Modeling autoimmune pathogenesis and beta cell destruction by T1D immune systems

模拟 T1D 免疫系统的自身免疫发病机制和 β 细胞破坏

• 批准号: 10762177
• 负责人: Mark S Anderson
• 金额: $ 42.5万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-20 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10762177
• 关键词: Address; Animal Model; Antigens; Autoimmune; Autoimmune Diseases; Beta Cell; CD8-Positive T-Lymphocytes; Cell Line; Development; Disease; Epithelial Cells; Family suidae; Genetic; Genetic Engineering; Genetic Risk; Genotype; HLA-A2 Antigen; HLA-DQ8 antigen; Hematopoietic stem cells; Heterogeneity; Human; Hybrids; Immune; Immune system; Immunodeficient Mouse; Immunotherapy; In Vitro; Inbred Mouse; Individual; Insulin; Insulin-Dependent Diabetes Mellitus; Lead; Mediating; Methodology; Methods; Modeling; Mus; Pathogenesis; Patients; Peptides; Research Design; System; T-Cell Development; T-Lymphocyte; Testing; Thymic Tissue; Thymic epithelial cell; Thymus Gland; Transgenic Organisms; autoimmune pathogenesis; autoreactive T cell; autoreactivity; diabetes pathogenesis; epithelial stem cell; fetal; genetic risk factor; genetic variant; human disease; human model; human pluripotent stem cell; humanized mouse; improved; in vivo; mouse model; novel; patient population; scaffold; stem cells; thymocyte; tool

Project Summary The study of Type 1 diabetes (T1D) pathogenesis has been limited by the insufficiency of animal models and the heterogeneity of patient populations, who derive genetic risk from HLA and different assortments of about 60 non-HLA genetic variants. Given this complexity, there is a need for improved models of human T1D pathogenesis involving diverse genetic backgrounds. We have developed humanized (HU) mouse models, including one in which T1D patient and healthy control (HC) immune systems are generated de novo from hematopoietic stem cells (HSCs) in Personalized Immune (PI) mice and models introducing transgenic (Tg) autoreactive TCRs into their HSCs and T cells. We have developed methods of generating thymic epithelial cell (TEC) progenitors and β cells from human pluripotent stem cells (hPSCs). We will use these tools to address the hypothesis that HSC-intrinsic and thymus-intrinsic genetic variants in T1D individuals lead to abnormal thymic selection of β cell antigen-autoreactive TCRs. We will: Aim 1: Determine the impact of T1D-prone genotypes on selection of autoreactive T cells in the human thymus. We have demonstrated negative selection of a Tg HLA-DQ8-restricted insulin B9-23 peptide-specific TCR in HLA-DQ8+ thymi of HU mice. We will assess additional β cell-autoreactive class I- and class II-restricted TCRs, determine the impact of both HSC and AIRE+ mTEC antigen expression and compare selection of T1D patient vs HC thymocytes bearing β cell-reactive TCRs; Aim 2: Determine the impact of T1D-prone genotypes of TECs on thymic selection of β cell-reactive T cells in humanized mice. We will utilize a novel model in which hPSC-TECs create a “hybrid thymus” on a living scaffold of fetal pig thymic tissue that supports human T cell development in HU mice. We will use this model with genetically engineered hPSCs to determine the impact of human TEC expression of TID-associated genetic variants on selection of autoreactive TCRs; Aim 3: Assess autoimmune interactions between hPSC-derived β cells and autoreactive T cells in humanized mice. We have developed models for rejection of hPSC-derived β cells in HU mice and for autoimmune β cell destruction by transducing autoreactive TCRs into their T cells. We have generated a hPSC cell line that lacks all HLA except HLA-A2, evading rejection in HLA-A2+ immune systems. These will be used in vivo and in vitro, with and without expression of an HLA Class II molecule, DQ8, to model autoimmune destruction by TCR Tg T1D patient and HC-derived T cells, assessing the requirement for Class II HLA and CD4 help for CD8 cell-mediated autoimmune β cell destruction. Our collaborative effort will generate novel and robust systems to model human T1D, enhancing understanding of its pathogenesis and providing a platform for testing of immunotherapies.

项目摘要 1型糖尿病（T1 D）发病机制的研究一直受到动物模型不足的限制， 患者人群的异质性，他们从HLA和大约100个基因的不同解释中获得遗传风险， 60种非HLA遗传变异。考虑到这种复杂性，需要改进的人类T1 D模型 发病机制涉及不同的遗传背景。我们已经开发了人源化（HU）小鼠模型， 包括其中T1 D患者和健康对照（HC）免疫系统从头产生自 在个体化免疫（PI）小鼠和引入转基因（Tg）的模型中的造血干细胞（HSC） 将自身反应性TCR导入其HSC和T细胞。我们已经开发了产生胸腺上皮细胞的方法 (TEC)在一些实施方案中，所述细胞可以是来自人多能干细胞（hPSC）的祖细胞和β细胞。我们将使用这些工具来解决 T1 D个体中HSC内在和胸腺内在遗传变异导致 β细胞抗原-自身反应性TCR的异常胸腺选择。我们将：目标1：确定 T1 D倾向基因型对人类胸腺中自身反应性T细胞选择的影响我们已经证明 HU的HLA-DQ 8+胸腺中Tg HLA-DQ 8限制性胰岛素B 9 -23肽特异性TCR的阴性选择 小鼠我们将评估额外的β细胞自身反应性I类和II类限制性TCR，确定其影响。 HSC和AIRE+ mTEC抗原表达，并比较T1 D患者与HC胸腺细胞的选择 目的2：确定TEC的T1 D倾向基因型对胸腺细胞增殖的影响。 在人源化小鼠中选择β细胞反应性T细胞。我们将利用一种新的模型，其中hPSC-TEC 在胎猪胸腺组织的活支架上创建“杂交胸腺”，支持人类T细胞发育， HU小鼠。我们将使用该模型与基因工程hPSC，以确定人类TEC的影响， TID相关遗传变异体的表达对自身反应性TCR的选择;目的3：评估自身免疫性 人源化小鼠中hPSC衍生的辟田胞与自身反应性T细胞之间的相互作用。我们有 开发了HU小鼠中hPSC衍生的β细胞的排斥和通过免疫抑制剂破坏自身免疫β细胞的模型。 将自身反应性TCR转导到T细胞中。我们已经产生了一种hPSC细胞系，其缺乏所有的HLA， HLA-A2，在HLA-A2+免疫系统中逃避排斥。这些将在体内和体外使用，有和没有 表达HLA II类分子DQ 8以模拟TCR Tg T1 D患者的自身免疫破坏，以及 HC衍生的T细胞，评估对II类HLA和CD 4的需求有助于CD 8细胞介导的自身免疫性疾病 β细胞破坏。我们的合作努力将产生新的和强大的系统来模拟人类T1 D， 增强了对其发病机制的理解，并为免疫疗法的测试提供了平台。