Lymph node stromal cells coordinate immune cell environments during Aspergillus fumigatus infection

烟曲霉感染期间淋巴结基质细胞协调免疫细胞环境

• 批准号: 10751936
• 负责人: Cheryl Mai
• 金额: $ 5.26万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10751936
• 关键词: ATAC-seq; Adoptive Transfer; Affect; Allergic Bronchopulmonary Aspergillosis; Anatomy; Antifungal Agents; Antigen-Presenting Cells; Antigens; Antimicrobial Resistance; Aspergillosis; Aspergillus; Aspergillus fumigatus; CCL20 gene; CCR6 gene; CD4 Positive T Lymphocytes; CXCL10 gene; CXCL9 gene; CXCR3 gene; Cell Communication; Cell Differentiation process; Cell physiology; Cells; Chronic; Competence; Critical Illness; Cues; Data; Development; Disease; Dissection; Dissociation; Effector Cell; Environment; Epigenetic Process; Exhibits; Gene Targeting; Genetic; Goals; Health; Host Defense; Human; Image Analysis; Immune; Immune System Diseases; Immune response; Immunity; Immunobiology; Immunofluorescence Immunologic; Immunofluorescence Microscopy; Immunologic Deficiency Syndromes; Infection; Inflammatory; Knowledge; Life; Ligands; Lymphocyte; Lymphoid Tissue; Maps; Mediastinal lymph node group; Methods; Microscopy; Modification; Molecular; Mus; Mycoses; Neighborhoods; Patients; Pattern; Physicians; Population; Positioning Attribute; Primary Infection; Process; Property; Proteins; Resolution; Respiratory Tract Infections; Reticular Cell; Role; Scientist; Signal Transduction; Source; Stromal Cells; System; T cell differentiation; T cell response; T-Lymphocyte; T-Lymphocyte Subsets; Testing; Therapeutic; Training; Vaccines; adaptive immune response; adaptive immunity; cell type; chemokine; chemokine receptor; clinically relevant; confocal imaging; cytokine; design; dimensional analysis; draining lymph node; effector T cell; experience; human disease; imaging approach; immunomodulatory strategy; improved outcome; insight; lymph node microenvironment; lymph nodes; member; memory retention; mortality; mouse model; novel; novel strategies; novel therapeutic intervention; pathogen; pathogenic fungus; receptor; response; secondary lymphoid organ; segregation; single-cell RNA sequencing; spatiotemporal; therapy development; tool; trafficking; transcriptomic profiling; transcriptomics; vaccine development

Project Summary The opportunistic fungal pathogen Aspergillus fumigatus presents a major health concern in immunodeficient and critically ill patients, with invasive disease contributing to high mortality rates. Infection with A. fumigatus elicits a diverse adaptive CD4 T cell response. Our preliminary data have demonstrated distinct TH1 and TH17 spatial neighborhoods within mediastinal lymph nodes (LNs) during A. fumigatus airway infection. This spatial organization positions activated CD4 T cells to receive tailored signals (e.g., antigens, cytokines) for optimal effector cell differentiation and function. Lymph node stromal cells (LNSCs) are known to provide guidance cues for immune cell trafficking during monotypic effector T cell responses. However, the mechanisms by which distinct TH1 and TH17 microenvironments are concurrently established within LNs remain unknown. Defining these mechanisms will offer insights into the functional importance of LN microenvironments in establishing human immunity to infection and inflammatory disorders for the identification of novel therapeutic strategies. Based upon our single-cell RNA-sequencing and flow cytometric analyses of FRC and effector T cell subsets during A. fumigatus challenge, we propose that (i) FRC subsets establish spatially distinct TH1 and TH17 neighborhoods by dynamically regulating chemotactic receptor-ligand axes, such as CXCR3:CXCL9/CXCL10 and CCR6:CCL20 and that ii) these neighborhoods can persist after pathogen clearance. We will address these hypotheses in a clinically relevant murine model of invasive pulmonary aspergillosis. In Specific Aim 1, we will characterize distinct spatiotemporal microenvironments formed by CD4 T cells and FRCs in A. fumigatus infection by applying spatial transcriptomic profiling followed by high-content immunofluorescence methods paired with a novel computational approach for image analysis. In Specific Aim 2, we will define the functional role of CD4 T cell spatial organization in A. fumigatus infection by perturbing FRC- dependent chemokine gradients in a cell type-dependent manner using FRC-specific gene targeting. We will also conduct adoptive co-transfers of A. fumigatus-specific CD4 T cells sufficient and deficient in CXCR3 and CCR6. In Specific Aim 3, we will investigate the durability of infection-driven spatial FRC diversity by studying epigenetic modifications in A. fumigatus-experienced FRCs. We will also employ high-content confocal imaging to determine whether prior infections affect FRC formation of T cell neighborhoods in subsequent infections. This study employs and develops novel genetic tools, microscopy methods, and computational approaches to generate a systems level understanding of secondary lymphoid organ immunobiology. Furthermore, this proposal is tailored for a physician-scientist in training as it investigates the mechanisms by which stromal cells induce adaptive immunity to the clinically relevant pathogen A. fumigatus, with implications for anti-fungal therapeutic strategies and vaccine development.

