新生大鼠心肌细胞通过STAT和Smads信号交叉对话介导心肌细胞肥大的体外研究

Study of cross-talk between stat family and smads pathway mediated cardiomyocyte hypertrophy in cultured neonatal rat in vitro

目的:通过研究新生大鼠体外培养的心肌细胞信号转导机制交叉对话,探讨心肌细胞肥大机制。方法:用血管紧张素Ⅱ(AngⅡ)和转化生长因子-β1(TGF-β1)及其相应的阻断剂Valsartan和Staurosporine刺激体外培养新生大鼠心肌细胞,以蛋白含量、搏动频率、细胞表面积等指标评价心肌细胞肥大。RT-PCR检测Smad3mRNA水平。Westernblotting检测磷酸化Stat1/Stat1,磷酸化Stat3/Stat3,Smad2/3。结果:AngⅡ(10-7mol/L)或者TGF-β1(3μg/L)刺激心肌细胞时,蛋白含量、搏动频率、细胞表面积呈现时间依赖方式的增加。与对照组相比,AngⅡ和TGF-β1刺激明显增加Smad3的mRNA水平。AngⅡ和TGF-β1刺激时,磷酸化Stat1/Stat1,磷酸化Stat3/Stat3,Smad2/3的增加可以被相应的阻断剂呈现时间依赖性逆转。结论:AngⅡ和TGF-β1通过Stat1,Stat3,和Samd2/3信号途径介导心肌细胞肥大。心肌细胞的信号转导交叉对话可以为心肌细胞肥大提供有效干预方法。

Objective：To study the signal transduction crosstalk between the stat family and smad pathway in cardiomyocyte of neonatal rat in vitro, and investigate the mechanism of cardiomyoctye hypertrophy. Methods： Cultured neonatal rat cardiomyocytes were used to evaluate the effects of Ang Ⅱ and TGF-β1 on cardiomyoctye hy pertrophy with or without their inhibitor valsartan and staurosporine, respectively. Protein content, beat frequen ey per minute, and cell surface area detection were used to evaluate the cardiomyocyte hypertrophy. The mRNA of Smad3 were measured by RT-PCR. Ratios of phospho-Statl/total Statl, phospiao-Stat3/total Star3 and Smad2/3 were detected by western blotting. Result：Protein content, beat frequency per minute, cell surface area were elevated in cardiomyocytes treated with Ang Ⅱ （10^-7 mol/L） or TGF-β1 （3 ng/ml） in a time-dependent manner compared with those control groups. The hallmark of cardiomyoctye hypertrophy such as Smad3 mRNA levels were elevated in the Ang Ⅱ or TGF-β1 treated with groups compared to those without drug interfered with groups. The augmentation of ratios of phospho-Statl/total Stat1, phospho-Stat3/total Stat3 and Smad2/3 in the two specific cells stimulated with Ang Ⅱ or TGF-β1 were reversed by the selective AT1 receptor antagonist, Valsartan, and Staurosporine, a PKC inhibitor in a time-dependent manner. Conclusion：These findings indicate that the Ang Ⅱ and TGF-β1 mediated the hypertrophy of cultured neonatal rat cardiomyocyte via the common Statl, Stat3, and Smad2/3 signal pathway. The cross talk between them in the cardiomyocyte may provide an effective means of ameliorating cardiomyoctye hypertrophy.