【摘要】 目的观察MDA-7/IL-24基因对肝癌的选择性杀伤作用,为肝癌的基因治疗提供理论基础。方法将携带人MDA-7/IL-24基因的腺病毒Ad.mda-7感染人正常肝细胞L02和肝癌细胞HepG2;用RT-PCR法观察MDA-7/IL-24基因的表达;ELISA方法检测细胞培养上清液中MDA-7/IL-24蛋白的浓度;4甲基偶氮唑蓝染色法(MTT)及Hoechst染色观察MDA-7/IL-24对肝癌细胞的生长抑制和杀伤作用;Annexin-V和PI双染后流式细胞仪检测2种细胞的凋亡;用流式细胞仪检测细胞周期。结果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HepG2和正常细胞L02中的高效表达;细胞培养上清液中有MDA-7/IL-24蛋白的表达;MDA-7/IL-24能明显抑制肝癌细胞生长并可促进肝癌细胞的凋亡;MDA-7/IL-24阻滞肝癌细胞于G2/M期,能选择性杀伤肝癌细胞而对正常的肝细胞无阻滞作用和毒性作用。结论复制缺陷型重组腺病毒载体Ad.mda-7能介导MDA-7/IL-24基因在人肝癌细胞中高效表达,促使细胞增殖阻滞及诱导肿瘤细胞凋亡,选择性地杀伤肝癌细胞HepG2,而对正常肝细胞L02无任何毒性作用。更多还原

【Abstract】 Objective To investigate the selective killing effect of MDA/IL-24 on human hepatocellular carcinoma line HepG2 in vitro and provide a theoretical basis for gene therapy of hepatocellular carcinoma.Methods The MDA-7/IL-24 gene was transfected into human hepatocullular carcinoma cell line HepG2 and normal liver cell line L02 with a replication-incompetent adenovirus vector.The mRNA and protein expression of MDA7/IL-24 in HepG2 and L02 cells was examined by RT-PCR and ELISA assay respectively.MTT assay and flow cytometry were used to study tumor cell proliferation and cell cycle in vitro.Hoechst and Annexin-V and PI staining were studied to indicate the apoptosis.Results RT-PCR confirmed that the exogenous MDA-7/IL-24 gene was expressed in HepG2 and L02 cells.The protein product was confirmed by assay of the supernatant with ELISA.MTT and apoptosis test indicated MDA-7/IL-24 induced growth suppression and cell apoptosis of the HepG2 cell in vitro but not in cell line L02,and cell cycle test revealed MDA-7/IL-24 could block HepG2 cell in G2/M but not in L02.Conclusions MDA-7/IL-24 selectively induces growth suppression and apoptosis in lines HepG2 in vitro but not in L02 cell,which indicates that adenovirus mediated MDA-7/IL-24 can be an excellent tool for gene therapy in hepatocellular carcinoma.更多还原