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Trafficking and the role of myelin specific Regulatory T cells in EAE

髓磷脂特异性调节性 T 细胞在 EAE 中的贩运和作用

基本信息

批准号：
8105664
负责人：
Mohamed Oukka
金额：
$ 29.25万
依托单位：
SEATTLE CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-20 至 2012-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8105664
关键词：
Activation Analysis Address Affect Animal Model Antigens Autoimmune Diseases Autoimmune Process Autoimmunity Back Behavior CD4 Positive T Lymphocytes Cell Separation Cells Commit Data Dendritic Cells Development Disease Disease remission Doctor of Philosophy Effector Cell Experimental Autoimmune Encephalomyelitis Frequencies Generations Genomics Green Fluorescent Proteins Hand Human IL2RA gene Immunization In Vitro Inflammation Inflammatory Injury Interferon Type II Interleukin-10 Interleukin-17 Interleukin-6 Knock-in Mouse Laboratories Measures Molecular Multiple Sclerosis Mus Myelin Neuraxis Organ Pathogenicity Patients Peptides Peripheral Phenotype Play Positioning Attribute Principal Investigator Production Reagent Regulation Regulatory T-Lymphocyte Reporter Research Personnel Resistance Role Source Specificity Staging Surface T-Lymphocyte T-Lymphocyte Subsets Testing Time Tissues Toxin base cytokine design feeding homologous recombination in vitro activity in vivo interest novel prevent programs protein expression receptor red fluorescent protein research study tool trafficking transcription factor

项目摘要

DESCRIPTION (provided by applicant): The molecular and cellular mechanisms by which CD4+CD25+FoxP3+ T cells are induced and by which they inhibit induction of autoimmunity have not been elucidated. The role of this subset of T cells in the regulation of EAE, an animal model of Multiple Sclerosis (MS), is just beginning to be investigated. To study FoxP3+ T cells at the single cell level in vivo, we specifically introduced the green fluorescent protein (GFP) into the endogenous FoxpS genomic locus by homologous recombination and generated a FoxPS knock-in mouse (FoxPSKI). In the FoxpSKI mice and with the use of MOG lAb specific tetramer, we followed in vivo the generation of MOG specific regulatory cells (T-regs) and found that MOG specific T-regs accumulate in the central nervous system (CNS) during the course of EAE, and produced IL-10 and unexpectedly IFN-g. To study the function of T-regs in EAE, we will take advantage of mice in which a DTR/GFP (Diphteria Toxin Receptor fused with the Green Fluorescent Protein) reporter has been introduced into the endogenous FoxPS locus, allowing in vivo conditional depletion of FoxPS expressing cells through administration of diphteria toxin. Using these mice we will be able to study the generation, trafficking and in vivo function of MOG specific T-regs, by deleting them at different stages of EAE. We also discovered that CD4+ T cells can differentiate in vitro into either pathogenic Th17 cells in the presence of TGF-b plus IL-6 or T-reg in the presence of TGF-b alone. These results suggest the existence of reciprocity in the function of these two subsets in vivo, and that they may regulate each other. However, whether the inflammatory milieu and pathogenic Th17 cells can modulate the generation and the suppressive activity of T-reg cells has not been elucidated. To address these important questions related to the interplay between Th17 and T-reg, we have generated a novel reporter mouse in which IL-17 expressing cells IL-17 can be followed by the Red Fluorescent Protein (RFP) and these cells can be conditionally depleted through administration of diphteria toxin. Based on our novel reagents and preliminary data, we have designed experiments to directly answer the following questions: Aim1: What is the origin and function of different subsets of MOG specific Tregs? Aim2: What is the role of IL-6 on Treg function in EAE? Aim3: What is the interplay between T-regs and Th17inEAE?

描述（由申请人提供）：诱导CD4+CD25+FoxP3+T细胞以及它们抑制自身免疫诱导的分子和细胞机制尚未阐明。 T 细胞亚群在 EAE（一种多发性硬化症 (MS) 动物模型）调节中的作用刚刚开始被研究。为了在体内单细胞水平研究FoxP3+ T细胞，我们通过同源重组将绿色荧光蛋白（GFP）特异性引入内源性FoxpS基因组位点，并产生FoxPS敲入小鼠（FoxPSKI）。在 FoxpSKI 小鼠中，通过使用 MOG 1Ab 特异性四聚体，我们在体内跟踪了 MOG 特异性调节细胞 (T-reg) 的生成，发现 MOG 特异性 T-reg 在 EAE 过程中在中枢神经系统 (CNS) 中积累，并产生 IL-10 和意外的 IFN-g。为了研究 EAE 中 T-reg 的功能，我们将利用将 DTR/GFP（与绿色荧光蛋白融合的白喉毒素受体）报告基因引入到内源性 FoxPS 基因座的小鼠，通过给予白喉毒素，允许体内有条件地耗竭 FoxPS 表达细胞。使用这些小鼠，我们将能够通过在 EAE 的不同阶段删除 MOG 特异性 T 调节细胞来研究它们的生成、运输和体内功能。我们还发现，在 TGF-b 加 IL-6 存在的情况下，CD4+ T 细胞可以在体外分化为致病性 Th17 细胞，在单独 TGF-b 存在的情况下，CD4+ T 细胞可以分化为 T-reg 细胞。这些结果表明这两个子集在体内的功能存在相互作用，并且它们可能相互调节。然而，炎症环境和致病性Th17细胞是否可以调节T-reg细胞的产生和抑制活性尚未阐明。为了解决这些与 Th17 和 T-reg 之间相互作用相关的重要问题，我们培育了一种新型报告小鼠，其中 IL-17 表达细胞 IL-17 可以跟随红色荧光蛋白 (RFP)，并且可以通过施用白喉毒素有条件地消除这些细胞。基于我们的新试剂和初步数据，我们设计了实验来直接回答以下问题：目的1：MOG特异性Tregs不同亚群的起源和功能是什么？目标2：IL-6对EAE中Treg功能的作用是什么？目标 3：T-regs 和 Th17inEAE 之间的相互作用是什么？