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树突状细胞(DC)是一群专职的抗原递呈细胞(APC),DC负载肿瘤多肽作为疫苗治疗肿瘤已经应用于临床.为此,我们利用人脐带/外周血,建立了体外扩增DC细胞的方法,并研究其特性,以期为临床应用提供物质基础.利用人脐带肝素抗凝血50～100ml,常规密度梯度离心,获取单个核细胞,贴壁2小时,用RPMI 1640+10%胎牛血清培养基培养,加人人粒/巨噬细胞集落刺激因子(rGM-CSF)300ng/ml,人肿瘤坏死因子(rTNF-α)1nd/ml培养,每隔一天半量换液.6～7天,细胞出现少量粘附集聚现象,10天左右,细胞呈梭形或不规则形.经电子显微镜分析,细胞表面有许多树突状突起,胞内具有丰富的线粒体.免疫表型分析证实,此群细胞高表达CDla、HLA-DR、B_7抗原,低表达或不表达CD14、CD22、CD4等分化抗原.经混合淋巴细胞反应(MLR),此群细胞能介导很强的T淋巴细胞增值作用(~3H掺入法).经10～14天培养,能获取1～2×10~7个DC细胞.人
Dendritic cells (DCs) are a group of dedicated antigen-presenting cells (APC), and DC-loaded tumor peptides have been used in clinic as vaccines for clinical treatment. To do this, we established in vitro expanded DCs using human umbilical cord blood / peripheral blood , And its characteristics were studied in order to provide a material basis for clinical application.Using umbilical cord blood anticoagulant 50 ~ 100ml, conventional density gradient centrifugation, mononuclear cells, adherent 2 hours, with RPMI 1640 + 10% fetal calf Serum culture medium was incubated with 300ng / ml of human granulocyte / macrophage colony stimulating factor (rGM-CSF) and 1nd / ml of human tumor necrosis factor (rTNF-α) , The cells appeared a small amount of agglutination phenomenon, about 10 days, the cells were spindle or irregular shape.Analysis by electron microscopy, there are many dendritic protrusions on the cell surface, the cell is rich in mitochondria.Immune phenotype analysis confirmed that this The population of cells express CDla, HLA-DR and B7 antigen at high level, low expression or no differentiation antigen such as CD14, CD22 and CD4. The mixed lymphocyte reaction (MLR) can induce strong T lymphocyte proliferation (~ 3H incorporation method) .After 10 ~ 14 days, 1 ~ 2 × 10 ~ 7 DC fine can be obtained . People