Self-glycolipid-reactive T cells: repertoire & function

描述（由申请人提供）：一些T细胞识别I类mhc样CD1分子呈递的脂质抗原。虽然活化NK T细胞的自体脂质抗原尚未被发现，但大多数NK T细胞被认为具有自身反应性，可以被替代糖脂配体α -半乳糖神经酰胺或α - galcer快速激活。为了表征自身糖脂反应性T细胞，它们的表型和生理功能，我们重点研究了髓磷脂来源的主要糖脂，硫脂，（3'-硫酸β -半乳糖神经酰胺）的识别。我们已经生成了硫脂- cd1 -四聚体，并在C57BL/6小鼠的肝脏、胸腺和脾脏中鉴定了硫脂反应性T细胞群。我们也产生了硫脂反应性T细胞杂交瘤。抗原诱导的实验性自身免疫性脑脊髓炎（EAE）是人类多发性硬化症（MS）的一种模型，在此过程中，中枢神经系统中的硫脂- cd1 -四聚体+ T细胞选择性地增加了数倍。在野生型小鼠中，硫脂治疗可以预防和逆转正在进行的EAE，但在cd1缺陷小鼠中则不然。基于我们的研究结果，我们提出硫脂是一种由cd1限制性T细胞识别的自身糖脂配体，可以被激活来调节自身免疫反应。我们将使用硫脂酰和a- galcer - cd1 -四聚体和针对NK T细胞上各种细胞表面标记物的抗体来表征硫脂反应性T细胞的表型。硫脂反应性T细胞杂交瘤将用于分析抗原精细特异性和TCR v基因库。使用糖脂四聚体和Elispot分析，我们将确定其激活的动力学，并研究不同脂质反应性T细胞群体之间的相互作用。我们将测定野生型和cd1缺陷小鼠注射硫脂后NK细胞、B细胞和髓磷脂蛋白反应性致病性CD4 T细胞的数量、表型和命运。在过继性转移实验中，我们将直接研究硫脂- cdld -四聚体+ T细胞的调控潜力。中和抗体和1型和2型细胞因子基因缺陷小鼠将被用来确定它们在硫脂调节EAE中的作用。我们将优化磺胺脂治疗持续性疾病，以探索其治疗潜力。这些研究不仅对理解自然发生的自糖脂反应性T细胞群的生物学很重要，而且因为CD1d分子在物种间的高度保守性，它们将形成人类自身免疫性脱髓鞘疾病（如多发性硬化症）的基础。

DESCRIPTION (provided by applicant): Some T cells recognize lipid antigens presented by class I MHC-like CD1 molecules. Although self-lipid antigens activating them have not yet been identified, most NK T cells are thought to be autoreactive and can be rapidly activated by a surrogate glycolipid ligand alpha-galactosyl ceramide or alpha-GalCer. To characterize self-glycolipid-reactive T cells, their phenotype and their physiological function, we have focused on the recognition of a major myelin-derived glycolipid, sulfatide, (3'-sulfated beta-galactosyl ceramide). We have generated sulfatide-CD1d-tetramers and have identified sulfatide-reactive T cell populations in the liver, thymus and spleen from naive C57BL/6 mice. We have also generated sulfatide-reactive T cell hybridomas. During the course of antigen-induced experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis (MS) in humans, sulfatide-CD1d-tetramer+ T cells are selectively increased several-fold within the central nervous system. Treatment with sulfatide can both prevent and reverse ongoing EAE in wild type but not in CD1d-deficient mice. Based on our findings we propose that sulfatide is a self-glycolipid ligand recognized by a distinct population of CD1d-restricted T cells that can be activated to modulate autoimmune responses. We will use sulfatide- and a-GalCer-CD1d-tetramers and antibodies against various cell surface markers on NK T cells to characterize the phenotype of sulfatide-reactive T cells. Sulfatide-reactive T cell hybridomas will be used to analyze the antigen fine specificity and the TCR V-gene repertoire. Using glycolipid tetramers and Elispot analysis we will determine the dynamics of their activation and investigate mutual interactions among different lipid-reactive T cell populations. We will determine the number, phenotype and the fate of NK cells, B cells and myelin protein-reactive pathogenic CD4 T cells following sulfatide injection of wild type and CD1d-deficient mice. In adoptive transfer experiments we will directly examine the regulatory potential of sulfatide-CD ld-tetramer+ T cells. Neutralizing antibodies and mice genetically deficient for type 1 and 2 cytokines will be used to determine their role in the modulation of EAE by sulfatide. We will optimize treatment of ongoing disease with sulfatide in order to explore its therapeutic potential. These studies are important not only for understanding the biology of a naturally occurring self-glycolipid-reactive T cell population, but also because of the highly conserved nature of CD1d molecules across species they will form the basis for manipulation of human autoimmune demyelinating diseases, such as MS.