Apoptin对宫颈癌Hela细胞的抑制效应

Anti-tumor effects of Apoptin gene on uterine cervix cancer cells

目的:探讨Apoptin基因对人宫颈癌Hela细胞的抑制作用和作用方式。方法:应用脂质体转染法将重组质粒pVAX1-Apoptin和载体质粒pVAX1分别体外转染Hela细胞,作为重组质粒pVAX1-Apoptin组和载体质粒pVAX1组,同时设空白细胞对照组。应用Western blotting检测Apoptin基因在Hela细胞中的表达;运用噻唑兰(MTT)分析法检测重组质粒对Hela肿瘤细胞的抑制作用;罗丹明123和DCFA染色测定线粒体跨膜电位(ΔΨm)和活性氧水平变化;底物染色反应检测caspase-3活性。结果:pVAX1-Apoptin组转染48h后,Hela细胞的抑制率为69.28%,明显高于对照组(0.74%)和pVAX1组(10.11%)(P<0.01)。与对照组比较,pVAX1-Apoptin组线粒体ΔΨm下降(P<0.01),细胞内活性氧水平升高(P<0.05),caspase-3活性增强(P<0.01)。结论:Apoptin可通过诱导Hela细胞凋亡,进而特异性杀伤Hela肿瘤细胞。

Objective cancer cells. Methods To investigate the anti-tumor effects and mode action of Apoptin on human uterine cervix Recombinant plasmid pVAX1-Apoptin and pVAX1 were transfected into Hela cells by application of liposome in vitro and used as pVAX1-Apoptin group and pVAX1 group, meanwhile control group （without cells） was set up. The expression of Apoptin in Hela cells was detected by Western blotting. Anti-tumor effect on Hela cells was measured through methyl thiazolyl tetrazolium （MTT） assay. The alteration of mitochondrial transmembrane potential and ROS level of the cells were detected by flow cytometry （FCM） with rhodamine 123 and DCFA staining. The activation of caspase-3 was assayed by its substrate color reaction. Results The inhibitory rate in pVAX1-Apoptin group 48 h after transfection （69.28%） was higher than those in control group （0.74%） and pVAX1 group （10.11%） . Compared with control group , the mitochondrial transmembrane potential was decreased （P〈0. 01）, ROS level of the cells was increased （P〈0.05）, and caspase-3 activity was increased （P〈0.01） in pVAX1-Apoptin group. Conclusion Anti-tumor effects of Apoptin on Hela cells may be resulted from the apoptosis-inducing function of Apoptin.