Dissection of cellular interactions in T1DM with integrated functional genomics

项目摘要 认识到系统方法对复杂疾病的作用，在最初的资助期间， 开发了高度可控的三色阵列和分析平台，专注于采集质量数据。 我们的目标是将这些工具应用于导致自身免疫的事件，使用的是一种独特的衍生物， 1型糖尿病（T1 DM）的生物育种（BB）大鼠模型。BB DRlyp/lyp大鼠是自发性糖尿病 由于Gimap 5基因突变，使它们缺乏调节性T（Treg）细胞。DR+/+大鼠 具有完整的Gimap 5基因，不会发生自发性T1 DM，但在消耗后会变成糖尿病 Treg细胞BB大鼠中的T1 DM还涉及细胞毒性T细胞，因为它们的耗竭具有保护作用。我们的基因 糖尿病前期胰腺淋巴结（PLN）的表达研究表明，肥大细胞终末 在DRlyp/lyp动物中激活与在DR+/+动物中负调控;我们已经证实了它们的 通过两种不同的肥大细胞抑制药物预防大鼠T1 DM的功能作用。因此我们 假设除了细胞毒性T效应细胞外，肥大细胞在DRlyp/lyp中起关键早期作用 糖尿病发生，尽管有疾病倾向，但Treg细胞足以预防DR+/+中的T1 DM 大鼠我们进一步假设与这些状态相关的是不同的细胞因子环境，因为我们发现 将来源于处于T1 DM风险或新近发作的T1 DM的人类个体的血清与健康人一起孵育 外周血单个核细胞（PBMC）诱导一种独特的基因表达特征， 正常对照组和长期存在的T1 DM。重要的是，我们发现观察这种分子特征 T1 DM发作前5年。BB大鼠的可预测病程使得能够纵向解剖 免疫细胞亚群的活动，并检查这些活动如何表现在免疫细胞亚群中。 外围因此，我们建议：1）时间和空间解剖抗原呈递的活动， 通过基因表达谱分析和组织学方法， 接近。2)将纵向评价WF、DR+/+和DRlyp/lyp大鼠的血清中的因素， 诱导健康PBMC中基因表达。存在候选介质和疾病时间点特异性 生物标志物将被确认。了解肥大细胞、T效应细胞和Treg细胞之间的关系， 自身免疫，并有可能建立一个新的免疫调节模式，并创造新的见解 人类疾病。

PROJECT SUMMARY Recognizing the power of a systems approach to complex disease, during the initial funding period we developed a highly-controlled three color array and analysis platform focused on acquisition of quality data. Our goal is apply these tools to the events leading to autoimmunity using a unique derivative of the BioBreeding (BB) rat model of type 1 diabetes mellitus (T1DM). BB DRlyp/lyp rats are spontaneously diabetic due to a mutation in the Gimap5 gene, rendering them deficient in regulatory T (TREG) cells. DR+/+ rats possess an intact Gimap5 gene and do not develop spontaneous T1DM, but become diabetic upon depletion of TREG cells. T1DM in BB rats also involves a cytotoxic T cells, since their depletion is protective. Our gene expression studies of prediabetic pancreatic lymph nodes (PLN) have shown that mast cells are terminally activated in DRlyp/lyp animals versus negatively regulated in DR+/+ animals; we have confirmed their functional role through preventing T1DM in rats with two different mast cell inhibiting drugs. Thus, we hypothesize that in addition to cytotoxic T effector cells, mast cells play a crucial early role in DRlyp/lyp diabetogenesis and in spite of their disease predisposition, TREG cells are sufficient to prevent T1DM in DR+/+ rats. We further hypothesize that associated with these states are distinct cytokine milieus, since we find incubation of sera derived from human individuals at risk for T1DM or recent onset T1DM with healthy peripheral blood mononuclear cells (PBMC) induce a characteristic gene expression signature that is distinct from normal controls and long standing T1DM. Importantly, we find observe this molecular signature as much as 5 years prior to T1DM onset. The predictable disease course of the BB rat enables longitudinal dissection of the activities of immune cell subpopulations and examining how these activities are manifested in the periphery. Thus we propose to 1) temporally and spatially dissect the activities of antigen presenting, regulatory, and effector cells at defined pre-onset timepoints through gene expression profiling and histological approaches. 2) The sera of WF, DR+/+ and DRlyp/lyp rats will be longitudinally evaluated for factors that induce gene expression in healthy PBMCs. Presence of candidate mediators and disease time point-specific biomarkers will be confirmed. Understanding the relationship between mast cells, T effector, and TREG cells in autoimmunity and has the potential to establish a new paradigm for immune regulation and create new insights to human disease.