Kinetics of cytokine gene expression in human CD4+ and CD8+ T-lymphocyte subsets using quantitative real-time PCR
Kinetics of cytokine gene expression in human CD4+ and CD8+ T-lymphocyte subsets using quantitative real-time PCR
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DOI：
10.1111/j.1365-3083.2003.01348.x
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发表时间：
2003-12-01
影响因子：
3.7
通讯作者：
Rabbani, H
中科院分区：
文献类型：
--
作者：
Abdalla, AO;Kiaii, S;Rabbani, H
在体外刺激人CD 4和CD 8 T淋巴细胞后，通过实时定量聚合酶链反应（PCR）分析了五种细胞因子[白细胞介素-2（IL-2）、IL-5、干扰素-γ（IFN-γ）、粒细胞巨噬细胞集落刺激因子（GM-CSF）和肿瘤坏死因子-α（TNF-α）]和一种细胞毒性效应蛋白（颗粒酶B）的时间动力学。使用两种刺激物，促分裂原[植物血凝素（PHA）]和回忆抗原[纯化蛋白衍生物（PPD）]。发现细胞因子mRNA表达的模式依赖于所使用的T细胞亚群和刺激。细胞因子基因表达模式的个体间差异很大。观察到两种表达模式。在PHA活化后，在亚群和PPD活化的CD 4 T细胞中观察到大多数细胞因子的钟形表达谱，而在PPD刺激后，在CD 8 T细胞中观察到双相/多相表达模式。对于大多数细胞因子，诱导的时间在活化的30分钟内，并且似乎在活化4-8小时后获得最大积累。然而，可以注意到持续的高水平长达24小时。颗粒酶B基因的表达也在活化30 min内诱导，但表现出连续的逐渐增加和晚期的最大积累（48-72 h）。本研究的结果是重要的，当设计研究使用细胞因子基因表达谱作为抗原特异性T淋巴细胞的标志物。可能建议在特异性活化4-8小时后测量细胞因子基因表达（IL-2、IL-5和IFN-γ），但刺激长达24小时也是可接受的。应优先在活化48-72小时后测量颗粒酶B。
The time kinetics of five cytokines [interleukin-2 (IL-2), IL-5, interferon-gamma (IFN-gamma), granulocyte macrophage-colony stimulating factor (GM-CSF) and tumour necrosis factor-alpha (TNF-alpha)] and one cytotoxic effector protein (granzyme B) was analysed by real-time quantitative polymerase chain reaction (PCR) following in vitro stimulation of human CD4 and CD8 T lymphocytes. Two stimuli were used, a mitogen [phytohemagglutinin (PHA)] and a recall antigen [purified protein derivative (PPD)]. The pattern of cytokine mRNA expression was found to be dependent on the T-cell subset and stimulus used. A wide interindividual variability in the cytokine gene expression pattern was demonstrated. Two expression patterns were observed. A bell-shaped expression profile was seen for most cytokines upon PHA activation in both subsets and PPD-activated CD4 T cells, whereas a biphasic/multiphasic expression pattern was noted in CD8 T cells upon PPD stimulation. For most cytokines, the time to induction was within 30 min of activation, and maximum accumulation seemed to be obtained after 4-8 h of activation. A sustained high level could, however, be noticed for up to 24 h. Granzyme B gene expression was also induced within 30 min of activation but showed a continuous gradual increase and late maximal accumulation (48-72 h). The findings of the present study are of importance when designing studies using the cytokine gene expression profile as a marker for antigen-specific T lymphocytes. It might be recommended that cytokine gene expression (IL-2, IL-5 and IFN-gamma) should be measured after 4-8 h of specific activation but also up to 24 h of stimulation is acceptable. Granzyme B should preferentially be measured after 48-72 h of activation.