Ad．mda-7高选择性杀伤不同转移潜能肝癌细胞的研究

Selective killing effects of Ad. mda-7 on hepatocellular carcinoma cell line with different metastatic potentials in vitro

目的利用携带人MDA-7／IL-24基因的复制缺陷型腺病毒（Ad．mda7）作为载体，感染不同转移潜能的肝癌细胞MHCC97H，MHCC97L，Hep3B和正常的肝细胞L02，观察该基因对肝癌的选择性杀伤作用和增殖抑制作用。方法将携带人MDA-7／IL-24基因的腺病毒Ad．mda-7感染正常肝细胞L02和肝癌细胞MHCC97H，MHCC97L和Hep3B，通过RT-PCR和ELISA方法观察MDA7基因的表达，通过四甲基偶氮哩蓝染色法（MTT）观察该基因对肝癌细胞的生长抑制，Hoechst染色和流式细胞仪观察MDA-7对肝癌细胞的杀伤作用，流式细胞仪检测细胞周期变化。结果RT-PCR提示复制缺陷型腺病毒能介导外源基因MDA-7／IL-24在肝癌细胞株MHCC97H，MHCC97L，Hep3B和正常细胞L02中高效表达；ELISA方法检测提示细胞培养上清液中MDA-7／IL-24蛋白的表达，而且随着感染时间延长表达量升高；MTT实验结果表明MDA7能明显抑制肝癌细胞的生长，Hoechst染色和流式细胞仪提示MDA7能促进肝癌细的凋亡；细胞周期检测提示肝癌细胞大量被阻滞在G2／M期，正常细胞没有增殖阻滞作用。各项结果提示MDA7对正常的肝细胞没有促进凋亡和增殖阻滞的作用。结论Ad．mda-7选择性的杀伤肝癌细胞，促进细胞增殖阻滞及诱导肿瘤细胞凋亡与肝癌细胞的转移潜能无关。

Objective To investigate the effects of MDA/IL-24 on hepatocellular carcinoma （HCC） cell lines MHCC97H, MHCC97L and Hep3B with different metastatic potentials and normal liver cell line L02 in vitro. Methods The 3 HCC cell lines and normal L02 were infected by Ad. mda- 7 and the mRNA expression of MDA 7/IL-24 in these cell lines was determined by RT-PCR. Protein expression was confirmed by ELISA. MTT assay and flow cytometry were used to study tumor cell proliferation and cell cycle in vitro. Heochst and Annexin-V and PI staining were performed to indicate the apoptotic effect. Results It was confirmed by RTPCR and ELISA that the exogenous MDA-7/ IL-24 gene expressed the 3 HCC cell lines and normal L02. MTT and cycle test showed that Ad. mda- 7 could induce HCC cell line growth suppression and block cancer cells in G2/M but not in L02. Hoechst staining and flow cytometry revealed that MDA-7/IL-24 could induce apoptosis in HCC cell lines but not in normal liver cells. Conclusions MDA-7/IL-24 can selectively induce growth suppression and apoptosis in HCC cell lines and this induction is independent of different metastatic potentials.