circ-HIPK3对急性心肌梗死的影响及作用机制研究  

The impact and mechanism of circ-HIPK3 on acute myocardial infarction

作　　者：徐鹏 杨建敏 钱宇峰 朱钢杰 谢剑昶 刘梅军 XU Peng;YANG Jianmin;QIAN Yufeng;ZHU Gangjie;XIE Jianchang;LIU Meijun(Department of Cardiovascular Medicine,Hangzhou First People's Hospital Affiliated to Xihu University School of Medicine,Hangzhou 310006,China)

机构地区：[1]西湖大学医学院附属杭州市第一人民医院心血管内科,310006

出　　处：《心电与循环》2024年第3期219-224,I0001,I0002,共8页Journal of Electrocardiology and Circulation

基　　金：浙江省医药卫生科技计划项目(2021KY245)。

摘　　要：目的探讨circ-HIPK3对急性心肌梗死(AMI)的影响及作用机制研究。方法将C57BL/6小鼠按随机数字表法分为假手术组和AMI组,每组10只;通过左冠状动脉前降支结扎法构建AMI模型。另分离乳鼠原代心肌细胞并构建体外缺氧模型。采用原位末端转移酶标记染色法检测心肌细胞凋亡情况,实时荧光定量聚合酶链反应检测circ-HIPK3、miR-215表达水平,蛋白质印迹法检测叉头框蛋白O1(FOXO1)以及凋亡相关蛋白B淋巴细胞瘤-2(Bcl-2)、Bcl-2关联X蛋白(Bax)、活化的胱天蛋白酶-3(Cleaved caspase-3)表达水平;采用双荧光素酶报告基因检测验证miR-215与FOXO1的相互作用。结果与假手术组比较,AMI组小鼠心脏组织病理学改变明显,细胞凋亡数量增多,circ-HIPK3和FOXO1表达水平均升高,miR-215水平降低,差异均有统计学意义(均P<0.01);在体外缺氧培养的心肌细胞中亦有相同变化(均P<0.01)。circ-HIPK3敲低后心肌细胞活力增加,细胞凋亡数量减少,Bcl-2表达水平升高,Bax、Cleaved caspase-3表达水平均降低,差异均有统计学意义(均P<0.01)。miR-215与FOXO1存在结合位点;双荧光素酶报告基因检测证实FOXO1是miR-215的下游靶基因,其表达受miR-215调控。与缺氧组比较,缺氧+si-FOXO1(敲低FOXO1)组细胞活力增加,细胞凋亡数量减少,Bcl-2表达水平升高,Bax、Cleaved caspase-3表达水平均降低,差异均有统计学意义(均P<0.01);转染circ-HIPK3过表达质粒后,si-FOXO1对缺氧心肌细胞的作用被逆转(均P<0.01)。结论circ-HIPK3通过调控mi R-215促进FOXO1表达,进而增强心肌细胞凋亡作用,促进AMI的发生、发展。Objective To explore the impact and mechanism of circ-HIPK3 on acute myocardial infarction(AMI).Methods C57BL/6 mice were randomly divided into sham operation group and AMI group,with 10 mice in each group.The AMI model was constructed by ligation of the left anterior descending branch of the left coronary artery.The primary cardiomyocytes were isolated and the hypoxia model was constructed in vitro.The apoptosis of cardiomyocytes was detected with the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay.Quantitative real-time polymerase chain reaction was used to measure the expression levels of circ-HIPK3 and miR-215.The expressions of forkhead box protein O1(FOXO1),B-cell lymphoma-2(Bcl-2),Bcl2-associated X protein(Bax)and Cleaved caspase-3 were detected by Western blot.Dual luciferase reporter gene assay was used to detect the interaction between miR-215 and FOXO1.Results Compared with sham operation group,the cardiac histopathological changes of AMI group were obvious,the number of apoptotic cells was increased,the expression levels of circ-HIPK3 and FOXO1 were elevated,and the level of miR-215 decreased,with all statistical significance(all P<0.01).In vitro hypoxic culture of cardiac myocytes also showed similar changes(all P<0.01).After circ-HIPK3 knockdown,the viability of cardiomyocytes increased,the number of apoptotic cells decreased,the expression levels of Bcl-2 increased,and the expression levels of Bax and Cleaved caspase-3 decreased,with all statistical significance(all P<0.01).There were binding sites between miR-215 and FOXO1.Dual luciferase reporter gene assay confirmed that FOXO1 was the downstream target gene of miR-215,and its expression was regulated by miR-215.Compared with the hypoxia group,the hypoxia+si-FOXO1(knockdown FOXO1)group exhibited increased cell viability,reduced apoptotic cell count,increased Bcl-2 expression,and decreased expression levels of Bax and Cleaved caspase-3,with all statistical significance(all P<0.01).After transfection with circ-HIPK3 over

关 键 词：circ-HIPK3 mi R-215 叉头框蛋白O1 急性心肌梗死 

分 类 号：R542.22[医药卫生—心血管疾病]