The cellular mechanisms of immunological memory development in COVID-19 patients

COVID-19患者免疫记忆发展的细胞机制

基本信息

批准号：
10465343
负责人：
WEIGUO CUI
金额：
$ 42.85万
依托单位：
VERSITI WISCONSIN, INC.
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-01 至 2022-08-26
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10465343
关键词：
2019-nCoV Acute Address Adoptive Transfer Affinity Age Algorithms Antibody Formation Antibody Response Antiviral Agents Autoimmunity Autologous B-Lymphocytes Bar Codes Biological Assay Biological Markers Blood CD4 Positive T Lymphocytes CD8-Positive T-Lymphocytes COVID-19 COVID-19 pandemic COVID-19 patient Cell Communication Cell physiology Cells Cellular Immunity Clinical Clone Cells Coculture Techniques Coronavirus Coronavirus Infections Cryopreservation Defect Development Disease Disease Outcome Effector Cell Elderly Epitopes Equilibrium Flow Cytometry Generations Genetic Transcription Health Helper-Inducer T-Lymphocyte Heterogeneity Human Humoral Immunities Immune response Immunity Immunoglobulin Class Switching Immunologic Memory In Vitro Infection Instruction Knowledge Measurement Measures Mediating Memory Memory B-Lymphocyte Modeling Molecular Genetics Monitor Mus Mutation Nature Pathway Analysis Pathway interactions Patients Peptide Library Peripheral Blood Mononuclear Cell Phase Phenotype Plasma Cells Population Positioning Attribute Production Regulator Genes Regulatory T-Lymphocyte Research SARS-CoV-2 antibody SARS-CoV-2 immune response SARS-CoV-2 immunity SARS-CoV-2 infection SARS-CoV-2 pathogenesis Sampling Severity of illness Signal Transduction T cell differentiation T cell receptor repertoire sequencing T cell response T memory cell T-Lymphocyte T-cell receptor repertoire Techniques Technology Testing Th1 Cells Tissues Vaccine Design Viral Viral Load result Viral Pathogenesis Virus Virus Diseases base biobank combat comorbidity cross reactivity cytokine effector T cell exhaust exhaustion experimental study immunopathology improved individual patient innovative technologies insight interest neutralizing antibody novel novel coronavirus pathogen patient stratification programs response single cell sequencing single-cell RNA sequencing tool transcription factor transcriptome sequencing vaccination strategy vaccine development

项目摘要

The recent unexpected emergence of the COVID-19 pandemic has spurred significant interest in an improved understanding of immunological memory to SARS-CoV-2, which consists of humoral (neutralizing antibodies) and cellular (T and B cells) memory. The generation of immunological memory to the SARS-CoV-2 virus critically depends on T cell responses. During a viral infection, CD4 helper T cells differentiate largely into either Th1 cells that orchestrate a type I antiviral immune response or follicular helper (Tfh) cells that enhance antibody production. CD8 T cells clonally expand and acquire effector function to directly kill virus-infected cells. Despite the heterogeneity and clonal diversity of virus-specific T cells, the majority of effector T cells die after viral clearance and only a small portion of them develop into memory T cells that provide long-lasting protection for the host. Similarly, B cells develop into memory B cells and long-lived plasma cells that produce neutralizing antibodies. Snapshot observations with multi-parameter flow cytometry-based assays along the course of infection has yielded abundant knowledge of T cell phenotypic and functional diversity. However, approaches as such fail to address the developmental trajectory of virus-specific T cells. This becomes more obvious in human studies given that lineage tracing by genetic alterations or adoptive transfer experiments, done easily in mice, are inherently difficult or impossible in humans. In this proposal, we will first combine newly developed single- cell RNA sequencing (scRNA-seq) and T cell receptor sequencing (TCR-seq) techniques on the same cells and use TCR sequences as natural barcodes to directly “lineage trace” each patient’s SARS-CoV-2-specific CD4 and CD8 T cell effector response and memory formation at the single-cell level throughout the course of natural infection. Furthermore, we will perform gene regulatory network (GRN) analysis to delineate which transcription factors collaboratively regulate virus-specific CD4 and CD8 T cell differentiation trajectories. Next, we will measure T cell clonal diversity and the quality of T cell memory from COVID-19 patients as well as healthy human controls. The latter will be used to gauge the possible presence of pre-existing immunity (PEI) in the form of memory T cells derived from cross-reactivity to common coronaviruses. These measurements will use high- throughput RNA-seq of TCR amplicons and scRNA-seq of memory T cells from recall cultures as inputs for computational TCR motif analysis. Lastly, successful vaccine development relies on an advanced understanding of the types of Tfh cells that are generated during natural infection and how they interact with B cells as well as T regulatory cells for anti-SARS-CoV-2 antibody production in humans. To this end, we propose to monitor circulating Tfh cells, including three major populations (Th1-, Th2- and Th17-like subsets), and T follicular regulatory (Tfr) cells in both SARS-Cov-2-infected and healthy human control subjects. In addition, we will perform T cell-B cell coculture assays to dissect the functional contributions of each subset of Tfh cells and how they interact with Tfr cells in regulating anti-SARS-CoV-2 neutralizing antibody responses.

