正常人周围血初始和记忆T细胞亚群及细胞因子表达的探讨

目的：利用多色流式检测技术探讨初始和记忆T细胞亚群与细胞因子表达之间的关系。方法：自正常人静脉血中分离PBMC,经超抗原（SEB）刺激5h后,加入多种抗细胞表面标记和抗细胞因子抗体进行染色,利用流式细胞术检测,并利用Flow Jo软件分析结果。结果：根据CD45RO表达与否,将CD4^＋和CD8^＋T细胞分为初始和记忆T细胞,再根据归巢受体（CD62L）和趋化因子受体（CCR7）的表达与否,将初始和记忆T细胞进一步分为不同的亚群。当T细胞受到SEB激活后,CD45RO^＋和CD45RO^-的CD4^＋或CD8^＋T细胞均表达IL-2、IFN-γ和TNF-α。进一步分析结果表明,CD62L^hi和CD62L^hiCCR7^＋细胞不表达细胞因子,而CD62L^loCCR7^lo和CCR7^＋T细胞均表达细胞因子,其中CD62L^loCCR7^lo细胞表达细胞因子的阳性率明显高于CCR7^＋细胞亚群。结论：只利用CD45RO表达与否区分初始和记忆T细胞是不够准确的,同时检测CD62L的表达,可明显地提高其准确性。

Subpopulations and cytokine expression of na(i)ve and memory T cells in normal human PBMCs

AIM: To determine subpopulations and cytokine expression of na?ve and memory T cells by polychromatic flow cytometry(PFC). METHODS: PBMCs were separated from normal human peripheral blood and stimulated by superantigen (SEB). After incubation for 5 hours, the cells were fixed, permeablized and stained with multiplecolor-labeled mAbs for cell surface and intracellular cytokines. Then they were detected by polychromatic flow cytometry and the data were analyzed by Flow Jo software. RESULTS: Based on the expression of CD45RO molecules, CD4(+) and CD8(+) T cells were classified as na?ve and memory cells, which were further devided into several subsets according to the expression of CD62L and CCR7. When PBMCs were stimulated by SEB for 5 hours, CD45RO(+) and CD45RO(-) CD4(+) or CD8(+) T cells expressed IL-2, IFN-gamma and TNF-alpha. Further analysis indicated that CD62L(lo) CCR7(lo) and CCR7(+) T cells but not CD62L(hi) and CD62L(hi) CCR7(+) T cells expressed cytokines. In addition, the percentage of cytokine expression in CD62L(lo) CCR7(lo) subsets was markedly higher than that in CCR7(+) subsets. CONCLUSION: It is not scientific to identify na?ve and memory cells in human T cells only based on the expression of CD45RO. Accurate determination of na?ve, effector and memory cell populations can be achieved not only based on the expression of CD45RO but also according to the expression of CD62L.