Follicular helper T cells as drivers of epitope spreading

滤泡辅助 T 细胞作为表位扩散的驱动因素

• 批准号: 10678864
• 负责人: Elliot Hideki Akama-Garren
• 金额: $ 5.27万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10678864
• 关键词: Adoptive Transfer; Antibody Formation; Antibody Response; Antigens; Autoantibodies; Autoantigens; Autoimmune; Autoimmune Diseases; B Cell Proliferation; B-Lymphocytes; Biological; Bone Marrow; CD4 Positive T Lymphocytes; Cell physiology; Cells; Chimera organism; Clinical; Clonal Evolution; Clonality; Clone Cells; Coculture Techniques; Development; Diagnostic; Disease; Epitope spreading; Epitopes; Event; Evolution; Functional disorder; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Goals; Hashimoto Disease; Helper-Inducer T-Lymphocyte; Immune Evasion; Immune response; Immunization; In Vitro; Individual; Length; Malignant Neoplasms; Measures; Mediating; Modeling; Molecular; Molecular Mimicry; Mus; Myasthenia Gravis; Organ failure; Pathogenesis; Pathway interactions; Peptides; Prevention; Production; Regulatory T-Lymphocyte; Research; Role; Severity of illness; Sjogren' s Syndrome; Sorting; Specificity; Structure of germinal center of lymph node; Systemic Lupus Erythematosus; T cell receptor repertoire sequencing; T-Lymphocyte; Testing; Therapeutic; Time; Tissue-Specific Gene Expression; Tissues; Untranslated RNA; Visualization; Wegener' s Granulomatosis; Wild Type Mouse; autoreactive B cell; autoreactivity; candidate identification; clinical diagnosis; clinical prognosis; cytokine; design; driving force; immune checkpoint blockade; in vivo; insight; novel therapeutic intervention; overexpression; pathogen; peripheral tolerance; receptor; response; retroviral transduction; screening; single-cell RNA sequencing

Project Summary: Autoantibodies mediate tissue damage, organ failure, and clinical decline in a range of autoimmune diseases. The reactivities of these autoantibodies may change over time leading to evolving autoimmune sequalae, termed epitope spreading. Epitope spreading correlates with disease severity and can be used for both clinical diagnosis and prognosis, but the driving forces of autoantibody responses and epitope spreading remain unclear. Characterization of these mechanisms might provide therapeutic insight to the prevention and treatment of autoantibody-mediated diseases. We have developed a model of epitope spreading in mice, in which mixed bone marrow chimeric mice develop autoantibodies to a diverse set of self-antigens. To characterize the cellular mechanisms contributing to this clonal evolution, I performed single cell RNA sequencing (scRNA-seq) of follicular T cells. Preliminary analyses have found that follicular helper T (Tfh) cells and follicular regulatory T (Tfr) cells from autoimmune chimeras expressed increased levels of several long non-coding RNAs (lncRNAs). In parallel, I have performed TCR sequencing of follicular T cells to provide paired clonotypic information, revealing T cell clones enriched in autoimmune chimeras as well as differential gene expression within individual clonotypes. These initial findings suggest that follicular T cells are transcriptionally and clonally distinct in B cell-driven autoimmune disease. I hypothesize that lncRNA expression in autoreactive Tfh cells promotes loss of peripheral tolerance, serving as a necessary and sufficient driver of epitope spreading in autoreactive germinal centers. This proposal seeks to determine whether lncRNA expression alters Tfh cell function (Aim 1), whether Tfh cells are autoreactive in B cell-driven autoimmune disease (Aim 2), and whether these Tfh cells are responsible for epitope spreading (Aim 3). Aim 1 will test if lncRNAs can alter Tfh functionality by retrovirally transducing primary T cells and characterizing their function in vitro and in vivo. Aim 2 will determine the reactivity of autoimmune-associated Tfh clonotypes by expressing these TCRs in vitro and screening their reactivity using MHC-TCR chimeric receptors. Aim 3 will determine the necessity and sufficiency of Tfh cells to promote epitope spreading by generating mixed bone marrow chimeric mice that lack Tfh cells, and adoptively transferring Tfh cells into these mice and measuring autoantibody production. The proposed project aims to provide insight into the cellular and molecular mechanisms of epitope spreading, while potentially revealing new therapeutic strategies for both autoantibody- mediated disease and other diseases of germinal center dysfunction.

项目概要： 自身抗体介导一系列自身免疫性疾病中的组织损伤、器官衰竭和临床衰退。 这些自身抗体的反应性可能随着时间的推移而改变，导致自身免疫性后遗症的发展， 称为表位扩散。表位扩散与疾病的严重程度相关，并且可以用于两种临床诊断。 诊断和预后，但自身抗体反应和表位扩散的驱动力仍然存在 不清楚这些机制的表征可能为预防和治疗提供治疗见解。 治疗自身抗体介导的疾病。我们已经开发了一种小鼠表位扩散模型， 这种混合骨髓嵌合小鼠产生了针对多种自身抗原的自身抗体。到 为了描述这种克隆进化的细胞机制，我进行了单细胞RNA 滤泡T细胞的测序（scRNA-seq）。初步分析发现，滤泡辅助性T细胞（Tfh） 来自自身免疫嵌合体的细胞和滤泡调节性T（Tfr）细胞表达增加水平的几种 长链非编码RNA（lncRNA）。同时，我对滤泡T细胞进行了TCR测序， 配对的克隆型信息，揭示了富含自身免疫嵌合体的T细胞克隆以及差异表达。 基因在个体克隆型中的表达。这些初步发现表明，滤泡T细胞是 在B细胞驱动的自身免疫性疾病中转录和克隆上不同。我假设lncRNA 自身反应性Tfh细胞中的表达促进外周耐受性的丧失，作为一种必要的和 自身反应性生发中心中表位扩展的足够驱动。该提案旨在确定 lncRNA表达是否改变Tfh细胞功能（目的1），Tfh细胞是否在B细胞驱动的 自身免疫性疾病（Aim 2），以及这些Tfh细胞是否负责表位扩散（Aim 3）。目的 1将测试lncRNA是否可以通过逆转录病毒转导原代T细胞并表征 它们在体外和体内的功能。目的2将确定自身免疫相关的Tfh克隆型的反应性 通过在体外表达这些TCR并使用MHC-TCR嵌合受体筛选它们的反应性。目标3将 确定Tfh细胞通过产生混合骨促进表位扩散的必要性和充分性 缺乏Tfh细胞的骨髓嵌合小鼠，并将Tfh细胞过继转移到这些小鼠中， 自身抗体产生。拟议的项目旨在提供深入了解细胞和分子 表位传播机制，同时可能揭示针对自身抗体和自身抗体的新治疗策略 介导的疾病和其他老年中枢功能障碍疾病。