The human B cell toll-like receptor 4 complex

描述（由申请人提供）：粘膜上的宿主-微生物相互作用需要一种微妙的平衡，尽管人们对这种平衡知之甚少，共生菌和致病菌的存在需要歧视性反应。虽然肠道的正常微生物如果停留在肠道微生态位内就不会致病，但如果它们穿过上皮屏障就会引起炎症反应。易位细菌的持续存在被认为在炎症性肠病（IBD）的病因和发病机制中起重要作用。文献中关于细菌感应基因在IBD中的作用的不一致表明，这种宿主-微生物相互作用的复杂性比人们所认识的要高。我们在知识上的一些差距可能是由于粘膜中最丰富的白细胞之一B细胞的作用定义不清。B细胞在IBD中表达toll样受体（TLR） 4，这是脂多糖（LPS）的受体，可能在感知微生物中起重要作用。矛盾的是，尽管TLR4表达，患者B细胞通常对大肠杆菌LPS反应不佳。相比之下，TLR4+ B细胞对LPS分子的反应具有低酰基化脂质A的特征，这是一组被证明对髓系TLR4具有拮抗作用的配体。本应用程序的目的是定义由人B细胞表达的潜在的新型动态TLR4复合物，以及B细胞TLR4反应在IBD中的功能意义。我们的中心假设是B细胞TLR4是一个动态复合体，允许通过TRIL（一种新的TLR4共受体）和结构修饰TLR4来识别低酰基化的LPS。我们的假设是基于我们关于B细胞TLR4复合物的疾病特异性变化的令人兴奋的数据，以及我们最近的报告，表明IBD患者的B细胞通过分泌IgA对低酰化LPS作出反应。因此，我们进一步假设TLR4刺激受限的IgA库，其表现出广泛的反应性并有助于保护粘膜。我们将通过两个具体目标来检验我们的假设。在第一个目标中，我们将通过使用患者原代B细胞进行消融和添加研究，确定所需的共受体，从而定义人类B细胞TLR4复合物。我们还将通过对已鉴定的B细胞TLR4变体的靶向测序和克隆，确定可能影响LPS识别的TLR4潜在结构修饰的影响。数据将与IBD的临床参数和TLR4多态性相关。在第二个目标中，我们将通过评估基因和激酶激活以及通过靶向测序IgA V区B细胞反应的克隆性来确定tlr4介导的IgA产生的机制。该提案中产生的数据将通过确定改善治疗方法发展的关键途径而具有直接的临床意义，从而改变疾病的进程。此外，通过定义B细胞如何识别LPS，我们也将推进全基因组关联研究领域，人类B细胞免疫学，以及涉及许多慢性疾病的人-微生物界面机制。

DESCRIPTION (provided by applicant): The host-microbial interactions at the mucosa require a delicate, although poorly understood, balance where the presence of commensal and pathogenic bacteria requires discriminatory responses. Although the normal microbes of the gut are not pathogenic if they remain within their gut micro-niche, they will induce an inflammatory response if they cross the epithelial barrier. The persistence of translocated bacteria is thought to play an important role in the etiology and pathogenesis of inflammatory bowel disease (IBD). Inconsistencies in the literature regarding the role of bacteria-sensing genes in IBD suggest a higher level of complexity to this host-microbe interaction than has been appreciated. Some of our gaps in knowledge may be explained by the poorly defined role of one of the most abundant leukocytes in the mucosa, the B cell. B cells express Toll-like receptor (TLR) 4, the receptor for lipopolysaccharide (LPS), in IBD and likely have an important role in sensing microbes. Paradoxically, despite TLR4 expression, patient B cells generally respond poorly to E. coli LPS. In contrast, TLR4+ B cells respond to LPS molecules with characteristics of hypo-acylated lipid A, a group of ligands shown to be antagonistic for myeloid TLR4. The objective of this application is to define the potentially novel and dynamic TLR4 complex expressed by human B cells and the functional significance of B cell TLR4 responses in IBD. Our central hypothesis is that B cell TLR4 is a dynamic complex that allows recognition of hypo-acylated LPS through TRIL, a novel TLR4 co-receptor, and structural modification TLR4. Our hypothesis is based on our exciting data on disease-specific changes in the B cell TLR4 complex and our recent report demonstrating that B cells from IBD patients respond to hypo-acylated LPS by secreting IgA. We thus further hypothesize that TLR4 stimulates a restricted IgA repertoire, which exhibits broad reactivity and helps to protect the mucosa. We will test our hypothesis through two Specific Aims. In the first aim, we will define the human B cell TLR4 complex by identifying the required co-receptors through ablation and addition studies using primary B cells from patients. We will also define the effect of potential structural modification(s) of TLR4 that may affect recognition of LPS through targeted sequencing and cloning of identified B cell TLR4 variants. Data will be correlated with clinical parameters of IBD and polymorphisms in TLR4. In the second aim, we will define the mechanism(s) of TLR4-mediated IgA production by evaluating gene and kinase activation and the clonality of the B cell response through targeted sequencing of the V region of IgA. The data generated in this proposal will have direct clinical relevance by identifying critical pathways for the development of improved treatments that change the course of disease. Further, by defining how B cells recognize LPS, we will also advance the field of genome-wide association studies, human B cell immunology, and mechanism(s) of the human-microbial interface implicated in many chronic diseases. PUBLIC HEALTH RELEVANCE: Inflammatory bowel disease (IBD) is a chronic gastrointestinal disease for which we have no cure. In this proposal, we hope to identify new ways in which our immune system interacts with the friendly bacteria of the intestines in hopes of developing treatments for patients with IBD.