Virulent Rickettsia species utilize the CD300f phosphatidylserine-binding receptor on macrophages for host colonization and pathogenesis

强毒立克次体利用巨噬细胞上的 CD300f 磷脂酰丝氨酸结合受体进行宿主定植和发病机制

• 批准号: 10526450
• 负责人: Mohammed Sayeedur Rahman
• 金额: $ 23.18万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10526450
• 关键词: ANXA5 gene; ARHGEF5 gene; Address; Affinity; Animals; Anti-Inflammatory Agents; Antibodies; Antigens; Apoptotic; Arthropod Vectors; Arthropods; Bacteria; Binding; Biological Assay; Bone Marrow; Boutonneuse Fever; Cell Surface Receptors; Cell surface; Cells; Central America; Data; Dendritic Cells; Dermis; Development; Dichloromethylene Diphosphonate; Disease Outbreaks; Distant; Eating; Endemic Flea-Borne Typhus; Endothelial Cells; Etiology; Europe; Excision; Extracellular Domain; Glycerophospholipids; Goals; Grant; Homeostasis; Host Defense; Human; Immunoglobulin Variable Region; Immunologic Surveillance; Impairment; Individual; Infection; Invaded; Knowledge; Lethal Dose 50; Ligands; Link; Lipopolysaccharides; Liposomes; Mediating; Membrane; Membrane Proteins; Middle East; Morbidity - disease rate; Mus; Parasites; Pathogenesis; Pathogenicity; Phagocytes; Phagocytosis; Phosphatidylserines; Process; Recombinants; Reporting; Research; Research Project Grants; Rickettsia; Rickettsia Infections; Rickettsia conorii; Rickettsia rickettsii; Rickettsia typhi; Rocky Mountain Spotted Fever; Role; Signal Transduction; South America; Sum; Surface; System; Testing; Typhus; Vaccines; Virulence; Virulent; Virus; Work; arthropod-borne; fighting; host colonization; in vivo; macrophage; member; mortality; mouse model; novel; pathogen; pathogenic bacteria; phosphatidylserine receptor; prevent; programs; receptor; receptor binding; spotted fever; therapeutically effective

PROJECT SUMMARY Rickettsia species are arthropod-borne obligate intracellular bacteria with both symbiotic and pathogenic lifecycles. The global impact of rickettsial infections is highlighted by the resurgence of human infections with R. rickettsii (etiologic agent of Rocky Mountain Spotted Fever) in Central and South America, reappearance of R. conorii (etiologic agent of Boutonneuse Fever) in Europe, Middle East, and Africa9 and the recent outbreaks of R. rickettsii and R. typhi (etiologic agent of murine typhus) in the USA. Unfortunately, our inadequate understanding of Rickettsia-host interaction, in particular at the level on rickettsial engulfment via receptor-ligand- mediated phagocytosis on professional host defense cells, like macrophages (MΦ), has significantly impaired the development of effective therapeutics against these pathogens. To address these knowledge gaps, we propose to: i) characterize the specific rickettsial outer membrane glycerophospholipid ligand(s) involved in the phagocytic process of SFG and TG rickettsiae by MΦ, ii) decipher the role of efferocytic receptor, CD300f, in facilitating host colonization of virulent Rickettsia species. Our preliminary work identified phosphatidylserine (PS), a well-characterized “eat-me” signal involved in the phagocytosis of apoptotic cells (aka efferocytosis), as a putative ligand on outer membrane of rickettsiae. We further validated PS as crucial ligand for engulfment of R. typhi and R. rickettsii [Shelia Smith], two virulent bacteria representing the TG and SFG, respectively, by using recombinant Annexin V, a molecule known to block PS-mediated efferocytosis, in bone marrow-derived macrophages (BMMΦ). As PS-dependent phagocytosis requires PS-receptor recognition, we focused on CD300f, a type I transmembrane cell surface receptors with a high binding affinity to PS. Using WT and CD300f- /- BMMΦ, we showed that engulfment of R. typhi and R. rickettsii was dependent on CD300f expression, which was further confirmed by αCD300f antibody-mediated neutralization assays on WT BMMΦ. We tested the role of CD300f in vivo, by establishing mouse models of rickettsiosis with ~LD50 for both R. typhi and R. rickettsii species in C57BL/6J WT mice and showed that, unlike WT mice, CD300f-/- animals were protected against Rickettsia-induced lethality. Furthermore, in vivo depletion of MΦ, via clodronate-liposomes, suggest that CD300f-expression on MΦ contributed to the protection against Rickettsia-induced lethality. Hence, we will test the hypothesis, that both SFG and TGs Rickettsia exploit efferocytic signaling to invade MΦ, using the following Aims: to identify rickettsial outer membrane glycerophospholipid [phosphatidylserine (PS) or other] ligand(s) involved in the engulfment and colonization of pathogenic Rickettsia by MΦ (Aim 1) and define the contributing role of glycerophospholipid-binding receptor, CD300f, in promoting engulfment and colonization of the host by pathogenic Rickettsia in MΦ (Aim 2). Under this “Exploratory/Developmental Research Grant Program” we will study the mechanisms by which rickettsiae from both SFG and TG exploit the normally efferocytic CD300f-ligand systems on MΦ to successfully colonize the host. In sum, the purpose of this grant is to unveil mechanisms of rickettsial invasion with the goal to identify more proficient targets for anti-virulence therapy.

