The roles of Lyme spirochete adhesins in hematogenous dissemination

莱姆病螺旋体粘附素在血行传播中的作用

• 批准号: 10570186
• 负责人: George Chaconas
• 金额: $ 71.46万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-10 至 2027-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10570186
• 关键词: Adhesives; Animals; Arthritis; Bacteria; Bacterial Adhesins; Binding; Biochemical; Biology; Blood Vessels; Borrelia afzelii; Borrelia burgdorferi; Borrelia garinii; CCL2 gene; Cell Line; Cells; Chemosensitization; Complex; DBL Oncoprotein; Endothelial Cells; Endothelium; Event; Extracellular Matrix; Gene Family; Generations; Genes; Genetic Transcription; Geography; Health system; Hematogenous; Hour; Infection; Infectious Skin Diseases; Integrin Binding; Interleukin-10; Intravenous; Invaded; Investigation; Joints; Laboratories; Left; Ligands; Lyme Disease; Measures; Mediating; Modeling; Monitor; Mus; Nature; Order Spirochaetales; Organism; OspC protein; Phase; Prevention; Principal Investigator; Process; Protein Array; Research Personnel; Role; Skin; TNF gene; Testing; Time; Tissues; Vector-transmitted infectious disease; Visualization; chronic infection; cytokine; decorin binding protein B; genetic analysis; genome wide screen; in vivo; inhibitor; insight; intravital microscopy; meetings; microbial colonization; migration; mouse model; mutant; neutrophil; novel; pathogenic microbe; tick bite; transmission process; vector tick; virtual

Upon transmission by a vector tick bite, Lyme disease spirochetes, primarily B. burgdorferi (Bb) in the US, establish a local skin infection, then disseminate to multiple tissues. Chronic infection by Bb is often associated with arthritis. Our laboratories have identified and/or characterized many Bb cell- or extracellular matrix (ECM)- binding adhesins using multiple approaches, overcoming the challenges of defining their roles in Bb biology. Our approaches include analyses of biochemical activities and generation of targeted mutants selectively defective for a single adhesive activity and analysis of the mutants in multiple murine infection models. To gain detailed mechanistic insight into interactions that may occur during Bb dissemination in vivo, following intravenous inoculation we used intravital microscopy to characterize vascular attachment and transmigration in skin and joint-proximal tissue. These studies revealed that adhesins BBK32 and VlsE account for virtually all of the transient Bb-endothelium binding occurring minutes after inoculation, termed “Meeting” interactions. A distinct set of adhesins, DbpB/A, OspC, and P66, mediate contacts required for invasion into extravascular joint space after 24 hours (hr), termed “Transmigrating” interactions. Using isogenic strain sets that have acquired or lost specific adhesive activities in multiple short-term and long-term infection models we showed roles for five of the six Meeting or Transmigrating adhesins in short-term tissue localization and/or long-term colonization in other murine infection models. While Transmigrating adhesins do not promote “Meeting” interactions, our discovery of enhanced adhesive capacity of the endothelium as infection progresses has helped clarify why different adhesins have roles at different stages of infection. Within hr, “Endothelial Activation” permits BBK32- and VlsE-independent (“Greeting”) interactions. After ~24 hr “Endothelial Potentiation” occurs, reflected by the ability of the joint vasculature to support Bb transmigration. Although both OspC and P66 function as Transmigrating adhesins, only P66, an integrin-binding adhesin that alters transcription in cultured endothelial cells, is also Potentiating, i.e., required to promote the rapid transmigration of a second Bb strain. Activation is mimicked by exogenous treatment of mice with several cytokines produced by infected mice, but potentiation is seen only with TNF-α, MCP-1 or IL-10. These findings reveal previously unrecognized steps that are critical for Bb spread and provide a means to distinguish roles for each adhesin in distinct infection stages: Meeting, Greeting, Potentiating, Transmigrating and Colonizing. In Aim 1 we will identify known adhesins that facilitate Greeting interactions; to better prioritize our adhesin list, we propose a genome-wide screen that may also identify novel adhesins. In Aim 2 we will clarify the roles of known (and, if applicable, novel) adhesins in endothelial Potentiation and Transmigration. Our use of rigorous genetic analyses in infection models from visualization of key interactions in vivo to quantitative analysis of Bb at different stages of infection will result in detailed understanding of a critical facet of Bb biology: dissemination.

在通过vector tick叮咬传播后，莱姆病螺旋体，美国的主要B. burgdorferi（BB）， 建立局部皮肤感染，然后多次传播。 BB的慢性感染通常是相关的 患有关节炎。我们的实验室已经确定和/或表征了许多BB细胞或细胞外基质（ECM） - 使用多种方法结合粘附，克服了定义其在BB生物学中角色的挑战。 我们的方法包括对生化活动的分析和靶向突变体的产生 多种鼠感染模型中突变体的单个粘合活性和分析的缺陷。获得 详细的机理洞察力洞察体内BB传播期间可能发生的相互作用 静脉接种，我们使用浸润显微镜表征血管附着和传播 在皮肤和关节组织中。这些研究表明，粘附于BBK32和VLSE几乎所有内容 接种后几分钟发生的瞬态BB - 内皮结合，称为“遇见”相互作用。一个 独特的粘附剂，DBPB/A，OSPC和P66，入侵血管外所需的介导接触 24小时后（HR）的关节空间称为“迁移”相互作用。使用具有的等源性应变集 在多个短期和长期感染模型中获得或失去了特定的粘合剂活动 六次会议中五个或在短期组织定位和/或长期内的六次会议中的五个角色 在其他鼠类感染模型中定植。同时移民信封并未促进“会议” 相互作用，随着感染的进展，我们发现内皮增强的粘附能力的发现 帮助阐明了为什么不同的粘合剂在不同感染阶段具有作用。在人力资源内，“内皮 激活”允许BBK32-和vlse独立（“问候”）相互作用。大约24小时“内皮” 增强”发生，反映出关节脉管系统支持BB传播的能力。 OSPC和p66起跨移民胶的功能，仅p66，p66，一种整合素结合粘合剂，可以改变 培养的内皮细胞中的转录也有增强的，即促进快速迁移所需 第二个bb菌株。通过产生的几种细胞因子对小鼠的外源治疗模仿激活 通过感染的小鼠，但仅用TNF-α，MCP-1或IL-10看到潜在。这些发现之前揭示了 对BB传播至关重要的未识别步骤，并提供了区分每种粘附素角色的手段 独特的感染阶段：会议，问候，增强，迁移和殖民。在目标1中，我们将 确定有助于问候互动的已知粘附；为了更好地确定我们的贴素清单，我们提出了一个 全基因组筛选也可能鉴定出新的粘附素。在AIM 2中，我们将阐明已知的作用（以及，如果 适用的，新颖的）粘附在内皮增强和移民中。我们使用严格的遗传 从体内关键相互作用的可视化到BB AT的定量分析的感染模型中的分析 不同的感染阶段将导致对BB生物学关键方面的详细了解：传播。