U50,488H抑制心肌细胞缺氧/复氧诱导Drp1线粒体转位的作用及机制  

Effect of U50,488H on Drp1 mitochondria translocation in cardiomyocytes subjected to hypoxia/reoxygenation and its mechanism

作　　者：刘振华 刘银姬 牛津 贾敏 付锋 裴建明 樊荣 LIU Zhen-hua;LIU Yin-ji;NIU Jin;JIA Min;FU Feng;PEI Jian-ming;FAN Rong(Department of Physiology and Pathophysiology,School of Basic Medical Sciences,Air Force Medical University,Xi’an 710032,Shaanxi,China)

机构地区：[1]空军军医大学基础医学院生理与病理生理学教研室,陕西西安710032

出　　处：《心脏杂志》2020年第6期565-571,共7页Chinese Heart Journal

基　　金：国家自然科学基金项目资助(81770243,81670354)。

摘　　要：目的研究κ-阿片受体(κ-OR)激动剂U50,488H对缺氧复氧(HR)原代心肌细胞Drp1线粒体转位的作用与机制。方法将心肌细胞共分为6组,分别为常氧(Control)组、HR组、HR+U50,488H组、Control+Scramble RNAi组、HR+Scramble RNAi组、HR+Mid51 RNAi组。利用CCK-8试剂盒检测细胞活力;AnnexinV、PI试剂盒检测细胞凋亡;采用激光共聚焦显微镜观察线粒体的形态变化和Drp1与线粒体共定位情况,Western blotting检测细胞内线粒体外膜Drp1相关结合蛋白(Fis1、Mff、Mid49和Mid51)的表达水平。结果与常氧组相比,HR组细胞凋亡与死亡率明显增加(P<0.01),线粒体分裂明显增加(P<0.01),Drp1的线粒体转位明显增多,Mid51表达显著上调(P<0.01),但Fis1、Mff与Mid49的表达无明显差异;而κ-OR激动剂U50,488H可降低HR后细胞凋亡与死亡率(P<0.01),抑制线粒体分裂(P<0.01),减少Drp1与线粒体的结合同时下调Mid51的表达水平(P<0.01),但Fis1、Mff与Mid49的表达无明显变化;进一步研究发现,敲低Mid51可降低HR后细胞凋亡与死亡率(P<0.01,P<0.05),抑制线粒体的分裂(P<0.01),减少Drp1的线粒体的转位。结论HR可引起线粒体分裂与心肌细胞损伤,激活κ-OR可通过下调Mid51抑制Drp1的线粒体转位,进而抑制线粒体分裂,从而减轻HR损伤。AIM To examine the effect and mechanism ofκ-opioid receptor(κ-OR)activation by U50,488H on Drp1 mitochondria translocation in primary cardiomyocytes subjected to hypoxia reoxygenation(HR).METHODS Cardiomyocytes were divided into 6 groups:Control group,HR group,HR+U50,488H group,control+Scramble RNAi group,HR+Scramble RNAi group,and HR+Mid51 RNAi group.Cell viability was measured by CCK-8 detecting kit.Cell apoptosis was determined by flow cytometry.Mitochondria morphology and the colocalization of Drp1 and mitochondria were observed by laser confocal microscope.Western blotting was used to examine the Drp1-related protein’s expression levels of Fis1,Mff,Mid49 and Mid51.RESULTS Compared with the control group,cell apoptosis,cell death and mitochondrial fission increased significantly in the HR group(P<0.01).In addition,the amount of Drp1 translocated to mitochondrial surface and Mid51 were significantly increased(P<0.01)in the HR group,while no significant changes were found in the expression of Fis1,Mff and Mid49.κ-OR stimulation by U50,488H reduced cell apoptosis and death rate(P<0.01),inhibited mitochondrial fission(P<0.01),reduced the colocalization level of Drp1 and mitochondria,and downregulated the expression of Mid51(P<0.01).There were no significant changes in the expression of Fis1,Mff and Mid49 in the HR group.Knockdown of Mid51 with small interfering RNA decreased the apoptosis and death rate(P<0.01,P<0.05),inhibited the mitochondrial fission(P<0.01)and reduced the mitochondrial translocation of Drp1.CONCLUSION HR triggers cardiomyocyte injury and increases mitochondrial fission.Activation ofκ-OR by U50,488H reduces Drp1 translocation to mitochondria by inhibiting Mid51,and suppresses the abnormal mitochondrial dynamics induced by HR,so as to alleviates cell injury.

关 键 词：Κ-阿片受体 心肌细胞 缺氧复氧 线粒体动力学 线粒体动力学蛋白51 

分 类 号：R364.1[医药卫生—病理学]