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: 10310477
• 负责人: Michael S Diamond
• 金额: $ 64.01万
• 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 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10310477
• 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; Interferons; Joints; Knowledge; Laboratories; Liposomes; LoxP-flanked allele; Lymphoid Tissue; 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; macrophage; monocyte; mouse model; neutralizing antibody; novel therapeutic intervention; recruit; response; reverse genetics; single-cell RNA sequencing; therapy development; 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细胞反应对于清除病毒感染至关重要。慢性病毒颠覆的机制 B细胞的免疫力知之甚少。 Chikungunya病毒（Chikv）是蚊子传播的RNAα病毒 这会导致衰弱的多关节症的爆炸性发作。关节肿胀，关节刚度和脊髓炎炎 可以持续数月到几年，并与持续的Chikv感染有关。如何机械基础 Chikv逃避B细胞免疫以建立慢性感染是未知的。我们发现chikv衰减 菌株，但没有从WT小鼠的关节清除父母CHIKV菌株（与5个氨基酸不同）。 在无法产生病毒特异性抗体（AB）的小鼠中恢复了衰减菌株的持续感染。 映射研究表明，Chikv E2糖蛋白的位置82处的单个精氨酸或甘氨酸保留率 分别决定病毒清除或持久性。与急性清除的chikv相比，持久 CHIKV感染与B细胞反应改变有关，这是未能在 排水淋巴结（DLN），质量较差的病毒特异性中和ABS以及独特的病毒定位和 淋巴组织中的炎症反应。值得注意的是，炎症性髓样细胞的耗竭改善了B细胞 对致病性CHIKV的反应。莫里森和钻石之间这个高度互动项目的目标 实验室要机械地定义致病性CHIKV菌株如何逃避B细胞免疫和 炎症单核细胞对此过程的贡献。在特定目标1中，特定的机制 CHIKV E2-82居住地决定了DLN中的炎症反应。病毒仅在 E2-82和糖胺聚糖（GAG）中具有遗传缺陷的新小鼠将用于确定如何 E2-GAG相互作用决定了DLN中的CHIKV定位。 LN巨噬细胞部署和单细胞RNASEQ 将定义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。在特定的目标2中，I型IFN信号传导如何调节急性清除期间的B细胞响应，并且 将定义持续的CHIKV感染。 I型IFN记者小鼠，流式细胞术和IHC将用于 定义在淋巴组织中产生I型IFN的细胞。 AB介导的IFNAR1或不同类型I的封锁 IFN亚型（PAN-α与β），并且I型IFN信号中具有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细胞反应的影响。在特定的目标3中，如何炎症 将阐明持续性CHIKV感染期间B细胞反应的髓样细胞将被阐明。的影响 多克隆抗肿瘤的炎症性髓样细胞在亲发，中和能力和表位库中 CHIKV AB响应将定义。使用新的NOS2F/F或NOX2F/F小鼠，以及一组CRE驾驶员应变，我们将 确定髓样细胞的特定亚群抑制最佳B细胞反应的机制。这项工作将 阐明病毒颠覆B细胞免疫并建立持久性的新机制。知识 获得的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。