Mechanisms of Cellular and Humoral Immune Evasion and Subsequent Polyomavirus Encephalitis

细胞和体液免疫逃避及随后的多瘤病毒脑炎的机制

• 批准号: 9760815
• 负责人: Heather Marie Ren
• 金额: $ 2.54万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY 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: 2019-09-01 至 2020-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9760815
• 关键词: AIDS/HIV problem; Acquired Immunodeficiency Syndrome; Adaptive Immune System; Allografting; Amino Acids; Animal Model; Antibodies; Antibody Affinity; Antibody Repertoire; Antibody Response; Antibody Specificity; Antiviral Agents; Autoimmune Diseases; Autoimmune Process; B-Lymphocytes; Biological Assay; Blood; Brain; Brain Diseases; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; CD8B1 gene; Capsid Proteins; Cells; Central Nervous System Diseases; Central Nervous System Infections; Chronic; Complication; Data; Defect; Development; Disease; Encephalitis; Enzyme-Linked Immunosorbent Assay; Epitopes; Genomic DNA; Goals; HIV; Haplotypes; Hematologic Neoplasms; Homeostasis; Host Defense; Humoral Immunities; Immune; Immune Evasion; Immune response; Immunocompromised Host; Immunoglobulin G; Immunohistochemistry; Immunologic Monitoring; Immunologics; Immunosuppression; Impairment; Incidence; Individual; Infection; Inflammatory; Integrin alpha4; Interleukin-10; Interleukins; JC Virus; Label; Laboratories; Lesion; Life; Link; Maintenance; Mediating; Memory; Messenger RNA; Modeling; Monoclonal Antibodies; Multiple Sclerosis; Mus; Mutant Strains Mice; Mutation; Neuraxis; Neuroglia; Pathogenesis; Patients; Plasma Cells; Polyomavirus; Polyomavirus Infections; Prevention; Preventive therapy; Preventive treatment; Production; Progressive Multifocal Leukoencephalopathy; Proteins; Reaction; Receptor Cell; Receptor Signaling; Residencies; Role; Serum; Signal Transduction; Source; Specificity; Structure of germinal center of lymph node; Systemic Lupus Erythematosus; T cell response; T-Lymphocyte; Testing; Time; Tissues; Transplantation; Tropism; Tumor-infiltrating immune cells; Variant; Viral; Virus; Virus Diseases; base; central nervous system demyelinating disorder; chemotherapy; cytokine; immunomodulatory therapies; immunosuppressed; improved; interleukin-21; interleukin-21 receptor; leukemia; mouse model; mouse polyomavirus; multiple sclerosis patient; mutant; natalizumab; neurotropic; neurotropic virus; neutralizing antibody; prevent; receptor; residence; response; transcriptome; transcriptome sequencing

ABSTRACT Progressive Multifocal Leukoencephalopathy (PML), a life-threatening demyelinating brain disease caused by JC polyomavirus (JCV), is a significant complication for patients with HIV/AIDS, hematologic malignancies, rheumatologic diseases, and those receiving long-term immunomodulating therapies. The immunomodulating therapy mostly closely associated with PML is natalizumab, an α4 integrin antibody (Ab) commonly used to treat multiple sclerosis (MS). Lack of a tractable animal model for PML is a widely recognized hurdle to defining the mechanisms of immunological control of JCV CNS infection. Using mouse polyomavirus (MuPyV), our laboratory developed a robust model of polyomavirus-associated demyelinating leukoencephalitis, with viral infection and T cell infiltration. This proposal seeks to use the MuPyV-CNS infection model to mechanistically understand the cellular and humoral immune mechanisms in place to control JCV CNS infection and how immunosuppression predisposes patients to JCV CNS infection. Increasing evidence supports the importance of CD4+ T cells in controlling JCV and thereby preventing PML. Recent studies in our laboratory show that CD4+ T cells are essential for differentiation and maintenance of MuPyV-specific tissue-resident memory CD8+ T cells (TRMs) in the brain. My preliminary data strongly supports the likelihood that IL-21 is a major component of the help provided by CD4+ T cells. I have further found that the serum from IL-21 receptor (IL21R)-deficient mice has a reduced ability to neutralize MuPyV carrying an amino acid mutation in its VP1 capsid protein that corresponds to a frequent VP1 mutation in PML-JCV. In Specific Aim 1, I hypothesize that IL-21 is required for brain-infiltrating CD8+ T cells to differentiate into TRMs. To test this hypothesis, I will define a time course for IL-21 production in the MuPyV-infected brain, its cellular sources, and when its receptor is expressed by MuPyV-specific CD8+ T cells. Using donor MuPyV-specific CD8+ T cells lacking IL21R, I will determine if signaling through the IL21R is intrinsically required by CD8+ T cells for their differentiation into TRMs. I will also apply RNA-seq to determine if the transcriptomes of IL21R-/- CD8+ T cells in the brain are similar to CD8+ T cells in CD4+ T cell-deficient mice. In Specific Aim 2, I hypothesize that IL21R signaling diversifies the VP1 Ab repertoire in a B cell-intrinsic manner to facilitate recognition of a MuPyV carrying a PML-synonymous VP1- mutation. To test this hypothesis, I will use MuPyV Ab neutralization assays, ELISAs, and germinal center immunohistochemistry to define the time course for developing a hole in the VP1 Ab specificity repertoire under T-independent (TCRα-/-) and IL-21-deficient (IL21R-/-) conditions. I also will determine plasma cell specificity against the PML-synonymous VP1 MuPyV mutant using fluorescently labeled MuPyVs in these aforementioned conditions. Finally, I will distinguish whether the mechanistic basis for the diminished VP1 neutralizing Ab repertoire is due to an intrinsic defect in IL21R signaling in B and/or T cells.

