Taguchi T, McGhee J R, Coffman R L, Beagley K W, Eldridge J H, Takatsu K, Kiyono H
Department of Oral Biology, University of Alabama, Birmingham 35294.
J Immunol. 1990 Jul 1;145(1):68-77.
After Ag and/or mitogen stimulation, cloned mouse Th1 and Th2 cells produce different cytokines that contribute to induction of particular B cell isotype responses. In this regard, IL-5 produced by Th2 cells has been shown to enhance IgA synthesis in LPS-triggered splenic (SP) B cell or in unstimulated Peyer's patch (PP) B cell cultures. This raises the possibility that Th2 cells may occur in higher frequency in gut-associated tissues, because B cells in these areas are committed to IgA synthesis. We have used an ELISPOT assay to detect individual T cells producing IFN-gamma or IL-5. For the IL-5 assay, the mAb TRFK-5 and biotinylated TRFK-4 were used in coating and detection, respectively, whereas the mAb R4-6A2 and biotinylated XMG 1.2 were similarly used for enumeration of IFN-gamma-specific spot forming cells (SFC). Specificity of each assay was tested by using Con A-activated, cloned Th1 (H66-61) or Th2 (CDC-25) cells, where the Th1 cells only produced IFN-gamma SFC and the Th2 cells only gave IL-5-specific spots. Further, preincubation of biotinylated TRFK-4 or XMG 1.2 with rIL-5 or IFN-gamma, respectively, abrogated the formation of specific spots when tested with Con A-activated SP CD4+ T cells. Both IFN-gamma and IL-5 were produced de novo, because treatment of T cells with cycloheximide inhibited both IFN-gamma and IL-5 SFC. We have assessed the numbers of T cells spontaneously secreting these cytokines in PP and in lamina propria and intraepithelial lymphocyte (LPL and IEL) populations. Moderate levels of IL-5 SFC occurred in the IEL subset, whereas higher levels existed in the LPL population. Although significant numbers of IFN-gamma SFC (Th1-type) were also seen in LPLs, the frequency of IL-5 SFC was always higher (Th1:Th2 in LPL = 1:3). In IELs, equal numbers of IFN-gamma and IL-5 SFC were seen. Interestingly, CD8+ IEL T cells produced these two cytokines. In contrast, T cells freshly isolated from PP, an IgA inductive site, contained smaller numbers of IL-5- or IFN-gamma-secreting cells and SP T cells had essentially no SFC. When PP or SP T cells were stimulated with Con A, significant and approximately equal numbers of IFN-gamma- and IL-5-producing cells appeared.(ABSTRACT TRUNCATED AT 250 WORDS)
在受到抗原和/或丝裂原刺激后，克隆的小鼠Th1和Th2细胞会产生不同的细胞因子，这些细胞因子有助于诱导特定的B细胞同种型反应。在这方面，Th2细胞产生的IL-5已被证明可增强LPS刺激的脾脏（SP）B细胞或未刺激的派伊尔结（PP）B细胞培养物中的IgA合成。这增加了Th2细胞可能在肠道相关组织中以更高频率出现的可能性，因为这些区域的B细胞致力于IgA合成。我们使用ELISPOT测定法来检测产生IFN-γ或IL-5的单个T细胞。对于IL-5测定，单抗TRFK-5和生物素化的TRFK-4分别用于包被和检测，而单抗R4-6A2和生物素化的XMG 1.2同样用于计数IFN-γ特异性斑点形成细胞（SFC）。通过使用Con A激活的克隆Th1（H66-61）或Th2（CDC-25）细胞来测试每种测定的特异性，其中Th1细胞仅产生IFN-γ SFC，而Th2细胞仅产生IL-5特异性斑点。此外，分别用重组IL-5或IFN-γ对生物素化的TRFK-4或XMG 1.2进行预孵育，在用Con A激活的SP CD4+ T细胞进行测试时，可消除特异性斑点的形成。IFN-γ和IL-5都是重新产生的，因为用放线菌酮处理T细胞会抑制IFN-γ和IL-5 SFC。我们评估了PP以及固有层和上皮内淋巴细胞（LPL和IEL）群体中自发分泌这些细胞因子的T细胞数量。IEL亚群中出现中等水平的IL-5 SFC，而LPL群体中的水平更高。尽管在LPL中也发现了大量的IFN-γ SFC（Th1型），但IL-5 SFC的频率总是更高（LPL中Th1:Th2 = 1:3）。在IEL中，IFN-γ和IL-5 SFC的数量相等。有趣的是，CD8+ IEL T细胞产生这两种细胞因子。相比之下，从PP（一个IgA诱导部位）新鲜分离的T细胞含有较少数量的分泌IL-5或IFN-γ的细胞，而SP T细胞基本上没有SFC。当用Con A刺激PP或SP T细胞时，出现了数量可观且大致相等的产生IFN-γ和IL-5的细胞。（摘要截短于250字）
