Tissue-engineered lymph node stroma to study peripheral tolerance in autoimmune diabetes

项目摘要： 1型糖尿病（T1 D）是由自身反应性T细胞引起的，其选择性地破坏胰岛中分泌胰岛素的B细胞。 胰腺对B细胞免疫耐受性的丧失导致T1 D，但其潜在机制尚不清楚 这是一个很好的理解，阻止了T1 D治疗的发展。外周耐受性的一个主要组成部分， T1 D是淋巴结（LN）中循环自身反应性T细胞通过与致耐受性Ag- 提呈细胞（APC）和调节性T细胞，这两者似乎在T1 D中有缺陷。 成纤维细胞网状细胞（FRC）形成LN网状结构，引导T细胞和APC的迁移和相互作用。 FRC还负责重塑LN并允许其在炎症期间扩张。有关 在自身免疫性疾病中，FRC是非专职APC，其可以通过以下方式诱导抗原特异性耐受： 在没有共刺激信号的情况下表达抗原并将抗原呈递给特异性T细胞，导致T细胞免疫应答。 接合（刺激和增殖），然后删除。 大多数研究使用转基因小鼠进行人工（与疾病无关） 因此，FRC在维持对疾病相关自身抗原的外周耐受性中的作用是 不太了解。我们发现在T1 D中，相对频率、抗原表达水平和网状组织 的LN FRC受到影响。该提案的一个目标是使用创新的组织工程化FRC网状物， 概括FRC网状组织在LN副皮质和T1 D已知自身抗原表达的研究 FRC网状特性（目标1.1）和抗原表达水平（目标2.1）如何影响体外T细胞接合 通过共同文化研究。 虽然专业APC可以转换为免疫原性表型并促进T1 D，但非专业APC如 FRC不太可能这样做。我们发现，促进表达T1 D抗原的FRC网状细胞的形成， 足以在非肥胖糖尿病（NOD）小鼠中诱导耐受性并预防T1 D。因此，本发明的第二个目标 我们的建议是在NOD小鼠中测试我们是否可以从体循环中删除自身反应性T细胞， 通过（i）将它们吸引到植入的工程化网状细胞和（ii）促进致耐受性T细胞来改善T1 D 通过调节FRC网状特性（目的1.2）和/或FRC抗原表达水平（目的2.2）进行接合。

Project Summary: Type-1 diabetes (T1D) is caused by autoreactive T cells that selectively destroy insulin-secreting b cells in the pancreas. Loss of immunological tolerance to b cells causes T1D, but the underlying mechanisms are poorly understood, preventing the development of a cure for T1D. A major component of peripheral tolerance to prevent T1D is the regulation of circulating autoreactive T cells in lymph nodes (LNs) by interactions with tolerogenic Ag- presenting cells (APCs) and regulatory T cells, both of which appear to be defective in T1D. Fibroblastic reticular cells (FRCs) form the LN reticula that guide migration and interactions of T cells and APCs. FRCs are also responsible for remodeling the LN and allowing its expansion during inflammation. Relevant to autoimmune diseases, FRCs are non-professional APCs that can induce antigen-specific tolerance by expressing and presenting antigens to specific T cells without co-stimulatory signals, leading to T cell engagement (stimulation and proliferation) followed by deletion. Most studies have been done using transgenic mice for overexpression of artificial (not disease relevant) antigens in FRCs; thus, the role of FRCs in maintaining peripheral tolerance to disease-relevant self-antigens is poorly understood. We found that in T1D, relative frequency, antigen expression levels and reticular organization of LN FRCs are affected. One goal of this proposal is to use innovative tissue-engineered FRC reticula that recapitulate FRC reticular organization in the LN paracortex and T1D known self-antigen expression to study how FRC reticular properties (aim 1.1) and antigens expression levels (aim 2.1) affect T cell engagement in vitro through co-culture studies. While professional APCs can switch to immunogenic phenotypes and promote T1D, non-professional APCs like FRCs are less likely to do so. We showed that promoting formation of FRC reticula expressing T1D antigens is sufficient to induce tolerance and prevent T1D in non-obese diabetic (NOD) mice. Thus, a second goal of this proposal is to test in NOD mice whether we can delete autoreactive T cells from the systemic circulation to ameliorate T1D by (i) attracting them to implanted engineered reticula and by (ii) promoting tolerogenic T cell engagement by modulating FRC reticular properties (aim 1.2) and/or FRC antigen-expression levels (aim 2.2).