树突状细胞瘤苗联合吉西他滨对肝癌干细胞的杀伤效应

Cytotoxic activity induced by dendritic cell vaccine plus gemcitabine to the hepatocellular carcinoma cancer stem cell

目的:探讨负载CD133+肝癌细胞抗原的树突状细胞(DC)联合吉西他滨(GEM)在体外对肝癌干细胞的杀伤效应。方法:取对数生长期的人肝癌细胞系Huh-7以CD133作为分子标志进行流式分选,得到肝癌干细胞。将人外周血单个核细胞(PBMC)在体外诱导分化为树突状细胞(DC)。DC负载Huh-7细胞和CD133+细胞抗原后与T细胞共育得到特异性细胞毒性T细胞,将CD133+细胞作为靶细胞进行细胞毒性试验。实验组按处理因素分为:GEM组,CD133+-CTL组,GEM+CD133+-CTL组,Huh7-CTL组和GEM+Huh7-CTL组。CCK-8法检测杀伤率,然后比较各组间差异。结果:对CD133+细胞的杀伤效应以GEM+CD133+-CTL组最强,与其他各组比较差异均有统计学意义(P<0.05)。单独就DC瘤苗杀伤率,CD133+-CTL组高于Huh7-CTL组(P<0.05)。结论:CD133+肝癌干细胞裂解产物致敏的DC瘤苗联合化疗药物可以有效杀伤肝癌干细胞,进而可能降低肝癌术后和肝癌肝移植后的转移和复发率。

Objective： To investigate the cytotoxicity of gemcitabine （GEM） combined with immunotherapy mediated by dendritic cells DCs） loaded with CD133＋ liver cancer cell antigens to hepatocellular carcinoma cancer stem cells （HCC CSCs） in vitro. Methods： The HCC CSCs were sorted from Huh-7 cells in logarithmic growth phase by flow cytometery via the CD133 molecular mark. Human peripheral blood mononuclear cells （PBMCs） were induced into dendritic cells in vitro.Then theDCs were sensitized by Huh-7 and CD133 cancer cell antigens. After antigen Ioading-DCs were co-cultured with T cells,specific cytotoxic T lymphocytes were obtained. CD133 cells were taken as target cells for cytotoxicity tests. HCC CSCs were divided into five groups according to treatment factors： GEM group, CD133-CTL group, GEM＋CD133＊-CTL group, Huh7-CTL group and GEM＋Huh7-CTL group. The cytotoxic effects were assessed by CCK-8 assay. Then variances were compared and analyzed. Results： The GEM＋CD133-CTL group have the strongest cytotoxic effect to target cells, when compared with other groups. And the differences were statistically significant P〈0.05）. In terms of the cytotoxic effect induced by the DC vaccine alone, the CD133＋-CTL group was better than Huh7-CTL group （P〈0.05）. Conclusion： GEM combined with immunotherapy mediated by DCs loaded with CD133 liver cancer cell antigen could kill the HCC CSCs effectively. So this therapy may reduce the metastasis and recurrence after hepatectomy and liver transplantation and prompt a new strategy for the treatment of hepatocellular carcinoma.