Objective: In this study, we investigated the regulation of resveratrol (RV) on miR-34a alteration due to ARI and further studied the involvement of miR-34a/Sirt1 signaling pathway in ROS generation and cell survival after ARI.
Materials and methods: In-vitro anoxia and reoxygenation injury (ARI) model based on rat heart-derived H9c2 cells was established. The expression of miR-34a and Sirt1 in H9c2 cells with or without RV pretreatment was measured. Flow cytometric analysis of intracellular reactive oxygen species (ROS) generation, CCK-8 assay of cell viability and Western blot analysis of active caspase-3 expression were performed to study the role of miR-34a/Sirt1 signaling pathway in RV modulated ARI injury protection.
Results: Pretreatment with RV substantially restored Sirt1 expression in cardiomyocytes in a dose-dependent manner in the in-vitro ARI model. MiR-34a level was significantly increased due to ARI. But pretreatment with RV significantly suppressed its upregulation. MiR-34a overexpression significantly reduced the effect of RV on restoring Sirt1 expression in ARI. Both miR-34a overexpression and Sirt1 knockdown significantly reduced the effect of RV on reducing ROS generation and also abrogated the effect of RV on enhancing cell viability and reducing cell apoptosis.
Conclusions: The present study demonstrated that RV has a suppressive effect on miR-34a upregulation in ARI and the miR-34a/Sirt1 axis is an important signaling pathway modulating the protective effect of RV on cardiomyocytes in ARI. Nonetheless, future in vivo studies are required to validate this mechanism.