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背景心肌细胞凋亡是心脏功能下降的主要原因,而胰岛素样生长因子可以保护心肌细胞免于凋亡。目的探讨胰岛素样生长因子对大鼠心肌细胞凋亡保护作用的基因调控机制。方法体外培养大鼠心肌细胞H9C2并用胰岛素样生长因子1(IGF-1,10nmol/L)刺激,用基因芯片观察实验及对照组基因表达的差异,从表达发生改变的基因中筛选出一个新的未被报道与心肌细胞凋亡相关的基因基础转录元件结合蛋白(BTEB)进行深入研究。通过半定量PCR及Western-blot观察IGF-1调节BTEB及其调控的基因细胞色素P4501A1(CYP1A1)的表达情况,同时进一步通过DNA片段化及半胱天冬蛋白酶3(caspase-3)酶活性观察IGF-1,BTEB及CYP1A1对心肌细胞凋亡的影响。结果H9C2细胞经IGF-1刺激60min后,BTEBmRNA和蛋白表达均明显下降;IGF-1刺激48h后,CYP1A1mRNA和蛋白表达均明显下降(均P<0.01)。与无血清组相比,用BTEB及CYP1A1特异的siRNA人为下调BTEB及CYP1A1的基因表达后,大鼠心肌细胞caspase-3酶活性降低,DNA片段化减少,细胞存活率增加(均P<0.01),与IGF-1的抗心肌细胞凋亡效应相似。结论IGF-1可以通过抑制BTEB、CYP1A1途径保护大鼠心肌细胞免于凋亡。
Background Cardiomyocyte apoptosis is a major cause of cardiac dysfunction, whereas insulin-like growth factor can protect cardiomyocytes from apoptosis. Objective To investigate the gene regulation mechanism of insulin-like growth factor (IGF) on cardiomyocyte apoptosis in rats. Methods Rat H9C2 cells were cultured in vitro and stimulated with insulin-like growth factor 1 (IGF-1, 10 nmol / L). The gene expression differences between the experimental and control groups were observed by gene chip, and a new The gene-based transcriptional element binding protein (BTEB), which is not reported to be involved in cardiomyocyte apoptosis, was studied in depth. The expression of cytochrome P4501A1 (CYP1A1) regulated by BTEB and its regulation by IGF-1 was observed by semi-quantitative PCR and Western-blot. Meanwhile, DNA fragmentation and caspase-3 activity were also observed Effects of IGF-1, BTEB and CYP1A1 on Cardiomyocyte Apoptosis. Results The mRNA and protein expressions of BTEB in H9C2 cells were significantly decreased after stimulated with IGF-1 for 60min. The mRNA and protein expressions of CYP1A1 in H9C2 cells were significantly decreased after IGF-1 stimulation for 48h (all P <0.01). Compared with the non-serum group, BTEB and CYP1A1-specific siRNA artificially down-regulated the gene expression of BTEB and CYP1A1, the activity of caspase-3 decreased, DNA fragmentation decreased and the cell survival rate increased (all P <0.01) , Similar to the anti-cardiomyocyte apoptosis effect of IGF-1. Conclusion IGF-1 can protect rat cardiomyocytes from apoptosis by inhibiting the BTEB and CYP1A1 pathways.