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目的 研究携带分化基因RA 53 8的重组腺病毒 (Ad RA53 8)对人肝癌细胞的抑制作用及机制。方法 Ad RA53 8感染人肝癌细胞系BEL 740 2和QSG 770 1 ,绘制生长曲线 ,克隆形成实验评价Ad RA53 8对肝癌细胞增殖的抑制作用 ,应用流式细胞术、DNA原位末端标记、梯形DNA研究细胞周期及细胞凋亡情况 ,逆转录 聚合酶链式反应 (RT PCR)和免疫印迹法 (Westernblot)检测相关基因的表达。结果 感染Ad RA53 8后 ,RA53 8在肝癌细胞内获得了表达 ,并且降低了c myc的表达。肿瘤细胞的增殖受到明显抑制 ,感染Ad RA53 85d后 ,BEL 740 2的活细胞数为(1 0 .0± 3 .7)× 1 0 4,对照组为 (1 72 .7± 1 1 .9)× 1 0 4(P <0 .0 0 1 ) ;QSG 770 1感染Ad RA53 83d后的活细胞数为 (2 3 .0± 4.3 )× 1 0 4,对照组为 (91 .7± 7.2 )× 1 0 4(P <0 .0 0 1 ) ;感染Ad RA53 8后BEL 740 2和QSG 770 1形成的克隆数分别为 1 3± 4和 1 5± 2 ,明显少于对照组的 1 88± 9和 1 75± 9(P <0 .0 0 1 ) ,而且还出现了细胞周期的阻滞和凋亡。结论 Ad RA53 8对肝癌细胞具有明显的抑制作用 ,是有前途的抗肿瘤基因药物
Objective To study the inhibitory effect and mechanism of recombinant adenovirus carrying the differentiation gene RA 53 8 (Ad RA53 8) on human hepatoma cells. METHODS: Ad RA53 8 was used to infect human hepatocellular carcinoma cell lines BEL 740 2 and QSG 770 1 , and the growth curve was drawn. The effect of Ad RA53 8 on the proliferation of hepatocellular carcinoma cells was evaluated by colony formation assay. Flow cytometry, in situ DNA end-labeling, ladder DNA were used. The cell cycle and apoptosis were studied. The expression of related genes was detected by reverse transcription polymerase chain reaction (RT PCR) and Western blot. Results After infection with Ad RA53 8 , RA53 8 was expressed in hepatocellular carcinoma cells and decreased the expression of c myc. The proliferation of tumor cells was significantly inhibited. After 85 days of infection with Ad RA53, the number of viable cells in BEL 740 2 was (1 0 . 0 ± 3 .7) × 1 0 4 and that in the control group was (1 22.7 ± 1 1 . 9 ) × 1 0 4 (P <0. 0 0 1 ); The number of viable cells after QSG 770 1 infection with Ad RA53 for 83 days was (2 3 .0 ± 4.3) × 1 0 4; for the control group (91.7 ± 7.2). The number of colonies formed by BEL 740 2 and QSG 770 1 after infection with Ad RA 8 8 were 1 3 ± 4 and 15 ± 2 , respectively, which was significantly less than that of the control group 1 (P <0. 0 0 1). 88 ± 9 and 175 ± 9 (P < 0.01), but also cell cycle arrest and apoptosis. Conclusion Ad RA53 8 has obvious inhibitory effect on hepatoma cells and it is a promising anti-tumor gene drug.