Impairment of B cell Responses by Pathogenic Chikungunya Viruses

致病性基孔肯雅病毒对 B 细胞反应的损害

• 批准号: 10540705
• 负责人: Michael S Diamond
• 金额: $ 63.94万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-01-25 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10540705
• 关键词: Acute; Alphavirus; Amino Acid Sequence; Amino Acids; Anabolism; Antibodies; Antibody Response; Antigens; Arginine; Attenuated; Avidity; B-Lymphocytes; Biological Assay; Cells; Chikungunya virus; Chronic; Chronic Disease; Cre driver; Culicidae; Data; Defect; Development; Diamond; Dichloromethylene Diphosphonate; Enzymes; Epidemic; Epitopes; Failure; Fever; Flow Cytometry; Genetic; Genetic study; Glycine; Glycoproteins; Glycosaminoglycans; Goals; Human; IFNAR1 gene; Immunity; Impairment; Infection; Inflammation; Inflammation Mediators; Inflammatory; Inflammatory Response; Innate Immune Response; Interferons; Joints; Knowledge; Laboratories; Liposomes; LoxP-flanked allele; Lymphoid Tissue; Macrophage; Maps; Mediating; Modeling; Mouse Strains; Mus; Myeloid Cells; Nitrogen; Oxygen; Pathogenicity; Phagocytes; Polyarthralgias; Positioning Attribute; Process; Production; RNA; Reporter; Role; Signal Transduction; Sinus; Site; Structure of germinal center of lymph node; Swelling; Tenosynovitis; Tissues; Vaccines; Viral; Virus; Virus Diseases; Work; adaptive immune response; chikungunya infection; chronic infection; confocal imaging; diphtheria toxin receptor; draining lymph node; improved; insight; joint stiffness; monocyte; mouse model; neutralizing antibody; novel therapeutic intervention; recruit; response; reverse genetics; single-cell RNA sequencing; therapy development; transmission process; vaccine development

PROJECT SUMMARY B cell responses are essential for clearance of viral infections. The mechanisms by which chronic viruses subvert B cell immunity are poorly understood. Chikungunya virus (CHIKV) is a mosquito-transmitted RNA alphavirus that causes explosive epidemics of debilitating polyarthralgia. Joint swelling, joint stiffness, and tenosynovitis can last for months to years and are associated with persistent CHIKV infection. The mechanistic basis for how CHIKV evades B cell immunity to establish chronic infection is unknown. We found that an attenuated CHIKV strain, but not the parental CHIKV strain (which differs by 5 amino acids), is cleared from joints of WT mice. Persistent infection of the attenuated strain was restored in mice unable to produce virus-specific antibody (Ab). Mapping studies revealed that a single arginine or glycine residue at position 82 in the CHIKV E2 glycoprotein dictates viral clearance or persistence, respectively. In comparison with acutely cleared CHIKV, persistent CHIKV infection was associated with altered B cell responses, a failure to develop germinal centers in the draining lymph node (DLN), poorer quality virus-specific neutralizing Abs, and distinct viral localization and inflammatory responses in lymphoid tissue. Remarkably, depletion of inflammatory myeloid cells improved B cell responses to pathogenic CHIKV. The goal of this highly interactive project between the Morrison and Diamond laboratories is to define mechanistically how pathogenic CHIKV strains evade B cell immunity and the contribution of inflammatory monocytes to this process. In Specific Aim 1, the mechanisms by which a specific CHIKV E2-82 residue dictates inflammatory responses in the DLN will be determined. Viruses differing only at E2-82, and new mice with genetic deficiencies in glycosaminoglycans (GAGs), will be used to determine how E2-GAG interactions dictate CHIKV localization in the DLN. LN macrophage depletion and single-cell RNAseq will define the role of LN myeloid cells in inflammatory and B cell responses to persistent and acutely cleared CHIKV. In Specific Aim 2, how type I IFN signaling modulates B cell responses during acutely cleared and persistent CHIKV infection will be defined. Type I IFN reporter mice, flow cytometry, and IHC will be used to define the cells that produce type I IFN in lymphoid tissue. Ab-mediated blockade of IFNAR1 or distinct type I IFN subtypes (pan-α versus β), and mice with B cell-specific defects in type I IFN signaling will be used to determine the effects of type I IFN on B cell, Th, and Tfh cell responses. In Specific Aim 3, how inflammatory myeloid cells antagonize B cell responses during persistent CHIKV infection will be elucidated. The impact of inflammatory myeloid cells on the avidity, neutralization capacity, and epitope repertoire of the polyclonal anti- CHIKV Ab response will be defined. Using new Nos2F/F or Nox2F/F mice, and a panel of Cre driver strains, we will determine mechanisms by which specific subsets of myeloid cells inhibit optimal B cell responses. This work will elucidate new mechanisms by which viruses subvert B cell immunity and establish persistence. The knowledge gained may aid the development of vaccines or therapies against CHIKV or other chronic viral infections.

