Regulation of T cell-derived cytokines in allergic airway inflammation

T 细胞衍生细胞因子在过敏性气道炎症中的调节

• 批准号: 10424606
• 负责人: Jennifer S Chen
• 金额: $ 5.18万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10424606
• 关键词: 3' Untranslated Regions; Affect; Affinity; Allergens; Allergic Disease; Alternaria; Antibodies; Antibody Affinity; Asthma; B-Lymphocytes; Basophils; Binding; Binding Sites; Cell Survival; Complementary DNA; Data; Development; Disease; Event; Extrinsic asthma; Genetic Transcription; Helper-Inducer T-Lymphocyte; HuR protein; IgE; Immune response; Immunization; Immunoglobulin Class Switching; Immunologics; Immunoprecipitation; Inflammation; Interleukin-4; Internal Ribosome Entry Site; Length; Lipopolysaccharides; Maps; Mediating; Mediator of activation protein; Messenger RNA; Modeling; Molecular; Mus; Mutate; Mutation; Pathogenesis; Pathogenicity; Patients; Play; Population; Post-Transcriptional Regulation; Prevalence; Production; Protein Isoforms; Proteins; Public Health; Pulmonary Inflammation; RNA-Binding Proteins; Regulation; Reporter; Role; Signal Transduction; Source; Structure of germinal center of lymph node; Surface; T cell regulation; T-Lymphocyte; Th2 Cells; Training; Transcript; Translations; airborne allergen; allergic airway disease; allergic airway inflammation; anti-IgE; asthma exacerbation; chemical release; crosslink; cytokine; improved; insight; mRNA Expression; mRNA Stability; mast cell; microbial; mouse model; prevent; response; targeted treatment; therapeutically effective

Project Summary The increasing prevalence of asthma over the past few decades signals an urgent need to understand disease pathogenesis and to develop more effective therapeutics. High affinity IgE plays a central role in the disease by mediating mast cell and basophil degranulation, which releases chemical mediators responsible for asthma exacerbation. T follicular helper (Tfh) cells promote the relevant high affinity antibodies for a given immune response through the cytokines they secrete. Tfh cell-derived IL-4 is required to induce high affinity IgE in response to allergens. However, IL-4 has also been considered a canonical Tfh cell cytokine, produced even during microbial immunizations that do not elicit IgE. Such studies largely rely on the use of an IL-4 cytokine reporter that indicates activation of Il4 transcription but not necessarily transcript stability or translation. Our lab has established murine models of Alternaria-induced allergic airway inflammation (AAI), which involves high affinity IgE production, as well as lipopolysaccharide (LPS)-induced lung inflammation, which does not. Using these two models, I observed Il4 mRNA expression in Tfh cells from both conditions; in contrast, I found that only Tfh cells in AAI produce IL-4 protein. My preliminary data suggest that Il4 mRNA in Tfh cells from AAI is more stable than that from LPS-induced inflammation, uncovering a post-transcriptional regulatory mechanism that governs IL-4 production in Tfh cells. My overall hypothesis is that post-transcriptional regulation of Il4 is a checkpoint dictating the high affinity IgE-promoting capacity of Tfh cells. My proposal therefore suggests a new paradigm for the regulation of IL-4 protein production in Tfh cells and the molecular basis of IgE-mediated AAI. My first aim is to identify the mRNA regulatory sequences responsible for Il4 post-transcriptional regulation in Tfh cells during AAI versus LPS-induced lung inflammation. To accomplish this, I will clone Il4 expression constructs mutated in regulatory sequences to determine the effect on Il4 mRNA stability, IL-4 protein production, and high affinity IgE responses. My second aim is to evaluate the role of an RNA-binding protein (RBP) on IL-4 production in Tfh cells during AAI. To do this, I will map RBP-Il4 binding sites, mutate these binding sites in Il4 expression constructs, and evaluate the effect of such mutations on Il4 mRNA stability, IL-4 protein production, and high affinity IgE responses. If successful, this project will define a previously undescribed mechanism of IgE regulation in asthma while also elucidating the immunologic rules that prevent the inappropriate induction of IgE in non-allergic responses. Such findings will offer valuable insight into new strategies for blocking IgE development in asthma and other allergic diseases.

项目摘要 在过去的几十年里，哮喘的发病率不断增加，这表明迫切需要了解疾病 发病机制和开发更有效的治疗方法。高亲和力IgE在疾病中起核心作用， 介导肥大细胞和嗜碱性粒细胞脱颗粒，从而释放导致哮喘的化学介质 加重滤泡辅助性T细胞（Tfh）促进针对给定免疫的相关高亲和力抗体 通过它们分泌的细胞因子进行反应。Tfh细胞来源的IL-4是诱导高亲和力IgE所必需的， 对过敏原的反应。然而，IL-4也被认为是典型的Tfh细胞细胞因子，甚至在Tfh细胞中产生。 在不引起IgE的微生物免疫过程中。这些研究主要依赖于使用IL-4细胞因子 指示Il 4转录激活但不一定是转录稳定性或翻译的报告基因。我们实验室 建立了链格孢菌诱导的过敏性气道炎症（AAI）小鼠模型， 亲和力IgE的产生，以及脂多糖（LPS）诱导的肺部炎症，而不是。使用 在这两种模型中，我观察了两种条件下Tfh细胞中IL 4 mRNA的表达;相反，我发现， AAI中只有Tfh细胞产生IL-4蛋白。我的初步数据表明，来自AAI的Tfh细胞中的IL 4 mRNA是 比LPS诱导的炎症更稳定，揭示了转录后调节机制 控制Tfh细胞中IL-4的产生。我的总体假设是，IL 4的转录后调控是一个重要的机制。 检查点指示Tfh细胞的高亲和力IgE促进能力。因此，我的建议提出了一个新的 Tfh细胞中IL-4蛋白产生的调节范例和IgE介导的AAI的分子基础。 我的第一个目标是确定mRNA调控序列负责IL 4转录后 AAI与LPS诱导的肺部炎症期间Tfh细胞的调节。为了实现这一点，我将克隆 在调节序列中突变的IL-4表达构建体以确定对IL-4 mRNA稳定性的影响， 蛋白质产生和高亲和力IgE应答。我的第二个目标是评估RNA结合的作用， 蛋白（RBP）对Tfh细胞在AAI期间产生IL-4的影响。为此，我将绘制RBP-I14结合位点， 这些结合位点在IL 4表达构建体中，并评估这些突变对IL 4 mRNA稳定性的影响， IL-4蛋白产生和高亲和力IgE应答。如果成功，该项目将定义一个以前 哮喘中IgE调节的未知机制，同时也阐明了预防哮喘的免疫学规则， 非过敏反应中IgE的不适当诱导。这些发现将为新的 在哮喘和其他过敏性疾病中阻断IgE发展的策略。