目的探讨双调蛋白(amphiregulin,AREG)对糖尿病心肌病大鼠心肌细胞凋亡和氧化应激的的影响。方法25 mmol/L葡萄糖和500μmol/L棕榈酸(HGHF)诱导大鼠心肌细胞(H9C2)18 h,建立体外糖尿病心肌病大鼠心肌细胞模型。采用实时定量聚合酶链反应...目的探讨双调蛋白(amphiregulin,AREG)对糖尿病心肌病大鼠心肌细胞凋亡和氧化应激的的影响。方法25 mmol/L葡萄糖和500μmol/L棕榈酸(HGHF)诱导大鼠心肌细胞(H9C2)18 h,建立体外糖尿病心肌病大鼠心肌细胞模型。采用实时定量聚合酶链反应(quantitative real time polymerase chain reaction,qRT-PCR)和蛋白质印迹(Western blotting,WB)检测AREG基因的表达,采用流式细胞分析和脱氧核糖核苷酸末端转移酶介导的缺口末端标记法(terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling,TUNEL)染色观察siRNA下调AREG后H9C2细胞凋亡和氧化应激情况。通过qRT-PCR检测心肌细胞肥厚和纤维化的分子水平。结果在HGHF处理的H9C2细胞中AREG表达上调。抑制AREG的表达,减轻了HGHF导致的大鼠心脏氧化应激的增强,且减少了心肌细胞的凋亡,改善了心肌肥厚和纤维化的分子指标。结论AREG下调通过抑制细胞氧化和细胞凋亡来缓解心肌损伤,AREG可以作为治疗糖尿病心肌病的分子靶标。展开更多
Background Myocardial ischemia/reperfusion(I/R)injury remains a great challenge in clinical therapy.Recent studies indicated that circularRNA(circRNA)play an important role in the regulation of atherosclerotic heart d...Background Myocardial ischemia/reperfusion(I/R)injury remains a great challenge in clinical therapy.Recent studies indicated that circularRNA(circRNA)play an important role in the regulation of atherosclerotic heart disease.However,as a new circRNA,the effect of circRNA regulating ischemia-reperfusion through microRNA-1(CRIRTM)on myocardial apoptosis during myocardial ischemia/reperfusion is still unknown.The purpose of this study was to investigate the effect of CRIRTM on I/R-induced apoptosis and the expression of microRNA-1(miR-1)and calmodulin(CaM)in cultured neonatal rat ventricular cardiomyocytes(NRVCs)in vitro and SD rat hypoxia-reoxygenation model in vivo.Methods NRVCs were subjected to hypoxia for 30 minutes and then oxygen was added for 12 hours to establish the model of cell ischemia reperfusion.The SD rats were subjected to the surgery of ligating the anterior descending branch of the left coronary artery for 4 hours and then reperfused for 48 hours to establish myocardial ischemia animal model.The expression of CRIRTM,miIR-1 and CaM was detected by quantitative real-time polymerase chain reaction(qRT-PCR).TUNEL was used to analyze the apoptosis level of cells and animal myocardium after small interfering RNA(siRNA)down-regulation of CRIRTM.The expression of CRIRTM in the blood of patients with myocardial infarction was detected,and the downstream of CRIRTM was analyzed by Target Scan.Results The specific upregulation of CRIRTM was detected in the plasma of patients with ST-segment elevation myocardial infarction after reperfusion,and multiple binding sites between CRIRTM and miR-1 were found by Target Scan analysis.The expression of CRIRTM and CaM was up-regulated and the expression of miR-1 was down-regulated in cells and rats I/R model.Both in vivo and in vitro,cardiomyocyte apoptosis induced by myocardial ischemia-reperfusion will increase after down-regulation of CRIRTM.It may be due to the decreased inhibition of miR-1 by down-regulated CRIRTM,which increases the degradation of CaM by miR-1 and aggravates cardiomyocyte apoptosis.Conclusions Our data demonstrated that CRIRTM alleviates myocardial ischemia-reperfusion injury via suppressing miR-1 leading to enhanced CaM levels.[S Chin J Cardiol 2021;22(1):30-37]展开更多
