B Cells and Type 1 Diabetes

描述（由申请人提供）：自身免疫影响近1000万美国人，并且由于未知原因，其频率正在增加。现有证据表明，自身免疫性疾病的发生是由遗传决定的易感性和先天免疫刺激共同作用的结果，它们共同刺激了对自身的适应性免疫。自身免疫的基础是T细胞和/或B细胞区室对自身免疫耐受的破坏。虽然我们对维持B细胞耐受性的机制了解很多，但我们对导致自身免疫失败的机制知之甚少。在这个应用中，我们建议研究小鼠和人类1型糖尿病中B细胞对胰岛素耐受性丧失的运作机制，这种疾病已知需要B细胞，被认为通过抗原呈递到CD4 T细胞起作用。我们的方法将建立在我们以前的工作定义信号通路在维持无能B细胞抗原无反应的运作。我们将通过流式细胞术分析胰岛素特异性B细胞状态的变化，这些细胞使用一种新的基于磁珠的方法从正常、前驱糖尿病和糖尿病小鼠和患者中分离出来。在某些情况下，免疫球蛋白重链转基因将用于产生多种抗原受体库，但在胰岛素特异性细胞中富集。目的1将探讨外周胰岛素特异性B细胞如何在正常小鼠中沉默，以及这种沉默是否在糖尿病小鼠中被打破。在Aim 2中，我们将研究抗原受体亲和胰岛素在决定沉默模式、命运和糖尿病发病潜力中的作用。这将涉及到一种反向转基因的方法，以产生具有胰岛素特异性抗原受体亲和力的抗原库。在Aim 3中，我们将把研究扩展到人类，比较糖尿病前期和糖尿病个体与非糖尿病一级亲属的胰岛素特异性B细胞。最后，在Aim 4中，我们将评估一种新型B细胞脱敏疗法对T1D的治疗效果，并将这种疗法与消耗B细胞的抗cd20疗法进行比较。该实验将解决1型糖尿病中高亲和胰岛素特异性B细胞沉默被打破的总体假设，这些细胞通过向CD4 T细胞呈递自身抗原来促进疾病。

DESCRIPTION (provided by applicant): Autoimmunity affects nearly ten million Americans and, for unknown reasons, is increasing in frequency. Available evidence indicates that autoimmune diseases arise due to a combination of genetically determined susceptibility and innate immune stimulation, which conspire to stimulate adaptive immunity to self. Underpinning autoimmunity is the breakdown of immune tolerance to self in T cell and/or B cell compartments. While a great deal is known about the mechanisms that maintain B cell tolerance, we understand little about the mechanisms that cause it to fail in autoimmunity. In this application we propose to study mechanisms operative in loss of B cell tolerance to insulin in murine and human Type 1 Diabetes, a disease known to require B cells that are thought to function by antigen presentation to CD4 T cells. Our approach will build upon our previous work defining signaling pathways operative in maintaining antigen unresponsiveness of anergic B cells. We will analyze by flow cytometry changes in the status of insulin-specific B cells isolated using a novel magnetic bead-based approach from normal, prediabetic and diabetic mice and patients. In some cases immunoglobulin heavy chain transgenesis will be used to generate a diverse antigen receptor repertoire that is nonetheless enriched in insulin-specific cells. Aim 1 will explore how peripheral insulin-specific B cells are silenced in normal mice and whether this silencing is breached in diabetic mice. In Aim 2 we will study the role of antigen receptor affinit for insulin in determining mode of silencing, fate and diabetogenic potential. This will involve a retrotransgenic approach to generate a repertoire with defined antigen receptor affinity for insulin. In Aim 3 we will extend studies to the human, comparing insulin-specific B cells in prediabetic and diabetic individuals to non-diabetic first- degree relatives. Finally, in Aim 4 we will assess the therapeutic efficacy in T1D of a novel B cell desensitizing therapy, comparing this therapy to anti-CD20, which depletes B cells. The experiments will address the overarching hypothesis that in Type 1 Diabetes the silence of high affinity insulin-specific B cells is broken and these cells promote disease by presentation of autoantigen to CD4 T cells.