Myeloid-Derived Regulatory Cells in Asthma

 描述(申请人提供)：目的：本提案的目的是探讨由产生自由基髓系来源的调节细胞(MDRC)产生的氧化修饰自体多肽可触发呼吸道高反应性(AHR)这一新概念。我们最近将多药耐药细胞描述为小鼠和人类呼吸道炎症的关键调节因子。MDRCs利用活性氧和活性氮(ROS和RNS)促进T细胞增殖，加重AHR。我们最近的研究表明，MDRCs可以诱导自体多肽的硝化和氧化修饰，而自体多肽是一种免疫原性新抗原，但尚未建立耐受性。因此，这些新抗原可以引发代表一种新的自身免疫形式的病理性炎症反应。因此，多药耐药细胞是耐受性和炎症之间平衡的调节者。在目标1中，我们将鉴定哮喘患者中促炎症的气道MDRCs产生的修饰的自身抗原/抗原肽。我们将确定从正常人和哮喘患者分离的产生O2的气道MDRCs的HLA-Class II分子结合的肽库，并确定ROS和RNS诱导的这些自肽的修饰。这些研究将通过从支气管肺泡灌洗(BAL)MDRC中洗脱II类结合多肽，并通过质谱仪鉴定这些多肽的硝化和氧化修饰来进行。在目标2中，我们将确定哮喘患者中产生ROS的MDRCs呈递的修饰的自身蛋白是否是真正的新抗原。从健康和哮喘患者中提纯的外周血T细胞和呼吸道MDRC，以及体外修饰的自身蛋白/肽将用于功能分析， 限制稀释分析和共培养，以研究T细胞增殖反应、克隆性增殖和Th极化。我们将使用哮喘小鼠模型来研究MDRC介导的Th极化的分子机制。这些研究将提供证据，证明MDRCs在哮喘的免疫耐受和炎症调节中发挥重要作用，并阐明哮喘的新致病范式。识别翻译后修饰的多肽新抗原有助于确定表征哮喘表型的生物标记物。这些研究还有可能开发新的和改进的治疗策略，以多药耐药细胞为目标，用于哮喘表型亚型的疾病控制(即，精确/个性化的药物方法)。我们将获得潜在的针对抗原特异性T细胞的新型多肽免疫治疗策略，这些多肽代表了修饰的T细胞表位。。

 DESCRIPTION (provided by applicant): Objective: The goal of this proposal is to investigate the novel concept that oxidant-modified self-peptides produced by free radical producing myeloid-derived regulatory cells (MDRCs) can trigger airway hyper- responsiveness (AHR). We recently characterized MDRCs as critical regulators of airway inflammation in both mice and humans. MDRCs use reactive oxygen and reactive nitrogen species (ROS and RNS) to enhance T cell proliferation and exacerbate AHR. Our recent studies show that MDRCs induce nitrative and oxidative modifications of self-peptides which are immunogenic neo-antigens for which tolerance has not been established. Consequently, these neo-antigens can elicit pathologic inflammatory responses that represent a novel form of autoimmunity. MDRCs thus are regulators of balance between tolerance and inflammation. In Aim 1, we will identify modified self-antigens/antigenic peptides produced by pro-inflammatory airway MDRCs in asthmatics. We will determine the peptide repertoire bound to HLA-Class II molecules of O2.-- producing airway MDRCs isolated from normal and asthmatic subjects, and define the ROS- and RNS-induced modifications of these self-peptides. These studies will be conducted by eluting the Class II-bound peptides from bronchoalveolar lavage (BAL) MDRCs, and identifying the nitrative and oxidative modifications of these peptides by mass spectrometry. In Aim 2, we will determine if modified self-proteins presented by ROS- producing MDRCs in asthmatics are true neo-antigens. Peripheral blood T cells and airway MDRCs purified from healthy and asthmatic subjects, and in-vitro modified self-proteins/peptides will be used in functional assays, limiting dilution analyses and co-cultures to investigate T cell proliferative responses, clonal proliferations and Th polarization. We will use a murine model of asthma to examine molecular mechanisms of MDRC-mediated Th polarization. These studies will provide evidence of a major role for MDRCs as regulators of immune tolerance and inflammation in asthma, and elucidate a new pathogenic paradigm for asthma. Identification of post- translational modified peptide neo-antigens can help define biomarkers to characterize asthma phenotypes. These studies also have the potential to enable development of new and improved therapeutic strategies to target MDRCs for disease control of subsets of asthma phenotypes (i.e., a precision/personalized medicine approach). We will obtain insight for potential novel peptide immunotherapy strategies targeting antigen- specific T cells with novel synthetic peptides representing modified T cell epitopes. .