EPCR, TAFI as Regulators of PMN/Endothelial Interaction

EPCR、TAFI 作为 PMN/内皮相互作用的调节剂

基本信息

批准号：
7939125
负责人：
FLOREA LUPU
金额：
$ 9.25万
依托单位：
OKLAHOMA MEDICAL RESEARCH FOUNDATION
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-09-30 至 2010-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7939125
关键词：
Active Sites Adherence Adopted Affect Alteplase Animals Antigens Appearance Attention Attenuated Binding Biological Assay Biological Markers Blood Coagulation Disorders Blood Pressure Complement 5a Complex Dose Elastases Electron Microscopy Endothelium Escherichia coli Event Exhibits Fibrin Fibrin split products Generations Half-Life Hemostatic Agents Hemostatic function Histopathology Imaging Techniques Immunologics Inflammatory Inflammatory Response Infusion procedures Intervention Ischemia Knock-out Learning Link Measurement Mediator of activation protein Messenger RNA Metalloproteases Methods Modeling Monitor Mus Neutrophil Activation Neutrophil Infiltration Papio Permeability Phase Physiological Plasma Production Protein C Protein C Inhibitor Regulation Reperfusion Therapy Role Sepsis Staging Stretching Temperature Thrombin Thrombin Receptor Thrombomodulin Thromboplastin Time Tissues Work Xai factor activated Protein C design improved inhibitor/antagonist neutrophil receptor response

项目摘要

DESCRIPTION (provided by applicant): These studies focus on the role of endothelial protein C receptor (EPCR) and thrombin activatable fibrinolytic inhibitor (TAFI) as regulators of the neutrophil/endothelial interaction induced by E. coli. All the inflammatory and hemostatic events studied in the baboon model of E. coil sepsis culminate in an aberrant neutrophil/endothelial interaction leading to increased permeability and coagulation disorders. EPCR and thrombomodulin (TM) are at the point of attack and therefore are both targets and regulators of this interaction through activation of protein C and TAFI by the TM/thrombin complex and through release of soluble EPCR by endothelial-derived metalloproteases. We postulate that TAFla (procarboxypeptidase beta) attenuates neutrophil activation by inactivating C5a, and that soluble EPCR attenuates subsequent tight binding of neutrophils to endothelium. The close association of EPCR and TAFla with the endothelium and neutrophils favor these actions. Two general questions are: What is the response and distribution of EPCR t and TAFI between endothelium and neutrophils? Can the information be used to design and time intervention using soluble EPCR and TAFla that would improve the efficacy of activated protein C? To study such questions we have adopted the sublethal model of E. coil sepsis, because the otherwise lethal events are stretched out over time into an initial host (stage 1) and ischemia reperfusion (stage 2) responses. This model allows one to track and intervene with the responses of these regulatory components at critical points of the response to E. coil. Specific questions include what are the timing and distribution of EPCR and TAFI between endothelium and neutrophils with respect to mediators (e.g., C5a) and adherence of neutrophils to the microvascular endothelium? Are there critical events involving these regulators in stage 1 that determine subsequent stage 2 events and whether the response becomes lethal? Can APC and either sEPCR or TAFI be used together to better regulate the neutrophil/endothelial response to E. coil? How critical is timing of intervention in determining whether it is beneficial or harmful? Changes in the expression and distribution of EPCR, TAFI, protein C, thrombomodulin, tissue factor (etc.) on the endothelium, perivascular tissues and neutrophils will be assessed using immunohistochemical and confocal imaging techniques. This includes FACS analysis, and determination of neutrophil half-life. The responses of plasma factors will be followed with ELISAs. Standard physiological parameters will be followed (e.g., temperature, CBC, blood pressure and global assays of hemostatic function)

描述（由申请人提供）：这些研究重点关注内皮蛋白 C 受体 (EPCR) 和凝血酶可激活纤溶抑制剂 (TAFI) 作为大肠杆菌诱导的中性粒细胞/内皮相互作用调节剂的作用。在狒狒败血症模型中研究的所有炎症和止血事件最终导致异常的中性粒细胞/内皮相互作用，导致通透性增加和凝血障碍。 EPCR 和血栓调节蛋白 (TM) 处于攻击点，因此通过 TM/凝血酶复合物激活蛋白 C 和 TAFI，以及通过内皮源性金属蛋白酶释放可溶性 EPCR，它们都是这种相互作用的目标和调节器。我们假设TAFla（羧肽酶β原）通过灭活C5a来减弱中性粒细胞的活化，并且可溶性EPCR减弱随后的中性粒细胞与内皮的紧密结合。 EPCR 和 TAFla 与内皮细胞和中性粒细胞的密切联系有利于这些作用。两个常见问题是：EPCR t 和 TAFI 在内皮细胞和中性粒细胞之间的反应和分布是什么？这些信息是否可以用于使用可溶性 EPCR 和 TAFla 来设计和计时干预，以提高活化蛋白 C 的功效？为了研究这些问题，我们采用了大肠杆菌脓毒症的亚致死模型，因为否则致命事件会随着时间的推移延伸到初始宿主（第 1 阶段）和缺血再灌注（第 2 阶段）反应。该模型允许人们在对大肠杆菌反应的关键点上跟踪和干预这些调节成分的反应。具体问题包括内皮细胞和中性粒细胞之间 EPCR 和 TAFI 相对于介质（例如 C5a）的时间和分布以及中性粒细胞对微血管内皮的粘附是什么？第一阶段中是否存在涉及这些监管机构的关键事件，这些事件决定了随后的第二阶段事件以及响应是否会致命？ APC 和 sEPCR 或 TAFI 能否一起使用以更好地调节中性粒细胞/内皮细胞对大肠杆菌的反应？干预的时机对于确定它是有益还是有害有多重要？使用免疫组织化学和共聚焦成像技术评估内皮、血管周围组织和中性粒细胞上EPCR、TAFI、蛋白C、血栓调节蛋白、组织因子等的表达和分布的变化。这包括 FACS 分析和中性粒细胞半衰期的测定。血浆因子的反应将通过 ELISA 进行跟踪。将遵循标准生理参数（例如，温度、CBC、血压和止血功能的整体测定）