Genetic regulation of intestinal epithelial cell innate immune signaling in human type 1 diabetes

项目摘要/摘要 1型糖尿病（T1 D）是一种遗传和环境因素共同作用的自身免疫性疾病， 一个单一的环境因素尚待确定。我们建议，而不是具体的身份， 在环境因素中，接收环境信号的方式对疾病最重要。之间 对于有发展为T1 D风险的个体，血清转化为胰岛自身抗体（IA）是最好和最早的 疾病进展的生物学预测因子。因此，导致破坏性β细胞自身免疫的关键事件 与T1 D相关的疾病与IA的发展有关。在具有高风险HLA等位基因的受试者中，9个基因区域 以单核苷酸多态性（SNP）为标志的基因与IA血清转化相关。我们的合作者 对这些IA风险基因座的分析揭示了仅调节三种细胞信号传导的基因/蛋白质的富集 途径（RAS/MAPK，PI 3 K/AKT和JAK/STAT），由此四种IA血清转化风险变体（PTPN 22， SH 2B 3、ERBB 3和UBASH 3A）分别影响所有三个。因此，我们建议， 感测至少部分地由与开发相关联的这些风险变量中的一个或多个控制 的IAs。 胃肠道（GI）与环境的适当沟通是局部和 全身免疫稳态，其部分由肠上皮细胞（IEC）通过其免疫调节。 与先天性和适应性免疫群体的物理和生化相互作用。为此， 先前已经证明了胃肠道内对远端部位自身免疫表达的免疫性。 因此，我们认为血清转换风险变异影响IEC内的免疫信号传导途径。 有助于T1 D自身免疫的维持，通过继发性扩张和/或极化， 自身反应性效应T细胞。 为了研究人类疾病，一个主要的需求是细胞特异性的人类同基因系统，可以用来 剖析个体T1 D风险基因变异等位基因的作用。我们的初步研究支持使用疾病- a-dish策略，将人类诱导多能干细胞（iPSC）与CRISPR/Cas9基因修饰配对， 满足这些需求。我们建议将iPSC和CRISPR/Cas9技术与我们的可重复的方法结合起来。 原代人IEC培养以获得单倍不足、单等位基因纯合性风险或保护性等位基因 评价有助于维持β细胞自身免疫的IEC免疫机制的改变。

Project Abstract/Summary Type 1 Diabetes (T1D) is an autoimmune disease with both genetic and environmental contributions, yet a single environmental factor has yet to be identified. We propose that rather than the specific identity of the environmental factor, the way in which an environmental signal is received is most important to disease. Among individuals at risk of developing T1D, seroconversion to islet autoantibody (IA) production is the best and earliest biological predictor of disease progression. Thus, critical events leading to the destructive β cell autoimmunity associated with T1D are linked to the development of IAs. In subjects with high risk HLA alleles, 9 gene regions marked by single nucleotide polymorphisms (SNPs) are associated with IA seroconversion. Our collaborator’s analysis of these IA risk loci reveals enrichment for genes/proteins that regulate only three cell signaling pathways (RAS/MAPK, PI3K/AKT and JAK/STAT), whereby four IA seroconversion risk variants (PTPN22, SH2B3, ERBB3 and UBASH3A) each impact all three. Thus, we propose that regulation of environmental sensing is at least in part controlled by the one or more of these risk variants associated with the development of IAs. Appropriate communication of the gastrointestinal tract (GI) with the environment is required for local and systemic immunologic homeostasis, which is in part regulated by the intestinal epithelial cell (IEC) through their physical and biochemical interactions with innate and adaptive immune populations. To this end, contribution of immunity within the GI tract to the expression of autoimmunity at distal sites has been previously demonstrated. Thus we propose that the seroconversion risk variants affect immune signaling pathways within the IEC contributing to the maintenance of autoimmunity in T1D, through the secondary expansion and/or polarization of autoreactive effector T-cells. To study human disease, a major need is cell-specific human isogenic systems that can be used to dissect the role of individual T1D risk gene variant alleles. Our preliminary studies support use of a disease-in- a-dish strategy that pairs human induced pluripotent stem cells (iPSCs) with CRISPR/Cas9 gene modification to fulfill these needs. We propose to couple iPSCs and CRISPR/Cas9 technology with our method of reproducible primary human IECs culture to achieve haploinsufficient, mono-allelic homozygosity at risk or protective alleles to evaluate alterations in IEC-immune mechanisms which contribute to the maintenance of βcell autoimmunity.