CD4+ T cell affinity for self and foreign antigens in the CNS

迄今为止，我们评估CD4+ T细胞频率和亲和力的最佳工具是肽：MHC （pMHC）四聚体或功能测定；这两种方法都不能有效地识别出针对特定抗原的所有应答细胞。这与CD8+ T细胞形成对比，这些技术被证明更准确。一般来说，II类抗原的pMHC四聚体很难产生，并且在应答的CD4+ T细胞中识别出一小部分，特别是当抗原来源于自身蛋白时。原因是四聚体是基于亲和力的，如果亲和力太低，那么四聚体在评估反应中的作用就有限了。功能性反应也低估了抗原反应性CD4+ T细胞的数量。例如，在针对髓鞘抗原的反应中，效应细胞因子反应通常使用强药理学试剂（如PMA和离子霉素）来确定，它们打击T细胞信号级联，可能与抗原本身反应中产生的细胞因子无关。为了更好地评估CD4+ T细胞的应答范围，我们已经开始使用基于微移液管的亲和试验。我们在此报告了CD4+ T细胞针对髓鞘或病毒抗原的频率和亲和力评估的重大进展。因此，首次有可能在疾病进展过程中跟踪CD4+ T细胞反应的亲和力范围。我们的初步研究结果导致了以下中心假设，即CD4+ T细胞亲和力直接影响疾病结局和免疫介导的中枢神经系统组织损伤。针对CD4+ T细胞自身抗原髓鞘少突胶质细胞糖蛋白和病原体相关淋巴细胞脉络丛脑膜炎病毒抗原，提出了三个特异性目标来验证这一假设。目的1-确定CD4+ T细胞在慢性自身免疫性疾病和病毒感染过程中的亲和力。目的2-定义T细胞亲和力和效应表型之间的联系。目的3-建立高亲和力T细胞和低亲和力T细胞对中枢神经系统中发现的抗原的反应。

DESCRIPTION (provided by applicant): To date, our best tools for assessing the frequency and affinity of CD4+ T cells have been peptide:MHC (pMHC) tetramers or functional assays; neither of which effectively identify all of the responding cells to a particular antigen. This is in contrast to CD8+ T cells where these techniques prove more accurate. Generally, pMHC tetramers for class II antigens are difficult to produce and identify a small percentage of responding CD4+ T cells especially when antigen is derived from self proteins. The reason for this is that tetramer is based on affinity and if affinity is too low, then the tetramer is of limited use in assessing the response Functional responses also underestimate the number of antigen reactive CD4+ T cells. In the case of responses directed against myelin antigens for instance, the effector cytokine response is often determined using strong pharmacologic agents such as PMA and ionomycin, which hammer the T cell signaling cascade and may have little relevance to the cytokines being produced in response to antigen itself. To better assess the range of responding CD4+ T cells, we have begun to use micropipette based affinity assay. We report here a major advancement in assessment in the frequency and affinity of CD4+ T cells directed against myelin or viral antigens. Therefore for the first time, it is possible to track the range of affinities of a CD4+ T cells response during disease progression. Our preliminary findings have led to the following central hypothesis that the CD4+ T cell affinity profile directly impacts disease outcome and immune mediated tissue damage in the CNS. Three specific aims are proposed to test this hypothesis focused on CD4+ T cells specific for self antigen myelin oligodendrocyte glycoprotein and pathogen associated lymphocytic choriomeningitis virus antigens. Aim 1- Identify the affinity of CD4+ T cells over the course of chronic autoimmune disease and viral infection. Aim 2- Define the connection between T cell affinity and effector phenotype. Aim 3- Establish the contribution of high versus low affinity T cells in response to antigens found in the CNS. PUBLIC HEALTH RELEVANCE: CD4+ T cells play a major role in the pathogenesis of autoimmune responses such as multiple sclerosis (MS) and are critical for proper response to chronic viral infections. The overall goal of this grant is to define the range of T cell affinities for myelin and viral antigens in the central nervous system (CNS. The proposed studies apply a novel means for assessing the affinity if the responding T cell response in the murine EAE and LCMV models, but our results should be rapidly transferable to patients. Therefore, our findings are highly relevant to human health and will prove to be a valuable asset for assessing disease prognosis and for the development of effective treatments.