Project 1 - Immunologic and Cellular Mechanisms of T1D Modifier Mutations

项目 1 - T1D 修饰突变的免疫学和细胞机制

• 批准号: 10642553
• 负责人: Lucienne CHATENOUD
• 金额: $ 26.17万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-13 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10642553
• 关键词: Acceleration; Activities of Daily Living; Adoptive Transfer; Affect; Agreement; Antigen Presentation; Attention; Autoantigens; Beta Cell; Bone Marrow Transplantation; Breeding; CRISPR/Cas technology; Cell Count; Cells; Cellular Immunology; Chromogranin A; Collaborations; DNA Sequence Alteration; Data; Development; Diabetes Mellitus; Disease; Ethylnitrosourea; Event; FOXP3 gene; Gene Targeting; Genes; Genetic; Genome; Goals; Health; Hematopoietic stem cells; Human; Immune; Immune system; Immunologics; Inbred NOD Mice; Incidence; Induced Mutation; Infiltration; Insulin; Insulin Resistance; Insulin-Dependent Diabetes Mellitus; Islet Cell; Islets of Langerhans; Islets of Langerhans Transplantation; Knockout Mice; Laboratories; Maps; Measurement; Mediating; Metabolic; Missense Mutation; Molecular; Mus; Mutagens; Mutation; Organoids; Orthologous Gene; Pathogenesis; Pathogenicity; Pathologic; Patients; Peptides; Phenotype; Phosphoric Monoester Hydrolases; Predisposition; Process; Proteins; Regulatory T-Lymphocyte; Research; T cell therapy; T-Cell Receptor; T-Lymphocyte; Techniques; Technology; Testing; Therapeutic Intervention; Time; Tissues; Transgenic Organisms; Work; cell type; conditional knockout; diabetogenic; dominant genetic mutation; experimental study; improved; in vivo; innovation; insulin sensitivity; insulitis; islet; lymphoid organ; molecular sequence database; neonate; new therapeutic target; novel; prevent; success; synergism; tool; transcriptome

PROJECT SUMMARY/ABSTRACT The goal of Project 1 is to understand in detail the pathological consequences of ENU-induced mutations implicated by Core B and Core C to modify the T1D phenotype of NOD/NckH mice, and to determine the cellular mechanisms by which those mutations impact the development of T1D. As we have done for Dusp10, a spontaneous mutation bred to homozygosity in our lab based on phenotypic selection, we will analyze each selected mutation to determine which salient immunopathological steps—recognized as hallmarks of T1D initiation and/or progression—are affected. To that end, we will use tools such as analysis of pancreatic islet infiltration (i.e. insulitis); measurement of the numbers and the in vivo functional capacity in various lymphoid organs of diabetogenic or regulatory CD4+FoxP3+ T lymphocytes; and analysis of islet antigen presentation using adoptive transfer of diabetogenic cells expressing the transgenic BDC2.5 T cell receptor (specific for an autoantigenic fusion peptide incorporating insulin and chromogranin A sequences) into 4-week-old recipients. We will pay particular attention to mechanisms of insulin resistance, and to the definition of markers of increased β-cell sensitivity witnessed with particular genetic mutations. In doing so it is plausible that novel immunopathological steps will be discovered. Second, we will investigate the contribution of particular tissues or cell types bearing each mutation to the development of T1D using bone marrow transplantation, adoptive transfer in neonates, islet transplantation, tissue-specific conditional knockout mice, and islet organoid cultures in the presence or absence of immune cells. Results will provide important guidance to Project 2, focused on molecular pathogenesis. We will perform cellular mechanistic studies to understand of the effects of mutations in Paqr8 (protective), and Dusp10, Rapgef1, and Xpnpep1 (exacerbating), identified in our initial collaboration. We expect to actively pursue mechanistic studies of four to six proteins at a time, adding one new protein per year by mutual agreement with Project 2. We expect that strong synergy between Projects will accelerate our progress. An important question we will address is the relevance of our findings to human health. To understand how mutations identified in NOD mice relate to human T1D, we will study the effects of selected mutations in NOD mice bearing a T1D-susceptible humanized immune system modified to contain or lack the orthologous mutations in human hematopoietic stem cells and/or human islet cells. We will also identify human T1D patients with mutations in genes orthologous to those we implicated in our screen of NOD mice. Our findings will uncover new therapeutic targets for treatment of T1D and help to improve prediction of T1D onset in humans.

项目摘要/摘要 项目1的目标是详细了解ENU诱导的突变的病理后果 由Core B和Core C实现以修改NOD/NCKH小鼠的T1D表型，并确定细胞 这些突变影响T1D发展的机制。就像我们为dusp10所做的那样 基于表型选择，我们的实验室中繁殖具有纯合性的赞助突变，我们将分析每个 选定的突变以确定哪种显着免疫病理学步骤 - 被认为是T1D的标志 启动和/或进步 - 受到影响。为此，我们将使用诸如胰岛分析之类的工具 浸润（即胰岛炎）；在各种淋巴机构中的数量和体内功能能力的测量 糖尿病或调节性CD4+ FOXP3+ T淋巴细胞的器官；和胰岛抗原表现的分析 使用表达转基因BDC2.5 T细胞接收器的糖尿病性细胞的自适应转移（特异 将胰岛素和染色体A序列掺入4周龄的受体中的自身抗原融合肽。 我们将特别关注胰岛素抵抗的机制，以及增加的标志物的定义 特定的遗传突变见证了β细胞灵敏度。这样做是合理的小说 将发现免疫病理学步骤。其次，我们将研究特定组织或 使用骨髓移植，适应性转移的细胞类型，将每种突变带有每种突变的T1D发育 在新生儿，胰岛移植，组织特异性的条件敲除小鼠和胰岛类器官培养物中 存在或不存在免疫细胞。结果将为项目2提供重要的指导，重点是分子 发病。我们将进行细胞机械研究以了解PAQR8突变的影响 （保护性），DUSP10，Rapgef1和XPNPEP1（恶化），在我们的最初协作中确定。我们期望 一次积极进行四到六种蛋白质的机械研究，每年通过相互添加一种新蛋白质 与项目2的同意。我们希望项目之间的强大协同作用将加速我们的进步。一个 我们将要解决的重要问题是我们的发现与人类健康的相关性。了解如何 在NOD小鼠中鉴定出的突变与人T1D有关，我们将研究选定突变在NOD中的影响 带有T1D具有T1D启示的人源性免疫系统的小鼠被修饰以包含或缺乏直系同源 人造血干细胞和/或人类胰岛细胞中的突变。我们还将确定人类T1D患者 与我们在点头小鼠屏幕上实现的基因的突变。我们的发现将发现 用于治疗T1D的新的治疗靶标，并有助于改善人类T1D发作的预测。