论文部分内容阅读
目的：建立体外诱导和扩增人外周血树突状细胞（ＤＣ） 的方法，并分析其生物学特性。方法：造血动员后外周血单个核细胞经贴壁去除悬浮细胞，加入细胞因子（ＩＬ－４、ＧＭ－ ＣＳＦ和ＴＮＦ－α）培养８ 天。用流式细胞仪分析细胞的表型，经ＥＬＩＳＡ法测定其培养上清中的ＩＬ－ １２ ，并将诱导的树突状细胞与脐带血原始Ｔ（ｎａｉｖｅ Ｔ） 细胞混合培养，用３Ｈ－ ＴｄＲ掺入法测定细胞的增殖指数。结果：贴壁的造血动员后外周血单个核细胞在体外经细胞因子诱导培养后，高表达人树突状细胞分化抗原ＣＤ１ａ（８９ ．１％ ）、ＣＤ４０（９９ ．８％ ）、ＣＤ８０（９５ ．１％ ）、ＣＤ８３（４５ ．７％ ）、ＨＬＡ－ ＤＲ（９７．６％ ），同时所获的ＤＣ能分泌ＩＬ－１２ 和可有效地激活脐带血原始Ｔ细胞并促其增殖（ＳＩ＝ ６．９２） 。结论：采用造血动员的外周血经体外贴壁去除非粘附细胞，再经过细胞因子序贯培养，无需纯化ＣＤ３４ ＋ 细胞，能获得高纯度（ ＞８０％ ）和具有功能性的ＤＣ细胞。
Objective: To establish a method for inducing and expanding dendritic cells (DCs) from human peripheral blood in vitro and to analyze their biological characteristics. Methods: Peripheral blood mononuclear cells (PBMCs) were removed by adherent cells after hematopoietic mobilization and cultured with cytokines (IL-4, GM-CSF and TNF-α) for 8 days. The phenotypes of the cells were analyzed by flow cytometry. IL-12 in the culture supernatant was measured by ELISA. The induced dendritic cells were mixed with umbilical cord blood naive T cells and stained with 3H-TdR Adulteration assay was used to determine the cell proliferation index. Results: Peripheral blood mononuclear cells were highly expressed in peripheral blood mononuclear cells induced by cytokines in vitro after differentiation. CD1a (89.1%), CD40 (99.8%) and CD80 CD83 (45.7%), HLA-DR (97.6%). Meanwhile, DCs secreted IL-12 and effectively activated cord blood progenitor T cells and promoted their proliferation (SI = 6.92). Conclusion: The hematopoietic mobilized peripheral blood can be used to remove non-adherent cells by in vitro adherent cells. After sequential cytokines culturing, high purity (> 80%) and functional DC cells can be obtained without purification of CD34 + cells.