脐血来源树突状细胞的体外诱导及扩增

Differentiation and Increase of Dendritic Cells from Umbilical Cord Blood in vitro

本研究的目的是分析脐血的细胞组成 ,研究加入细胞因子培养前后脐血树突状细胞的变化 ,探索体外诱导、扩增树突状细胞的方法并进行表型鉴定。选择正常成人外周血 9份 ,脐血 12份 ,分离单个核细胞。在脐血单个核细胞中加入细胞因子GM CSF、IL 3、SCF和EPO ,培养 4周。应用流式细胞仪和CD4、CD8、CD19、CD34、CD38、CD1a、CD11c及CDw12 3单克隆抗体测定正常成人外周血、培养前后 1,2 ,3,4周脐血细胞表面抗原及树突状细胞情况。结果表明 :正常成人外周血CD34+ 细胞 0 .0 2× 10 5 ml,CD1a+ 细胞 0 .0 1× 10 5 ml,CD11c+ 细胞 4 .32×10 5 ml,CD83+ 细胞 1.31× 10 5 ml,CDw12 3+ 细胞 1.4 1× 10 5 ml。新鲜脐血中CD34+ 细胞 0 .2 2× 10 5 ml,CD1a+ 细胞 0 .2 7× 10 5 ml,CD11c+ 细胞 5 .87× 10 5 ml,CD83+ 细胞 1.94× 10 5 ml,CDw12 3+ 细胞 2 .73× 10 5 ml。加入细胞因子GM CSF ,IL 3,SCF ,EPO后培养 1- 4周的脐血单个核细胞分化为CD1a+ ,CD11c+ ,CD83+ ,CDw12 3+ 树突状细胞 ,在培养的 2 - 4周 ,脐血树突状细胞数量明显增多 ,此后逐渐减少。通过培养 ,树突状细胞数量增加 ,CD1a+ 细胞达 11.0 2× 10 5 ml,CD11c+ 细胞 2 8.2 4× 10 5 ml,CD83+ 细胞 10 .5 7× 10 5 ml,CDw12 3+ 细胞 18.7× 10

The aims of this study were to analyze the composition of umbilical cord blood c ells (UCBC), to examine the characterist ics of dentritic cells (DC) before and after culture, to search the method of di fferentiation and increase of DC in vitro and to appraise surface antigen from UCBC. Twelve units of umbilical cord blood were collected from May 2002 to September 2002. Peripheral blood mononuclear cells of 9 cases were collected from healthy adult donors. The nature of UCBC wa s freshly determined and then UCBC were cultured for 1,2,3 and 4 weeks with granu locyte- monocyte colony-stimulating factor (GM-CSF), interleukin 3 (IL-3), r ecombinant human stem cell factor (SCF) and EPO. Method of flow cytometry was us ed to d etermine the number of DC and cell surface antigens before and after culture by using monoclonal antibodies. The monoclonal antibodies included CD4, CD8, CD19, CD34, CD38, CD83, CD1a, CD11c and CDw123. The results showed that amounts of C D34+ progenitor s in peripheral blood cells were 0.02×105/ml, and amounts of CD34+ progeni tors in human UCBC were 0.22×105/ml. UCBC cultured for 1, 2, 3 and 4 weeks wi th GM-CSF, IL-3, EPO and SCF were shown to differentiate into CD1a+ CD11c+ CD83+ CDw123+ D C. Numbers of DC from UCBC remarkably generated in 2-4 weeks and then decrease d in number. By culture with cytokines DC increased up to (10.6-28.2)×105/ml in actual numbers. It is concluded that the mononuclear cells of UCB are able to differentiate into CD1a+, CD83+, CD11c+ and CDw123+ DC when UCBC are cu ltured with proper cytokines of GM-CSF, SCF, EPO and IL-3 for 2-4 weeks. These DCs as antigen presenting cells are possibly effective in cancer immunotherapy.