Lymphatic endothelial cells as inducers of systemic peripheral tolerance

描述(申请人提供)：外周耐受可防止逃避胸腺阴性选择的自身反应性T细胞引起自身免疫。在内源性外周耐受的流行模型中，静止的组织内树突状细胞(DC)迁移到淋巴结节(LN)，在那里它们将自身抗原转移到T细胞，然后T细胞发生缺失或无能。我们已经证明，CD8T细胞对酪氨酸酶产生的表位的耐受性不是由胸腺负选择或交叉耐受介导的，酪氨酸酶是一种通常仅表达于黑素细胞和视网膜色素上皮细胞的蛋白质。相反，CD8T细胞在LN驻留的淋巴管内皮细胞(LEC)的介导下经历了异常增殖和缺失，这些LEC外向表达酪氨酸酶。我们和其他小组已经证实，至少另外两个LN基质亚群也可以诱导CD8T细胞耐受。这些亚群中的每一个都表达许多外周组织AGs的不同亚群，这表明它们代表了系统性内在外周耐受的一个主要的、以前未被认识的机制。本申请的目的是提供证据支持这一假说，并特别关注LEC，并探索影响LEC执行这一功能的因素。在目标1中，我们将使用一个新开发的模型系统来确定表达MHC-II分子的LEC耐受CD4T细胞的机制，并探索抗原表达水平对CD4和CD8T细胞耐受结果的作用。在目的2中，我们将探讨PD-L1在不同抗原提呈细胞诱导的耐受中的意义。我们的初步研究表明，PD-L1信号可以在没有同时参与Ag的情况下发生，我们假设LEC作为守门人来加强免疫反应的质量。我们还将建立诱导T细胞增殖的机制，以及LEC在这一过程中的作用。增殖似乎是缺失过程中的一个重要因素，但也可以使自身反应性T细胞积累。在目标3中，我们将根据周围组织AGS和包括PD-L1在内的一系列免疫调节分子的表达，确定LN中的LEC(相对于组织淋巴管中的LEC)能够诱导耐受的机制。我们将定义影响LN驻留LEC中这些分子表达的细胞和分子成分，并使用基因表达分析来确定控制这些分子和外周组织AGS在LEC中表达的转录因子。总而言之，我们期待由这笔资金支持的研究将对建立和维持外周自我耐受的机制产生重大的新见解，这将与理解自身免疫性疾病的发展以及自我耐受如何限制对来自组织特异性蛋白质的癌细胞AGs的免疫反应有关。

DESCRIPTION (provided by applicant): Peripheral tolerance prevents self-reactive T cells that escape thymic negative selection from causing autoimmunity. In the prevailing model of intrinsic peripheral tolerance, quiescent tissue-resident dendritic cells (DC) migrate to lymph nodes (LN) where they cross present self-Ag to T cells, which then undergo deletion or anergy. We have shown that CD8 T cell tolerance to an epitope derived from tyrosinase, a protein whose expression is normally confined to melanocytes and retinal pigment epithelial cells, is not mediated by either thymic negative selection or cross-tolerance. Rather, CD8 T cells undergo abortive proliferation and deletion mediated by LN resident lymphatic endothelial cells (LEC) that adventitiously express tyrosinase. We and other groups have established that at least two other LN stromal subsets can also induce CD8 T cell tolerance. These subpopulations each express distinct subsets of many peripheral tissue Ags, suggesting that they represent a major, previously unrecognized mechanism of systemic intrinsic peripheral tolerance. The goal of the present application is to provide evidence to support this hypothesis, with a specific focus on LEC, and to explore factors that influence the ability of LEC to carry out this function. In Aim 1 we will use a newly developed model system to determine the mechanisms by which LEC, which express MHC-II molecules, tolerize CD4 T cells, and explore the role of antigen expression levels on tolerogenic outcome in both CD4 and CD8 T cells. In Aim 2 we will explore the significance of PD-L1, which LEC express at a higher level than any other LN cell population, on tolerance inducedby different antigen presenting cells. Our preliminary studies suggest that PD-L1 signaling can occur without simultaneous Ag engagement, and we hypothesize that LEC act as gatekeepers to enforce the quality of immune responses. We will also establish the mechanisms by which proliferation of T cells destined to undergo deletion is induced, and the role of LEC in this process. Proliferation appears to be an essential element of the deletion process, but also can enable accumulation of autoreactive T cells. In Aim 3 we will define the mechanisms by which LEC that reside in LN, as opposed to those in tissue lymphatic vessels, are able to induce tolerance based on their expression of peripheral tissue Ags and a set of immunomodulatory molecules, including PD-L1. We will define the cellular and molecular components that influence expression of these molecules in LN resident LEC, and use gene expression analyses to identify the transcription factors that control expression of these molecules and peripheral tissue Ags in LEC. Collectively, we expect the studies supported by this grant to lead to significant new insights into the mechanisms by which peripheral self-tolerance is established and maintained, which will be relevant in understand the development of autoimmune disease and the ways in which self-tolerance limits immune responses to cancer cell Ags that are derived from tissue specific proteins.