摘要
本文应用大鼠心肌细胞缺氧/复氧损伤模型,探讨microRNA-21(miR-21)在大鼠心肌缺氧复氧损伤中的作用及其对细胞自噬的影响.缺氧复氧后,RT-PCR检测发现心肌miR-21表达上调(P<0.05),流式细胞术检测表明细胞凋亡增加,RT-PCR及蛋白质印迹(Western blot)检测发现p62显著下调而beclin-1显著上调(P<0.05),提示缺氧复氧诱导心肌细胞凋亡和自噬异常.脂质体转染miR-21 mimic后,细胞凋亡显著增加(P<0.05),p62显著上调而beclin-1显著下调(P<0.05),而转染miR-21抑制剂引起相反结果,提示miR-21在心肌缺氧复氧损伤中具有促进细胞凋亡、抑制细胞自噬的作用.生物信息学预测显示,Rab11a的3′-UTR含有miR-21的结合位点,双荧光素酶基因报告系统及Rab11a过表达实验表明Rab11a是miR-21的靶基因之一.心肌过表达Rab11a能减少缺氧复氧后miR-21介导的细胞凋亡及自噬.由此表明,在大鼠心肌缺氧复氧损伤中,miR-21可能通过负调控Rab11a促进心肌细胞凋亡,抑制心肌细胞自噬.本研究可能为预防和治疗心肌缺血再灌注损伤提供新策略.
The hypoxia/reoxygenation injury model was established in rat cardiomyocytes to investigate the influence of microRNA-21 on apoptosis and autophagy in rat cardiomyocytes exposed to hypoxia/reoxygenation injury. According to RT-PCR, miR-21 was up-regulated（P〈0.05） in myocardial cells after hypoxia/reoxygenation injury. Moreover, myocardial apoptosis was aggravated according to flow cytometry. According to RT-PCR and Western blot, p62 was down-regulated whereas beclin-1 was up-regulated in cardiomyocytes（P〈0.05） after hypoxia/reoxygenation injury. These consequences inditicated that hypoxia reoxygenation induced abnormal myocardial apoptosis and autophagy. miR-21 mimic or inhibitor were transfected into cardiomyocytes via liposome. Mi R-21 mimic transfection significantly enhanced myocardial apoptosis（P〈0.05）, up-regulated expression of p62, and down-regulated of expression of beclin-1in myocardial cells in rat cardiomyocytes（P〈0.05）,while miR-21 inhibitor transfection caused opposite effects. These data suggested that miR-21 in rat cardiomyocytes exposed to myocardial hypoxia reoxygenation injury can accelerate cell apoptosis and inhibit cell autophagy. Bioinformatics projections shown that Rab11 a 3＇-UTR contains a binding site for miR-21. Dual luciferase report gene assay system, coupled with the overexpression of Rab11 a assay validated that Rab11 a is one of miR-21 target genes. Overexpression of Rab11 a significantly attenuated myocardial apoptosis,up-regulation of p62 and down-regulation of beclin-1 induced by miR-21 in hypoxia/reoxygenation injury. In conclusion, miR-21 can promote myocardial apoptosis and inhibit myocardial autophagy by negative regulating Rab11 a in rat cardiomyocytes exposed to hypoxia/reoxygenation injury.This research proposes a new strategy for the prevention and treatment of myocardial ischemia-reperfusion injury.
出处
《生物化学与生物物理进展》
SCIE
CAS
CSCD
北大核心
2016年第2期150-156,共7页
Progress In Biochemistry and Biophysics
