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: 8903501
• 负责人: JEROME W BRESLIN
• 金额: $ 1.57万
• 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/8903501
• 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（由申请人提供）：微血管高渗透性是93%的重症监护患者出现全身性炎症反应综合征（SIRS）的标志。SIRS严重复杂化了创伤患者的复苏和恶化的临床结果，经常进展为多器官衰竭。据估计，美国每年与SIRS相关的医疗保健费用负担为167亿美元。尽管对炎症诱导的微血管高通透性的了解有所进展，但在炎症攻击后微血管通透性恢复正常的机制尚不清楚。此外，一旦炎症级联发生，现有的临床治疗方法对于恢复微血管屏障的完整性是无效的。我们提出了一种新的信号机制范式，负责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，在失血性休克大鼠模型中，多胺包被的Rnd3 （Rho家族小GTPase）细胞渗透性可降低微血管高通透性。我们还通过内皮细胞的活细胞成像观察到，板足的形成和转换代表了一种以前未被表征的细胞行为，对正常内皮屏障的完整性很重要。我们假设Rnd3通过抑制RhoA/ rock介导的细胞收缩和激活rac1介导的细胞间连接完整性增强来促进炎症挑战后内皮屏障的恢复。我们的具体目的是：1)验证Rnd3降低微血管高通透性需要丝氨酸磷酸化和膜靶向的预测；2)验证Rnd3促进rock介导的细胞收缩负反馈抑制，从而增强内皮屏障完整性的预测；3)验证Rnd3增强rac1介导的板足形成和VE-cadherin在连接处的稳定，促进内皮屏障完整性的预测。提出的新颖、综合的方法利用了一种完善的、相关的大鼠失血性休克模型，结合体内肠系膜微循环的活体显微镜和分离小静脉方法来评估微血管的通透性。我们将通过采用一种创新的方法将细胞渗透性Rnd3蛋白递送到肠系膜微循环来探索Rnd3的核心作用。培养内皮细胞单层通透性模型、活内皮细胞中GFP-actin和GFP-VE-cadherin动力学成像以及生化研究将支持体内和离体小静脉研究。这项研究的结果将使我们能够发展一种新的理论，即内皮屏障功能如何在炎症期间恢复，这将成为新疗法的基础。发现可用于解决微血管高渗透性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。