Differential mechano-signaling in vascular endothelium by varying degrees of mechanical stretch - Resubmission 01

不同程度机械拉伸对血管内皮细胞不同力学信号的影响 对微血管内皮细胞施加过大的机械力会导致微血管的增加 通透性伴随着危及生命的情况，如中风、肺动脉高压或呼吸机 诱发性肺损伤，仅举几例。我们之前已经描述了血管中诱导的信号通路。 并描述了Rho GTP酶在高循环拉伸中的关键作用。 诱导内皮细胞屏障功能障碍。然而，细胞机械转导复合体转化为 对细胞反应的机械信号仍有待表征。我们未发表的试点研究表明 RAP1信号被生理上相关的低幅度(5%)CS激活，并促进重组 在病理上相关的高幅度(18%)CS中，紧密连接被细胞预适应破坏。这个 初步数据还表明，低CS诱导的EC屏障恢复与 紧接连接处的接头蛋白cinglin形成cinglin-gef-h1复合体。中环 本应用验证的假说是病理机械作用后血管内皮细胞屏障的恢复 应激可能通过细胞暴露于生理性CS水平而加速，并涉及RAP1依赖的重组 血管内皮细胞紧密连接的发生、cinglin向紧密连接的募集和cinglin-gef-h1的刺激 互动。这些事件导致了全球环境基金-H1核苷酸交换活性的抑制，Rho-H1的抑制。 依赖的屏障破坏机制，以及血管内皮屏障的加速恢复。这个 将解决以下问题：AIM-1将研究内皮屏障的cinglin依赖机制 通过生理机械拉伸进行调节。AIM-2将研究cinglin在Rho下调中的作用 病理性向生理性转变后机械刺激微血管内皮细胞的途径 CS幅度。AIM-3将在Cinglin依赖的血管保护机制的小鼠模型中评估 机械通风。

Differential mechano-signaling in vascular endothelium by varying degrees of mechanical stretch Excessive mechanical forces imposed on the microvascular endothelium lead to increased microvascular permeability accompanying life-threatening conditions such as stroke, pulmonary hypertension, or ventilator induced lung injury, to name a few. We have previously characterized signaling pathways induced in vascular endothelium by high magnitude cyclic stretch (CS) and described a key role of Rho GTPase in high CS- induced endothelial barrier dysfunction. However, cellular mechanotransduction complexes which transform mechanical signals to cellular responses remain to be characterized. Our unpublished pilot studies indicate that Rap1 signaling is activated by physiologically relevant low magnitude (5%) CS and promotes re-assembly of tight junctions disrupted by cell preconditioning at pathologically relevant high magnitude (18%) CS. The preliminary data also suggest that low CS-induced EC barrier restoration is associated with accumulation of adaptor protein cingulin at the tight junctions and formation of cingulin-GEF-H1 complex. The central hypothesis tested in this application is that recovery of vascular endothelial barrier after pathologic mechanical stress may be accelerated by cell exposure to physiologic CS levels and involves Rap1-dependent reassembly of endothelial tight junctions, recruitment of cingulin to the tight junctions, and stimulation of cingulin - GEF-H1 interaction. These events lead to inhibition of GEF-H1 nucleotide exchange activity, suppression of Rho- dependent barrier disruptive mechanisms, and accelerated recovery of the vascular endothelial barrier. The following questions will be addressed: Aim-1 will study cingulin-dependent mechanisms of endothelial barrier regulation by physiologic mechanical stretch. Aim-2 will study the role of cingulin in downregulation of the Rho pathway in mechanically stimulated microvascular endothelium after a switch from pathologic to physiologic CS amplitude. Aim-3 will evaluate cingulin-dependent vascular protective mechanisms in a mouse model of mechanical ventilation.