【摘要】 目的构建人PTEN基因RNAi慢病毒载体,观察PTEN基因敲减前后人子宫内膜癌HEC-1A细胞生长和信号通路的变化,为进一步研究子宫内膜癌的发病机制提供基础。方法利用慢病毒载体系统,构建人PTEN基因RNAi慢病毒载体,转染HEC-1A细胞,建立PTEN基因敲减细胞模型HEC-1A-ND;WesternBlot方法检测PTEN基因敲减前后PTEN蛋白的表达和AKT通路的活化情况,MTT法检测PTEN敲减前后细胞增殖的变化,流式细胞术检测PTEN敲减前后细胞周期的变化。结果慢病毒介导的RNAi能有效下调PTEN基因的表达,PTEN基因表达下调后,HEC-1A细胞生长受到促进,S期细胞增多,G0-G1期细胞和G2/M期细胞减少,PI3K/AKT通路激活。结论成功构建人PTEN基因RNAi慢病毒载体。PTEN基因敲减后,可以通过激活PI3K/AKT通路,促进细胞生长。更多还原

【Abstract】 Objectives Construct lentiviral RNA interference vector of human PTEN gene,to observe the change of the growth and signal pathway after PTEN gene knocked down in human endometrial carcinoma cell HEC-1A.To provide an experimental foundation of the research of the pathogenesis of human endometrial carcinomas.Methods Using lentivirus vector system to construct lentiviral RNA interference vector of human PTEN gene,then HEC-1A cells were transfected with the lentivirus to establish the cell model with PTEN gene knocked down(named HEC-1A-ND);Using Western Blot to detect the expression of PTEN and the activation of AKT signal pathway;Using MTT to detect the growth;Using flow cytometry to detect the cell cycle.Results The RNAi mediated by lentivirus can down regulate the expression of the human PTEN gene effectively.After the down regulation the HEC-1A-ND cells grow faster.The percent of the cells in phase S increase and The percent of the cells in phase G0-G1 and G2/M decrease.The PI3K/AKT signal pathway is activated.Conclusions Construct lentiviral RNA interference vector of human PTEN gene successfully.PI3K/AKT signal pathway can be activated after PTEN gene knocked down to promote the cell growth.更多还原