T Cell Tolerance & Autoimmunity to Nuclear Antigen La

描述（由申请人提供）：RNA结合核抗原是系统性自身免疫性疾病（包括系统性红斑狼疮（SLE）和干燥综合征（SS））中靶向的一类主要自身抗原。通常介导对RNA结合核抗原的免疫耐受的机制和在系统性自身免疫的背景下被破坏的特异性耐受途径还不清楚，尽管已知T辅助细胞（Th）对鼠狼疮模型中的疾病至关重要。我们已经开发了第一个也是唯一的转基因小鼠的RNA结合核抗原特异性TCR T细胞受体（TCR），以描绘特定的机制，导致T细胞耐受性。通过将特异于临床相关的人La/SS- B抗原（hLa）的T细胞转移到生理表达hLa的转基因小鼠中，并跟踪它们的数量、表型和功能，我们将描绘导致外周T细胞耐受的机制（目的1）。初步研究表明，这些T细胞采用的耐受模式取决于受体中抗原呈递的水平，并且在该系统中促进Foxp 3+调节性T细胞的体内线索与先前描述的不同。在没有其他狼疮易感基因座的情况下，对轻度（2X）过表达RNA结合Toll样受体7（TLR 7）的小鼠中的抗原呈递细胞（APC）和T细胞表型进行分析，表明一种线索可能是核酸结合TLR的组成性刺激。这一假设及其机制将在目标1中研究。在不存在其他狼疮易感性基因座的情况下过表达TLR 7 2X的小鼠是健康的并且具有耐受性T细胞，而过表达TLR 7高于正常值>4X的小鼠发展为T细胞依赖性的严重的、致命的狼疮。将在Aim 2中评估在CD 4+抗hLa特异性T细胞转移到表达TLR 7>4X的hLa转基因小鼠中以及随后分化成不同的Th和T效应细胞表型后在CD 4+抗hLa特异性T细胞中消除的精确外周耐受机制。我们还将确定这些命运是否与特定耐受机制（如缺失和Foxp 3 + Treg转换）的丧失相关，并确定IL-6和IL-21在该模型中促进致病性Th细胞分化的作用。很少有研究已经研究了体内DC在稳态下促进CD 4 + T细胞耐受中的作用，并且还没有描述针对核酸结合核抗原的此类研究。在目标3中，我们将确定是否B细胞，传统的树突状细胞（DC）或浆细胞样DC，是必不可少的促进外周耐受性的CD 4 + T细胞特异性的这种代表性的RNA结合核抗原，通过以下的数量，表型和功能的抗hLa特异性T细胞在hLa转基因小鼠，已选择性耗尽这些细胞类型。相反，我们将确定B细胞和DC在致病性（>4X）TLR 7表达的情况下促进耐受性丧失和抗hLa特异性T细胞分化成致病性Th和T效应细胞类型中的相对重要性。这些研究将确定CD 4 + T细胞耐受的机制及其在系统性自身免疫性疾病中靶向的主要抗原类别的丧失。系统性红斑狼疮（SLE）和干燥综合征（SS）是影响免疫系统的慢性自身免疫性疾病。人类和小鼠的免疫系统具有天然机制，旨在维持自身耐受性并对感染和炎症做出反应。我们试图了解这些元素如何控制CD 4 + T细胞的命运和表型在稳态和炎症条件下，使我们可以更好地了解自身免疫性疾病的发展。

DESCRIPTION (provided by applicant): RNA-binding nuclear antigens are a major class of autoantigen targeted in systemic autoimmune diseases, including systemic lupus erythematosus (SLE) and Sjvgren's syndrome (SS). The mechanisms that normally mediate immunologic tolerance to RNA-binding nuclear antigens and the specific tolerance pathways that are breached in the setting of systemic autoimmunity are not well understood, although it is known that T helper (Th) cells are crucial for disease in murine lupus models. We have developed the first and only mouse transgenic for an RNA-binding nuclear antigen-specific TCR T cell receptor (TCR) in order to delineate specific mechanisms leading to T cell tolerance. By transferring T cells specific for the clinically relevant human La/SS- B antigen (hLa) into mice transgenic for physiologic expression of hLa and tracking their numbers, phenotype and function, we will delineate the mechanisms leading to peripheral T cell tolerance (Aim 1). Preliminary studies have shown that the mode of tolerance adopted by these T cells depends on the level of antigen presentation in the recipients and that the in vivo cues promoting Foxp3+ regulatory T cells in this system differ from those previously described. Analysis of antigen presenting cell (APC) and T cell phenotypes in mice mildly (2X) over-expressing RNA-binding Toll-like receptor 7 (TLR7) in the absence of other lupus susceptibility loci suggest that one cue could be constitutive stimulation of nucleic acid-binding TLR. This hypothesis and its mechanism will be investigated in Aim 1. Mice that over-express TLR7 2X in the absence of other lupus susceptibility loci are healthy and harbor tolerant T cells, whereas mice that over-express TLR7 >4X above normal develop severe, fatal lupus that is T cell dependent. The precise peripheral tolerance mechanisms that are abrogated in CD4+ anti-hLa specific T cells following their transfer into hLa transgenic mice that express TLR7 >4X and their subsequent differentiation into distinct Th and T effector cell phenotypes will be evaluated in Aim 2. We will also determine whether these fates associate with the loss of particular tolerance mechanisms such as deletion and Foxp3+ Treg conversion, and determine the roles of IL-6 and IL-21 in promoting pathogenic Th cell differentiation in this model. Very few studies exist that have investigated the role of in vivo DC in promoting CD4+ T cell tolerance in the steady state, and no such studies for nucleic acid- binding nuclear antigens have been described. In Aim 3, we will determine whether B cells, conventional dendritic cells (DC) or plasmacytoid DC, are essential for promoting the peripheral tolerance of CD4+ T cells specific for this representative RNA-binding nuclear antigen by following the numbers, phenotype and function of anti-hLa-specific T cells in hLa transgenic mice that have been selectively depleted of these cell types. Conversely, we will determine the relative importance of B cells and DC in promoting loss of tolerance and differentiation of anti-hLa specific T cells into pathogenic Th and T effector cell types in the setting of pathogenic (>4X) TLR7 expression. These studies will define the mechanisms of CD4+ T cell tolerance and its loss for a major class of antigen that is targeted in systemic autoimmune disease. PUBLIC HEALTH RELEVANCE Systemic lupus erythematosus (SLE) and Sjogren's syndrome (SS) are chronic, debilitating autoimmune diseases that affect the immune system. The immune systems of humans and mice have natural mechanisms designed to maintain self-tolerance and respond to infection and inflammation. We seek to understand how these elements control CD4+ T cell fate and phenotype under both steady-state and inflammatory conditions so that we can better understand the development of autoimmune disease.