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A3734 - Signaling in Vascular Endothelium by Varying Degrees of Mechanical Stretch
Author Block: Y. Ke1, C. Zhang2, Y. Li2, T. Nguen2, P. Karki2, K. G. Birukov1, A. Birukova2; 1Dept of Anesthesiology, University of Maryland School of Medicine, Baltimore, MD, United States, 2Dept of Medicine, University of Maryland School of Medicine, Baltimore, MD, United States.
Mechanical ventilation remains an imperative treatment for the patients with acute respiratory distress syndrome (ARDS), but can also exacerbate lung injury. We have previously characterized signaling pathways induced in vascular endothelium by high magnitude cyclic stretch (CS) related to regions of suboptimal mechanical ventilation in inflammation-affected lung compartments and described a key role of RhoA GTPase in high CS-induced endothelial (EC) barrier dysfunction. However, cellular mechanotransduction complexes which transform mechanical signals to cellular responses remain to be characterized. This study tested a hypothesis that recovery of vascular EC barrier after pathologic mechanical stress may be accelerated by cell exposure to physiologic CS levels and involves Rap1-dependent rearrangement of endothelial tight junctions. Using biochemical, molecular, and imaging approaches we found that EC preconditioning at physiologically relevant low magnitude 5% CS promotes resealing of tight junctions disrupted by pathologic, high magnitude 18% CS. Moreover, the EC barrier recovery rate was higher in EC exposed to low CS vs. EC culture left at static conditions. Inhibitory analysis showed that low CS-induced TJ remodeling was dependent on small GTPase Rap1. Protective effects of EC preconditioning at low CS were abolished by an inhibitor of Rap1 processing GGTI-298 or siRNA-mediated Rap1 depletion. In a clinically-relevant two-hit model of EC exposed to bacterial particles followed by mechanical stimulation, low magnitude CS was also protective against deleterious effects by high magnitude CS. Using Rap1 knockout mice exposed to mechanical ventilation we found that protective effect of mechanical ventilation at low tidal volume (LTV) against lung injury caused by i.t. injection of LPS and mechanical ventilation at high tidal volume (HTV) was suppressed by molecular inhibition of Rap1, whereas administration of Rap1 activator significantly reduced parameters of ALI in wild type group. Taken together our results demonstrate a prominent role of signaling mechanisms activated by low CS in stimulation of the Rap1-mediated acceleration of lung vascular EC barrier restoration.