RNAi沉默PDCD4基因对人喉癌Hep-2细胞增殖及β-catenin表达的影响

Effect of silencing PDCD4 gene on proliferation of Hep-2 cells and the expression of β-catenin by RNA interference technique

目的应用RNAi技术下调人喉癌Hep-2细胞中PDCD4基因的表达,探讨其对Hep-2细胞增殖及β-catenin表达的影响。方法设计并合成针对PDCD4基因的shRNA质粒与阴性对照质粒,分别转染Hep-2细胞。实时定量RT-PCR和Western blotting检测转染前后阴性对照组与干扰组PDCD4、β-catenin基因mRNA和蛋白的表达。克隆形成实验观察Hep-2细胞增殖能力的变化。结果与对照组相比,PDCD4干扰组的Hep-2细胞克隆形成率显著升高(P=0.01),PDCD4干扰组PDCD4mRNA和蛋白均显著减低(P<0.01)、β-catenin mRNA和蛋白均较对照组显著增加(P<0.01)。结论成功应用RNAi技术下调人喉癌Hep-2细胞中PDCD4基因的表达,Hep-2细胞克隆形成能力增强,β-catenin mRNA和蛋白表达显著升高。

Objective To study the effect of silencing PDCD4 gene on proliferation of Hep-2 cells and the expression ofβ-catenin by RNA interference technique.Methods Specific shRNA plasmid and negative control plasmid of PDCD4 gene were designed and synthesized.Hep-2 cells were transfected with shRNA plasmid and negative control plasmid. Real-time RT-PCR and Western blotting analysis were performed to determine the expressions of PDCD4 and β-catenin mRNA and protein after transfection,respectively.The proliferation of Hep-2 cells was observed by colony-forming as-say.Results Compared to the control group,the colony formation rate of Hep-2 cells was significantly increased in the shRNNA plasmid group（P =0.01）;the expressions of PDCD4 mRNA and protein were significantly reduced （P 〈0.01）;the expressions of β-catenin mRNA and protein were significantly increased（P 〈0.01）.Conclusion RNA in-terference technique can effectively silence PDCD4 gene expression of Hep-2 cells ,enhance the colony forming ability of Hep-2 and increase the expressions of β-catenin mRNA and protein.