Self-glycolipid-reactive T cells: repertoire & function

描述（由申请人提供）：一些T细胞识别由I类MHC样CD 1分子呈递的脂质抗原。虽然激活它们的自身脂质抗原尚未被鉴定，但大多数NK T细胞被认为是自身反应性的，并且可以被替代糖脂配体α-半乳糖基神经酰胺或α-GalCer快速激活。为了表征自身糖脂反应性T细胞、它们的表型和它们的生理功能，我们集中于识别主要的髓磷脂衍生的糖脂硫苷脂（3 '-硫酸化β-半乳糖基神经酰胺）。我们已经产生了硫酸脂-CD 1d-四聚体，并已确定在肝脏，胸腺和脾脏从幼稚C57 BL/6小鼠的硫酸脂反应性T细胞群。我们还产生了硫酸酯反应性T细胞杂交瘤。在抗原诱导的实验性自身免疫性脑脊髓炎（EAE）（人类多发性硬化症（MS）的模型）的过程中，硫酸脂-CD 1d-四聚体+ T细胞在中枢神经系统内选择性地增加数倍。硫苷脂治疗可以预防和逆转野生型EAE，但不能在CD 1d缺陷小鼠。基于我们的研究结果，我们提出，硫苷脂是一个自我糖脂配体识别的一个独特的人口的CD 1d限制性T细胞，可以被激活，以调节自身免疫反应。我们将使用硫苷脂和α-GalCer-CD 1d-四聚体和针对NK T细胞上各种细胞表面标志物的抗体来表征硫苷脂反应性T细胞的表型。将使用硫酸脂反应性T细胞杂交瘤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，并研究不同的脂质反应性T细胞群之间的相互作用。我们将确定野生型和CD 1d缺陷小鼠注射硫苷脂后NK细胞、B细胞和髓鞘蛋白反应性致病性CD 4 T细胞的数量、表型和命运。在过继转移实验中，我们将直接检查硫酸脂-CD ld-四聚体+ T细胞的调节潜力。中和抗体和1型和2型细胞因子遗传缺陷的小鼠将用于确定它们在硫苷脂调节EAE中的作用。我们将优化治疗正在进行的疾病与硫苷脂，以探索其治疗潜力。这些研究不仅对于理解天然存在的自身糖脂反应性T细胞群体的生物学很重要，而且由于CD 1d分子在物种间的高度保守性，它们将形成操纵人类自身免疫性脱髓鞘疾病（如MS）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.