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， 嗜酸性粒细胞在食管粘膜中积聚。单细胞RNA测序（scRNA-Seq）表明， 致病组织Th 2细胞特异性地富集在EoE组织中，具有大量Th 2细胞因子 生产和独特的签名。值得注意的是，通过批量和单细胞CD 3 + T细胞测序，我们确定， 短链脂肪酸（SCFA，例如，丁酸）受体FFAR 3是IL 5的最高追踪基因， 其中两种基因在组织Th 2细胞中以单细胞水平唯一表达。FFAR 3可以被IL-4诱导， 循环的幼稚T细胞。我们还发现，丁酸，一个假定的配体FFAR 3和代谢产物产生的 食管微生物群，显著增强人和小鼠细胞系中Th 2细胞因子的产生， 原代T细胞和小鼠实验性哮喘模型。这些集体发现促使我们的中央 假设我们的总体假设是FFAR 3，连接到食管中微生物组来源的SCFA， 在组织效应器Th 2细胞中，IL-4诱导粘膜，并增强食管T细胞中的2型反应。 局部参与EoE发病机制的细胞。拟议的研究将描绘独特的人口， 组织Th 2细胞并阐明FFAR 3-SCFA轴在Th 2活化、Th 2细胞因子 EoE的分子发病机制。我们的初步数据表明FFAR 3是IL-4诱导的SCFA， 在体内和离体调节Th 2细胞产生Th 2细胞因子的受体。我们将证实 丁酸盐在循环T细胞中的增强作用发生在组织驻留T细胞中， 正常对照和EoE之间的差异;测量EoE和对照中的SCFA浓度 食管组织;并评估EoE与非EoE饮食中的SCFA和膳食纤维含量。我们还将 使用T细胞特异性FFAR 3缺陷小鼠研究SCFA在鼠Th 2免疫中的参与。通过 ChIP-Seq和ATAC-Seq，我们将定义SCFA诱导的表观遗传学改变，探索FFAR 3在 改变Th 2细胞因子位点的染色质结构，鉴定新的Th 2促进转录因子驱动 分析Th 2转录因子GATA 3与人T细胞基因组DNA区域的结合， 细胞在丁酸盐暴露的情况下。为了以高分辨率分析FFAR 3+组织Th 2细胞，我们将 通过scRNA-Seq表征5000个驻留食管淋巴细胞的队列，分析Th 2特异性 标记物和转录因子，仔细检查组织Th 2细胞的TCR克隆型，并将组织Th 2 细胞的关键参数EoE临床特征。总的来说，这笔赠款的重点是了解人类 组织驻留T细胞及其在FFAR 3-SCFA轴背景下对EoE的贡献。立即 这项研究的意义在于它有可能揭示组织Th 2细胞因子的产生机制， SCFA在Th 2炎症中的功能和新鉴定的FFAR 3+组织Th 2的表征 细胞，所有这些都可能为改善过敏性疾病的治疗策略提供新的理论基础。