Resveratrol protects H9c2 cells against hypoxia-induced apoptosis through miR-30d-5p/SIRT1/NF-κB axis

ABSTRACT

Previous studies have demonstrated the cardioprotective role of resveratrol (Res). However, the underlyingmolecular mechanisms involved in the protective role of Res are still largely unknown. H9c2 cells weredistributed into five groups normal condition (Control), DMSO, 20 mMRes (dissolved with DMSO), hypoxia(Hyp), and Res?Hyp. Cell apoptosis was evaluated using flow cytometry and protein analysis of cleavedcaspase 3 (cle-caspase 3). qRT-PCR assay was performed to measure the expression of microRNA-30d-5p(miR-30d-5p). MTT assay was performed to evaluate the cell proliferation. The relationship between miR-30d5p and silent information regulator 1 (SIRT1) was confirmed by luciferase reporter, RNA immunoprecipitation(RIP), and western blot assays. Western blot was performed to analyze NF-jB/p65 and I-jBa expressions. Ourdata showed that hypoxia enhanced apoptosis and NF-jB signaling pathway, which was alleviated by Restreatment. Hypoxia increased the expression of miR-30d-5p while decreased the SIRT1expression, which wasalso attenuated by Res treatment. Furthermore, miR-30d-5p depletion inhibited the proliferation, reducedapoptosis and decreased the expression of cle-caspase 3 in H9c2 cells with hypoxia treatment. Luciferasereporter, RIP, and western blot assays further confirmed that miR-30d-5p negatively regulated the expression ofSIRT1. Interestingly, the rescue-of-function experiments further indicated that knockdown of SIRT1 attenuatedthe effect of miR-30d-5p depletion on proliferation, apoptosis NF-jB signaling pathway inH9c2 cells withhypoxia treatment. In addition, the suppression of NF-jB signaling pathway increased cell viability whiledecreased cell apoptosis in hypoxia-mediatedH9c2 cells. Our data suggested Res mayprotectH9c2 cells againsthypoxia-induced apoptosis through miR-30d-5p/SIRT1/NF-jB axis