Role of End Binding 3 in Mechanism of vascular permeability

描述（由申请方提供）：肺血管通透性增加导致富含蛋白质的组织水肿，这是成人呼吸窘迫综合征的一个重要特征。微管（MT）细胞骨架在内皮细胞通透性增加机制中的作用还不清楚。已知MT响应于促炎介质而经历重组，并且因此可能有助于增加内皮渗透性的机制。拟议的研究将解决末端结合蛋白-3（EB 3），一种MT加末端结合因子，在调节肺血管通透性增加中的核心作用。我们将检验以下假设：（i）EB 3是血管内皮（VE）-钙粘蛋白粘附复合物和MT细胞骨架之间串扰的主要成分;（ii）EB 3介导的MT动力学控制对于维持肺微血管的基础渗透性和炎症介质引起的渗透性增加至关重要。这些研究将解决以下特定目的：（1）VE-钙粘蛋白介导的信号传导在EB 3磷酸化机制和MT生长抑制中的作用，从而“由外向内”信号传导在建立肺内皮细胞基础渗透性中的作用;和（2）EB 3在调节Ca 2+信号传导中的关键作用，从而介导内皮细胞渗透性增加和肺水肿的发展。我们期望通过了解VE-钙粘蛋白粘附如何信号传导EB 3磷酸化以及EB 3如何由此促进屏障通透性增加，将为肺液稳态失调的机制提供新的见解。我们将利用最先进的技术，包括活细胞成像，突变体构建体的表达，基因转移和肺部炎症的小鼠模型，以实现特定的目标。 公共卫生相关性：这项工作的重点是肺内皮，其中计划的研究将建立EB 3（一种微管结合蛋白）与ALI/ ARDS病理生理学的相关性。导致这一提议的工作使我们能够设计一种潜在的治疗肽，通过抑制EB 3功能来预防脓毒症中的肺水肿和致死性。

DESCRIPTION (provided by applicant): Increases in lung vascular permeability result in protein rich tissue edema, an important feature of adult respiratory distress syndrome. The role of the microtubule (MT) cytoskeleton in the mechanism of increased endothelial permeability is not well understood. MTs are known to undergo re-organization in response to pro- inflammatory mediators, and may thus contribute to the mechanism of increased endothelial permeability. The proposed studies will address the central role of End Binding protein-3 (EB3), a MT plus-end binding factor, in regulating increased lung vascular permeability. We will test the hypotheses that (i) EB3 is a major component of cross-talk between Vascular Endothelial (VE)-cadherin adhesion complexes and the MT cytoskeleton and (ii) EB3-mediated control of MT dynamics is critical for maintenance of basal permeability of lung microvessels and for permeability increase caused by inflammatory mediators. These studies will address the following Specific Aims: (1) role of VE-cadherin-mediated signaling in the mechanism of EB3 phosphorylation and inhibition of MT growth and, thereby the role of "outside-in" signaling in establishing basal permeability of lung endothelia; and (2) critical role of EB3 in regulating Ca2+ signaling, thus in mediating increased endothelial permeability and development of lung edema. It is our expectation that by understanding how VE-cadherin adhesion signals EB3 phosphorylation and how EB3 thereby elicits the barrier permeability increase will provide novel insights into the mechanisms of dysregulation of lung fluid homeostasis. We will exploit state of the art technologies including live cell imaging, expression of mutant constructs, gene transfer, and murine models of lung inflammation to accomplish the specific aims. PUBLIC HEALTH RELEVANCE: The focus of the work is the lung endothelium, in which the planned studies will establish the relevance of EB3, a microtubule-binding protein, to the pathophysiology of ALI/ ARDS. The work leading up to this proposal has allowed us to design a potential therapeutic peptide that by inhibiting EB3 function prevents lung edema and lethality in sepsis.