人外周血单核细胞体外诱导树突状细胞及鉴定

Inducing of Dendritic Cells from Human Peripheral Blood Monocytes and Its Identification

目的 :建立从人外周血单核细胞体外诱导培养树突状细胞 ( dendritic cell,DC)的方法。方法 :分离人外周血单个核细胞 ,以细胞因子粒细胞 -巨噬细胞集落刺激因子 ( GM- CSF)、白细胞介素 - 4 ( IL- 4 )、肿瘤坏死因子 α( TNF- α)诱导培养获得 DC,电镜及共聚焦显微镜观察其形态 ,流式细胞术检测细胞表面抗原 ,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 :从正常外周血分离得到的单核细胞 ,体外经重组人 GM- CSF、IL - 4、TNF-α的共同诱导培养 ,得到大量成熟 DC。形态学观察可见典型 DC特征 ;荧光激活细胞分离器 ( FACS)检测表明 ,诱导的 DC高表达 HL A- DR( 96 % )、CD83( 94.2 % )、CDla( 94.2 % )分子 ,同时也高表达 CD40 ( 97.7% )、CD80 ( 98.2 % )分子。同种混合淋巴细胞反应显示 ,诱导的 DC具有很强的激发同种 T细胞增殖的能力。结论 :人外周血单核细胞体外经细胞因子贯序诱导培养 ,可以生成大量功能成熟的 DC,为进一步开展 DC的基础研究和临床应用提供了可能。

Objective:To estabilish the method of inducing dend ri tic cell from human peripheral blood minocytes in vitro. Methods:Monocytes were isolated from normal human peripheral blood mononuclear cells (P BMC s) and cultured with the cytokines (GM-CSF, IL-4, TNF-α). The morpholog y was observed by electron microscope and confocal analysis. The surface antigen of the induced cells were analysed by fluorescence-activated cell sorting(FAC S). T cell proliferating activity was determined by allo-MLR (mixed lymphocyte reaction) in vitro. Results:Large amount of mature DCs could be ob tained from monocytes by culture in presence of GM-CSF?IL-4 and TNF-α. DCs expressed high level of HLA-DR, CD83, CD1a, CD40, CD80, and were potent to stim ulate the proliferation of allo-T cells. Conclusion:Large amount of m ature DCs could be generated in vitro by culture of human peripheral blood m onocytes with the cytokines.