Mechanisms of Human Basophil-Mediated Allergic Inflammation

描述(申请人提供)：在过去的20年里，该实验室的总体目标是阐明调节过敏性炎症的机制，从而确定治疗过敏性疾病的新靶点。这项建议将集中在调节人类嗜碱性细胞激活和变态反应性疾病的效应器功能的机制上。嗜碱性粒细胞是血液循环中最不丰富的粒细胞，直到最近才有证据表明，嗜碱性粒细胞具有强大的效应功能，可以连接先天免疫和获得性免疫，并在变态反应性和自身免疫性疾病的发病机制中发挥作用。初步研究表明，IL-3是使嗜碱性粒细胞极化的关键细胞因子，其效应功能包括FcεR1交联反应中的脱颗粒、细胞表面分子表达的调节、细胞因子的表达和免疫调节。主要的初步发现包括：1)对人嗜碱性粒细胞IL-3激活的详细分析，证明了IL-3诱导的具有不同功能特征的嗜碱性粒细胞的表型亚群；2)首次直接证据表明，人嗜碱性粒细胞表达胸腺基质淋巴生成素受体(TSLPR)并对由IL-3和IL-33调节的TSLP做出反应；以及3)有证据表明IL-3调节碱性粒细胞的“功能耐受”。这项建议将调查核心假设，即以嗜碱性粒细胞IL-3激活为靶点是治疗维甲酸嗜碱性粒细胞介导的变态反应性疾病的有效策略。研究将使用来自健康对照和过敏性疾病患者的人类嗜碱性粒细胞，以及来自IL-3缺陷和对照小鼠的小鼠嗜碱性粒细胞，以及最先进的细胞和分子技术。中心假说将通过检验以下四个推论来验证：目的1：调查假设，在缺乏IL-3的情况下，嗜碱性粒细胞处于“功能耐受”状态，并且嗜碱性粒细胞IL-3的激活打破了“功能耐受”，导致异质亚群，可能处于顺序激活状态，也可能是随机不同的亚群；目的2：研究IL-3激活的嗜碱性粒细胞对TSLP的反应，并获得极化的功能表型，以诱导Th2免疫应答；目的3：探讨嗜碱性粒细胞IL-3激活状态决定嗜碱性粒细胞激活试验对临床特应性与“功能耐受”的敏感性的假说；目的4：探讨IL-3拮抗或缺乏IL-3将导致嗜碱性粒细胞“功能耐受”的假说[一种特应性小鼠模型和一种新的体外特应性人类模型]。这些研究有可能描述嗜碱性粒细胞被激活以实现不同效应功能的机制，为临床评估嗜碱性粒细胞激活提供新的分析方法，并为潜在地将IL-3作为一种新的过敏性疾病治疗方法提供概念验证。

DESCRIPTION (provided by applicant): The overall goal of this laboratory over the past 20 years has been the elucidation of mechanisms regulating allergic inflammation and thereby identification of novel targets for treating allergic diseases. This proposal will focus on mechanisms regulating human basophil activation and effector function in allergic diseases. Basophils are the least abundant granulocytes in the circulation, and only recently has evidence demonstrated that basophils have potent effector functions that bridge innate and adaptive immunity and contribute to the pathogenesis of both allergic and autoimmune diseases. Preliminary studies demonstrate that IL-3 is a key cytokine for polarizing basophils for effector functions that include degranulation in response to FcεR1 crosslinking, regulation of cell surface molecule expression, cytokine expression, and immune modulation. Key preliminary findings include: 1) detailed analysis of IL-3 activation of human basophils that demonstrates IL-3-inducible phenotypic subsets of basophils with distinct functional characteristics; 2) the first direct evidence for human basophil expression of the thymic stromal lymphopoietin receptor (TSLPR) and signaling in response to TSLP, which is regulated by IL-3 and IL-33; and 3) evidence for basophil "functional tolerance' regulated by IL-3. This proposal will investigate the central hypothesis that targeting IL-3 activation of basophils is an effective strategy for treatin basophil-mediated allergic diseases. Studies will use human basophils from healthy controls and patients with allergic diseases, as well as murine basophils from IL-3 deficient and control mice, and state-of-the-art cellular and molecular techniques. The central hypothesis will be investigated by testing the following four corollary hypotheses: Aim 1: Investigate the hypothesis that in the absence of IL-3, basophils are in a state of "functional tolerance", and that IL-3 activation of basophils breaks "functional tolerance" and results in heterogeneous subpopulations which may be either in sequential states of activation or stochastically distinct subpopulations; Aim 2: Investigate the hypothesis that IL-3 activated basophils respond to TSLP and acquire a functional phenotype that is polarized for inducing Th2 immune responses; Aim 3: Investigate the hypothesis that the IL-3 activation state of basophils determines the sensitivity o basophil activation testing for assessment of clinical atopy versus "functional tolerance"; and Aim 4: Investigate the hypothesis that antagonism, or lack, of IL-3 will result in basophil "functional tolerance" in [a murine model of atopy and a novel in vitro human model of atopy.] These studies have the potential to delineate the mechanisms by which basophils are activated for distinct effector functions, lead to novel assays for clinical evaluation of basophil activatio, and provide proof-of- concept for potentially targeting IL-3 as a novel treatment of allergic disorders.