Inactivation of hyperpermeability after ischemia-reperfusion induced inflammation

描述（由申请方提供）：微血管通透性增加是炎症的标志，是缺血-再灌注初始事件的特征。以前的研究主要集中在如何预防发病和维持渗透性升高。我们建议研究在缺血再灌注的炎症阶段抑制高通透性并使其恢复到横纹肌基线水平的机制。我们推测，在高通透性阶段后，缺血后肌肉开始一个过程，以消除高通透性和恢复微血管壁的生理屏障特性。我们进一步提出Epac和Rap-1作为“屏障增强因子”参与高渗透性失活过程。微血管屏障特性的维持主要通过控制形成细胞间粘附的蛋白质的因子来调节。在许多细胞中，细胞-细胞粘附又部分地通过小GTP结合蛋白和连接蛋白之间的反馈信号传导来调节。具体目的1：研究缺血-再灌注诱导炎症后高通透性的定时失活。具体目的2：确定Epac/Rap-1的激活是否使缺血-再灌注中的高通透性失活。具体目标3：测试内皮Epac/Rap-1信号传导是否负责屏障完整性的恢复。我们将应用显微镜，计算机辅助图像分析和分子生物学方法在横纹肌（在体内）和内皮细胞，以阐明这些相关的cAMP刺激因子在缺血后肌肉通透性过高的失活阶段的作用。我们将使用暴露于氧合和再氧合的内皮细胞来探索细胞机制。身体对炎症和缺血再灌注的反应是高度复杂的，涉及精心策划的分子机制。了解缺血再灌注中高渗透性与其“生理”失活之间的时间，应提供一个机会之窗，用于促进高渗透性失活的干预措施，以防止过度水肿、间室综合征和组织损伤。我们的研究结果对择期和急诊血管手术的血运重建具有临床意义。公共卫生相关性：我们的研究结果将促进目前的分子机制的了解，在失活增加的渗透性。新出现的数据将在血运重建的背景下具有临床相关性。这些结果可以作为新的辅助治疗方法的基础，以帮助血管外科医生预防损伤和恢复缺血后肌肉的功能。

DESCRIPTION (provided by applicant): Increased microvascular permeability is a hallmark of inflammation and characterizes the initial events in ischemia-reperfusion. Previous research has focused on how to prevent the onset and the maintenance of the elevated permeability. We propose to investigate the mechanisms that inactivate hyperpermeability in the inflammatory phase of ischemia-reperfusion and return it to baseline levels in striated muscle. We hypothesize that after a hyperpermeability phase, the postischemic muscle begins a process to inactivate hyperpermeability and restore the physiological barrier properties of the microvascular wall. We further propose that Epac and Rap-1 serve as `Barrier Enhancing Factors' and participate in the hyperpermeability-inactivation process. Maintenance of microvascular barrier properties is mainly regulated through factors controlling proteins that form intercellular adhesions. Cell-cell adhesion, in turn is regulated in many cells in part through feed-back signaling between small GTP-binding proteins and junctional proteins. The Specific Aims designed to test the hypothesis are: Specific Aim 1: To investigate the timed inactivation of hyperpermeability after ischemia- reperfusion induced inflammation. Specific Aim 2: To determine whether activation of Epac/Rap-1 inactivates hyperpermeability in ischemia-reperfusion. Specific Aim 3: To test whether endothelial Epac/Rap-1 signaling is responsible for restoration of barrier integrity. We will apply microscopy, computer-assisted image analysis and molecular biology approaches in striated muscle (in vivo) and in endothelial cells to elucidate the role of these relevant cAMP-stimulated factors in the inactivating phase of hyperpermeability in postischemic muscle. We will use endothelial cells exposed to oxygenation and reoxygenation to explore the cellular mechanisms. The body response to inflammation and to ischemia reperfusion is highly complex and involves well orchestrated molecular mechanisms. Knowledge of the timing between hyperpermeability and its "physiological" inactivation in ischemia-reperfusion should provide a window of opportunity for interventions promoting inactivation of hyperpermeability to prevent excessive edema, compartment syndrome and tissue damage. Our results should be of clinical relevance to revascularization in elective and emergency vascular surgery. PUBLIC HEALTH RELEVANCE: Our results will advance current understanding of the molecular mechanisms involved in the inactivation of increased permeability. The emerging data will be clinically relevant in the settings of revascularization. The results could serve as a basis for new adjuvant therapeutic approaches to assist vascular surgeons in preventing damage and returning function to postischemic muscles.