ACE-shRNA质粒预处理减轻缺氧/复氧诱导的H9C2心肌细胞凋亡  

Silencing of Angiotensin Converting Enzyme by RNA Interference Prevents H9c2 Cardiomyocytes from Apoptosis Induced by Anoxia/Reoxygenation

作　　者：万为国[1] 李晓艳[1] 张翠[1] 蒋学俊[1] 

机构地区：[1]武汉大学人民医院心内科,湖北武汉430060

出　　处：《武汉大学学报（医学版）》2016年第3期363-366,385,共5页Medical Journal of Wuhan University

基　　金：国家自然科学基金资助项目(编号:81170307)

摘　　要：目的:探讨通过RNA干扰(RNAi)技术沉默H9C2心肌细胞内的血管紧张素转换酶(ACE)基因对缺氧/复氧(A/R)诱导的细胞凋亡的影响及机制。方法:H9C2心肌细胞随机分为正常对照组,A/R组,阴性对照ACE-shRNA质粒处理组(NC组)和ACE-shRNA质粒处理组(shRNA组)。检测心肌细胞的存活率、凋亡、ACE、Bcl-2及Bax表达水平和细胞培养液中AngⅡ的浓度。结果:A/R能诱导H9C2心肌细胞存活率明显降低、细胞内ACE表达明显增加、细胞培养液中AngⅡ水平明显升高、细胞凋亡明显增加、心肌细胞Bcl-2和Bax蛋白水平及Bax/Bcl-2比值明显增加(P<0.05),ACE-shRNA质粒处理能逆转这些改变(P<0.05)。结论:心肌细胞内ACE基因沉默能调节细胞内的肾素-血管紧张素系统,进而调节细胞内凋亡途径,保护A/R诱导的心肌细胞损伤。Objective：To demonstrate whether the apoptosis in H9C2 cardiomyocytes following anoxia/reoxygenation（A/R）would be improved viasilencing of intracellular ACE by RNAi.Methods：The cultured H9C2 cardiomyocytes were randomly divided into different groups：the control（Con）group,the A/R group,the negative control（NC）group,and the ACE-shRNA plasmidtreated group（shRNA groupt）.Cell viability and apoptosis were analyzed by the CCK-8assay and flow cytometry,respectively.The mRNA and protein of ACE were detected by quantitative real-time RT-PCR and Western blot analysis.Bcl-2and Bax proteins were also examined by Western blot analysis.The levels of AngⅡ in the culture medium were assessed by enzymelinked immunosorbent assay.Results：Compared with control group,in A/R group cell viability was significantly decreased,the expression of ACE,the level of AngⅡand apoptotic H9C2 cardiomyocytes were significantly increased,and the Bax,Bcl-2,and Bax/Bcl-2ratios were also significantly increased after A/R（P〈0.05）.Treatment of ACE-shRNA plasmids reversed the previous change（P〈0.05）.Conclusion：Gene silencing of intracellular ACE holds great potential intreatment of cardiomyocyte apoptosis after I/R injury by regulating the intracellular RAS,consequently regulating the intrinsic pathway of apoptosis.

关 键 词：血管紧张素转换酶 缺氧/复氧 凋亡 心肌细胞 RNA干扰 

分 类 号：R541.4[医药卫生—心血管疾病]