Interplay of Innate and Adaptive Immunity in Lupus Pathogenesis

描述（由申请人提供）：系统性红斑狼疮（SLE）是一种复杂的多基因疾病。系统性红斑狼疮的迟发性和多变的发病率支持疾病发病是一个多步骤过程的假设。人类系统性红斑狼疮的高度可变的临床表现使得在患者中研究这一过程变得困难。在这里，我们利用一个易于处理和简化的遗传系统，CD 45 E613 R小鼠，询问SLE的发病机制。这些小鼠表达导致组成性CD 45磷酸酶活性的单个氨基酸取代。与人类SLE的不同表现相似，CD 45 E613 R表型对遗传背景极其敏感。在杂交129-C57 B1/6（B6）背景下，CD 45 E613 R小鼠产生抗dsDNA抗体和免疫复合物介导的肾小球肾炎（GN）。然而，在B6遗传背景下，尽管对受体刺激具有生化高反应性，但小鼠缺乏自身抗体，而BALB/c CD 45 E613 R小鼠产生高滴度抗dsDNA，但无GN。我们利用细胞高反应性、自身抗体产生和终末器官损伤的遗传分离来寻找疾病所需的遗传修饰剂。在这里，我们集中在SLE发病机制的前两个步骤的机制：失去对自身和外周扩增这种自身反应性的耐受性。目的1的目标是确定我们最近发现的两个新基因座Wam 1和Wam 2对自身抗体产生的贡献。将产生同类小鼠以测试这些基因座如何在过度活跃的CD 45磷酸酶活性的背景下调节耐受性阈值。在目的2中，我们评估了Wam 1中一个有前途的候选基因Toll样受体（TLR）9对疾病发病机制的贡献。TLR 9多态性的分子建模表明它们映射到DNA配体结合结构域中富含亮氨酸的重复序列。我们的初步数据表明B6和BALB/c小鼠之间的TLR 9刺激的差异反应。这些差异进一步通过CD 45 E613 R突变来调节，表明CD 45和TLR 9信号网络之间存在新的相互作用。我们将使用遗传和混合骨髓嵌合体的方法来确定TLR 9对自身抗体产生的影响，并确定它必须在CD 45 E613 R小鼠中发挥作用的细胞区室。目的3的重点是询问TLR 9多态性和CD 45 E613 R突变调节耐受性的细胞和分子机制。我们将评估所观察到的差异是否是由于与配体的相互作用改变，细胞内运输的扰动，和/或信号转导网络改变。这些研究的结果不仅有助于阐明耐受性的机制，而且应该为免疫系统的适应性和先天性武器如何在系统性自身免疫中相互作用提供新的见解。这是在人类自身免疫性疾病的发病机制中具有显著重要性的主题，并且鉴于在治疗自身免疫性疾病中靶向TLR以及作为疫苗策略中的佐剂的兴趣日益增长，具有广泛的影响。

DESCRIPTION (provided by applicant): Systemic Lupus Erythematosus (SLE) is a complex, polygenic disease. The delayed onset and variable pentetrance of SLE support the hypothesis that disease pathogenesis is a multi-step process. The highly variable clinical presentation of human SLE makes the study of this process difficult in patients. Here, we take advantage of a tractable and simplified genetic system, the CD45E613R mouse, to interrogate SLE pathogenesis. These mice express a single amino acid substitution that results in constitutive CD45 phosphatase activity. Mirroring the variable presentation of human SLE, the CD45E613R phenotype is extremely sensitive to genetic background. On a hybrid 129-C57Bl/6 (B6) background, CD45E613R mice develop anti-dsDNA antibodies and immune complex-mediated glomerulonephritis (GN). However, on a B6 genetic background mice lack autoantibodies despite biochemical hyper-responsiveness to receptor stimulation while BALB/c CD45E613R mice develop high-titer anti-dsDNA but no GN. We have taken advantage of the genetic separation of cellular hyper-responsiveness, autoantibody production, and end-organ damage to search for genetic modifiers required for disease. Here, we focus on the mechanisms governing the first two steps in SLE pathogenesis: Loss of tolerance to self and peripheral amplification of this autoreactivity. The goal of Aim 1 is to define the contributions of two novel loci we recently identified, Wam1 and Wam2, to autoantibody production. Congenic mice will be generated to test how these loci regulate the threshold for tolerance in the context of hyperactive CD45 phosphatase activity. In Aim 2, we evaluate the contributions of a promising candidate gene in Wam1, Toll Like Receptor (TLR) 9, to disease pathogenesis. Molecular modeling of the TLR9 polymorphisms indicate they map to leucine rich repeats in the DNA ligand binding domain. Our preliminary data indicate differential responses to TLR9 stimulation between B6 and BALB/c mice. These differences are further modulated by the CD45E613R mutation, indicating a novel interaction between CD45 and TLR9 signaling networks. We will use genetic and mixed bone-marrow chimera approaches to define the impact of TLR9 on autoantibody production and identify the cellular compartments in which it must function in CD45E613R mice. The focus of Aim 3 is to interrogate the cellular and molecular mechanisms by which the TLR9 polymorphisms and the CD45E613R mutation regulate tolerance. We will evaluate whether the observed differences are due to altered interaction with ligand, perturbations in intracellular trafficking, and/or altered signal transduction networks. Results of these studies will not only help elucidate the mechanisms governing tolerance, but should provide new insights into how the adaptive and innate arms of the immune system interact in systemic autoimmunity. This is a topic of significant importance in the pathogenesis of autoimmune disease in humans and has broad impact given the growing interest in targeting TLRs in the treatment of autoimmune disease and as an adjuvant in vaccine strategies.