鸡尾酒法体外培养人外周血来源的树突状细胞  

In vitro generation of mature dendritic cells from peripheral blood mononuclear cell by cocktail of cytokines

作　　者：李丽[1] 刘宝瑞[1] 钱晓萍[1] 

机构地区：[1]南京大学医学院附属鼓楼医院肿瘤中心,210008

出　　处：《江苏医药》2008年第1期53-55,I0005,共4页Jiangsu Medical Journal

基　　金：国家自然科学基金(30471701)

摘　　要：目的探讨从健康人外周血中体外诱导培养成熟树突状细胞(DC)的方法。方法用密度梯度离心法分离人外周血单个核细胞(PBMC),再用贴壁法获取单核细胞后加入人粒细胞-巨噬细胞集落刺激因子(GM-CSF)和IL-4,培养6d后分组:Ⅰ组为对照组,仅含上述细胞因子;Ⅱ组加入多聚次黄嘌呤胞嘧啶核苷酸(PolyI∶C);Ⅲ组加入钙离子载体(CI)A23187;Ⅳ组加入PolyI∶C和CIA23187。第8天收获细胞,显微镜下观察形态,流式细胞术检测DC表型,体外同种混合淋巴细胞反应检测DC刺激T细胞的增殖活性。结果Ⅳ组细胞形态学观察可见典型DC特征,荧光激活细胞分离器(FACS)检测高表达CD83、CD80、CD86和CD40;混合淋巴细胞反应(MLR)表明Ⅳ组细胞具有较强的刺激T细胞增殖能力。结论PBMC体外经过细胞因子序贯、联合诱导培养能够获得大量功能成熟的DC。Objective To explore the generating mature dendritic ceils （DC） from human peripheral blood minocytes in vitro. Methods Monocytes were isolated from normal human peripheral blood mononuclear cells （PBMCs）and were separated into four groups after cultured with GM-CSF and IL-4 for 6 days. The cells in group A were cultured with GM-CSF and IL-4 only as control group. The cells in group B were cultured with additional polyriboinosinic polyribocytidylic acid （Poly（Ⅰ ： C）） ,in group C with additional calcium Ionophore （CI）,and in group D with additional Poly（Ⅰ ： C） and CI. The morphology was observed by microscope. The surface antigens of the induced cells were analyzed by fluorescence-activated cell sorting （FACS）. T cell proliferating activity was determined by allo-MLR （mixed lymphocyte reaction） in vitro. Results Large amount of mature DCs could be obtained from monocytes by culture in presence of GM-CSF,IL-4,Poly（Ⅰ ： C） and CI. DCs expressed high level of CD83, CD80, CD86, CD40, and were potent to stimulate the proliferation of allo- lymphocytes. Conclusion Large amount of mature DCs could be generated in vitro by culturing human peripheral blood monocytes with the cocktail cytokines.

关 键 词：树突状细胞 多聚次黄嘌呤胞嘧啶核苷酸 钙离子载体A23187 

分 类 号：R73[医药卫生—肿瘤]