Piezo1 Mediated Adjustments in Lung Fluid Balance

项目总结/摘要 粘附连接（AJs）水平的肺内皮屏障完整性是肺液体稳态所必需的。 导致肺动脉高压等疾病中内皮屏障完整性丧失的一个关键机制是， 水肿是肺毛细血管响应高压的"应力衰竭"。虽然已知AJs， 由VE-钙粘蛋白和相关的连环蛋白组成，限制内皮通透性，关于 机械力，特别是血管壁张力，如何控制内皮通透性和肺水肿。 我们的支持性数据描述了微血管中的静水压力在激活微血管中的潜在重要作用。 机械传感器Piezo1在内皮细胞（EC）和增加内皮屏障通透性。我们 观察到Piezo1的激活诱导细胞内Ca2+信号传导，这反过来又引起磷酸化， VE-钙粘蛋白和微血管通透性增加。这些发现首次将增加的 EC暴露于Piezo1激活和AJs分解的张力，导致 基本问题"Piezo1如何在EC的质膜上感测到张力激活VE- 钙粘蛋白磷酸化，从而破坏AJs？"在目标1中，我们将确定微血管的作用 激活肺EC中机械传感器通道Piezo1的压力和Piezo1在调节 内皮渗透性和肺液体平衡。我们将确定Src依赖的磷酸化 Piezo1激活的Ca2+信号传导需要Piezo1，以及这是否因此介导增加 内皮通透性在目标2中，我们将确定Piezo1信号传导在介导拆卸中的作用 通过VE-钙粘蛋白的磷酸化，以及增加内皮通透性。在这里，我们将 鉴定诱导VE-钙粘蛋白磷酸化的Piezo1激活下游的信号传导途径， VE-钙粘蛋白内吞作用，从而分解AJs。在目标3中，我们将确定Piezo 1在以下方面的作用： 介导与左心衰相关的肺血管通透性过高（"应激衰竭"）和水肿 （LHF）。这些研究将阐述Piezo 1在肺动脉高压机制中的病理生理学相关性。 由LHF诱导的肺微血管压力增加导致的水肿。上述研究将至关重要 了解Piezo 1在增加肺微血管通透性中的作用，目的是确定新的 治疗目标的高压力引起的肺水肿。

PROJECT SUMMARY / ABSTRACT Lung endothelial barrier integrity at the level of adherens junctions (AJs) is required for lung fluid homeostasis. A crucial mechanism contributing to the loss of endothelial barrier integrity in conditions such as pulmonary edema is “stress failure” of pulmonary capillaries in response to high pressure. While it is known that AJs, comprised of VE-cadherin and associated catenin proteins, restrict endothelial permeability, little is known about how mechanical forces, specifically vessel wall tension, control endothelial permeability and pulmonary edema. Our Supporting Data describe the potentially important role of hydrostatic pressure in microvessels in activating the mechanosensor Piezo1 in endothelial cells (ECs) and in increasing endothelial barrier permeability. We observed that activation of Piezo1 induced intracellular Ca2+ signaling, which in turn, caused phosphorylation of VE-cadherin and increased microvascular permeability. These findings have for the first time linked increased tension to which ECs are exposed to the activation of Piezo1 and disassembly of AJs, leading to the fundamental question “how does tension sensed at the plasma membrane of ECs by Piezo1 activate VE- cadherin phosphorylation and thereby disrupt AJs?” In Aim 1, we will determine the role of microvessel pressure in activating the mechanosensor channel Piezo1 in lung ECs and Piezo1’s role in regulating endothelial permeability and lung fluid balance. We will determine whether Src dependent phosphorylation of Piezo1 is required for Piezo1 activated Ca2+ signaling in ECs and whether this thereby mediates increased endothelial permeability. In Aim 2, we will determine the role of Piezo1 signaling in mediating disassembly of AJs through phosphorylation of VE-cadherin, and in increasing endothelial permeability. Here, we will identify the signaling pathway downstream of Piezo1 activation that induces phosphorylation of VE-cadherin and VE-cadherin endocytosis and thus disassemble the AJs. In Aim 3, we will determine the role of Piezo1 in mediating lung vascular hyper-permeability (“stress failure”) and edema associated with left heart failure (LHF). These studies will address the pathophysiological relevance of Piezo1 in the mechanism of pulmonary edema resulting from LHF-induced increases in lung microvessel pressure. The above studies will be essential for understanding the role of Piezo1 in increasing lung microvessel permeability, with the goal of identifying new therapeutic targets for high pressure-induced pulmonary edema.