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目的观察黑色素瘤分化相关基因7(MDA-7/IL-24)基因对人肝癌细胞Hep3B和正常的肝细胞L02的作用,并且探讨其该作用机制。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HeP3B,逆转录-聚合酶链式反应(RT-PCR)和ELISA方法观察MDA-7/IL-24基因的表达,噻唑蓝染色法(MTT)观察MDA-7/IL-24对肝癌细胞的生长抑制,Hoechst染色和Annexin-V和PI双染后流式细胞仪检测二种细胞的凋亡,利用PI染色后流式细胞仪检测细胞周期,RT-PCR方法检测bcl-2的表达变化。结果Ad.mda-7能介导外源基因MDA-7/IL-24在肝癌细胞株Hep3B和正常细胞L02中高效表达,细胞培养上清液中MDA-7/IL-24蛋白的表达(Hep3B:L02分别为790:810ng/L)。MDA-7/IL-24能明显抑制肝癌细胞的生长(抑制率分别是83%和1.2%),能促进肝癌细胞的凋亡(58%:2.2%),阻滞肝癌细胞在G_2/M期(48.29%:7.95%)。而对正常的肝细胞没有促凋亡和增殖阻滞作用;能明显的抑制Hep3B的凋亡抑制基因bcl-2的表达。结论Ad.mda-7能介导MDA-7/IL-24基因在人肝癌细胞中高效表达,选择性的杀伤肝癌细胞Hep3B,促进细胞增殖阻滞,其机制是通过抑制bcl-2的表达诱导肿瘤细胞凋亡。
Objective To observe the effect of MDA-7 / IL-24 gene on human hepatocellular carcinoma cell line Hep3B and normal liver cell line L02, and to explore its possible mechanism. Methods The adenovirus Ad.mda-7 carrying human MDA-7 / IL-24 gene was transfected into human normal hepatocytes L02 and Hep3B cells by reverse transcriptase-polymerase chain reaction (RT-PCR) 7 / IL-24 gene expression, MTT assay MDA-7 / IL-24 on the growth inhibition of liver cancer cells, Hoechst staining and Annexin-V and PI double staining by flow cytometry two kinds of cells The cell cycle was detected by flow cytometry after PI staining. The expression of bcl-2 was detected by RT-PCR. Results Ad.mda-7 could induce the expression of MDA-7 / IL-24 protein in hepatocellular carcinoma cell line Hep3B and normal cell L02, and the expression of MDA-7 / IL-24 protein in cell culture supernatant : L02 respectively 790: 810ng / L). MDA-7 / IL-24 can significantly inhibit the growth of hepatocellular carcinoma cells (the inhibition rate is 83% and 1.2% respectively), and promote the apoptosis of hepatocellular carcinoma cells (58% vs 2.2% (48.29%: 7.95%). However, there was no apoptosis-promoting and proliferation-arresting effect on normal hepatocytes. The expression of bcl-2, an inhibitor of Hep3B apoptosis, was significantly inhibited. Conclusion Ad.mda-7 can induce the high expression of MDA-7 / IL-24 gene in human hepatocellular carcinoma cells and selectively kill Hep3B hepatocarcinoma cells and promote cell proliferation arrest by inducing the expression of bcl-2 Tumor cell apoptosis.