人γhG—CSF动员的外周血采集物来源的树突状细胞成熟诱导的研究  

The study of maturation induction of γhG-CSF mobilized peripheral blood grafts-derived dendritic cells

作　　者：魏静[1] 蔡秋妍 陈家萍[1] 潘健[1] 应美爱[1] 完晓菊[1] 周志风 王保龙[2] 

机构地区：[1]安徽医科大学附属省立医院输血科,合肥230001 [2]安徽医科大学附属省立医院检验科,合肥230001

出　　处：《国际免疫学杂志》2013年第2期132-137,共6页International Journal of Immunology

基　　金：国家自然科学基金面上项目（81172172）;安徽省卫生厅医学科研课题（09A082）

摘　　要：目的探讨优化人γhG-CSF动员的外周血采集物来源的树突状细胞（DC）成熟诱导条件。方法利用密度梯度离心法从重组人粒细胞集落刺激因子（吖hG-CSF）动员的外周血采集物分离出单个核细胞，以重组人粒细胞-巨噬细胞集落刺激因子（GM—CSF）和重组人白细胞介素-4（IL4）体外诱导成不成熟树突状细胞（imDC），分别采用传统细胞因子鸡尾酒（IL-1β、IL-6、TNF—α、PGE2）和改良细胞因子鸡尾酒（IL-1β、IL-6、TNF-α、polyI：C、CpGODN）刺激成熟。3d后收获成熟DC，观察细胞形态、流式细胞仪检测DC的内吞能力通过异硫氰酸荧光素标记的葡聚糖（FITC—Dextran）、检测DC表面CD83、CD1a、CD80、HLA—DR、CD86和CCR-7，酶联免疫吸附法（ELISA）检测其IL一12的分泌，M1Tr法检测其刺激淋巴细胞增殖活性。结果两种方法均能诱导DC成熟，以改良细胞因子鸡尾酒效果更佳，FITC—Dextran的内吞能力明显下降；CD83、CD1a的表达率分别是84．44％、87．94％（P〈0．05）；IL-12分泌量明显增高，成熟DC能有效的刺激T淋巴细胞增殖。结论改良细胞因子鸡尾酒法可能是诱导DC成熟的最佳方法。Objective To investigate the most effective strategy for mature induction of ~/h G-CSF mo- bilized peripheral blood grafts-derived dendritic ceils. Methods The peripheral blood mononuclear ceils （PBMCs） were isolated from γh G-CSF mobilized peripheral blood grafts, by using density gradient centrifugation method,then imDC was induced in the presence of cytokines GM-CSF and IL-4. The immature DC was pulsed with each of a traditional cocktail of cytokines （TNF-α,IL-β,IL-6 ,PGE2） or a modified cocktail of cyto- kines （IL-1β,IL-6 ,TNF-α,polyI：C ,CpG ODN）. DC was harvested after 3d induction, Cells morphology was Observed, FITC-Dextran endocytic activity was measured by FCM. Detected the surface markers for DC matura- tion CD83 ,CDla , CD80,HLA-DR,CD86 and CCR-7 by FCM. IL-12 production was detected by ELISA. The capacity of DC for T cell activation was detected by MTF assay. Results The two means all induce DC mature. The most effective method for induction of maturation was modified cocktail of cytokines. FITC-Dextran endocytic activity was clearly declined, the expression rate of CD83 and CDla were up to 84.44% and 87.94% （ P 〈 0.05）. The amount of IL-12 production increased significantly,mature DC can effective stimulation T lympho cyte proliferation. Conclusion The most effective method for inducing DC＇ maturation was modified cocktail of cytokines.

关 键 词：树突状细胞 细胞因子鸡尾酒 改良 成熟诱导 

分 类 号：R[医药卫生]