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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：
7778249
负责人：
Mohamed Oukka
金额：
$ 42.61万
依托单位：
SEATTLE CHILDREN'S HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-04-15 至 2012-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7778249
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）：尚未阐明诱导CD 4 + CD 25 + FoxP 3 + T细胞及其抑制自身免疫诱导的分子和细胞机制。该T细胞亚群在EAE（多发性硬化症（MS）的动物模型）调节中的作用刚刚开始研究。为了在体内单细胞水平上研究FoxP 3 + T细胞，我们通过同源重组将绿色荧光蛋白（GFP）特异性地引入内源性FoxpS基因组位点，并产生FoxPS敲入小鼠（FoxPSKI）。在FoxpSKI小鼠中并使用MOG lAb特异性四聚体，我们在体内跟踪MOG特异性调节细胞（T-IFN）的产生，并发现MOG特异性T-IFN在EAE过程中在中枢神经系统（CNS）中积累，并产生IL-10和出乎意料的IFN-γ。为了研究T-GFP在EAE中的功能，我们将利用其中DTR/GFP（与绿色荧光蛋白融合的白喉毒素受体）报告基因已被引入内源性FoxPS基因座的小鼠，允许通过施用白喉毒素在体内条件性耗尽FoxPS表达细胞。使用这些小鼠，我们将能够通过在EAE的不同阶段删除它们来研究MOG特异性T-binding的产生、运输和体内功能。我们还发现，CD 4 + T细胞可以在体外分化成致病性Th 17细胞在TGF-β和IL-6的存在下或T-reg在TGF-β单独存在下。这些结果表明，在体内这两个子集的功能存在互惠性，它们可以相互调节。然而，炎症环境和致病性Th 17细胞是否可以调节T-reg细胞的产生和抑制活性尚未阐明。为了解决与Th 17和T-reg之间的相互作用相关的这些重要问题，我们已经产生了一种新的报告小鼠，其中表达IL-17的细胞IL-17可以被红色荧光蛋白（RFP）跟随，并且这些细胞可以通过施用白喉毒素而有条件地耗尽。基于我们的新试剂和初步数据，我们设计了直接回答以下问题的实验：目的1：MOG特异性TdR的不同子集的起源和功能是什么？目的2：IL-6在EAE中对Treg功能的作用是什么？目的3：T-IFN和Th 17 inEAE之间的相互作用是什么？