2O2 also induced a decrease in the expression of heme oxygenase-1 (HO-1) protein in P2 primary rat cardiomyoblasts in a time-dependent manner. Moreover, treatment with H2O2 resulted in an increase in the proportion of cells in the sub-G0/G1 phase, indicating that H2O2 induces the apoptotic death of P2 primary rat cardiomyoblasts. However, the intracellular level of calcium was markedly decreased under the same experimental conditions. CONCLUSION: An in vitro culture system is useful for investigating the mechanism underlying the beating capability of rat heart cells and the mechanism underlying apoptotic damage to primary rat cardiomyoblasts induced by ischemic/reperfusion injury, including ROS-induced damage.