HAGLROS靶向miR-20a-5p调控高糖诱导的心肌细胞增殖和凋亡  

HAGLROS regulates the proliferation and apoptosis of high glucose-induced cardiomyocyte by targeting miR-20a-5p

作　　者：刘晓东 刘晓琴 党宏伟[1] 赵玲[2] Liu Xiaodong;Liu Xiaoqin;Dang Hongwei;Zhao Ling(Department of Cardiology,Baoji Central Hospital,Baoji721000,China;Department of Cardiology,The Affiliated Hospital of Xi'an Medical College,Xi'an 721000,China)

机构地区：[1]陕西宝鸡市中心医院心内科,宝鸡721000 [2]陕西西安医学院附属医院心内科,西安721000

出　　处：《广西医科大学学报》2020年第12期2159-2165,共7页Journal of Guangxi Medical University

基　　金：陕西省教育厅项目资助(No.16JK1648)。

摘　　要：目的:探讨长链非编码RNA(lncRNA)HAGLROS对高糖(HG)诱导的心肌细胞H9c2增殖和凋亡的影响及作用机制。方法:H9c2细胞分为对照组(NC组,细胞正常培养)、HG组(33 mmol/L葡萄糖作用细胞24 h)、HG+si-NC组(33 mmol/L葡萄糖作用转染si-NC的细胞24 h)、HG+si-HAGLROS组(33 mmol/L葡萄糖作用转染si-HAGLROS的细胞24 h)、HG+miR-NC组(33 mmol/L葡萄糖作用转染miR-NC的细胞24 h)、HG+miR-20a-5p组(33 mmol/L葡萄糖作用转染miR-20a-5p mimics的细胞24 h)、HG+si-HAGLROS+anti-miR-NC组(33 mmol/L葡萄糖作用于共转染si-HAGLROS与anti-miR-NC的细胞24 h)和HG+si-HAGLROS+anti-miR-20a-5p组(33 mmol/L葡萄糖作用于共转染si-HAGLROS与anti-miR-20a-5p的细胞24 h),实时荧光定量PCR(qPCR)检测细胞中HAGLROS和miR-20a-5p表达,四甲基噻唑蓝染色法(MTT)、流式细胞术和蛋白印迹(Western blotting)分别检测细胞存活率、凋亡率及细胞周期蛋白D1(CyclinD1)、活化的半胱天冬酶-3(cleaved-caspase-3)、磷酸化磷脂酰肌醇3激酶(p-PI3K)和磷酸化蛋白激酶B(p-AKT)的蛋白表达的影响。双荧光素酶报告基因实验验证HAGLROS与miR-20a-5p调控关系。结果:与NC组比较,HG组HAGLROS水平升高(P<0.05),miR-20a-5p水平降低(P<0.05),H9c2细胞存活率和CyclinD1蛋白水平降低(P<0.05),而细胞凋亡率和cleaved-caspase-3蛋白水平升高(P<0.05)。与HG+si-NC组比较,HG+si-HAGLROS组H9c2细胞存活率和CyclinD1、p-PI3K和p-AKT蛋白水平升高(P<0.05),而细胞凋亡率和cleaved-caspase-3蛋白水平降低(P<0.05)。与HG+miR-NC组,HG+miR-20a-5p组H9c2细胞存活率和CyclinD1蛋白表达升高(P<0.05),而细胞凋亡率和cleaved-caspase-3蛋白表达降低(P<0.05)。与HG+si-HAGLROS+anti-miR-NC组比较,HG+si-HAGLROS+anti-miR-20a-5p组H9c2细胞存活率和CyclinD1、p-PI3K和p-AKT蛋白表达降低(P<0.05),而细胞凋亡率和cleaved-caspase-3蛋白表达升高(P<0.05)。HAGLROS在H9c2细胞中靶向负调控miR-20a-5p表达。结论:低表达HAGLROS可促进HG诱导的心肌细胞Objective:To investigate the effects of lncRNAHAGLROS on the proliferation and apoptosis of high glucose(HG)-inducedcardiomyocyte H9c2and its mechanism.Methods:H9c2 cells were divided into a control group(NC group,cells were cultured normally),a HG group(33 mmol/L glucose-treated cells for 24 h),a HG+si-NC group(33 mmol/L glucose-treated transfected si-NC cells for 24 h),a HG+si-HAGLROS group(33 mmol/L glucose-treated transfected si-HAGLROS cells for 24 h),a HG+miR-NC group(33 mmol/L glucose-treated transfected miR-NC cells for 24 h),a HG+miR-20a-5p group(33 mmol/L glucose-treated transfected miR-20a-5p mimics cells for 24 h),a HG+si-HAGLROS+anti-miR-NC group(33 mmol/L glucose-treated co-transfected with si-HAGLROS and anti-miR-NC cells for 24 h),and a HG+si-HAGLROS+anti-miR-20a-5p group(33 mmol/L glucose-treated co-transfected si-HAGLROS and anti-miR-20a-5p cells 24 h).RT-qPCRwas used to detect the expression levels of HAGLROS and miR-20a-5p.MTT,flow cytometry,and Western blotting were used to detect the survival rate,apoptosis rate,and the expressionlevels ofCyclinD1,cleaved-caspase-3 protein,p-PI3K,and p-AKT.The double luciferase reporter gene experiment verified the regulatory relationship between HAGLROS and miR-20a-5p.Results:Compared with the NC group,the level of HAGLROS in the HG group was increased,while the level of miR-20a-5p was decreased(P<0.05).H9c2 cell survival rate and the protein level of CyclinD1 were decreased,while cell apoptosis rate and the protein level of cleaved-caspase-3 were increased(P<0.05).Compared with the HG+si-NC group,H9c2 cell survival rate and the protein level of CyclinD1,p-PI3K,and p-AKT in the HG+si-HAGLROS group were increased,while cell apoptosis rate and the protein level of cleaved-caspase-3 was decreased(P<0.05).Compared with the HG+miR-NC group,H9c2 cell survival rate and the protein level of CyclinD1 in the HG+miR-20a-5p group were increased,while cell apoptosis rate and the protein level of cleaved-caspase-3 were decreased(P<0.05).Compared with the HG+si-HAGLROS+an

关 键 词：HAGLROS miR-20a-5p 高糖 心肌细胞 增殖 凋亡 

分 类 号：R542.2[医药卫生—心血管疾病]