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目的:研究肿瘤特异性凋亡蛋白基因(apoptin)在诱导人结肠癌细胞凋亡过程中的信号转导机制。方法:用含有apoptin基因的真核表达载体瞬间转染体外培养的人结肠癌细胞;采用逆转录-聚合酶链反应(RT-PCR)、DNA凝胶电泳、流式细胞术检测人结肠癌细胞的凋亡;Westernblot检测Bcl-2/Bax蛋白表达,SPSS13.0统计软件进行数据分析。结果:Apoptin基因瞬间转染的结肠癌细胞可出现典型的细胞凋亡所具有的DNA梯状带;流式细胞术检测发现,实验组细胞凋亡率〔(40.45±0.76)%〕明显高于正常对照组〔(3.25±0.16)%〕和载体对照组〔(3.55±0.12)%〕(P<0.01);Bax的活性升高,但bcl-2蛋白含量无明显变化。结论:Apoptin基因可通过激活Bax诱导结肠癌细胞凋亡。
Objective: To study the signal transduction mechanism of apoptin in inducing human colon cancer cell apoptosis. Methods: Human colon cancer cells were transiently transfected with the eukaryotic expression vector containing apoptin gene. The human colon cancer cells were detected by reverse transcription polymerase chain reaction (RT-PCR), DNA gel electrophoresis and flow cytometry The protein expression of Bcl-2 / Bax was detected by Western blot and SPSS 13.0 software was used for data analysis. Results: Apoptosis DNA ladder was observed in typical Apoptin transfected colon cancer cells. Flow cytometry showed that the apoptotic rate in the experimental group was (40.45 ± 0.76)%, which was significantly higher than that in the experimental group (3.25 ± 0.16)% in the normal control group and (3.55 ± 0.12)% in the vehicle control group (P <0.01). The activity of Bax increased but the content of bcl-2 protein did not change significantly. Conclusion: Apoptin gene can induce apoptosis of colon cancer cells by activating Bax.