脐血CD_(34)^+细胞和外周血单核细胞两种来源的树突状细胞特性的比较研究

Studies on the properties of dendritic cells from cord blood CD_34^+ cells and peripheral blood monocytes

目的 体外大量扩增和纯化具有典型表型、形态和功能的树突状细胞（ＤＣ），以进行相关基础研究和临床应用。方法 采用免疫磁珠法分离脐血ＣＤ３４ ＋ 细胞及外周血去Ｂ、去Ｔ淋巴细胞的单个核细胞（ 单核细胞） ，然后以ＧＭＣＳＦ、ＩＬ４、ＴＮＦα、Ｆｌｔ３ 配基（ＦＬ）、ＳＣＦ等不同的细胞因子配伍，分别诱生ＤＣ，通过流式细胞仪、电镜、光镜分析其特性，同时检测其刺激同种Ｔ细胞增殖的能力。结果 脐血与外周血诱生ＤＣ的方案不同，由脐血ＣＤ３４＋ 细胞诱导ＤＣ 时，ＧＭＣＳＦ＋ ＴＮＦα＋ ＳＣＦ＋ ＦＬ 组合可使ＣＤ１ａ＋ 细胞比例增至（２７ ．１８ ±１－５６）％ ，明显高于单独应用ＧＭＣＳＦ组［（０．６５±０ ．３８）％ ］ 。外周血单核细胞诱导的ＤＣ，则ＧＭＣＳＦ＋ 高剂量ＩＬ４（１０００ Ｕ／ｍｌ） 组合效率最高，诱生的ＣＤ１ａ ＋ 细胞可达（２１ ．８０ ±０－３２） ％ 。两种来源的ＤＣ在表型及形态上差异无显著性，两者同样具有刺激同种异体淋巴细胞增殖的能力。结论 从脐血和外周血均可诱生ＤＣ，根据应用目的的不同对其进行选择，这为ＤＣ用于临床治疗选择不同细胞来源提供了实验基础。

Objective Ex vivo expansion and purification of dendritic cells(DC) with typical phenotype, morphology and function are critical to the further studies on DC and their clinical application. Methods CD 34 + cells were isolated from umbilical cord blood by using a high gradient magnetic cell sorting system (MACS), and peripheral blood mononuclear cells (PBMC) were depleted of T and B cells by mixing them with T/CD 2 and B/CD 19 magnetic beads. DC were induced and expanded with different combinations of cytokines. They were identified for their properties by FACS, electronic microscopy, microscopy, and mixed lymphocyte reaction (MLR). Results Cultures of cord blood CD 34 + cells with the combination of GM CSF+TNF α+SCF+FL yielded (27.18±1.56)% CD 1a + cells, much higher than that (0.65±0.38)% with GM CSF alone. The combination of GM CSF and high dose IL 4(1?000?U/ml) was the most potential for expanding CD 1a + cells [(21.8±0.32)%] for PBMC. Both of the DC from different sources were similar in phenotype and morphology, and had the capacity to stimulate proliferation of allogeneic T lymphocytes. Conclusion Both of cord blood and peripheral blood are the source for generation of a large number of typical DC, and can be adopted for the application according to different purposes. These results lay foundation for the immunotherapy with DC.