Fuctional Analyses of Autoimmune Disease Variants

这项名为“自身免疫性疾病变异的功能分析”的项目主要关注人类和小鼠中的三个1型糖尿病(T1D)易感基因：CTLA4、PTPN22和CD226。T1D是儿童的一种主要疾病，发病率不断上升，5岁以下儿童确诊人数不断增加，原因不明。拟议的研究完全融入了PPG的活动和目标，并提供了一个平台，以继续剑桥和波士顿的PIs之间的高生产力、相互依赖和协同合作，旨在了解T1D和自身免疫性疾病基因变异的生物学影响(我们已经使用遗传图谱确定了这些基因变异)。TLD基因变异将使用来自当地可遗传选择的健康志愿者(剑桥生物资源)的主要来源的新鲜血液样本进行体外研究，并在体内通过精确设计的T1D的NOD小鼠模型进行研究。 CTLA4、PTPN22和CD226KO等位基因将被用于开发NOD菌株来模拟人类T1D。增加T1D的小鼠PTPN22变体将成为PTPN22基因机制研究的重点，PTPN22基因是影响多种人类自身免疫性疾病的分子途径的一部分。具有PTPN22易感等位基因的T细胞具有更高的激活阈值，在群体水平上，当体外刺激时，产生IL-2的细胞较少。由于人和小鼠IL-2途径中的基因变异都影响T1D的易感性，因此提出了研究两个物种中PTPN22和IL-2途径之间的基因-基因相互作用的实验。初步数据表明，CTLA4的易感等位基因降低了可溶性CTLA-4的表达，降低了Tregs被激活的可能性。可溶性CTLA-4在原代T细胞和细胞系中的过表达将是研究可溶性CTLA-4影响T细胞活化早期事件的机制之一。由于PTPN22和CTLA-4基因的变异被认为可以改变T细胞的激活，因此将在人类和小鼠中研究PTPN22和CTLA-4途径之间的基因-基因相互作用。

The project, Functional Analyses of Autoimmune Disease Variants, is focused on three type 1 diabetes (T1D) susceptibility loci, CTLA4, PTPN22 and CD226 in humans and in mice. T1D is a major disease of children with an unexplained steady rise in incidence and increasing numbers of children diagnosed under age 5 years. The research proposed is fully integrated into the PPG activities and goals, and provides a platform to continue the highly productive, interdependent and synergistic collaboration amongst the PIs in Cambridge and Boston aimed at understanding the biological effects of T1D and autoimmune disease gene variants (that we have identified using genetic mapping). TlD gene variants will be studied ex vivo using fresh blood samples from a major resource of genetically-selectable, local healthy volunteers (the Cambridge BioResource) and in vivo, in precisely engineered NOD mouse models of T1D. The Ctla4, Ptpn22 and Cd226 KO alleles will be used to develop NOD strains to model human T1D. A mouse Ptpn22 variant that increases T1D will be a focus of mechanistic studies on the PTPN22 gene, which is part of a molecular pathway that affects multiple human autoimmune diseases. T cells having the susceptibility allele at Ptpn22 have a higher threshold of activation and at a population level fewer of the cells produce IL-2 when stimulated ex vivo. Since both human and mouse gene variants in the IL-2 pathway affect T1D susceptibility, experiments to study gene-gene interactions between the PTPN22 and IL-2 pathways in both species are proposed. Preliminary data indicate that the susceptibility allele at CTLA4, which decreases the expression of soluble CTLA-4, reduces the probability that Tregs will be activated. Overexpression of soluble CTLA-4 in primary T cells and cell lines will be one approach used to study the mechanism by which soluble CTLA-4 affects early events in T cell activation. Since variants at both the PTPN22 and CTLA-4 genes are proposed to alter T cell activation, gene-gene interactions between the PTPN22 and CTLA-4 pathways in both humans and mice will be investigated.