Roles of FFAR 3-SCFA axis in Th2 cytokine production by tissuelymphocytes in EoE

FFAR 3-SCFA 轴在 EoE 组织淋巴细胞产生 Th2 细胞因子中的作用

• 批准号: 10307578
• 负责人: Marc E. Rothenberg
• 金额: $ 39.75万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-12-01 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10307578
• 关键词: ATAC-seq; Acetylation; Agonist; Allergic; Allergic Disease; Allergic inflammation; Asthma; Binding; Biopsy; Blood; Butyrates; CD3 Antigens; Cell Line; Cells; ChIP-seq; Characteristics; Chromatin; Chromatin Structure; Chronic; Clinical; Data; Diet; Dietary Fiber; Disease; Enhancers; Eosinophilic Esophagitis; Epigenetic Process; Esophageal Tissue; Esophageal mucous membrane; Esophagus; Exhibits; FFAR3 gene; Food; Food Hypersensitivity; G-Protein-Coupled Receptors; GATA3 gene; Gene Chips; Genes; Genetic Transcription; Genomic DNA; Grant; Helper-Inducer T-Lymphocyte; Histones; Human; IL5 gene; Immune; Immune response; Immunity; Inflammation; Inflammatory; Interleukin-13; Interleukin-4; Interleukin-5; Knowledge; Leukocytes; Ligands; Lymphocyte; Lysine; Measures; Mediating; Messenger RNA; Modeling; Molecular; Mouse Cell Line; Mus; Pathogenesis; Pathogenicity; Patients; Population; Production; Proteins; Regulation; Regulatory Element; Resolution; Rodent; Role; Source; T-Cell Receptor; T-Lymphocyte; Testing; Therapeutic; Tissues; Transcript; Volatile Fatty Acids; base; clinical care; cohort; cytokine; eosinophil; gastrointestinal; gastrointestinal bacteria; gastrointestinal symptom; genetic variant; genome-wide; human tissue; immunoregulation; improved; in vivo; innovation; interest; knockout gene; microbiome; microbiota; mouse model; novel; overexpression; receptor; response; selective expression; single-cell RNA sequencing; therapeutic target; transcription factor

Eosinophilic esophagitis (EoE) is a chronic, immune-mediated, allergic disease characterized by remarkable eosinophil accumulation in the esophageal mucosa. Single-cell RNA sequencing (scRNA-Seq) indicated that the pathogenic tissue Th2 cells are specifically enriched in EoE tissue, with a large amount of Th2 cytokine production and a unique signature. Notably, by bulk and single-cell CD3+ T cell sequencing, we identified that the gene of the short-chain fatty acid (SCFA, e.g., butyrate) receptor FFAR3 is the top tracking gene with IL5, with both genes uniquely expressed in tissue Th2 cells at the single-cell level. FFAR3 can be induce by IL-4 in circulating naïve T cells. We also found that butyrate, a putative ligand of FFAR3 and a metabolite produced by esophageal microbiota, significantly enhanced Th2 cytokine production in a human and mouse cell line, primary T cells, and a model of murine experimental asthma. These collective findings prompt our central hypothesis Our overall hypothesis is that FFAR3, ligated to microbiome-originated SCFA in esophageal mucosa, is induced by IL-4 in tissue effector Th2 cells and enhances type 2 responses in esophageal T cells that locally contribute to EoE pathogenesis. The proposed study will delineate unique populations of tissue Th2 cells and elucidate a novel and pivotal role of FFAR3-SCFA axis in Th2 activation, Th2 cytokine production and EoE molecular pathogenesis. Our preliminary data show that FFAR3 is an IL-4–induced SCFA receptor that regulates Th2 cytokine production by Th2 cells in vivo and ex vivo. We will substantiate whether the boosting effect of butyrate in circulating T cells occurs in tissue-resident T cells and whether there is a difference between normal control and EoE; measure the SCFA concentrations in the EoE and control esophageal tissue; and assess the SCFA and dietary fiber content in an EoE vs. non-EoE diet. We will also employ a T cell–specific FFAR3-deficient mouse to study the involvement of SCFA in murine Th2 immunity. By ChIP-Seq and ATAC-Seq, we will define epigenetic alterations induced by SCFAs, explore FFAR3's roles in altering chromatin structure at the Th2 cytokine locus, identify new Th2-promoting transcription factors driven by FFAR3 and analyze binding of the Th2 transcription factor GATA3 to genomic DNA regions in human T cells in the context of butyrate exposure. To profile the FFAR3+ tissue Th2 cells with high resolution, we will characterize a cohort of 5000 residential esophageal lymphocytes by scRNA-Seq, analyze Th2-specific markers and transcription factors, scrutinize the TCR clonotypes of tissue Th2 cells and correlate tissue Th2 cells' key parameters to EoE clinical characteristics. Collectively, this grant focuses on understanding human tissue-resident T cells and their contribution to EoE in the context of FFAR3-SCFA axis. The immediate significance of this study is its potential to uncover tissue Th2 cytokine production mechanisms, the novel functionality of SCFA in Th2 inflammation and the characterization of the newly identified FFAR3+ tissue Th2 cells, all of which may provide new rationales for improved therapeutic strategies for allergic disorders.

