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目的研究人甲胎蛋白(AFP)启动子控制表达HIV-1 vpr基因的重组腺病毒对AFP(+)肝癌细胞G2期阻滞和细胞凋亡的作用,评估以该策略利用vpr进行肿瘤特异性基因治疗的可行性。方法将以AFP启动子控制HIV-1vpr表达的重组腺病毒rvAdAFP-vpr和空白载体rvAd-null分别感染AFP(+)的肝癌细胞BEL-7402和AFP(-)的肝癌细胞SMMC-7721,利用流式细胞仪检测Vpr的特异性表达和细胞周期分布,用细胞荧光染色和线粒体膜电位检测等方法观察细胞凋亡。结果与对照病毒rvAd-null相比,重组腺病毒rvAdAFP-vpr只在AFP(+)肝癌细胞中表达HIV-1Vpr,并诱导肝癌细胞的细胞周期G2期阻滞和细胞凋亡,而在AFP(-)肝癌细胞中不明显表达Vpr,不能显著诱导肝癌细胞G2期阻滞和细胞凋亡。结论重组腺病毒rvAdAFP-vpr对于Vpr的表达是AFP表达特异性的,可有效诱导肝癌细胞G2期阻滞和细胞凋亡,有望用于AFP(+)肝癌的基因治疗研究。
Objective To investigate the effect of human AFP promoter on the G2 arrest and apoptosis of AFP (+) hepatoma cells under the control of recombinant adenovirus expressing HIV-1 vpr gene. We evaluated the tumor-specific Feasibility of gene therapy. Methods AFP (+) hepatoma BEL-7402 and AFP (-) hepatocellular carcinoma cells SMMC-7721 were infected by recombinant adenovirus rvAdAFP-vpr and rvAd-null expressing HIV-1vpr under the control of AFP promoter. Cytometry was used to detect the expression of Vpr and cell cycle distribution. Cell apoptosis was observed by fluorescence staining and mitochondrial membrane potential assay. Results Compared with control virus rvAd-null, recombinant adenovirus rvAdAFP-vpr expressed HIV-1Vpr only in AFP (+) hepatoma cells and induced cell cycle G2 arrest and apoptosis in hepatoma cells. However, -) hepatoma cells did not express Vpr significantly, can not significantly induce G2 arrest and apoptosis in hepatoma cells. Conclusion The recombinant adenovirus rvAdAFP-vpr is specific for AFP expression and can induce G2 arrest and apoptosis in hepatocellular carcinoma cells. It may be used for the gene therapy of AFP (+) hepatocellular carcinoma.