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目的:研究下调VEGF基因的表达对人乳腺癌细胞MCF-7的凋亡及相关基因表达的影响,探讨敲减VEGF后乳腺癌细胞MCF-7凋亡的机制。方法:体外化学合成针对VEGF基因的siRNA序列,在脂质体Li-pofectamine2000TM介导下转染MCF-7细胞,RT-PCR检测VEGF、Survivin及wtP53表达水平,比色法检测Caspase-3的活性,免疫组织化学法检测Survivin蛋白的表达,并用图像分析软件分析蛋白表达强度。结果:靶向VEGF的siRNA转染MCF-7后,明显促进了细胞凋亡,VEGF mRNA表达明显减少;Survivin基因mRNA及蛋白水平表达下调(P<0.01);p53的mRNA表达上调,Caspase-3活性增高。结论:敲减VEGF基因可促进MCF-7细胞的凋亡,其主要途径可能是下调Survivin的表达,活化Caspase-3来实现。
OBJECTIVE: To study the effect of downregulation of VEGF gene expression on the apoptosis and related gene expression of human breast cancer cell line MCF-7 and to explore the mechanism of MCF-7 apoptosis after knockdown of VEGF. Methods: siRNA targeting VEGF gene was synthesized in vitro and transfected into MCF-7 cells by liposome Li-pofectamine2000TM. The expression of VEGF, Survivin and wtP53 was detected by RT-PCR. The activity of Caspase-3 The expression of Survivin protein was detected by immunohistochemical method and the protein expression intensity was analyzed by image analysis software. Results: The apoptosis of MCF-7 cells transfected with VEGF targeting siRNA significantly decreased the expression of VEGF mRNA and the mRNA and protein expression of Survivin (P <0.01). The mRNA expression of p53 was up-regulated while the expression of Caspase-3 Increased activity. Conclusion: The knockdown of VEGF gene can promote the apoptosis of MCF-7 cells. The main pathway may be to down-regulate the expression of Survivin and activate Caspase-3.