项目摘要 B细胞应答对于清除病毒感染是必不可少的。慢性病毒破坏的机制 对B细胞免疫了解甚少。基孔肯雅病毒（CHIKV）是一种蚊子传播的RNA甲病毒 会导致多关节痛的爆发性流行关节肿胀、关节僵硬和腱鞘炎 可持续数月至数年，并与持续性CHIKV感染有关。机械基础如何 CHIKV逃避B细胞免疫以建立慢性感染是未知的。我们发现一个减弱的CHIKV 从WT小鼠的关节中清除CHIKV毒株，而不是亲本CHIKV毒株（其相差5个氨基酸）。 在不能产生病毒特异性抗体（Ab）的小鼠中恢复了减毒株的持续感染。 定位研究显示，CHIKV E2糖蛋白中82位的单个精氨酸或甘氨酸残基 分别决定了病毒的清除率或持久性。与急性清除的CHIKV相比， CHIKV感染与B细胞反应的改变有关，即在细胞中不能发育成生发中心。 引流淋巴结（DLN），质量较差的病毒特异性中和抗体，以及不同的病毒定位， 淋巴组织的炎症反应。值得注意的是，炎性骨髓细胞的去除改善了B细胞 对致病性CHIKV的反应。这个莫里森和戴蒙德之间高度互动的项目的目标是 实验室的目的是从机制上定义致病性CHIKV毒株如何逃避B细胞免疫， 炎症单核细胞对这一过程的贡献。在具体目标1中， 将确定CHIKV E2-82残基决定DLN中的炎症反应。不同的病毒仅在 E2-82和糖胺聚糖（GAG）遗传缺陷的新小鼠，将用于确定 E2-GAG相互作用决定DLN中的CHIKV定位。LN巨噬细胞耗竭和单细胞RNAseq 将确定LN髓样细胞在炎症和B细胞对持续和急性清除的反应中的作用。 CHIKV。在具体目标2中，I型IFN信号传导如何在急性清除和急性排斥期间调节B细胞应答。 将定义持续性CHIKV感染。I型IFN报告小鼠、流式细胞术和IHC将用于 定义淋巴组织中产生I型IFN的细胞。Ab介导的IFNAR 1或不同I型的阻断 IFN亚型（泛-α对β）和I型IFN信号传导中具有B细胞特异性缺陷的小鼠将用于 确定I型IFN对B细胞、Th和Tfh细胞应答的影响。在具体目标3中， 将阐明在持续CHIKV感染期间骨髓细胞拮抗B细胞应答。的影响 炎性骨髓细胞的亲和力，中和能力，和表位库的多克隆抗 将定义CHIKV Ab应答。使用新的Nos 2F/F或Nox 2F/F小鼠和一组Cre驱动株，我们将 确定骨髓细胞的特定亚群抑制最佳B细胞应答的机制。这项工作将 阐明病毒破坏B细胞免疫并建立持久性的新机制。知识 获得的基因可能有助于开发针对CHIKV或其他慢性病毒感染的疫苗或疗法。