COVID-19大流行最近出现的意外出现引起了人们对改善的重大兴趣对SARS-COV-2的免疫记忆的理解，该记忆由体液组成（中和抗体）和细胞（T和B细胞）记忆。批判性的SARS-COV-2病毒产生免疫记忆取决于T细胞反应。在病毒感染期间，CD4辅助T细胞主要区分为TH1细胞协调I型抗病毒免疫响应或滤泡辅助器（TFH）细胞，以增强抗体生产。 CD8 T细胞克隆扩展并获取效应子功能，以直接杀死病毒感染的细胞。尽管病毒特异性T细胞的异质性和克隆多样性，大多数效应T细胞在病毒后死亡清除率，只有一小部分发展为记忆T单元，可为您提供长期保护主人。同样，B细胞发展为记忆B细胞和长期寿命的浆细胞，产生中和抗体。沿着多参数流式细胞术测定的快照观察感染已经获得了T细胞表型和功能多样性的丰富知识。但是，AS AS 这种未能解决病毒特异性T细胞的发育轨迹。这在人类中变得更加明显鉴于通过遗传改变或自适应转移实验进行的谱系追踪，在小鼠中很容易进行。在人类中本质上是困难或不可能的。在此提案中，我们将首先结合新开发的单一在同一细胞和使用TCR序列作为天然条形码直接“谱系跟踪”每个患者的SARS-COV-2特异性CD4 在整个自然过程中感染。此外，我们将执行基因调节网络（GRN）分析来描述哪个转录因素协作调节病毒特异性CD4和CD8 T细胞分化轨迹。接下来，我们会的测量T细胞克隆多样性和COVID-19患者的T细胞记忆质量以及健康人类控制。后者将用于衡量形式可能存在预先存在的免疫（PEI）来自交叉反应性到常见冠状病毒的记忆T细胞的。这些测量将使用高从回忆培养物中的TCR扩增子的吞吐量RNA-seq和记忆T细胞的SCRNA-SEQ作为输入计算TCR图案分析。最后，成功的疫苗开发依赖于先进的理解自然感染过程中产生的TFH细胞类型以及它们如何与B细胞相互作用以及人类抗SARS-COV-2抗体产生的T调节细胞。为此，我们建议监视循环TFH细胞，包括三个主要种群（Th1-，Th2-和Th17样子集）和T卵泡 SARS-COV-2感染和健康的人类对照受试者中的调节性（TFR）细胞。此外，我们将执行T细胞-B细胞共培养分析，以剖定每个子集的每个子集的功能贡献以及如何它们在确定抗SARS-COV-2中和抗体反应时与TFR细胞相互作用。