干扰UCP2保护H9C2心肌细胞免受缺氧诱导的细胞凋亡并抑制细胞自噬  

Interference with UCP2 in protection of H9C2 cardiomyocytes from hypoxia-induced apoptosis and inhibition of autophagy

作　　者：张莉[1] 李峰[2] 刘悦雁[2] 王艳梅[1] 章辰琛 Zhang Li;Li Feng;Liu Yueyan;Wang Yanmei;Zhang Chenchen(Department of Human Anatomy,Wanxi Health Vocational College,Lu'an 237005,China;Department of Physiology,Wanxi Health Vocational College,Lu'an 237005,China)

机构地区：[1]皖西卫生职业学院解剖教研室,安徽六安237005 [2]皖西卫生职业学院生理教研室,安徽六安237005

出　　处：《山东第一医科大学（山东省医学科学院）学报》2021年第9期651-656,共6页Journal of Shandong First Medical University ＆ Shandong Academy of Medical Sciences

摘　　要：目的研究干扰解偶联蛋白2(uncoupling protein 2,UCP2)表达对缺氧诱导的细胞凋亡及自噬的影响。方法通过RT-qPCR检测缺氧后H9C2心肌细胞UCP2表达水平,活性氧簇(reactive oxygen species,ROS)荧光实验检测干扰UCP2对细胞内ROS水平的影响,通过流式细胞实验检测细胞凋亡率,通过蛋白免疫印迹实验进一步检测干扰UCP2对cleaved-caspase 3的影响,并通过ELISA检测细胞损伤相关因子:乳酸脱氢酶(lactate dehydrogenase,LDH)、肌酸激酶(creatine kinase,CK)、白细胞介素-6(interleukin-6,IL-6)和肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α),最后检测细胞自噬相关因子:膜型LC3/胞浆型LC3(LC3-Ⅱ/LC3-Ⅰ)、自噬基因Beclin-1以及p62的表达水平。结果缺氧环境下H9C2细胞中UCP2的表达量明显升高,干扰UCP2表达可以降低缺氧引起的细胞内ROS水平升高。干扰UCP2表达可以降低缺氧环境下H9C2细胞凋亡率,并且降低LDH、CK、IL-6和TNF-α的表达水平。在蛋白免疫印迹实验中,干扰UCP2表达可降低LC3-Ⅱ/LC3-Ⅰ和Beclin-1表达,并增加p62的表达水平。结论干扰UCP2可降低缺氧引起的细胞内ROS水平,并抑制缺氧引起的细胞凋亡及自噬。Objective:To study the effect of interfering uncoupling protein 2 expression on apoptosis and autophagy induced by hypoxia.Methods:The expressions of UCP2 in H9C2 myocardial cells after hypoxia were detected by RT-q PCR.Reactive oxygen species(ROS)fluorescence assay was used to detect the effect of UCP2 interference on intracellular ROS level.The apoptotic rate was detected by flow cytometry.The effects of UCP2 interference on cleaved-caspase 3 were further detected by Western blotting.The cell damage related factors were detected by ELISA:lactate dehydrogenase(LDH),creatine kinase(CK),interleukin-6(IL-6)and tumor necrosis factor-alpha(TNF-a).The expression levels of membrane LC3/cytoplasmic LC3(LC3-Ⅱ/LC3-Ⅰ),autophagy gene Beclin-1 and p62 were detected.Results:The expression of UCP2 was significantly increased in H9C2 cells under hypoxia.Interference with UCP2 expression could reduce the increase of ROS level in H9C2 cells induced by hypoxia.Interference with UCP2 expression could reduce the apoptotic rate of H9C2 cells and the expression levels of LDH,CK,IL-6 and TNF-alpha in hypoxic environment.In Western blotting,interfering with UCP2 expression could decrease the expression of LC3-Ⅱ/LC3-Ⅰand Beclin-1,and increase the expression of p62.Conclusion:Interference with UCP2 can reduce ROS level in cells induced by hypoxia,and inhibit apoptosis and autophagy induced by hypoxia.

关 键 词：UCP2 H9C2 细胞凋亡 细胞自噬 ROS 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]