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NKG2D激活和LAIR-1抑制受体所起的作用感兴趣。对T细胞克隆和激活的T细胞株的研究表明，NKG2D是TCR介导的信号的共刺激受体。我们纯化了CD8+T细胞，并通过CD62L+CD45RO-表型分选获得了一个原始群体。分析表明，NKG2D是TCR诱导新鲜分离的初始CD8+T细胞钙动员的共刺激受体。此外，NKG2D共刺激导致效应细胞产生大量的IFNpha和TNFpha。在这些细胞中，这些反应与CD28共同刺激的反应相同，有时甚至更高。在增殖细胞中观察到NKG2D配体MICA、MICB和ULBP-1、-2和-3的表达，我们推测这是NKG2D在激活的细胞中表达下调的原因。在培养液中加入稳态细胞因子IL-7和IL-15，不仅能促进细胞增殖，而且能拮抗NKG2D的下调作用。这些结果表明，NKG2D是人初始CD8+T细胞中重要的共刺激分子，其表达由IL-7和IL-15维持。 LAIR-1在大多数人单核白细胞上有结构性表达，是人NK细胞、B细胞、巨噬细胞和树突状细胞的抑制性受体。它在T细胞中的表达及其在T细胞中所起的作用尚未得到很好的描述。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。CD8T细胞表达的LAIR-1水平高于CD4。幼稚的CD4和CD8T细胞比记忆T细胞表达更高水平的LAIR-1。交联型抗CD3单抗激活外周血T细胞可增加增殖细胞表面LAIR-1的表达。最重要的是，我们发现LAIR-1在培养的和新分离的CD4和CD8T细胞中都是TCR介导的激活的有效抑制物。我们还确定了LAIR-1是由肥大细胞表达的，并正在确定连接这种抑制性受体是否可以用来控制过敏反应。

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.