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: 8496100
• 负责人: JEROME W BRESLIN
• 金额: $ 35.58万
• 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/8496100
• 关键词: Abbreviations; Actins; Adhesions; Admission activity; Adult Respiratory Distress Syndrome; Binding; Biochemical; 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; trauma care; 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（由申请方提供）：微血管通透性过高是全身炎症反应综合征（SIRS）的标志，93%的重症监护患者存在这种症状。SIRS使创伤受害者的复苏严重复杂化，并影响临床结果，经常进展为多器官衰竭。估计每年与SIRS相关的美国医疗保健费用负担为167亿美元。尽管对炎症诱导的微血管通透性过高的理解有所进展，但在炎症激发后使微血管通透性恢复正常的机制尚不清楚。此外，一旦炎症级联反应已经启动，现有的临床治疗对于恢复微血管屏障完整性是无效的。我们提出了一种新的范式的信号机制负责恢复微血管的完整性后，炎症引起的创伤性损伤。我们的初步数据表明，在失血性休克大鼠模型中，给予多胺包被的、细胞可渗透形式的Rnd 3（Rho家族小GTd 3）可降低微血管通透性过高。我们还观察到，与内皮细胞的活细胞成像，板状伪足的形成和营业额代表了以前的未表征的细胞行为的重要正常内皮屏障的完整性。我们推测Rnd 3通过抑制RhoA/ROCK介导的细胞收缩和激活Rac 1介导的细胞间连接完整性增强来促进炎症刺激后内皮屏障的恢复。我们的具体目标是：1）测试Rnd 3的Ser磷酸化和膜靶向是Rnd 3降低微血管高通透性所需的预测; 2）测试Rnd 3促进ROCK介导的细胞收缩的负反馈抑制的预测，这增强了内皮屏障完整性; 3）为了测试Rnd 3增强Rac 1介导的片状伪足形成和VE-钙粘蛋白在连接处的稳定性的预测，促进内皮屏障完整性。所提出的新的，综合的方法利用了一个完善的和相关的大鼠模型失血性休克结合活体显微镜在体内肠系膜微循环和孤立的小静脉方法来评估微血管通透性。我们将探索Rnd 3的核心作用，采用创新的方法，提供细胞可渗透的Rnd 3蛋白质的肠系膜微循环。培养的内皮细胞单层渗透性模型，在活的内皮细胞中的GFP-肌动蛋白和GFP-VE-钙粘蛋白动力学的成像，和生物化学研究将支持在体内和分离的小静脉研究。这项研究的结果将使我们能够开发一种新的理论，即在炎症过程中如何恢复内皮屏障功能，这将成为新疗法的基础。发现可用于解决微血管通透性过高的靶点将彻底改变创伤患者的治疗，也将为治疗与多种疾病相关的水肿创造新的机会。