CHEMOKINE REGULATION OF NKT CELLS, AND ACAID

趋化因子与多种炎性和自身免疫性疾病相关，因为它们在将细胞募集到免疫诱导和炎症部位中起作用。 然而，很少有研究探讨了趋化因子在外周耐受诱导中的作用，也没有关于NKT细胞特异性趋化因子的研究报道。 这项研究计划将探讨特定趋化因子在免疫炎症反应的负调控中的作用，特别是在招募NKT细胞到耐受诱导部位中的作用。 前房相关免疫偏离（ACAID）与抗原进入眼内后迟发型超敏反应（DTH）的选择性负调节有关。 脾NKT细胞的积累是ACAID发展所必需的。初步数据显示MIP-2与NKT细胞募集相关。 我们将使用分子、细胞和形态学技术来鉴定和确定趋化因子在耐受诱导过程中的作用。 目的通过RNase保护试验和多色流式细胞术检测趋化因子的表达谱。 第二个目标将使用经典的趋化性测定来确定NKT细胞对ACAID相关趋化因子的迁移反应，沿着分子技术来确认这些细胞上趋化因子受体的表达。 第三个目标是通过用来自趋化因子受体敲除小鼠的NKT细胞重建NKT细胞敲除小鼠来探索趋化因子-NKT细胞相互作用在ACAID诱导过程中的机制。 局部过继转移测定将测量是否产生调节性T细胞。 最后，第四个目标将探讨这一假设，即招募到脾脏诱导ACAID的细胞必须形成相互作用的细胞簇。 在识别出这些簇和细胞后，我们将调节趋化因子和潜在的粘附分子，并通过簇的质量和数量来评估结果。 NKT细胞缺陷和/或缺乏与小鼠和人类中的多种自身免疫性疾病（狼疮、1型糖尿病、系统性硬皮病）相关。 因此，NKT细胞介导的调节性T细胞诱导机制可能在除了免疫特权部位如眼睛之外的多种器官和组织中维持自身耐受中起作用。 该提案中提出的关于趋化因子、先天细胞和外周耐受性的新假设代表了迄今为止完全未探索的研究领域。

Chemokines are associated with a variety of inflammatory and autoimmune diseases for their role in recruitment of cells to the sites of immune induction and inflammation. However, few studies have explored a role for chemokines in peripheral tolerance induction and no studies are reported for specific chemokines for NKT cells. This research plan will explore the role of specific chemokines in negative regulation of an immune inflammatory response and specifically in the recruitment of NKT cells to the site of tolerance induction. Anterior Chamber Associated Immune Deviation (ACAID) is associated with the selective negative regulation of delayed type hypersensitivity (DTH) after the introduction of antigen into the eye. An accumulation of splenic NKT cells is absolutely required for the development of ACAID. Preliminary data show that MIP-2 is associated with NKT cell recruitment. We will use molecular, cellular and morphological techniques to identify and define the role of the chemokines in the tolerance inducing process. Aim One will define the chemokine profile by RNase Protection Assay and multi-color flow cytometry. The second aim will use classic chemotaxis assays to define the migratory response of NKT cells to ACAID associated chemokines, along with molecular techniques to confirm the expression of chemokine receptors on these cells. The third aim will explore the mechanisms of chemokine-NKT cell interaction during ACAID induction by reconstituting NKT cell knockout mice with NKT cells from chemokine receptor knockout mice. A local adoptive transfer assay will measure if regulator T cells were generated. Finally, the fourth aim will explore the hypothesis that the cells recruited to the spleen for the induction of ACAID must necessarily form interactive cell clusters. After identifying such clusters and cells within, we will modulate chemokines and potential adhesion molecules and evaluate the outcome by quality and quantity of clusters. NKT cell defects and/or deficiency is associated with a variety of autoimmune disorders in both mice and humans (lupus, type 1 diabetes, systemic scleroderma). Thus, the mechanisms of NKT cell mediated induction of T regulatory cells might function in the maintenance of self tolerance in a variety of organs and tissues in addition to immune privileged sites such as the eye. The novel postulates about chemokines, innate cells and peripheral tolerance presented in this proposal, represent a heretofore totally unexplored area of research.