项目概要 机会性真菌病原体烟曲霉是免疫缺陷患者的主要健康问题 以及重症患者，侵袭性疾病导致高死亡率。烟曲霉感染 引发多种适应性 CD4 T 细胞反应。我们的初步数据显示了 TH1 和 TH17 的不同 烟曲霉气道感染期间纵隔淋巴结（LN）内的空间邻域。这个空间 组织定位激活的 CD4 T 细胞以接收定制信号（例如抗原、细胞因子），以获得最佳效果 效应细胞的分化和功能。众所周知，淋巴结基质细胞 (LNSC) 可以提供指导线索 用于单型效应 T 细胞反应期间的免疫细胞运输。然而，其机制 LN 内同时建立的不同 TH1 和 TH17 微环境仍然未知。定义 这些机制将深入了解液氮微环境在建立 人类对感染和炎症性疾病的免疫力，以确定新的治疗策略。 基于我们对 FRC 和效应 T 细胞亚群的单细胞 RNA 测序和流式细胞术分析 在烟曲霉挑战期间，我们建议 (i) FRC 子集建立空间上不同的 TH1 和 TH17 通过动态调节趋化受体-配体轴来调节邻域，例如 CXCR3:CXCL9/CXCL10 和 CCR6:CCL20 以及 ii) 这些邻域在病原体感染后仍能持续存在 清除。我们将在临床相关的侵袭性肺鼠模型中解决这些假设 曲霉病。在具体目标 1 中，我们将描述 CD4 形成的不同时空微环境的特征 通过应用空间转录组分析和高内涵分析烟曲霉感染中的 T 细胞和 FRC 免疫荧光方法与图像分析的新型计算方法相结合。特定目标 2，我们将通过干扰 FRC- 来定义 CD4 T 细胞空间组织在烟曲霉感染中的功能作用 使用 FRC 特异性基因靶向以细胞类型依赖性方式产生依赖性趋化因子梯度。我们将 还对 CXCR3 充足和缺乏的烟曲霉特异性 CD4 T 细胞进行过继共转移 CCR6。在具体目标 3 中，我们将通过研究感染驱动的空间 FRC 多样性的持久性 烟曲霉经历的 FRC 中的表观遗传修饰。我们还将采用高内涵共焦成像 以确定先前的感染是否影响后续感染中 T 细胞邻域的 FRC 形成。这 研究采用并开发了新颖的遗传工具、显微镜方法和计算方法 产生对次级淋巴器官免疫生物学的系统水平理解。此外，这 该提案是为正在接受培训的医生科学家量身定制的，因为它研究了基质细胞的机制 诱导对临床相关病原体烟曲霉的适应性免疫，对抗真菌具有影响 治疗策略和疫苗开发。