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目的探讨从脐血CD34+造血细胞诱导DC2的方法及CD40分子在DC2分化诱导中的作用。方法运用磁珠从脐血中分离出CD34+造血干细胞,以rhIL3(10ngmL)、rhFlt3L(100ngmL)和rhGMCSF(100ngmL)诱导其向DC2分化,采用流式细胞仪分析DC2表型,并观察抗人CD40单克隆抗体诱导DC2分化成熟的作用。结果人脐血CD34+造血细胞在rhIL3、rhFlt3L和rhGMCSF联合诱导培养12d,获得具有DC2样(淋巴样DC)形态的细胞,然后分别加入rhIL3+抗人CD40mAb(Ⅰ组)或rhIL3+TNFα(Ⅱ组)诱导DC2的分化成熟,流式细胞仪分析细胞表型,发现Ⅰ组和Ⅱ组Lineage-(CD3、CD14、CD16、CD19、CD56)CD123+细胞的HLADR、CD83、CD86、CD80表达率分别为88.78%、37.38%、32.83%和99.08%;78.87%、32.29%、29.57%和98.86%。结论体外联合多种细胞因子从CD34+造血细胞成功地诱导出富集DC2表型的细胞,抗人CD40mAb可以促进DC2的分化成熟。
Objective To investigate the method of inducing DC2 from cord blood CD34 + hematopoietic cells and the role of CD40 in induction of DC2 differentiation. Methods CD34 + hematopoietic stem cells were isolated from umbilical cord blood by magnetic beads. The DCs were induced to differentiate into DC2 by rhIL3 (10ngmL), rhFlt3L (100ngmL) and rhGMCSF (100ngmL). The DC2 phenotype was analyzed by flow cytometry and the anti-human CD40 Monoclonal antibody induces DC2 differentiation and maturation. Results Human umbilical cord blood CD34 + hematopoietic cells were induced by rhIL3, rhFlt3L and rhGMCSF for 12 days to obtain DC2-like (lymphoid DC) cells, and then rhIL3 + anti-human CD40 mAb (group Ⅰ) or rhIL3 + TNFα (group Ⅱ) The differentiation and maturation of DC2 were induced and the cell phenotypes were analyzed by flow cytometry. The expression rates of HLADR, CD83, CD86 and CD80 of Lineage- (CD3, CD14, CD16, CD19, CD56) CD123 + cells in group I and group II were 88.78% , 37.38%, 32.83% and 99.08%; 78.87%, 32.29%, 29.57% and 98.86% respectively. Conclusion In vitro and in combination with various cytokines, cells enriched in DC2 phenotype were successfully induced from CD34 + hematopoietic cells. Anti-human CD40 mAb could promote the differentiation and maturation of DC2.