Lu Min, Qin Xinglei, Yao Jungong, Yang Yuanyuan, Zhao Minghu, Sun Lin
Department of Cardiologry, Henan Provincial People's Hospital, People's Hospital of Zhengzhou University, School of Clinical Medicine, Zhengzhou, Henan, China.
Department of Hepatobiliary Surgery, Henan Provincial People's Hospital, People's Hospital of Zhengzhou University, School of Clinical Medicine, Zhengzhou, Henan, China.
IUBMB Life. 2020 Oct;72(10):2154-2166. doi: 10.1002/iub.2351. Epub 2020 Aug 14.
MicroRNA-134-5p (MiR-134-5p) has been proposed as a promising novel biomarker for the diagnosis of acute myocardial infarction (AMI). However, the biological role of miR-134-5p in ischemic cardiomyocytes has been little disclosed yet. Expression of miR-134-5p and X-linked inhibitor of apoptosis protein (XIAP) was detected using RT-qPCR and western blot. Oxidative stress and cell apoptosis were determined by enzyme-linked immunosorbent assays, 3-(4, 5-dimethylthiazole-2-y1)-2, 5-biphenyl tetrazolium bromide assay, flow cytometry, western blot, and terminal-deoxynucleoitidyl transferase-mediated nick end labeling (TUNEL). The interaction between miR-134-5p and XIAP was confirmed by luciferase reporter assay. Expression of miR-134-5p was upregulated in serum of AMI patients and hypoxia/reoxygenation (H/R)-induced cardiomyocytes (AC16 and HCM). MiR-134-5p downregulation could inhibit H/R-mediated release of lactic dehydrogenase enzyme (LDH) and malondialdehyde (MDA), and promote superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) levels. Meanwhile, cell viability was increased, while the apoptosis rate and TUNEL positive cells were inhibited by miR-134-5p downregulation in H/R-treated AC16 and HCM cells. Mechanically, XIAP was downregulated and targeted by miR-134-5p in H/R-induced cardiomyocytes in vitro. Overexpression of XIAP inhibited oxidative stress and cell apoptosis in H/R-treated AC16 and HCM cells, which was similar to miR-134-5p knockdown. Moreover, downregulation of XIAP could partially reverse the effect of miR-134-5p knockdown in H/R-induced cardiomyocytes. Knockdown of miR-134-5p protected cardiomyocytes from H/R-induced oxidative stress and apoptosis in vitro through targeting XIAP.
微小RNA-134-5p（MiR-134-5p）已被提议作为急性心肌梗死（AMI）诊断中一种有前景的新型生物标志物。然而，miR-134-5p在缺血性心肌细胞中的生物学作用尚未完全阐明。采用逆转录定量聚合酶链反应（RT-qPCR）和蛋白质免疫印迹法检测miR-134-5p和X连锁凋亡抑制蛋白（XIAP）的表达。通过酶联免疫吸附测定、3-（4,5-二甲基噻唑-2-基）-2,5-二苯基溴化四氮唑（MTT）法、流式细胞术、蛋白质免疫印迹法和末端脱氧核苷酸转移酶介导的缺口末端标记法（TUNEL）测定氧化应激和细胞凋亡。通过荧光素酶报告基因测定证实miR-134-5p与XIAP之间的相互作用。在AMI患者血清以及缺氧/复氧（H/R）诱导的心肌细胞（AC16和HCM）中，miR-134-5p的表达上调。下调miR-134-5p可抑制H/R介导的乳酸脱氢酶（LDH）和丙二醛（MDA）释放，并提高超氧化物歧化酶（SOD）和谷胱甘肽过氧化物酶（GSH-PX）水平。同时，在H/R处理的AC16和HCM细胞中，下调miR-134-5p可提高细胞活力，同时抑制凋亡率和TUNEL阳性细胞。机制上，在体外H/R诱导的心肌细胞中，XIAP被miR-134-5p下调并靶向。XIAP过表达抑制了H/R处理的AC16和HCM细胞中的氧化应激和细胞凋亡，这与敲低miR-134-5p相似。此外，下调XIAP可部分逆转敲低miR-134-5p对H/R诱导的心肌细胞的影响。在体外，敲低miR-134-5p通过靶向XIAP保护心肌细胞免受H/R诱导的氧化应激和凋亡。
