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Host Ca2+, actin, and ATP production in rickettsia-endothelial cell dysfunction

立克次体内皮细胞功能障碍中宿主 Ca2、肌动蛋白和 ATP 的产生

基本信息

批准号：
10509838
负责人：
JOHN STEPHEN Dumler
金额：
$ 22.78万
依托单位：
HENRY M. JACKSON FDN FOR THE ADV MIL/MED
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-05 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10509838
关键词：
1-Phosphatidylinositol 3-Kinase ATP phosphohydrolase Actin-Binding Protein Actins Actomyosin Adenine Nucleotide Translocase Affect Bacteria Blood Vessels Brain Brazil Buffers Ca(2+)-Transporting ATPase Calcium Calcium Channel Calcium Channel Blockers Calmodulin Case Fatality Rates Cation Pumps Cell Death Cell Line Cell physiology Cells Cellular Structures Cessation of life Chelating Agents Colombia Communicable Diseases Consequentialism Consumption Cytoskeleton Cytosol Data Disease Endoplasmic Reticulum Endothelial Cells Endothelium Energy-Generating Resources Event Fatal Outcome Functional disorder Genetic Transcription Genome Germ Cells Goals Growth Human Hypotension Impairment In Vitro Infection Intervention Investigation Link Lung Lyme Disease Measures Membrane Metabolic Mexico Microscopy Molecular Monitor Multiple Organ Failure Myosin Light Chain Kinase Nutrient Nutritional Organ Organism PMCA1 protein Parasites Pathogenicity Pathologic Permeability Pharmacology Phospholipase Phosphorylation Phosphotransferases Preventive therapy Process Production Property Proteins Pump Regulation Research Resolution Resources Rickettsia Rickettsia Infections Rickettsia parkeri Rickettsia rickettsii Rocky Mountain Spotted Fever Role Sepsis Shock Signal Pathway Signaling Molecule Small Interfering RNA System Tick-Borne Infections Time Transcriptional Regulation Vascular Permeabilities Virulence Work adverse outcome antagonist base blood perfusion cadherin 5 calcium-dependent protein kinase cell motility chelation design electric impedance fitness hypoperfusion improved inhibitor ischemic injury microbial monolayer parasitism pathogen prevent receptor recruit release of sequestered calcium ion into cytoplasm response spotted fever temporal measurement voltage

项目摘要

Project Summary/Abstract Spotted fever group rickettsioses (SFGR) are in aggregate the second most common tick-borne infections in the U.S. and account for considerable severe disease and death, with case fatality rates over 10% in many regions of the world. Rocky Mountain spotted fever is the prototypical disease in this group for which the major pathophysiologic adverse consequence is increased vascular permeability, leading to hypotension, hypoperfusion, and ischemic injury to the lungs, brain and other organs. SFG rickettsiae infect endothelial cells and parasitize host ATP for growth and spread. Our preliminary studies showed that regulation of calcium flux in infected endothelial cell barriers using calcium chelators or specific calcium channel blockers abrogates vascular permeability. We propose that spotted fever rickettsiae utilize a nutritional virulence strategy to subvert host calcium handling – through an acquired “channelopathy” - that drives increases in access to cellular metabolic substrates, but that in turn damages key functions of endothelial cells. Our hypothesis is that increasing competition for cellular ATP by rickettsiae impairs ATP-dependent Ca2+ pumps, which in turn leads to increased intracellular calcium concentrations, activation of Ca2+ channels, Ca2+-dependent kinases, signaling pathways and transcriptional regulation that increase cellular content of key nutrients for rickettsial growth. The consequences of improved rickettsial fitness ultimately leads to reorganization of cellular cytoskeleton, disassembly of inter-endothelial junctions, and inevitably, increased vascular permeability. The proposal will measure intracellular calcium and ATP concentrations in the presence or absence of calcium channel-blocking agent, while monitoring for endothelial barrier dysfunction in both early and late stages of Rickettsia parkeri infection. Evidence of a role for calcium flux in rickettsial vascular permeability includes localized calcium “puffs” with transient focal cytosolic ATP depletion and local cytoskeletal restructuring as early post-invasion events. This is predicted to be followed by global increases in intracellular calcium, ATP depletion, cytoskeleton restructuring, and inter-endothelial junction disassembly with massive rickettsial proliferation, sensitive to buffering cytosolic calcium concentrations through chelation or blocking or silencing of key calcium channels. Rickettsial load will also be determined to understand the effect of these manipulations on microbial fitness. This work will identify key mechanisms that permit changes in vascular permeability with spotted fever rickettsia infection, perhaps driven by rickettsial effector proteins, and potential targets for host-based pharmacologic intervention to prevent severe and fatal outcomes of SFGR and potentially other infectious diseases.

项目摘要/摘要斑点热群立克次体病(SFGR)总体上是美国第二种最常见的壁虱传播感染，造成相当严重的疾病和死亡，在世界许多地区病死率超过10%。洛基山斑热是本组的典型疾病，其主要病理生理不良后果血管通透性增加，导致低血压、低灌流，以及对肺、脑和其他组织的缺血性损伤器官。SFG立克次体感染内皮细胞，寄生宿主三磷酸腺苷以促进生长和传播。我们的初步研究研究表明，使用钙离子螯合剂或特定的钙离子来调节感染的内皮细胞中的钙流通道阻滞剂可消除血管通透性。我们建议斑点热立克次体利用一种营养毒力颠覆宿主钙处理的策略--通过获得性“通道病”--推动细胞通透性的增加代谢底物，但这反过来又损害内皮细胞的关键功能。我们的假设是，不断增加的立克次体对细胞三磷酸腺苷的竞争损害了三磷酸腺苷依赖的钙泵，这反过来又导致增加细胞内钙离子浓度，激活钙离子通道，钙依赖的激酶，信号通路和转录调控，增加立克次体生长所需的关键营养物质的细胞含量。改进的后果立克次体的健康最终导致细胞细胞骨架的重组，内皮细胞间连接的分解，以及不可避免地，血管通透性增加。该提案将测量细胞内钙和ATP浓度。存在或不存在钙通道阻滞剂，同时监测两者早期内皮屏障功能障碍晚期帕氏立克次体感染。钙离子通量在立克次体血管通透性中作用的证据包括局部钙“喷发”，伴有一过性局灶性胞浆ATP耗竭和局部细胞骨架重构入侵后的事件。据预测，紧随其后的是全球细胞内钙的增加，ATP消耗，细胞骨架重构和内皮间连接解离伴随立克次体大量增殖，对通过螯合或阻断或沉默关键钙通道来缓冲细胞内钙浓度。立克次体负荷也将被确定，以了解这些操作对微生物适应性的影响。这项工作将确定斑点热立克次体感染时允许血管通透性改变的关键机制，可能是由立克次体效应蛋白和以宿主为基础的药物干预预防严重和致命的潜在靶点 SFGR和潜在的其他传染病的后果。