嗜酸性粒细胞性食管炎 (EoE) 是一种慢性、免疫介导的过敏性疾病，其特征是显着的 单细胞 RNA 测序 (scRNA-Seq) 表明食管粘膜中嗜酸性粒细胞积聚。 致病组织Th2细胞在EoE组织中特异富集，含有大量Th2细胞因子 值得注意的是，通过批量和单细胞 CD3+ T 细胞测序，我们发现了这一点。 短链脂肪酸（SCFA，例如丁酸）受体基因 FFAR3 是与 IL5 一起的顶级追踪基因， 这两个基因在单细胞水平上在组织 Th2 细胞中独特表达，可被 IL-4 诱导。 我们还发现了丁酸，FFAR3 的假定配体和 FFAR3 产生的代谢物。 食道微生物群，显着增强人类和小鼠细胞系中 Th2 细胞因子的产生， 原代 T 细胞和小鼠实验性哮喘模型这些集体发现提示了我们的中心。 总体假设 我们的假设是 FFAR3，与食管中微生物组来源的 SCFA 连接 粘膜，由组织效应 Th2 细胞中的 IL-4 诱导，增强食管 T 细胞的 2 型反应 拟议的研究将描述独特的 EoE 发病机制的细胞。 组织 Th2 细胞并阐明 FFAR3-SCFA 轴在 Th2 激活、Th2 细胞因子中的新颖且关键的作用 我们的初步数据表明 FFAR3 是 IL-4 诱导的 SCFA。 体内和离体调节 Th2 细胞产生 Th2 细胞因子的受体 我们将证实是否存在。 丁酸盐对循环 T 细胞的促进作用发生在组织驻留 T 细胞中，以及是否存在 正常对照和 EoE 之间的差异；测量 EoE 和对照中的 SCFA 浓度 我们还将评估 EoE 与非 EoE 饮食中的 SCFA 和膳食纤维含量。 使用 T 细胞特异性 FFAR3 缺陷小鼠来研究 SCFA 在小鼠 Th2 免疫中的作用。 ChIP-Seq 和 ATAC-Seq，我们将定义 SCFA 诱导的表观遗传改变，探索 FFAR3 在 改变 Th2 细胞因子基因座的染色质结构，识别新的 Th2 促进转录因子驱动 通过 FFAR3 并分析 Th2 转录因子 GATA3 与人类 T 基因组 DNA 区域的结合 为了以高分辨率分析 FFAR3+ 组织 Th2 细胞，我们将在丁酸盐暴露的情况下对细胞进行分析。 通过 scRNA-Seq 表征 5000 个住宅食管淋巴细胞队列，分析 Th2 特异性 标记物和转录因子，检查组织 Th2 细胞的 TCR 克隆型并关联组织 Th2 总的来说，这项资助的重点是了解人类的 EoE 临床特征。 组织驻留 T 细胞及其在 FFAR3-SCFA 轴背景下对 EoE 的贡献。 这项研究的意义在于它有可能揭示组织 Th2 细胞因子的产生机制， SCFA 在 Th2 炎症中的功能以及新鉴定的 FFAR3+ 组织 Th2 的表征 细胞，所有这些都可能为改进过敏性疾病的治疗策略提供新的原理。