Cell Surface Molecules Involved in Immune Function

本研究的目的是确定调节人幼稚T细胞活化的因素;特别是我们对NKG 2D活化和LAIR-1抑制性受体所起的作用感兴趣。T细胞克隆和活化T细胞系的研究表明，NKG 2D是TCR介导信号的共刺激受体。我们已经纯化了CD 8 + T细胞，并通过分选CD 62 L + CD 45 RO-表型获得了初始群体。分析显示，NKG 2D作为TCR诱导的新鲜分离的初始CD 8 + T细胞中Ca++动员的共刺激受体。此外，NKG 2D共刺激导致产生大量IFN α和TNF α的效应细胞。这些反应与在这些细胞中对CD 28共刺激观察到的反应相当或有时更大。在增殖细胞中观察到NKG 2D配体云母、MICB和ULBP-1、-2和-3的表达，我们推测这解释了在活化细胞中观察到的NKG 2D下调。向培养基中添加稳态细胞因子IL-7和IL-15不仅增强了增殖，而且抵消了NKG 2D的下调。这些结果表明，NKG 2D是人初始CD 8 + T细胞中的一种重要共刺激分子，其表达通过IL-7和IL-15的存在而得以维持。 LAIR-1在大多数人单核白细胞上组成型表达，并作为人NK细胞、B细胞、巨噬细胞和树突细胞上的抑制性受体发挥作用。它在T细胞中的表达及其在这些细胞中的作用尚未得到很好的表征。我们发现，在新鲜分离的外周血细胞中，LAIR-1在T细胞亚群中差异表达。CD 8 T细胞比CD 4表达更高水平的LAIR-1。初始CD 4和CD 8 T细胞比记忆T细胞表达更高水平的LAIR-1。通过交联抗CD 3 mAb激活外周血T细胞增加增殖细胞中LAIR-1的细胞表面表达。最重要的是，我们表明LAIR-1是培养的和新鲜分离的CD 4和CD 8 T细胞中TCR介导的活化的有效抑制剂。我们还确定LAIR-1由肥大细胞表达，并且正在确定这种抑制性受体的连接是否可以用于控制过敏反应。

The purpose of this research is to determine factors that regulate the activation of human naive T cells; in particular we are interested in the roles played by the NKG2D activation and LAIR-1 inhibitory receptors. Studies with T cell clones and activated T cell lines have shown that NKG2D is a co-stimulatory receptor for TCR mediated signals. We have purified CD8+ T cells and obtained a naive population by sorting for CD62L+ CD45RO- phenotype. Analyses showed that NKG2D serves as a co-stimulatory receptor for TCR induced Ca++ mobilization in freshly isolated naive CD8+ T cells. In addition, NKG2D co-stimulation results in effector cells that produce high amounts of IFNalpha and TNFalpha. The responses are equivalent or sometimes more than that seen for CD28 co-stimulation in these cells. The expression of the NKG2D ligands MICA, MICB and ULBP-1, -2, and -3 was observed in the proliferating cells, which we speculate accounts for the observed down-regulation of NKG2D in the activated cells. The addition of the homeostatic cytokines IL-7 and IL-15 to the culture medium not only enhanced proliferation but counteracted the down-regulation of NKG2D. These results indicate that NKG2D is an important co-stimulatory molecule in human naive CD8+ T cells and its expression is maintained by the presence of IL-7 and IL-15. LAIR-1 is constitutively expressed on the majority of human mononuclear leukocytes and functions as an inhibitory receptor on human NK cells, B cells, macrophages, and dendritic cells. Its expression by T cells and the role that it plays in these cells has not been welll characterized. We show that, in freshly isolated peripheral blood cells, LAIR-1 is differentially expressed in T cell subsets. CD8 T cells express higher levels of LAIR-1 than CD4. Naive CD4 and CD8 T cells express higher levels of LAIR-1 than memory T cells. Activation of peripheral blood T cells by cross-linking anti-CD3 mAb increases the cell surface expression of LAIR-1 in proliferating cells. Most importantly, we showed that LAIR-1 is a potent inhibitor of TCR mediated activation both in cultured and freshly isolated CD4 and CD8 T cells. We have also determined that LAIR-1 is expressed by mast cells and are in the process of determining if ligation of this inhibitory receptor can be used to control allergic reactions.