Department of Cardiology, Renmin Hospital of Wuhan University, Cardiovascular Research Institute of Wuhan University, Hubei Key Laboratory of Cardiology, Jiefang Road 238, Wuchang, 430060 Wuhan, PR China.
Department of Cardiology, Renmin Hospital of Wuhan University, Cardiovascular Research Institute of Wuhan University, Hubei Key Laboratory of Cardiology, Jiefang Road 238, Wuchang, 430060 Wuhan, PR China.
Eur J Pharmacol. 2017 Nov 15;815:343-350. doi: 10.1016/j.ejphar.2017.08.041. Epub 2017 Sep 1.
To explore the potential protective effect of Tanshinone ⅡA on myocardial cell apoptosis and elucidate the underlying molecular mechanisms. The rat heart cell H9c2 was treated by either HO or doxorubicin (DOX) to mimic oxidative stress and DNA damage conditions in vivo. Cell growth was monitored by optical microscope observation or CCK-8 counting kit. The relative expression of miR-133 and U6 snoRNA was semi-quantitated by RT-PCR or real-time PCR. Cell apoptosis was analyzed by flow cytometry with Annexin V/PI double staining. The microRNA binding sites were predicted by online bioinformatics tools. The regulatory effect of miR-133 on caspase-9 was measured by luciferase reporter assay. Apoptosis pathway factors were analyzed by immunoblotting. Our data demonstrated that Tanshinone ⅡA significantly ameliorated myocardial apoptosis induced by either HO or DOX. The protective effect was likely mediated by up-regulation of miR-133. We further identified Caspase-9 as the target of miR-133. Tanshinone ⅡA treatment significantly reversed down-regulation of miR-133 under harsh conditions and in turn suppressed evoking of Caspase-9 and related apoptotic effectors, which consequently contributed to the improvement of myocardial injury. In conclusion, Tanshinone ⅡA ameliorated myocardial apoptosis via restoration of miR-133 and suppression Caspase-9 signaling cascade, which underlies its well-proven clinical benefit and warrants larger scale clinical applications.
探讨丹参酮Ⅱ A 对心肌细胞凋亡的潜在保护作用，并阐明其潜在的分子机制。
采用过氧化氢（HO）或阿霉素（DOX）处理大鼠心肌细胞 H9c2，模拟体内氧化应激和 DNA 损伤条件。通过光学显微镜观察或 CCK-8 计数试剂盒监测细胞生长。采用 RT-PCR 或实时 PCR 半定量检测 miR-133 和 U6 snoRNA 的相对表达。采用 Annexin V/PI 双染流式细胞术分析细胞凋亡。采用在线生物信息学工具预测 microRNA 结合位点。通过荧光素酶报告实验测定 miR-133 对 caspase-9 的调控作用。采用免疫印迹法分析凋亡途径相关因子。
丹参酮Ⅱ A 显著改善了 HO 或 DOX 诱导的心肌细胞凋亡。这种保护作用可能是通过上调 miR-133 介导的。我们进一步鉴定出 Caspase-9 是 miR-133 的靶基因。在恶劣条件下，丹参酮Ⅱ A 治疗可显著逆转 miR-133 的下调，并抑制 Caspase-9 和相关凋亡效应因子的激活，从而改善心肌损伤。
丹参酮Ⅱ A 通过恢复 miR-133 和抑制 Caspase-9 信号级联反应改善心肌细胞凋亡，这是其在临床上得到充分证实的益处的基础，值得进一步进行更大规模的临床应用。
