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: 10410348
• 负责人: Jennifer S Chen
• 金额: $ 3.08万
• 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/10410348
• 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; Lung Inflammation; 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; 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，哮喘的患病率越来越高，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通过以下途径在疾病中发挥核心作用 介导肥大细胞和嗜碱性粒细胞脱颗粒，释放与哮喘有关的化学介质 病情恶化。T滤泡辅助细胞(TFH)促进与特定免疫相关的高亲和力抗体 通过它们分泌的细胞因子来做出反应。TFH细胞来源的IL-4需要诱导高亲和力IgE 对过敏原的反应。然而，IL-4也被认为是一种典型的TFH细胞因子，甚至产生 在微生物免疫期间，不会产生IgE。这种研究在很大程度上依赖于IL-4细胞因子的使用 提示IL4转录激活但不一定稳定转录或翻译的报告。我们的实验室 已经建立了链格孢霉诱导的过敏性呼吸道炎症(AAI)的小鼠模型，这涉及到高水平的 亲和力IgE的产生，以及脂多糖(LPS)诱导的肺炎症，这不是。vbl.使用 在这两个模型中，我观察到两种情况下TFH细胞中IL4mRNA的表达；相比之下，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IL4 mRNA是 比内毒素诱导的炎症更稳定，揭示了一种转录后调控机制 它控制着TFH细胞中IL-4的产生。我的总体假设是，IL4的转录后调控是一种 决定TFH细胞高亲和力IgE促进能力的检查点。因此，我的建议提出了一个新的 TFH细胞中IL-4蛋白产生的调节模式和IgE介导的AAI的分子基础。 我的第一个目标是确定负责IL4转录后转录的mRNA调控序列 AAI与脂多糖诱导的肺炎症过程中TFH细胞的调节。为了做到这一点，我将克隆 IL-4表达结构在调控序列中突变以确定对IL-4 mRNA稳定性、IL-4的影响 蛋白质的产生和高亲和力的IgE反应。我的第二个目标是评估RNA结合的作用 蛋白(RBP)对AAI过程中TFH细胞产生IL-4的影响。为此，我将绘制RBP-IL4结合位点图，突变 IL-4表达中的这些结合位点，并评估这些突变对IL-4 mRNA稳定性的影响， IL-4蛋白的产生和高亲和力的IgE反应。如果成功，这个项目将定义一个以前的 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 在非过敏性反应中不适当地诱导IgE。这些发现将为新的 阻断哮喘和其他过敏性疾病中IgE发展的策略。