Alterations of leukocyte integrin signaling leading to diabetes and autoimmunity

白细胞整合素信号的改变导致糖尿病和自身免疫

基本信息

批准号：
10502136
负责人：
Mark S Anderson
金额：
$ 64.67万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-12 至 2026-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10502136
关键词：
Actins Adaptor Signaling Protein Address Adhesions Adoptive Cell Transfers Adoptive Transfer Affect Animal Model Animals Antigen Presentation Antigens Autoantibodies Autoimmune Autoimmune Diseases Autoimmunity B-Lymphocytes Biochemical Biological Breeding CRISPR/Cas technology CTLA4 gene Cell Adhesion Cell Communication Cell Line Cell surface Cells Clone Cells Critical Pathways Data Defect Dendritic Cells Detection Development Diabetes Mellitus Disease Engineering Enrollment Enzyme-Linked Immunosorbent Assay Enzymes Etiology Event Experimental Models Family Female Frequencies Generations Genes Genetic Genetic Polymorphism Genetic Risk Genetic study Guanosine Triphosphate Phosphohydrolases Histologic Human Cell Line Human Genome Hyperactivity Hyperglycemia ITGB2 gene Image Immune Immune Tolerance Inbred BALB C Mice Inbred NOD Mice Incidence Individual Inflammation Inherited Insulin Insulin-Dependent Diabetes Mellitus Integrin Signaling Pathway Integrins Islets of Langerhans Kinetics Knock-in Knock-in Mouse Lead Leukocytes Life Link Lipid Binding Mediating Modeling Mouse Strains Mus Mutagenesis Mutation Myeloid Cells Neutrophil Activation Non obese Open Reading Frames Pathogenicity Pathway interactions Patients Phenotype Phosphatidylinositols Phosphotransferases Pre-Clinical Model Precision therapeutics Protein Region RNA Sequence Analysis Registries Regulator Genes Reporting Risk Factors STAT3 gene Signal Transduction Signaling Molecule T-Lymphocyte T-cell receptor repertoire Testing Therapeutic Transgenic Mice Transgenic Organisms Tumor-infiltrating immune cells Variant adhesion receptor autoreactive T cell diabetic draining lymph node early onset exome sequencing experimental study gain of function gain of function mutation genetic linkage genetic risk factor genome wide association study immune activation immune function insulin dependent diabetes mellitus onset insulitis islet knockin animal leukocyte activation macrophage male mouse model mutant neutrophil prevent two photon microscopy

项目摘要

Project Summary/Abstract The study of the genetic basis for type 1 diabetes (T1D) has benefited tremendously from examination of rare individuals with likely monogenic forms of the disease. Combined with GWAS, a number of polymorphisms in immune regulatory genes have been defined that contribute to genetic risk for T1D. Using whole exome sequencing of individuals with T1D enrolled in a monogenic diabetes registry, we identified an individual with a gain-of-function mutation in the SKAP2 gene, as well as several other T1D patients with potentially pathogenic variants in other leukocyte integrin signaling genes. These patients tend to have a number of autoimmune manifestations in addition to T1D, indicating defects in critical pathways of immune tolerance. Multiple GWAS studies have identified a strong genetic linkage between SKAP2 polymorphisms and T1D (at a frequency of ~20%), however the mechanisms by which alteration of SKAP2 could lead to autoimmune T1D are unknown. SKAP2 is expressed primarily in myeloid cells, where it functions as an adapter protein in the integrin signaling pathway, linking cell surface integrins to WASP and actin rearrangements that occur following leukocyte adhesion. The SKAP2 G153R mutation in our patient resulted in constitutive association of SKAP2 with WASP leading to a hyperadhesive phenotype in macrophages cultured from the patient or macrophages engineered to contain the SKAP G153R substitution. To understand how activation of leukocyte integrin signaling may contribute to T1D, we have generated knock-in (KI) mice containing the G153R substitution in murine Skap2, on the NOD genetic background. Female NOD.SKAP2 KI mice have a higher incidence and earlier onset of T1D than do NOD.WT animals; male NOD.SKAP2 also develop T1D (incidence ~50%) while male NOD.WT do not develop frank hyperglycemia. Initial analysis of these mice reveals evidence of ongoing inflammation early in life with development of a broad spectrum of auto-reactive antibodies. Dendritic cells from NOD.SKAP2 KI mice have increased antigen presenting activity to islet-specific transgenic T-cells while neutrophils from these mice show evidence of increased integrin signaling. These observations demonstrate that the NOD.SKAP2 KI mice appropriately model the autoimmune T1D disease observed in our patient. The project proposes to complete the analysis of these mice, under the hypothesis that increased cell adhesion in dendritic cells leads to prolonged DC-T cell interactions, which drives selection of auto-reactive T-cell clones leading to development of T1D, associated with broad spectrum autoimmunity. We will test this hypothesis in a variety of adoptive cell transfer experiments, by generation of conditional knock-in mice and by imaging of DC-T cell interactions in the inflamed islets. Similar studies will be performed for other candidate leukocyte integrin signaling mutations identified in the monogenic T1D registry. This study will address whether dysregulation of leukocyte integrin signaling may constitute an unrecognized genetic risk factor for T1D, suggesting potential alterative therapeutic approaches for these patients.

