肝癌细胞对树突状细胞线粒体功能的影响-第三军医大学学报.DOC

肝癌细胞对树突状细胞线粒体功能的影响 曾 柱1，2，龙金华3 （550004 贵阳，贵阳医学院基础医学院生物技术教研室1；100083北京，北京大学基础医学院生物物理系血液流变学研究中心2；550004 贵阳，贵州省肿瘤医院头颈肿瘤科3） 【摘 要】 目的 探索肝癌细胞分泌的可溶性细胞因子营造的微环境对树突状细胞（Dendritic cells，DCs）线粒体功能状态的影响，以进一步理解肿瘤的免疫逃逸机制。方法 用免疫磁珠从人外周血分离CD14＋单核细胞，加入粒-巨噬细胞集落刺激因子（Recombinant human granulocyte-macrophage CSF, rhGM-CSF）、白介素4（Recombinant human interleukins 4, IL-4）将单核细胞诱导分化为未成熟DCs（immature DCs，imDCs），利用肿瘤坏死因子α（Recombinant human Tumor necrosis factor-α, rhTNF-α）将imDCs诱导为成熟DCs（mature DCs, mDCs），分别将imDCs和mDCs与肝癌细胞（Hepatocellular carcinoma cells, HCCs）在Transwell中共培养48 h，正常培养和撤生长因子培养的DCs作为对照，利用免疫荧光和MTT比色等方法研究HCCs对DCs的线粒体膜电位、酶活力和胞内钙离子浓度的影响。结果 与HCCs共培养后，流式细胞仪的检测结果显示DCs的线粒体膜电位显著下降（P0.05），胞内钙离子浓度明显增加（P0.01, P0.05），MTT比色法的检测结果显示DCs酶活力受到了显著的抑制（P0.001）。结论 HCCs可能能够通过抑制DCs的线粒体功能来损伤其免疫功能。 【关键词】 树突状细胞；肝癌细胞；线粒体功能 【中图分类号】 Q6-3 【文献标志码】 A 【基金项目】 贵州省省长基金（黔省专字2009-79）；贵州省自然科学基金（黔省合J字2008-2274） 【通信作者】曾 柱，E-mail：zengzhu100@，电话：0851-6909969 The Effect of Hepatocellular Carcinoma Cells on the Mitochondrial Function of Dendritic Cells Zeng Zhu1,2, Long Jinhua3 （1Department of Biotechnology, School of Basic Medical Science, Guiyang Medical College, Guiyang, Guizhou, 550004；2Hemorheological Research Center, Department of Biophysics, School of Basic Medical Science, Peking University, Beijing, 100083,；3Department of Head and Neck Tumor, Tumor Hospital of Guizhou, Guiyang, Guizhou, 550004） 【Abstract】Objective This study focused on the influence of microenvironment created by dissoluble cytokines derived from hepatocellular carcinoma cells (HCCs) on the functional status of mitochondrions in DCs in order to further understand the mechanisms of tumor immune escape. Methods The CD14+ monocytes，which were isolated with immune magnetic beads from fresh peripheral blood of healthy human , were then cultured in RPMI 1640/10% FBS supplemented with 100 ng/ml rhGM-CSF and 100 ng/ml rhIL-4 for 7 days to develop into immature DCs (imDCs). Maturation was induced by addition of 20 ng/ml rhTNF-α to imDCs for another 3 days of