CXCR4 siRNA对AngⅡ诱导的H9c2心肌细胞凋亡的影响

Role of CXCR4 siRNA in Ang Ⅱ induced cardiomyocytes H9c2 apoptosis

目的:观察趋化因子受体-4(CXCR4)基因在血管紧张素Ⅱ(AngⅡ)诱导的心肌细胞凋亡中的作用。方法:体外常规培养心肌细胞H9c2,分为4组:对照组(不予任何处理);AngⅡ组(给予100nmol/LAngⅡ培养24h);siRNA-con+AngⅡ组(转染对照siRNA24h后,给予100nmol/LAngⅡ继续培养24h);siRNA-CXCR4+AngⅡ组(转染CXCR4siRNA24h后,给予100nmol/LAngⅡ继续培养24h)。采用RT-PCR检测各组心肌细胞中CXCR4基因的表达;采用流式细胞术检测各组心肌细胞凋亡情况;Westernblot检测各组心肌细胞中CleavedCaspase-3和Bax蛋白水平;采用ELISA法检测各组细胞上清液中TNF-α含量。结果:与对照组比较,AngⅡ组心肌细胞中CXCR4mRNA表达水平升高,细胞凋亡率升高,CleavedCaspase-3和Bax蛋白水平上调,培养上清中TNF-α含量增加(P<0.05)。与AngⅡ组比,siRNA-CXCR4+AngⅡ组心肌细胞中CXCR4mRNA表达水平降低,心肌细胞凋亡率降低,CleavedCaspase-3和Bax蛋白水平下调,培养上清中TNF-α含量减少(P<0.05)。结论:下调CXCR4基因对AngⅡ诱导的心肌细胞凋亡具有一定的抑制作用。

Aim :To observe the role of CXCR4 siRNA in angiotensin Ⅱ(Ang Ⅱ) induced cardiomyocyte apoptosis. Methods :Cardiomyocytes H9c2 were cultured in vitro . H9c2 cells were divided into control group(no treatment), Ang Ⅱ group(100 nmol/L Ang Ⅱ cultured for 24 hours);siRNA-con+Ang Ⅱ group(transfected with siRNA control for 24 hours,100 nmol/L Ang Ⅱ further cultured for 24 hours);siRNA-CXCR4+Ang Ⅱ group(after transfecting CXCR4 siRNA for 24 hours, 100 nmol/L Ang Ⅱ cultured for 24 hours).The effects of transfecting CXCR4 siRNA in cardiomyocytes was detected by RT-PCR. The apoptosis of cardiomyocytes was detected by flow cytometry. The levels of Cleaved Caspase-3 and Bax protein in cardiomyocytes were detected by Western blot. The content of TNF-α in the supernatant was detected by ELISA. Results :Compared with the control group, the expression level of CXCR4 mRNA in the Ang Ⅱ group was significantly increased, the apoptosis rate was increased, the levels of Cleaved Caspase-3 and Bax proteins were up-regulated, and the content of TNF-α in the supernatant was increased( P <0.05). Compared with the Ang Ⅱ group, the expression level of CXCR4 mRNA in the siRNA-CXCR4+Ang Ⅱ group was significantly decreased, the apoptosis rate of cardiomyocytes was decreased, the levels of Cleaved Caspase-3 and Bax proteins were down-regulated, and the content of TNF-α in supernatant was decreased( P <0.05). Conclusion :Down-regulation of CXCR4 gene has a certain inhibitory effects on Ang Ⅱ induced cardiomyocyte apoptosis.