摘要 进行性多灶性白质脑病(PML)，一种威胁生命的脱髓鞘脑病，由 JC多瘤病毒(JCV)是HIV/AIDS，血液系统恶性肿瘤， 风湿病，以及接受长期免疫调节治疗的人。免疫调节剂 与早幼粒白血病关系最密切的治疗方法是那他珠单抗，一种α4整合素抗体(Ab)，通常用于 治疗多发性硬化(MS)。缺乏可驯化的PML动物模型是公认的定义PML的障碍 JCV中枢神经系统感染的免疫调控机制使用小鼠多瘤病毒(MuPyV)，我们的 实验室开发了一种强大的多瘤病毒相关性脱髓鞘白质脑炎模型， 感染和T细胞浸润。这项建议寻求使用MuPyV-CNS感染模型来机械性地 了解控制JCV CNS感染的细胞和体液免疫机制以及如何控制 免疫抑制使患者易患JCV CNS感染。越来越多的证据支持这一重要性 CD4+T细胞在控制JCV从而预防PML方面的作用。我们实验室最近的研究表明 CD4+T细胞对MuPyV特异性组织驻留记忆CD8+的分化和维持至关重要 大脑中的T细胞(TRMS)。我的初步数据有力地支持了IL-21是主要成分的可能性 由CD4+T细胞提供的帮助。我还进一步发现，来自白介素21受体(IL21R)的血清缺乏 小鼠中和MuPyV的能力降低，其VP1衣壳蛋白携带氨基酸突变 与PML-JCV中频繁的VP1突变相对应。在具体目标1中，我假设IL-21是必需的 使脑内浸润性CD8+T细胞分化为TRMS。为了检验这一假设，我将定义一个时间进程 IL-21在感染MuPyV的脑中的产生，其细胞来源，以及其受体通过 MuPyV特异性CD8+T细胞。使用缺乏IL21R的供体MuPyV特异性CD8+T细胞，我将确定 CD8+T细胞通过IL21R信号转导是其向TRMS分化所必需的。我也会 应用RNA-seq技术确定脑内IL21R-/-CD8+T细胞的转录水平是否与CD8+T细胞相似 CD4+T细胞缺陷小鼠的细胞。在特定的目标2中，我假设IL21R信号使VP1抗体多样化 以B细胞固有的方式，以促进识别携带PML同义VP1的MuPyV- 突变。为了验证这一假设，我将使用MuPyV抗体中和试验、ELISA和生发中心 免疫组织化学方法确定VP1Ab特异性谱系出现空洞的时间进程 在T非依赖性(TCRα-/-)和IL-21缺乏(IL21R-/-)条件下。我还将确定浆细胞 在这些细胞中使用荧光标记的MuPyV对PML同义VP1 MuPyV突变体的特异性 上述条件。最后，我将区分VP1减小的机制基础 中和抗体谱系是由于B和/或T细胞中IL21R信号的固有缺陷。