ENDOTHELIAL MODIFICATIONS THAT REDUCE T CELL ACTIVATION

研究人员先前已经证明，培养的人内皮细胞（EC）可以激活同种异体CD4+和CD8+ T细胞分泌细胞因子，显示效应分子（如CD40L， FasL）和受体（如CD25， il - 2rα链），并在IL-2的作用下增殖。T细胞对培养EC的反应取决于EC中MHC分子、共刺激物和粘附分子的显示。这些观察结果导致了一个由两部分组成的假设。首先，研究者提出人类EC在血管化异体移植物中可以引发排斥反应。其次，减少移植物EC共刺激剂和/或粘附分子的表达或功能的修饰将减少移植物排斥反应。在目前的申请中，他们建议在体外扩展特异性共刺激分子（LFA-3和CD40）和粘附分子（ICAM-1和ICAM-2）在这些和其他早期T细胞反应（例如受体ITAM、ZAP 70和连接子分子磷酸化、转录因子激活、基因表达和原代和继代培养中的增殖）中的作用的研究。他们还将研究共刺激剂和粘附分子以及选定的其他蛋白质（如间隙连接蛋白）在同种异体反应性T细胞和EC之间免疫突触形成中的作用。他们将使用已建立和开发的新的限制性稀释方法来评估通过直接和间接（即树突状细胞依赖）途径对异体EC的识别程度，评估免疫调节细胞因子（IL-10和IL-11）对这些反应的影响，并评估获取和显示异体抗原的新方法（例如，膜转移或从吞噬细胞核中合并DNA）。最后，他们将把人类EC介导的T细胞激活的研究扩展到体内环境，通过检查EC共刺激剂和粘附分子在huPBL-SCID小鼠嵌合动物中所起的作用，这些嵌合动物植入了血管化的皮肤或用培养的人类EC形成的合成血管。这些研究的结果可能会导致保存同种异体移植物的新治疗策略，也可能适用于T细胞与血管内皮相互作用的其他临床环境，例如动脉粥样硬化和受体以及急性冠状动脉综合征。

The investigators have previously shown that cultured human endothelial cells (EC) can activate allogeneic CD4+ and CD8+ T cells to secrete cytokines, display effector molecules (e.g., CD40L, FasL) and receptors (e.g., CD25, the IL-2Ralpha chain), and proliferate in response to IL-2. The response of T cells to cultured EC depends upon EC display of MHC molecules, of costimulators and of adhesion molecules. These observations have led to a two part hypothesis. First, the investigators propose that human EC in vascularized allografts can initiate rejection reactions. Second, that modifications to reduce expression or function of graft EC costimulators and/or adhesion molecules will reduce graft rejection. In the current application, they propose to extend their studies of the role of specific costimulator molecules (LFA-3 and CD40) and adhesion molecules (ICAM-1 and ICAM-2) on these and other early T cell responses (e.g., receptor ITAM, ZAP 70 and linker molecule phosphorylation, transcription factor activation, gene expression, and proliferation in primary and secondary cultures) in vitro. They also will examine the role of costimulators and adhesion molecules and of selected other proteins (e.g., gap junction proteins) in the formation of immune synapses between alloreactive T cells and EC. They will use established and develop new limiting dilution approaches to evaluate the extent of recognition of allogeneic EC by the direct and indirect (i.e., dendritic cell-dependent) pathways, evaluate the effects of immunoregulatory cytokines (IL-10 and IL-11) on these responses, and evaluate novel methods of acquiring and displaying alloantigens (e.g., membrane transfer or DNA incorporation from phagocytosed nuclei). Finally, they will extend their study of human EC-mediated activation of T cells to in vivo settings by examining the roles played by EC costimulators and adhesion molecules in huPBL-SCID mice chimeric animals engrafted with vascularized skin or with synthetic vessels formed with cultured human EC. The results of these studies may lead to new therapeutic strategies for allograft preservation and may also apply to other clinical settings where T cells interact with vascular endothelium, e.g., in atherosclerosis and receptors and acute coronary syndromes.