干扰iASPP基因对转染MCF-7细胞凋亡变化的观察

目的:观察iASPP基因干扰RNA转染乳腺癌细胞MCF-7后的干扰效果及细胞凋亡变化。方法:设计特异性siRNA序列,将序列克隆至PGCsilencerTM H1/Neo/GFP质粒中,用脂质体将重组子转染至MCF-7细胞中,用RT-PCR方法检测i ASPP的表达,蛋白质印迹法检测蛋白表达的变化,流式细胞仪检测细胞凋亡的情况。结果:iASPP干扰质粒转染MCF-7细胞后,iASPP的mRNA表达和蛋白表达减少40%～50%;p53蛋白相对表达量由转染阴性质粒的0.37增加到转染干扰质粒的0.64;细胞凋亡率由原来的17.8%和16.2%分别增加到53.5%和51.3%。结论:抑制内源性i ASPP能有效地恢复乳腺癌细胞MCF-7中p53的抑癌功能,为乳腺癌的治疗提供新的思路。

Effect of iASPP RNAi vector transfection on apoptosis of MCF-7

OBJECTIVE: To observe the effect of the iASPP RNAi vector transfection on the apoptosis of breat cancer cell MCF-7.METHODS: The specific siRNA sequence was designed according to the genebank.The sequence was cloned into PGCsilencerTM H1/Neo/GFP.The recombinant plasmid was transfected into MCF-7 by LipofectamineTM 2000.The iASPP expression was analyzed by RT-PCR and Western blot.The cell apoptosis was detected by FCM.RESULTS: The iASPP expression in MCF-7 cell lines was descended from 40% to 50% after the RNAi transfection.The expression of p53 protein increased from 0.37 to 0.64,and the apoptosis rate increased from 17.8% and 16.2% to 53.5% and 51.3%.CONCLUSION:The inhibition of endogenous iASPP may resume the apoptosis activity of p53 in MCF-7.