体外诱导扩增的CD1d限制性人外周血NKT细胞克隆过早凋亡的机理研究  

Finite proliferation capacity of human CD1d-restricted natural killer T cells due to activation-induced apoptosis

作　　者：高跞[1] 杨洁[1] 杨懿铭[1] 任亚娜[1] 范华骅[1] 

机构地区：[1]上海市血液中心,上海200051

出　　处：《中国输血杂志》2016年第1期33-36,共4页Chinese Journal of Blood Transfusion

基　　金：上海市自然科学基金面上项目(132R1438900);上海市卫生局青年科研项目资助;No.024Y37;上海市血液中心科技发展基金项目资助;No.L(9)

摘　　要：目的针对NKT细胞培养扩增和培养过程中遇到的过早凋亡现象,在凋亡信号层面进行初步的描述和探讨。为今后的NKT相关研发提供数据参考。方法自健康人浓缩白细胞制品(白膜)分离有外周血单个核细胞(peripheral blood mononuclear cells PBMNCs),利用其中的抗原递呈细胞,以α-Gal Cer的刺激NKT细胞克隆产生,记录3-4周NKT的表型变化,和Caspase-3酶活化及细胞凋亡状况。结果刺激7 d后,NKT克隆开始产生,持续扩增3周左右后开始呈现凋亡状态,Caspase3酶在第2周即开始维持高活性状态。人NKT细胞对α-Gal Cer的刺激的反应具有很大的个体差异。结论 NKT细胞克隆可以再最初的3周内如同常规T细胞克隆一样生长,其后则会因α-GalCer刺激引发活化后凋亡。Objective To present the phenomena and to analyze the possible mechanism of the apoptosis during NKT induction and proliferation,which should be considered in future NKT cell development. Methods NKT cells were induced from PBMNC and proliferated in the presence of alpha-Gal Cer and IL-2 for 3-4 weeks in vitro. The various phenotypes of the NKT were analyzed through flow cytometry. During the entire culture process,the cells were harvested to analyze caspase-3activity by Western Blot and kinase activity assays every day. Results NKT clone arose after seven days of culture and mainly expanded during the later 2 weeks,but turned to apoptosis in a large scale from then,which was indicated by the flow cytometry assay on the cell surface markers and picture records. Western Blot demonstrated that the highest Caspase-3 kinase activity was achieved during the second week. Moreover,the sensitivity and the reaction time point of NKT to alpha-Gal Cer stimulation on proliferation and caspase-3 kinase activity was varied in different individuals. Conclusion According to the data,human NKT cells could be induced and proliferate excellently during the first three weeks,although NKT could not be expanded to a larger scale,due to the activation-induced apoptosis stimulated by alpha-Gal Cer.

关 键 词：NKT细胞 凋亡 Caspase-3 

分 类 号：R446.6[医药卫生—诊断学] R392.12[医药卫生—临床医学]