项目摘要 立克种类是节肢动物传播的义务，具有共生和致病性的细胞内细菌 生命周期。人类感染的全球感染对全球感染的影响。 Rickettsii（落基山斑点发烧的病因学代理）在中美洲和南美，重新出现。 欧洲，中东和非洲的Conorii（Boutonneuse Fever的病因学推动者）以及最近的爆发 R. Rickettsii和R. Typhi（鼠Typhus的病因学剂）在美国。不幸的是，我们的不足 理解立克求主-host相互作用，尤其是通过接收器配体的立克吞噬的水平 巨噬细胞（Mφ）等专业宿主防御细胞上的介导的吞噬作用显着受损 开发针对这些病原体的有效疗法。为了解决这些知识差距，我们 提示：i）表征与参与该的特定风险的外膜外膜配体 SFG和TG Rickettsiae的吞噬过程，由Mφ，II）解密efferocytic受体CD300F的作用 促进有毒人力体物种的宿主定植。我们的初步工作确定了磷脂酰丝氨酸 （PS），一种涉及凋亡细胞吞噬作用（又称输液性细胞增多症）的特征良好的“饮食”信号，AS Rickettsiae外膜上的假定配体。我们进一步验证PS是吞噬的关键配体 R. Typhi和R. Rickettsii [Shelia Smith]，分别通过使用TG和SFG的两种有毒细菌 重组膜联蛋白V（一种已知可以阻断PS介导的胞吞作用的分子）在骨髓来源 巨噬细胞（BMMφ）。由于依赖PS的吞噬作用需要PS受体识别，我们专注于 CD300F，一种I型跨膜细胞表面受体，与PS具有高结合亲和力。使用WT和CD300F- /-bmmφ，我们表明鼠伤寒河和R. rickettsii的吞噬依赖于CD300F表达， 通过αCD300F抗体介导的神经化测定在WTBMMφ上进一步证实。我们测试了角色 CD300F体内的of Vivo，通过建立R. Typhi和R. Rickettsii的Rickettsios的小鼠模型 C57BL/6J WT小鼠中的物种，表明与WT小鼠不同，CD300F - / - 动物受到保护 人力车引起的致死性。此外，通过氯膦酸酯脂质体在体内耗竭，表明 Mφ上的CD300F表达有助于防止立克诱导的致死性。因此，我们将测试 假设SFG和TGS Rickettsia使用以下 目的：鉴定立克外膜外膜甘油磷脂[磷脂酰丝氨酸（PS）或其他]配体（S） 涉及Mφ致病性人力车的吞噬和定殖（AIM 1）并定义了贡献 甘油磷脂结合受体CD300F的作用在促进宿主吞噬和定植中的作用 Mφ中的致病性人力赛（AIM 2）。在这个“探索/发展研究赠款计划”下，我们将 研究来自SFG和TG的Rickettsiae的机理 Mφ上的系统成功定居宿主。总而言之，这笔赠款的目的是揭露 立克入侵的目标是确定更熟练的抗病毒治疗靶标。