项目摘要/摘要 1型糖尿病（T1D）的遗传基础的研究从罕见的检查中受益匪浅具有可能是单基因疾病形式的个体。结合GWAS，许多多态性已经定义了有助于T1D的遗传风险的免疫调节基因。使用整个外显子在单基因糖尿病注册表中注册T1D的个体的测序，我们确定了一个患有A的个体 SKAP2基因的功能性突变以及其他可能致病性的其他T1D患者其他白细胞整联蛋白信号基因中的变体。这些患者往往有多种自身免疫性除T1D外，表现表明免疫耐受性的临界途径缺陷。多个GWAS 研究已经确定了SKAP2多态性与T1D之间的固有稳定的联系（以〜20％），但是，SKAP2改变可能导致自身免疫性T1D的机制尚不清楚。 SKAP2主要在髓样细胞中表达，在整联蛋白信号传导中，它充当衔接蛋白途径，将细胞表面整合素与白细胞后发生的WASP和肌动蛋白重排粘附。我们患者中的SKAP2 G153R突变导致SKAP2与WASP的组成型关联导致由患者或巨噬细胞培养的巨噬细胞中的高粘着表型包含SKAP G153R替代。了解白细胞整合素信号的激活如何有助于T1D，我们已经产生了含G153R替代的Murine Skap2中的敲入（Ki）小鼠，在点头遗传背景上。雌性点头skap2 ki小鼠的发病率更高，发病较早 T1D比点头动物；男性点头。SKAP2还会发展T1D（发病率〜50％），而男性点头。不要发展Frank Hyperglycemia。对这些小鼠的初步分析揭示了持续炎症的证据生命的早期，随着各种自身反应性抗体的发展。树突状细胞来自 NOD.SKAP2 KI小鼠具有增加对胰岛特异性转基因T细胞的抗原活性，而这些小鼠的中性粒细胞显示了整联蛋白信号增加的证据。这些观察结果证明了对我们患者观察到的自身免疫性T1D疾病适当地模拟了点头。这项目提议完成对这些小鼠的分析，这是假设增加了细胞粘附的假设树突状细胞会导致长时间的DC-T细胞相互作用，该相互作用驱动自动反应性T细胞克隆的选择导致T1D的发展，与广泛自身免疫有关。我们将在通过有条件的敲击小鼠以及成像的各种收养细胞转移实验发炎胰岛中的DC-T细胞相互作用。其他候选白细胞将进行类似的研究单基因T1D注册表中确定的整合素信号传导突变。这项研究将解决是否白细胞整联蛋白信号传导失调可能构成T1D的未识别遗传危险因素为这些患者提出潜在的改变治疗方法。