iASPP对表达野生型p53基因的乳腺癌细胞株MCF-7凋亡的影响

iASPP on apoptosis in breast cancer cells which expressed wild type p53

目的:P53凋亡刺激蛋白(ASPP)家族成员能够调节P53诱导的细胞凋亡。其中ASPP家族抑制成员(iASPP)主要通过特异性地和P53蛋白结合抑制肿瘤细胞的凋亡,本实验通过构建iASPP的RNAi质粒,研究在体内外抑制iASPP基因后对表达野生型p53基因的乳腺癌细胞MCF-7凋亡的影响。方法:采用分子生物学方法构建iASPP的腺病毒RNAi质粒pAd-iASPP-RNAi;体外转染MCF-7细胞,同时建立MCF-7细胞的裸鼠移植瘤模型;分别采用RT-PCR和Western blotting检测转染后瘤细胞iASPP mRNA和蛋白质表达的变化;采用流式细胞术检测转染前后瘤细胞凋亡的变化。结果:转染iASPPRNAi后,MCF-7细胞的iASPP mRNA及蛋白表达量降低(抑制率分别为95.4%和96.8%,P<0.01);MCF-7细胞的凋亡百分率、坏死百分率与对照组相比具有显著差异(P<0.01)。裸鼠移植瘤模型经pAd-iASPP-RNAi处理后,iASPP mRNA及蛋白表达量也明显降低(抑制率分别为87.4%和89.2%,P<0.01);移植瘤细胞的凋亡百分率和坏死百分率显著上升(P<0.01和P<0.05)。结论:在体内外抑制iASPP基因的表达能够通过p53途径诱导表达野生型p53的乳腺癌细胞凋亡。

AIM： To investigate the RNAi effect of the inhibitory member of the ASPP family （iASPP） on the apoptosis of human breast cancer cell MCF - 7 which expressed the wild type p53 gene. METHODS ： The recombinant plasmid pad - iASPP - RNAi was transfected into MCF - 7 cells. The expression of iASPP mRNA and protein was analyzed by RT-PCR and Western blotting, respectively. The cell apoptosis was detected by FCM, and then the MCF-7 cells were transplanted into nude mice to set up transplantation model. The expression of iASPP RNA and protein in transplanted neoplasm were determined by RT - PCR and Western blotting, the apoptosis index was detected by FCM at the same time. RESULTS： The results showed that the expression of iASPP descended in MCF -7 cells （mRNA 95.4% and protein 96. 8%, respectively, P 〈 0.01 ） and the apoptosis rate and necrosis rate of MCF - 7 cells increased （ P 〈 0. 01 ） after transfection. As treated with pAd - iASPP - RNAi, the expression of iASPP in transplantation tumor cells descended 87.4% （mRNA） and 89. 2% （protein）, respectively （P 〈0. 01 ）, and the apoptosis rate and necrosis rate increased accordingly （P 〈 0. 01, P 〈 0. 05）. CONCLUSION： The inhibition of iASPP may resume the ability of p53 to induce apoptosis in breast cancer cells which is able to express wild type p53.