Role of Innate Immunity and Stat3 Activation in Colitis Mouse Model

描述（由申请人提供）： 炎症性肠病（IBD）、克罗恩病或溃疡性结肠炎影响估计140万美国人，并且表现为胃肠道的慢性炎症，具有显著的发病率，并且有时具有危及生命的并发症。IBD被认为是由遗传易感个体对结肠植物群的粘膜免疫（先天性和适应性）应答失调引起的。脆弱拟杆菌是一种常见的人类肠道寄生菌。B的子集。fragilis，肠促炎性B。脆弱性链球菌（ETBF）与炎性结肠炎、IBD和结肠直肠癌有关。我们的ETBF鼠模型数据显示，ETBF定植导致促炎反应，在C57 BL/6小鼠中表现出由信号转导子和转录激活子-3（Stat 3）促进的主要Th 17（分泌白细胞介素17）CD 4 + T细胞浸润。活化的Stat 3以及结肠炎的存在在用ETBF口胃接种的数小时内是明显的。然而，迄今为止还没有关于先天免疫系统在ETBF结肠炎中的作用的研究。IL-17偏斜最近已在涉及含核苷酸结合寡聚化结构域（NOD）先天免疫途径的下游产物的不同感染模型中得到证实。NOD 2基因突变与克罗恩病有很强的相关性。因此，该数据支持先天免疫因子和信号转导子和转录激活子3（Stat 3）在结肠炎的ETBF小鼠模型中起主要作用，提供了对炎性肠病的发生的深入了解。该研究项目的目的是解决先天免疫信号通路，特别是NOD/Rip 2和Toll样受体（TLR）/MyD 88在ETBF诱导的结肠炎模型中的作用。我们将测试MyD 88-/-和Rip 2-/-小鼠将表现出ETBF诱导的结肠炎和Stat 3激活增强的假设。为此，提出了以下具体目的：具体目的1：使用各种遗传操作的小鼠评价NOD/Rip 2和TLR/MyD 88先天免疫途径在ETBF诱导的结肠炎应答中的作用;具体目的2：在NOD/Rip 2和TLR/MyD 88缺陷小鼠中确定Stat 3活化在ETBF诱导的结肠炎发病机制中的作用;具体目的3：通过创建和利用骨髓（BM）移植衍生的嵌合小鼠，确定TLR/MyD 88和NOD/Rip 2缺陷在造血与非造血衍生细胞中对Stat 3活化和ETBF诱导的结肠炎发病机制的作用。该项目将使用ETBF诱导的结肠炎小鼠（C57 BL/6）模型和各种免疫学测定、基因敲除小鼠、骨髓移植、免疫组织化学、流式细胞术、细胞因子和RNA分析。这项研究的意义在于，它将使人们能够深入了解与IBD相关的结肠细菌的免疫反应失调，IBD是一种发病率很高的疾病，希望对这种流行疾病有新的治疗见解。 公共卫生相关性： 本研究探讨了先天免疫系统的作用，特别是TLR和NOD受体信号通路，以及信号转导和转录激活因子3（Stat 3）在细菌诱导的结肠炎小鼠模型中的作用。从这项研究中获得的信息将允许深入了解与IBD相关的免疫反应失调，IBD是一种发病率很高的疾病。

DESCRIPTION (provided by applicant): Inflammatory bowel disease (IBD), Crohn's disease or ulcerative colitis, affects an estimated 1.4 million Americans and is manifested by chronic inflammation of the gastrointestinal tract with significant morbidity and, at times, life-threatening complications. IBD is proposed to result from a dysregulated mucosal immune (innate and adaptive) response to the colonic flora in genetically susceptible individuals. Bacteroides fragilis is a common human commensal residing in the colon. A subset of B. fragilis, enterotoxigenic B. fragilis (ETBF), has been associated with inflammatory diarrheal disease, IBD, and colorectal cancer. Our ETBF murine model data shows that colonization with ETBF leads to a proinflammatory response exhibiting a predominant Th17 (interleukin 17-secreting) CD4+ T cell infiltrate promoted by signal transducer and activator of transcription-3 (Stat3) in C57BL/6 mice. The presence of activated Stat3 as well as colitis is evident within hours of orogastric inoculation with ETBF. However, there have been no investigations to date regarding the role of the innate immune system in ETBF colitis. IL-17 skewing has been recently demonstrated in a different infectious model involving downstream products of the Nucleotide-binding oligomerization domain containing (NOD) innate immune pathway. There is a strong association of NOD2 gene mutations and Crohn's disease. Hence, this data supports that innate immune factors and signal transducer and activator of transcription 3 (Stat3) plays a major role in the ETBF mouse model of colitis providing insight into the genesis of Inflammatory Bowel Disease. The objective of this research project is to address the role of the innate immune signaling pathways, in particular the NOD/Rip2 and toll like receptor (TLR)/MyD88, in the ETBF-induced colitis model. We will test the hypothesis that MyD88-/- and Rip2-/- mice will exhibit enhanced ETBF-induced colitis and Stat3 activation. To that end, the following Specific Aims are proposed: Specific Aim 1: To evaluate the role of NOD/Rip2 and TLR/MyD88 innate immune pathways in response of ETBF induced colitis using various genetically manipulated mice; Specific Aim 2: To define the role of Stat3 activation in the pathogenesis of ETBF-induced colitis in the NOD/Rip2- and TLR/MyD88- deficient mice; Specific Aim 3: To determine the role of TLR/MyD88 and NOD/Rip2 deficiency in hematopoietic versus non-hematopoietic derived cells on Stat3 activation and the pathogenesis of ETBF-induced colitis by creating and utilizing chimeric mice derived via bone marrow (BM) transplantation. The project will use the murine (C57BL/6) model of ETBF-induced colitis and various immunologic assays, knock-out mice, bone marrow transplantation, immunohistochemistry, flow cytometery, cytokine and RNA analysis. The significance of this research is that it will allow insight into the dysregulated immune response to colonic bacteria associated with IBD, a disorder with substantial morbidity, with hopes for new therapeutic insights to this prevalent disease. PUBLIC HEALTH RELEVANCE: This study examines the role of innate immune system, specifically TLR and NOD receptor signaling pathways, and the role of signal transducer and activator of transcription 3 (Stat3) in a bacteria induced colitis mouse model. Information obtained from this study will allow insight into the dysregulated immune response associated with IBD, a disorder with substantial morbidity.