应用流式细胞术分析CIK细胞胞内Th1/Th2细胞因子

Analysis of Intracellular Th1/Th2 Cytokines of CIK Cells by Flow Cytometry

目的:建立细胞因子诱导的杀伤细胞(cytokine-inducedkiller,CIK)体外扩增的方法,并从单细胞水平分析CIK细胞的胞内Th1/Th2类因子产生的情况,进一步明确CIK细胞的免疫学特性及其参与的免疫效应机制。方法:取健康人外周血单个核细胞(PBMC),按一定的时间顺序分别加入IFN-γ、IL-2和CD3单抗,体外扩增CIK细胞,再应用流式细胞术,经刺激、阻断、固定、穿透和标记等过程测定CIK细胞内细胞因子IFN-γ和IL-4水平。结果:外周血单个核细胞加细胞因子培养两周后CD3+CD56+增加,培养至第21天、28天未见明显下降;诱导后CD3+CD8+细胞量与CD3+CD4+量相比明显增多;单细胞胞内细胞因子测定显示扩增后的CIK细胞Th1/Th2因子状态明显向Th1偏移。结论:采用多种因子组合刺激外周血单个核细胞,可大量诱导出CD3+CD56+双阳性细胞,具高度增殖性,是一类新型的免疫过继疗法细胞;其偏移的Th1/Th2因子状态可作为解释其极强的细胞毒作用的理论依据。

Objective:To investigate the TH1/TH2 cytokine profiles of CIK cells. Methods: Expansion of CIK cells from peripheral blood mononuclear cells ( PBMNCs) by the timed addition of interferon-g (IFN-g) , interleukin-2 (IL-2) , and the monoclonal antibody (McAb) OKT3. Intracellular cytokine production was assayed by fluorescence-activated cell sorter ( FACS) , Results: The expanded T-cell cultures produced IFN-γbut not IL-4. Both the CD4 + and CD8 + subsets secreted this cytokine profile. Conclusion: CIK cells may regulate the immune response by Th1 cytokine profile.