Human T cell-mediated immunity to viruses in tissues and circulation

项目1：项目概要 了解人类的抗病毒免疫需要研究人类免疫系统的复杂性。 响应采样不同群体的个人，并研究免疫反应，他们的功能和 维持在全身的组织部位。T细胞通过自身的免疫系统来协调抗病毒适应性免疫。 淋巴组织中的活化和分化，随后迁移到感染的组织部位，以及它们的 作为病毒特异性记忆T细胞亚群在循环和不同组织部位中的长期维持。的 全身的大多数记忆T细胞是非循环的、组织驻留的记忆T（TRM）细胞 如在小鼠模型中所证明的，其可以介导保护性免疫，因此对于 了解并瞄准抗病毒免疫。研究TRM在血液仍然存在的人类中具有挑战性 最容易取样和研究的地点。十多年前，我的实验室建立了一个人体组织资源库 从单个器官供体中获得多种淋巴和粘膜组织，并优化了 来自500多名捐赠者的免疫细胞，揭示了精确的组织调节区室化和位点特异性 适应在我们目前的HIPC奖下的研究中，我们确定了T细胞特异性的普遍存在的 病毒在组织中分布和发挥作用，最初的重点是巨细胞病毒（CMV）-后来扩展到 流感特异性和最近的SARS-CoV-2特异性免疫。我们的研究结果表明， 适应性免疫反应在多个层面上受到病毒本身的控制，包括其发病机制 和组织靶点，影响T细胞功能的组织部位，与T细胞功能相关的年龄， 分化和传播，以及性别特异性变异。我们的核心假设是病毒，组织， 年龄和性别对抗病毒T细胞应答的直接特异性和可测量的影响， 其功能性和保护功效。在拟议的研究中，我们将采用全系统的方法， 通过全面评估分析人类T细胞应答的复杂性和异质性 它们在血液和组织中的分布。在目标1中，我们将量化病毒类型，组织，年龄， 和性别在抗病毒T细胞表型中的作用，并了解这些因素如何在预测模型中相互作用， 人T细胞免疫在目标2中，我们将分析病毒特异性T细胞的功能状态和克隆特征 以解决病毒特异性T细胞如何整合位点特异性和病毒特异性特征 克隆和功能特征。我们将通过CITE-Seq使用组合单细胞分析， 确定CMV、流感和SARS-CoV-2应答的配对T细胞受体克隆序列 用病毒表位库刺激后的CD 4+和CD 8 +T细胞。功能和组织特异性特征 将通过细胞因子分析和免疫测定核心的组织成像进行验证，以识别细胞-细胞 在一些实施方案中，病毒特异性T细胞的相互作用是组织中病毒特异性T细胞的额外特征。拟议的研究将共同 提供跨多种组织和病毒特异性的抗病毒T细胞免疫的功能图谱。

PROJECT 1: PROJECT SUMMARY Understanding anti-viral immunity in humans requires investigating the complexity of the human immune response—sampling diverse groups of individuals, and studying immune responses where they function and are maintained in tissue sites throughout the body. T cells coordinate anti-viral adaptive immunity through their activation and differentiation in lymphoid tissues, subsequent migration to tissue sites of infection, and their long-term maintenance as virus-specific memory T cell subsets in circulation and in diverse tissue sites. The majority of memory T cells throughout the body are non-circulating, tissue-resident memory T (TRM) cells which can mediate protective immunity, as demonstrated in mouse models, and are therefore important for understanding and targeting anti-viral immunity. Studying TRM is challenging in humans where blood remains the most readily sampled and studied site. Over 10 years ago, my laboratory set up a human tissue resource to obtain multiple lymphoid and mucosal tissues from individual organ donors, and have optimized the study of immune cells from over 500 donors, revealing precise tissue-regulated compartmentalization and site-specific adaptations. In our studies under the current HIPC award, we identified how T cells-specific for ubiquitous viruses are distributed and function in tissues, with an initial focus on cytomegalovirus (CMV)--later extended to influenza-specific and more recently to SARS-CoV-2-specific immunity. Our results reveal virus-specific adaptive immune responses are controlled on multiple levels—by the virus itself, including its pathogenesis and tissue targets, the tissue site which influences T cell functionality, age which correlates with T cell differentiation and dissemination, and sex-specific variations. Our central hypothesis is that the virus, tissue, age and sex direct specific and measurable influences on the anti-viral T cell response that together influence its functionality and protective efficacy. In the proposed study, we will apply systems-wide approaches to analyze the complexity and heterogeneity of the human T cell response through a comprehensive assessment of their profiles across blood and tissues. In Aim 1 we will quantify the contribution of virus type, tissue, age, and sex in anti-viral T cell phenotypes, and understanding how these factors interact in predictive models for human T cell immunity. In aim 2, we will analyze virus-specific T cell functional states and clonal signatures across diverse tissue sites to address how virus-specific T cells integrate both site- and virus-specific features into their clonal and functional signatures. We will use combinatorial single cell profiling by CITE-Seq with determination of paired T cell receptor clone sequences of CMV-, influenza-, and SARS-CoV-2-responding CD4+ and CD8+T cells following stimulation with viral epitope pools. Functional and tissue-specific signatures will be validated by cytokine analysis and tissue imaging by the immune assay core, to identify cell-cell interactions as additional features of virus-specific T cells in tissues. Together, the proposed studies will provide a functional map of anti-viral T cell immunity across multiple tissues and virus specificities.