微小RNA-29a-5p通过Endoglin改善心肌梗死后心肌纤维化  

MicroRNA-29a-5p improves myocardial fibrosis after myocardial infarction through Endoglin

作　　者：李恩[1] 余海彬[1] 法宪恩[1] 汪涛[2] 周小翠 刘士超[1] 简立国[1] 张丽华[1] Li En;Yu Haibin;Fa Xianen;Wang Tao;Zhou Xiaocui;Liu Shichao;Jian Liguo;Zhang Lihua(Heart Center of the Second Affiliated Hospital of Zhengzhou University,Intervention Center of the Second Affiliated Hospital of Zhengzhou University,Zhengzhou 450014,China;Department of Geriatrics,the Second Affiliated Hospital of Zhengzhou University,Zhengzhou 450014,China)

机构地区：[1]郑州大学第二附属医院心脏中心,郑州大学第二附属医院介入中心,450014 [2]郑州大学第二附属医院老年医学科,450014

出　　处：《中华实验外科杂志》2020年第10期1907-1911,共5页Chinese Journal of Experimental Surgery

基　　金：河南省医学科技攻关计划项目(201802160)。

摘　　要：目的:观察微小RNA(miRNA,miR)-29a-5p在心肌梗死后心室重塑心肌纤维化中的作用,探讨miR-29a-5p通过靶基因Endoglin调控心肌梗死后心肌纤维化的作用机制。方法:选取2019年1月至2019年10月在郑州大学第二附属医院收治的30例心肌梗死患者和20例非急性冠脉综合征的患者,取血液标本,分别进行血清miR-29a-5p和NTPRO-脑钠肽(BNP)的检测。体内实验,建立小鼠心肌梗死模型,尾静脉分别注射miR-29a-5p mimics和阴性对照试剂。4周后处死小鼠,取部分心肌组织用实时定量聚合酶链反应(Real-time PCR)和蛋白质印迹法(Western blot)分别对miR-29a-5p、Endoglin、BNP进行检测。体外实验,处死1日龄小鼠,分离获得心肌成纤维细胞并进行培养。将培养的细胞用脂质体2000对细胞进行转染miR-29a-5p mimics和阴性对照试剂,转染后加入50μmol/L血管紧张素Ⅱ(AngⅡ)或等体积的磷酸盐缓冲液(PBS)持续48 h。用图像J软件检测各组成纤维细胞的表面积。另取细胞用miR-29a-5p mimics或antagomir,pRL-TK-endoglin-3’端非编码区(3’UTR)载体,pGL3-basic质粒进行共转染。转染48 h,获得细胞,用双荧光素酶报告试验系统检测细胞相对荧光素酶活性,并用Real-time PCR和Western blot分别检测Endoglin在各组的表达。采用单因素方差分析和LSD检验。结果:急性心肌梗死(AMI)患者血清中miR-29a-5p含量[(3.57±0.53)ng/ml]明显降低(F=935.899,P<0.01),NTPRO-BNP含量明显升高[(9.47±5.06)ng/ml,F=65.440,P<0.01]。小鼠MI组miR-29a-5p表达水平明显降低(0.51±0.12,t=4.315,P<0.01),在AngⅡ诱导的心肌纤维化模型中同样发现miR-29a-5p的表达水平明显降低(0.65±0.26,t=4.511,P<0.01)。miR-29a-5p mimics处理心肌成纤维细胞中Endoglin蛋白含量和mRNA表达水平降低(0.49±0.09,t=352.312,P<0.01),在miR-29a-5p antagomir处理心肌成纤维细胞中Endoglin蛋白含量和mRNA表达水平升高(2.15±0.14,t=7.814,P<0.05)。结论:miR-29a-5p是心肌纤维化的重要调节Objective To study the role of microRNA(miRNA,miR)-29a-5p in myocardial fibrosis after myocardial infarction,and further study the mechanism of miR-29a-5p regulating myocardial fibrosis after myocardial infarction through target gene Endoglin.Methods Thirty patients with myocardial infarction and 20 patients with non acute coronary syndrome were recruited.Blood samples were collected to detect miR-29a-5p and NTpro-brain natriuretic peptide(BNP).In vivo,the myocardial infarction model of mice was established,and the tail vein was injected with miR-29a-5p agomir and negative control reagent respectively.Four weeks later,After the mice were killed,some of the myocardial tissues were detected by real-time quantitative polymerase chain reaction(Real-time PCR)and Western blotting for miR-29a-5p,Endoglin and BNP respectively.In vitro experiments,1-day-old mice were killed,myocardial fibroblasts were isolated and cultured.The cultured cells were transfected with miR-29a-5p MICs and negative control reagent with liposome 2000.After transfection,50μmol/L angiotensinⅡ(AngⅡ)or phosphate buffer(PBS)of equal volume were added for 48 hours.The surface area of each fiber cell was measured by Image J software.The cells were co transfected with miR-29a-5p mimics or antigomir,pRL-tk-endoglin-3’untranslated regions(3’UTR)vector and pGL3 basic plasmid.After 48 hours of transfection,the cells were obtained,and the relative luciferase activity was detected by double Luciferase Report System.The expression of Endoglin in each group was detected by Real-time PCR and Western blotting respectively.Results The content of miR-29a-5p[(3.57±0.53)ng/ml]in serum of acute myocardial infarct(AMI)patients was significantly reduced(F=935.899,P<0.01),and the content of NTPRO-BNP[(9.47±5.06)ng/ml]was significantly increased(F=65.440,P<0.01).The expression level of miR-29a-5p(0.51±0.12)in the MI group of rats was significantly reduced(t=4.315,P<0.01),and the expression level of miR-29a-5p(0.65±0.26)was also found to be significantly reduce

关 键 词：微小RNA-29a-5p 心肌梗死 ENDOGLIN 心肌纤维化 

分 类 号：R542.22[医药卫生—心血管疾病]