Pitx2对缺氧诱导的心肌细胞H9c2凋亡的影响

Effect of Pitx2 on Apoptosis of Cardiomyocytes H9c2 Induced by Hypoxia

目的观察Pitx2对缺氧诱导的心肌细胞H9c2凋亡的影响。方法将心肌细胞H9c2分为对照组、模型组(缺氧处理)、Vector组(转染阴性对照载体,缺氧处理)、Pitx2组(转染Pitx2过表达载体,缺氧处理)。采用实时荧光定量逆转录-聚合酶链式反应(RT-qPCR)和免疫印迹法分别测定细胞Pitx2 mRNA和蛋白表达变化,四唑盐(MTT)法测定细胞增殖能力,流式细胞术测定细胞凋亡情况,免疫印迹法测定细胞C-Caspase-3、C-Caspase-12和C/EBP同源蛋白(CHOP)表达变化。将CHOP和Pitx2过表达载体共转染到心肌细胞H9c2中,测定细胞增殖、凋亡和C-Caspase-3、C-Caspase-12、CHOP表达变化。结果与对照组比较,模型组心肌细胞Pitx2 mRNA和蛋白表达水平下降,细胞增殖能力降低,细胞凋亡水平增加,细胞C-Caspase-3、C-Caspase-12和CHOP蛋白表达水平增加(P<0.05)。与Vector组比较,Pitx2组心肌细胞Pitx2mRNA和蛋白表达水平均升高,细胞增殖能力升高,细胞凋亡水平降低,细胞C-Caspase-3、C-Caspase-12和CHOP蛋白表达水平降低(P<0.05)。CHOP过表达载体可抑制Pitx2缺氧心肌细胞增殖,诱导细胞凋亡。结论上调Pitx2可抑制缺氧诱导的心肌细胞H9c2凋亡,其作用机制与调控CHOP影响内质网应激有关。

Objective To observe the effect of Pitx2 on apoptosis of cardiomyocytes H9c2 induced by hypoxia.Methods Cardiomyocytes H9c2 were divided into control group,model group(hypoxia treatment),Vector group(transfected with negative control Vector,hypoxia treatment),and Pitx2 group(transfected with Pitx2 overexpression vector,hypoxia treatment).The mRNA and protein expressions of Pitx2 were detected by real-time fluorescence quantitative reverse transcription-polymerase chain reaction(RT-qPCR)and Western Blotting,the cell proliferation ability were detected by methyl thiazolyl tetrazolium(MTT)assay,and the cell apoptosis ability was detected by flow cytometry.The protein expressions of C-Caspase-3,C-Caspase-12,and C/EBP homologous protein(CHOP)proteins were detected by Western Blotting.CHOP and Pitx2 overexpression vectors were co-transfected into cardiomyocytes H9c2.The cell proliferation,apoptosis,and expression of C-Caspase-3,C-Caspase-12,and CHOP proteins were detected.Results Compared with the control group,the mRNA and protein expressions of Pitx2 in the model group was decreased,cell proliferation ability was decreased.The levels of apoptosis,and the protein expressions of C-Caspase-3,C-Caspase-12,and CHOP were increased significantly(P<0.05).Compared with the Vector group,the mRNA and protein expressions of Pitx2,and cell proliferation capacity in the Pitx2 group was increased significantly.The levels of apoptosis,and the protein expressions of C-Caspase-3,C-Caspase-12,and CHOP significantly were decreased(P<0.05).The CHOP overexpression vector could inhibit the proliferation of Pitx2 hypoxia cardiomyocytes and induce apoptosis.Conclusion Upregulation of Pitx2 could inhibit hypoxia-induced apoptosis of cardiomyocyte H9c2,and its mechanism might be related to the regulation of CHOP on endoplasmic reticulum stress.