Molecular mechanisms of hydrogen peroxide preconditioning in cultured cardiomyocytes

In cardiac cells a short-timed stress, also called preconditioning (PR), may exert a protective role against a subsequent prolonged stress. In this study, we explored whether H2O2-PR protects against H2O2-induced oxidative stress in cardiac cells and the roles of phase II enzymes and MAPK signaling pathways in this adaptive protection. Primary cultures of cardiomyocytes were grown at confluence. PR was simulated with 1-100 µM H2O2 for 10 min, while oxidative stress was induced by 100 µM H2O2 for 30 min. Cell viability was evaluated by MTT and cytofluorimetric assays. Enzyme activities were determined by spectrophotometric methods. Phosphorylation of p38, Akt and ERK1/2 was analyzed by immunoblotting. MTT and cytofluorimetric analysis indicate a significant protective effect of PR against oxidative stress. PR was able to modulate the activities of different phase II enzymes and to increase the phosphorylation of ERK1/2, p38 and Akt protein kinases. Specific kinase inhibitors demonstrated that only p38 and Akt are involved in the protection elicited by PR.