DIRECT DEMONSTRATION OF CYTOKINE SYNTHESIS HETEROGENEITY AMONG HUMAN MEMORY/EFFECTOR T-CELLS BY FLOW-CYTOMETRY
DIRECT DEMONSTRATION OF CYTOKINE SYNTHESIS HETEROGENEITY AMONG HUMAN MEMORY/EFFECTOR T-CELLS BY FLOW-CYTOMETRY
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DOI：
10.1182/blood.v86.4.1408.bloodjournal8641408
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发表时间：
1995-08-15
期刊：
影响因子：
20.3
通讯作者：
MAINO, VC
中科院分区：
文献类型：
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作者：
PICKER, LJ;SINGH, MK;MAINO, VC
效应部位的T细胞产生的一系列细胞因子是这些细胞介导宿主防御的主要手段。众所周知，克隆的T细胞在细胞因子合成方面是异质性的，因此在它们介导特异性免疫反应的能力方面是不同的，但在克隆细胞中观察到的细胞因子分泌模式在多大程度上反映了体内存在的记忆/效应T细胞的实际群体，在很大程度上尚不清楚。在此，我们使用多参数流式细胞术同时检测单个T细胞在激活刺激和分泌抑制剂Brefeldin A体外孵育短时(4至8小时)后产生多种细胞因子的能力及其表型。该分析显示白细胞介素2(IL-2)、IL-4、在辅助性细胞非依赖性刺激(佛波酯13-醋酸酯[PMA]+离子霉素[I])或辅助性细胞依赖性(葡萄球菌肠毒素[SE]A和B)T细胞激活刺激刺激下，CD4(+)细胞胞浆中的γ-干扰素和干扰素(γ-干扰素)的产生，进一步分析表明，γ-干扰素和IL-4的产生主要(如果不是唯一的)限于CD45RO(高)记忆/效应T细胞亚群，而IL-2可能由CD45RO(高)和CD45RO(低)亚群产生。同时测定CD45RO(高)/CD4(+)T细胞产生IL-2和γ-干扰素的结果显示，单独产生这两种细胞因子的亚群不同(平均单独产生IL-2的30%，单独产生伽玛-干扰素的8%)，两者都产生(16%)，或者都不产生(46%)。对小的产生IL-4的记忆/效应T细胞亚群(仅占全部CD4(+)/CD45RO(高)T细胞的4.3%)的类似分析表明，这些产生IL-4的细胞中平均有51%也合成IL-2，23%只合成IL-4，16%合成所有三种细胞因子，9.6%合成IL-4和伽马-干扰素。这些细胞因子的合成模式被发现与PMA+I和SEA/SEB刺激相似，并在外周血记忆/效应CD4(+)T细胞和从皮肤迟发性超敏部位获得的表型相似的T细胞中观察到。综上所述，这些数据有力地支持了在体内存在具有预先编程的细胞因子合成潜力的人类记忆/效应器T细胞亚群，尽管它们表明这些亚群可能比最初在TH1/TH2假设中提出的更复杂。(C)1995年由美国血液病学会主办。
The array of cytokines produced by T cells in effector sites is a primary means by which these cells mediate host defense. It is well recognized that cloned T cells are heterogeneous with regard to cytokine synthesis and, thus, in their ability to mediate specific immune responses, but the extent to which the patterns of cytokine secretion observed in cloned cells reflect actual populations of memory/effector T cells existing in vivo is largely unknown. Here, we report our findings using a multiparameter flow cytometric assay that allows simultaneous determination of an individual T-cell's ability to produce multiple cytokines and its phenotype after only short (4 to 8 hours) in vitro incubation with an activating stimulus and the secretion inhibitor Brefeldin A. This assay shows a rapid accumulation of interleukin-2 (IL-2), IL-4, and gamma-interferon (gamma-IFN) in the cytoplasm of CD4(+) cells after stimulation with either accessory cell-independent (phorbol 12-myristate 13-acetate [PMA] + ionomycin [I]) or accessory cell-dependent (staphylococcal enterotoxins [SE] A and B) T-cell-activating stimuli, Further analysis showed that production of gamma-IFN and IL-4 is predominantly, if not exclusively, restricted to the CD45RO(high) memory/effector T-cell subset, whereas IL-2 may be produced by both the CD45RO(high) and CD45RO(low) subsets. Simultaneous determination of IL-2 and gamma-IFN production among CD45RO(high)/CD4(+) T cells showed distinct subsets that produce each of these cytokines alone (an average of 30% for IL-2 alone, 8% for gamma-IFN alone), both (16%), or neither (46%). Similar analyses with the small IL-4-producing memory/effector T-ceIl subset (only 4.3% of total CD4(+)/CD45RO(high) T cells) showed that an average of 51% of these IL-4-producing cells also synthesize IL-2, 23% synthesize only IL-4, 16% synthesize all three cytokines, and 9.6% synthesize IL-4 and gamma-IFN. These patterns of cytokine synthesis were found to be similar with both PMA + I and SEA/SEB stimulation and were observed in both peripheral blood memory/effector CD4(+) T cells and in T cells of similar phenotype obtained from cutaneous delayed-type hypersensitivity sites. Taken together, these data strongly support the in vivo existence of human memory/effector T-cell subsets with ''preprogrammed'' cytokine synthesis potential, although they suggest that these subsets may be more complex than originally proposed in the TH1/TH2 hypothesis. (C) 1995 by The American Society of Hematology.