Immunogenetic Control of Autoimmune Liver Disease

描述(由申请人提供)： 我们已经培育了几个非肥胖糖尿病(NOD)同源株(一般称为“NOD”小鼠)，它们可以产生自发的自身免疫性胆道疾病(ABD)和抗丙酮酸脱氢酶复合体(PDC)自身抗体，并作为人类原发性胆汁性肝硬变(PBC)的模型。它们为确定PBC的早期免疫遗传机制提供了一个亟需的模型系统，这一点至关重要，因为人类疾病有一个很长的“沉默”阶段。我们现在已经培育出一种新的同源基因小鼠，命名为NOD.Abd3，在结节背景上具有1个B6间隔的1.0兆碱基的染色体；该品系在雄性和雌性小鼠中发育严重的自发ABD，外显率为100%。这笔赠款将研究NOD.Abd3小鼠疾病的免疫学机制。我们的初步数据表明，ABD可以通过NOD.ABD转移，但不能通过NOD，CD8+细胞转移，这表明NOD.Abd3CD8细胞参与了发病过程。我们的初步数据还表明，适应性免疫系统是ABD疾病过程的核心。最后，共转移ABD T调节细胞可阻止CD8细胞介导的SCID受者ABD。这项资助将通过以下具体目的剖析NOD.ABD CD8细胞增强致病性的免疫学基础，以及NOD.Abd3Tregs的保护能力：目的#1.CD8+T细胞介导的自身免疫性胆道疾病的机制。我们已经证明，NOD.ABD，而不是NOD，CD8+T细胞足以将胆道病理转移到ABD-SCID接受者。本研究的目的是通过1)转移和骨髓嵌合研究，2)详尽分析CD8+T细胞标志物在NOD.ABD小鼠中的表达，以及3)过继转移后CD8+T细胞细胞因子在体外和体内的表达，剖析CD8+T细胞介导疾病的主要机制。目的#2.肝内NOD.ABD CD8T细胞的T辅助细胞依赖性和抗原特异性。这一目标将解决两个主要问题：1)NOD CD8 T细胞反应是依赖还是独立的？2)肝内CD8 T细胞是抗原特异性的？我们将通过研究有无CD4或CD8细胞的ABD，以及限制CD8+T细胞谱系和评估对疾病的影响来测试这些问题。目的#3：T调节细胞在ABD中的作用。我们已经证明，在我们的过继转移模型中，共转移NOD.Abd3脾树可以预防疾病。为什么NOD.Abd3Tregs不能在体内预防自发性疾病？我们将详尽地研究肝脏和外周Tregs的表型，并通过转移和耗竭研究来论证Tregs在ABD中的功能作用。这些研究是重要和有意义的，因为它们将使我们能够研究免疫介导的胆管损伤的机制，而以前由于缺乏自发的动物疾病模型而无法研究这些机制。了解胆汁免疫改变的早期机制将导致关于人类自身免疫性胆道疾病的病因和治疗的新假说。

DESCRIPTION (provided by applicant): We have produced several Nonobese diabetic (NOD) congenic strains (generically designated "NOD.ABD" mice) that develop spontaneous, autoimmune biliary disease (ABD) and anti-pyruvate dehydrogenase complex (PDC) autoantibodies, and serve as a model of human primary biliary cirrhosis (PBC). They provide a much needed model system for determining early immunogenetic mechanisms of PBC, which is critically important because the human disease has a long "silent" phase. We have now produced a novel congenic mouse, termed NOD.Abd3, with a 1.0 megabase chromosome one B6 interval on the NOD background; this strain develops severe, spontaneous ABD with 100% penetrance in male and female mice. This grant will study immunological mechanisms of disease in NOD.Abd3 mice. Our preliminary data show that ABD can be transferred by NOD.ABD, but not NOD, CD8+ cells, implicating NOD.Abd3 CD8 cells in pathogenesis. Our preliminary data also show that the adaptive immune system is central to the ABD disease process. Finally, co- transfer of ABD T regulatory cells prevented CD8 cell mediated ABD in scid recipients. This grant will dissect the immunological basis for this enhanced pathogenicity of NOD.ABD CD8 cells, and protective capacity of NOD.Abd3 Tregs, through the following specific aims: Aim #1. Mechanism of CD8+ T cell mediated autoimmune biliary disease. We have demonstrated that NOD.ABD, but not NOD, CD8+ T cells are sufficient to transfer biliary pathology to ABD-scid recipients. This aim dissects major mechanisms of CD8+ T cell mediated disease, using 1) transfer and bone marrow chimeric studies; 2) an exhaustive analysis of CD8+ T cell marker expression in NOD.ABD mice; and 3) a thorough analysis of CD8+ T cell cytokine expression in vitro, and in vivo following adoptive transfer. Aim #2. T helper dependence and antigen specificity of intrahepatic NOD.ABD CD8 T cells. This aim will address two major issues: 1) Are the NOD CD8 T cell responses CD4+ T cell dependent or independent? 2) Are intrahepatic CD8 T cells antigen specific? We will test these issues by studying ABD with and without either CD4 or CD8 cells and by restricting the CD8+ T cell repertoire and assessing effect on disease. Aim #3: Role of T regulatory cells in ABD. We have shown that co-transfer of NOD.Abd3 splenic Tregs can prevent disease in our adoptive transfer model. Why don't NOD.Abd3 Tregs prevent spontaneous disease in vivo? We will exhaustively study the phenotype of hepatic vs. peripheral Tregs and demonstrate the functional role of Tregs in ABD by employing transfer and depletion studies. These studies are important and significant because they will allow us to study mechanisms of immune mediated biliary damage that previously could not be studied due to a lack of a spontaneous animal model of disease. Understanding early mechanisms of altered biliary immunity will lead to new hypotheses regarding the etiology and treatment of human autoimmune biliary disease.