Regulatory Mechanisms for Resolution of Inflammatory Microvascular Leakage

解决炎症性微血管渗漏的调节机制

• 批准号: 8574448
• 负责人: JEROME W BRESLIN
• 金额: $ 30.38万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8574448
• 关键词: Abbreviations; Actins; Adhesions; Admission activity; Adult Respiratory Distress Syndrome; Binding; Biochemical; Caring; Cells; Clinical; Clinical Treatment; Coupled; Data; Development; Disease; Edema; Endothelial Cells; Endothelium; Extravasation; Family; Family member; Feedback; Foundations; Functional disorder; Glycocalyx; Health Care Costs; Hemorrhagic Shock; Hospitals; Image; Incidence; Infection; Inflammation; Inflammation Mediators; Inflammatory; Injury; Intensive Care; Intercellular Junctions; Lead; Life; Maintenance; Measurement; Mediating; Membrane; Mesentery; Methods; Microcirculation; Microvascular Permeability; Modeling; Molecular; Molecular Biology; Molecular Target; Monomeric GTP-Binding Proteins; Multiple Organ Failure; N-terminal; Outcome; Pathway interactions; Patients; Permeability; Phosphorylation; Physiological; Plasma Proteins; Polyamines; Rattus; Recovery; Resolution; Resuscitation; Rho-associated kinase; Rnd3 protein; Role; Sepsis Syndrome; Serine; Serine Phosphorylation Site; Severities; Signal Transduction; Site; Stimulus; Testing; Text; Therapeutic Intervention; Tissues; Trauma; base; cadherin 5; cell behavior; cellular imaging; cost; farnesylation; in vivo; innovation; intravital microscopy; knowledge of results; meetings; monolayer; mortality; mutant; neutrophil; novel; restoration; rho; theories; therapy development; venule

DESCRIPTION (provided by applicant): Microvascular hyperpermeability is a hallmark of the systemic inflammatory response syndrome (SIRS) present in 93% of intensive care patients. SIRS severely complicates resuscitation of trauma victims and worsens clinical outcomes, frequently progressing to multiple organ failure. The estimated annual US health care cost burden associated with SIRS is $16.7 billion. Despite advances in understanding of inflammation- induced microvascular hyperpermeability, the mechanisms that restore microvascular permeability to normal following an inflammatory challenge are unknown. Moreover, existing clinical treatments are not effective for restoration of microvascular barrier integrity once the inflammatory cascade has been initiated. We propose a novel paradigm of signaling mechanisms responsible for restoration of microvascular integrity following inflammation caused by traumatic injury. Our preliminary data show that administration of a polyamine-coated, cell permeable form of Rnd3, a Rho family small GTPase, reduces microvascular hyperpermeability in a rat model of hemorrhagic shock. We also have observed with live cell imaging of endothelial cells that lamellipodia formation and turnover represent a previously uncharacterized cell behavior important for normal endothelial barrier integrity. We hypothesize that Rnd3 promotes endothelial barrier restoration after inflammatory challenges by inhibiting RhoA/ROCK-mediated cell contraction and activating Rac1-mediated enhancement of intercellular junction integrity. Our specific aims are to: 1) Test the prediction that Ser phosphorylation and membrane targeting of Rnd3 are required for Rnd3 to reduce microvascular hyperpermeability; 2) To test the prediction that Rnd3 promotes negative feedback inhibition of ROCK-mediated cell contraction, which enhances endothelial barrier integrity; 3) To test the prediction that Rnd3 enhances Rac1-mediated lamellipodia formation and stabilization of VE-cadherin at junctions, promoting endothelial barrier integrity. The proposed novel, integrated approach capitalizes on a refined and relevant rat model of hemorrhagic shock combined with intravital microscopy of the in vivo mesenteric microcirculation and isolated venule methods to assess microvascular permeability. We will explore the central role of Rnd3 by employing an innovative method to deliver cell permeable Rnd3 protein to the mesenteric microcirculation. Cultured endothelial cell monolayer permeability models, imaging of GFP-actin and GFP-VE-cadherin dynamics in live endothelial cells, and biochemical studies will support the in vivo and isolated venule studies. The results of this study will enable us to develop a new theory of how endothelial barrier function can be restored during inflammation, which will serve as the foundation for novel therapies. Discovery of targets that can be used to resolve microvascular hyperpermeability will revolutionize the treatment of trauma patients, and will also create new opportunities to treat edema associated with a wide range of diseases.

描述（由申请人提供）：微血管高温性是93％的重症监护患者中存在的全身性炎症反应综合征（SIRS）的标志。 Sirs严重使创伤受害者复苏并使临床结果恶化，经常发展为多器官衰竭。估计与SIRS相关的美国医疗保健成本负担为167亿美元。尽管了解炎症引起的微血管高温性方面的进步，但在炎症挑战中恢复微血管渗透率正常的机制尚不清楚。此外，一旦启动了炎症性级联反应，现有的临床治疗对于恢复微血管屏障完整性无效。我们提出了一种新型的信号传导机制范式，负责恢复由创伤性损伤引起的炎症后微血管完整性的恢复。我们的初步数据表明，在大鼠出血性休克大鼠模型中，RHO家族小型GTPase的多胺涂层，可渗透性RND3的施用可降低微血管高透明性。我们还通过内皮细胞的活细胞成像观察到，层状脂蛋白层的形成和周转代表了先前未表征的细胞行为对于正常内皮屏障完整性很重要。我们假设RND3通过抑制RhoA/岩石介导的细胞收缩并激活Rac1介导的细胞间连接完整性增强，从而促进了炎症挑战后的内皮屏障恢复。我们的具体目的是：1）测试RND3需要RND3的Ser磷酸化和膜靶向的预测，以降低微血管高度过度性； 2）测试RND3促进岩石介导的细胞收缩的负反馈抑制的预测，从而增强了内皮屏障完整性； 3）测试RND3增强RAC1介导的薄片层状形成和VE-cadherin在交界处的稳定的预测，从而促进内皮屏障完整性。提出的新颖的综合方法利用了出血性休克的精致大鼠模型，结合了体内肠系膜微循环和分离的静脉静脉静脉静脉内显微镜，以评估微血管渗透性。我们将通过采用创新方法将可渗透的RND3蛋白传递给肠系膜微循环的细胞渗透性RND3的核心作用。活内皮细胞中培养的内皮细胞单层通透性模型，GFP-肌动蛋白和GFP-VE-钙粘蛋白动力学的成像，生化研究将支持体内和孤立的网状网络研究。这项研究的结果将使我们能够开发出一种新的理论，即在炎症过程中如何恢复内皮屏障功能，这将作为新疗法的基础。发现可用于解决微血管高温性的靶标将彻底改变创伤患者的治疗，还将创造新的机会来治疗与广泛的疾病相关的水肿。