文摘目的探讨双调蛋白(amphiregulin,AREG)对糖尿病心肌病大鼠心肌细胞凋亡和氧化应激的的影响。方法25 mmol/L葡萄糖和500μmol/L棕榈酸(HGHF)诱导大鼠心肌细胞(H9C2)18 h,建立体外糖尿病心肌病大鼠心肌细胞模型。采用实时定量聚合酶链反应(quantitative real time polymerase chain reaction,qRT-PCR)和蛋白质印迹(Western blotting,WB)检测AREG基因的表达,采用流式细胞分析和脱氧核糖核苷酸末端转移酶介导的缺口末端标记法(terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling,TUNEL)染色观察siRNA下调AREG后H9C2细胞凋亡和氧化应激情况。通过qRT-PCR检测心肌细胞肥厚和纤维化的分子水平。结果在HGHF处理的H9C2细胞中AREG表达上调。抑制AREG的表达,减轻了HGHF导致的大鼠心脏氧化应激的增强,且减少了心肌细胞的凋亡,改善了心肌肥厚和纤维化的分子指标。结论AREG下调通过抑制细胞氧化和细胞凋亡来缓解心肌损伤,AREG可以作为治疗糖尿病心肌病的分子靶标。
基金supported by National Natural Science Foundation of China(No.81670334/No.81800262)Science and Technology Planning Project of Guangzhou(No.201903010005)+1 种基金Science and Technology Planning Project of Guangzhou(No.201906010089)Natural Science Foundation of Guangdong Province(No.2018A030313029)
文摘Background Myocardial ischemia/reperfusion(I/R)injury remains a great challenge in clinical therapy.Recent studies indicated that circularRNA(circRNA)play an important role in the regulation of atherosclerotic heart disease.However,as a new circRNA,the effect of circRNA regulating ischemia-reperfusion through microRNA-1(CRIRTM)on myocardial apoptosis during myocardial ischemia/reperfusion is still unknown.The purpose of this study was to investigate the effect of CRIRTM on I/R-induced apoptosis and the expression of microRNA-1(miR-1)and calmodulin(CaM)in cultured neonatal rat ventricular cardiomyocytes(NRVCs)in vitro and SD rat hypoxia-reoxygenation model in vivo.Methods NRVCs were subjected to hypoxia for 30 minutes and then oxygen was added for 12 hours to establish the model of cell ischemia reperfusion.The SD rats were subjected to the surgery of ligating the anterior descending branch of the left coronary artery for 4 hours and then reperfused for 48 hours to establish myocardial ischemia animal model.The expression of CRIRTM,miIR-1 and CaM was detected by quantitative real-time polymerase chain reaction(qRT-PCR).TUNEL was used to analyze the apoptosis level of cells and animal myocardium after small interfering RNA(siRNA)down-regulation of CRIRTM.The expression of CRIRTM in the blood of patients with myocardial infarction was detected,and the downstream of CRIRTM was analyzed by Target Scan.Results The specific upregulation of CRIRTM was detected in the plasma of patients with ST-segment elevation myocardial infarction after reperfusion,and multiple binding sites between CRIRTM and miR-1 were found by Target Scan analysis.The expression of CRIRTM and CaM was up-regulated and the expression of miR-1 was down-regulated in cells and rats I/R model.Both in vivo and in vitro,cardiomyocyte apoptosis induced by myocardial ischemia-reperfusion will increase after down-regulation of CRIRTM.It may be due to the decreased inhibition of miR-1 by down-regulated CRIRTM,which increases the degradation of CaM by miR-1 and aggravates cardiomyocyte apoptosis.Conclusions Our data demonstrated that CRIRTM alleviates myocardial ischemia-reperfusion injury via suppressing miR-1 leading to enhanced CaM levels.[S Chin J Cardiol 2021;22(1):30-37]