MICROBIAL ANTIGENS IN CROHN'S DISEASE

克罗恩病候选微生物抗原的发病机制及免疫保护。共生菌是人类克罗恩病和小鼠IBD模型的重要致病因子。这一见解为新兴研究提出了两个关键问题：乳糜泻的免疫发病机制是否由对特定肠道微生物的反应介导？并且，这些反应可以通过操纵产生现在的炎症或抗炎免疫。在之前的资助期间，我们的实验室成功地应用免疫学和减法克隆方法鉴定了一组与UC和CD相关的细菌种类和微生物抗原。这次更新申请的重点是这些候选基因之一I2，这是一种发现定位于CD病变的细菌来源的新基因。I2编码蛋白的重组表达在人CD中显示出高度疾病特异性抗体水平。在小鼠中，对I2的天然记忆T细胞免疫被证实，效应谱在结肠炎易感和耐药小鼠菌株中分别极化为TH1和TH2。这些发现支持该细菌作为乳糜泻的候选病原体，并为在实验室小鼠菌株中评估候选人类乳糜泻病原体的微生物学和免疫反应提供了独特的机会。我们的更新项目测试了两个假设。首先，我们预测I2细菌是导致克罗恩病的微生物亚群之一，因为它能够定殖相关粘膜部位，引发局部组织破坏性粘膜部位，并在易感宿主中引发局部组织破坏性免疫反应。我们将通过分离和表征I2细菌的分类、毒力特征和肠组织分布来实验接近这一假设。从免疫学角度，我们将评估I2抗原特异性T细胞群和细胞系的肽特异性、个体发生、解剖起源和致病性。其次，我们预测这种或其他抗原共生微生物和粘膜自身抗原是抗炎T细胞的目标，这是对结肠炎的自然和治疗保护所必需的。我们将通过表征这些T细胞群的个体发生和定位来实验验证这一假设；分离抗原特异性抗炎T细胞群并评估其在结肠炎转移模型中的保护作用；并且，测试抗原转移策略作为治疗性结肠炎的免疫调节剂。这些目标将取决于与项目4（小鼠致病性和保护性粘膜T细胞群的细胞和拓扑发育），项目2（项目3衍生微生物抗原特异性的人类B细胞和T细胞克隆群体的表征）和项目1（与这些抗微生物反应相关的人类遗传位点）的共享研究。

Immune mechanisms of Pathogenesis and Protection for Crohn's Disease Candidate Microbial Antigens. Commensal bacteria are an important disease factor in human Crohn's disease and murine IBD models. This insight raises two critical issues for emerging research: Is immune pathogenesis of CD mediated by responses to particular enteric microorganisms? And, Can these responses by manipulated to yield now- inflammatory or anti-inflammatory immunity. In the previous grant period, our laboratory successfully applied immunologic and subtractive cloning approaches identify a set of bacterial species and microbial antigens associated with UC and CD. This renewal application focuses on one of these candidates, I2, a novel gene of bacterial origin found to be localized in CD lesions. Recombinant expression of the I2-encoded protein revealed highly disease-specific antibody levels in human CD. In the mouse, native memory T cell immunity to I2 was demonstrated, and the effector profiles were polarized to TH1 and TH2 in colitis susceptible and resistant mouse strains, respectively. These finding support the bacterium as a candidate pathogen in CD, and present the unique opportunity to evaluate the microbiology and immune response for a candidate human CD pathogen in laboratory mouse strains. Our renewal project tests two hypotheses. First, we predict that the I2 bacterium exemplifies one of the subset of microorganisms pathogenic in Crohn's disease, due to its capacity to colonize relevant mucosal sites and elicit a local tissue-destructive mucosal sites and elicit a local tissue-destructive immune response in susceptible hosts. We will experimentally approach this hypothesis by isolating and characterizing the I2 bacterium with regard to taxonomy, virulence traits, and intestinal tissue distribution. From an immunologic perspective, we will evaluate the peptide specificity, ontogeny, anatomic origin, and pathogenicity of T cell populations and cell lines specific for the I2 antigen. Second, we predict that this or other antigen commensal microorganisms and mucosal autoantigens are targets of anti-inflammatory T cells necessary for the natural and therapeutic protection to colitis. We will experimentally test this hypothesis by characterizing the ontogeny and localization of these T cell populations; isolating antigen-specific anti-inflammatory T cell populations and evaluating their protective effect in colitis transfer models; and, testing antigen-transfer strategies as therapeutic immunomodulators of model colitis. These aims will depend on shared studies with Project 4 (cellular and topologic development of pathogenic and protective mucosal T cell populations in the mouse), Project 2 ( characterization of human B and T cell clonal populations specific for Project 3-derived microbial antigens), and Project 1( human genetic loci associated with these anti-microbial responses).