摘要
目的:探讨从健康人外周血中体外诱导培养出成熟树突状细胞(dendritic cell,DC)的方法。方法:用密度梯度离心法分离人外周单个核细胞(PBMC),再用贴壁法获取单核细胞后加入GM-CSF和IL-4,培养5天后分5组:第1组为对照组,仅含有上述细胞因子;第2组,加入多聚次黄嘌呤胞嘧啶核苷酸[Poly(I:C)];第3组,加入钙离子载体(A23187);第4组,加入Poly(I:C)和CI;第5组,加入TNF-α。显微镜下观察培养细胞形态,流式细胞术检测DC表型,体外同种混合淋巴细胞反应(MLR)检测DC刺激T细胞的增殖活性。结果:第四组细胞具有典型的树突状细胞的特征,且高表达CD83、CD80、CD86和CD40分子,具有更强的刺激T细胞增殖能力。结论:经过组合细胞因子诱导能够培养出PBMC来源的DC,且经CI和Poly(I:C)进一步诱导后获得更成熟和功能更强DC。
Objective:To explore the generation of mature dendritic cells (DCs) induced from human peripheral blood monocytes in vitro. Methods: Monocytes were isolated from normal human peripheral blood mononuclear cells (PBMCs)and were separated into five groups after cultured with GM-CSF and IL-4 5 days later. The control group were added only GM-CSF and IL-4. One group were added polyriboinosinic polyribocytidylic acid[Poly(I:C)], one group were added calcium ionophore(CI),one group were added Poly (I:C) and CI, and the other group were added TNF-α The morphology of cells was observed by microscope. The surface antigens of the induced cells were analyzed by fluorescence-activated cell sorting (FACS). T cell proliferating activity was determined by allo- MLR (mixed lymphocyte reaction) in vitro. Results:Large amount of mature DCs could be obtained from monocytes by culture in presence of GM-CSF,IL-4,Poly (I:C) and CI. DC expressed high level of CD83, CD80, CD86 and CD40, and were potent to stimulate the proliferation of allo-lymphocytes. Conclusion:The phenotypes and functions of DC generated from PBMC by the treatment of cytokine combination can be improved by calcium ionophore and Poly(I:C).
出处
《南京医科大学学报(自然科学版)》
CAS
CSCD
北大核心
2007年第7期676-679,F0002,共5页
Journal of Nanjing Medical University(Natural Sciences)
基金
南京市医学科技发展项目(ZKX0112)