Introduction: Ischemic cardiomyopathy induced by ischemia/reperfusion injury is a major cause of morbidity and mortality around the world; therefore it is important to seek the new strategy that can reduce myocardial ischemia/reperfusion injury. Our recent study reported that the natural flavone acacetin effectively protected myocardium against ischemia/reperfusion injury; however, the detailed mechanism was poorly understood. The present study investigates the potential molecular mechanism of acacetin in myocardioprotection against ischemia/reperfusion injury.
Methods: An in vitro hypoxia/reoxygenation (H/R) model was established in primary cultured neonatal rat cardiomyocytes and H9C2 cardiomyoblasts, and potential molecular targets of acacetin in cardioprotection against H/R injury were determined in multiple biochemical and molecular biological approaches.
Results: It was found that acacetin (0.3-3 μM) significantly decreased the apoptosis and ROS production induced by H/R injury in both cardiomyocytes and H9C2. Acacetin reduced the pro-apoptotic proteins Bax and cleaved-caspase-3 and increased the anti-apoptotic protein Bcl-2 expression. In addition, acacetin not only suppressed the release of pro-inflammatory cytokines TLR4 and IL-6 induced by H/R injury, but also increased the secretion of anti-inflammatory cytokine IL-10. Moreover, acacetin increased the anti-oxidants Nrf2 and HO-1 in a concentration-dependent manner, and rescued the reduction of SOD1 and SOD2 induced by H/R injury. Interestingly, in cells with silenced Nrf2 by siRNA, acacetin (3 μM) no longer decreased the ROS production, the Bax and cleaved-caspase-3 expression, the TLR4 and IL-6 release, and no longer increased the Bcl-2 and antioxidants (HO-1,SOD-1,SOD-2) and the IL-10 release induced by H/R injury, suggesting that the cardiomyocyte protection of acacetin against H/R injury disappeared.
Conclusion: Our results demonstrate for the first time that acacetin is a novel Nrf2 activator and confers myocardioprotection against H/R injury by stimulating a series of intracellular signals related to anti-oxidation, anti-inflammation, and anti-apoptosis.