Role for P-Rex1 in Mediating Acute Lung Injury

Rationale: Maintenance of vascular endothelial integrity is of great importance to homeostasis of vital organ functions. The small GTPase Rac is one of the key signaling mediators for vascular endothelial functions, but how Rac activation is regulated under various pathophysiological conditions remains incompletely understood. Objective: The primary objective of this study was to investigate a role for P-Rex1 (PIP3 dependent Rac exchanger 1) a guanine nucleotide exchange factor for Rac, previously known for G protein-coupled receptor signaling, in mediating TNF-alpha-induced vascular hyper-permeability. Methods and Results: Using gene deletion and knockdown approaches, we investigated the potential role of the phosphoinositide- and G protein beta gamma subunits-regulated guanine nucleotide exchange factor P-Rex1 in TNF-alpha-induced lung vascular injury. P-Rex1, previously found in neutrophils and neurons, is also expressed in endothelial cells. In cultured human lung microvascular endothelial cells (HLMVECs), TNF-alpha exposure led to endothelial junctional disruption but, small interference (si) RNA-mediated knockdown of P-Rex1 prevented intercellular gap formation. Silencing P-Rex1 markedly attenuated TNF-alpha-induced Rac activation and reactive oxygen species (ROS) production. TNF-alpha stimulated P-Rex1 membrane translocation and the resulting Rac activation in a PI3K dependent manner. Both, in vivo and in vitro, absence of P-Rex1 resulted in significantly less ICAM-1 induction and neutrophil transendothelial migration. Moreover, endothelial P-Rex1 plays a predominant role over neutrophil P-Rex1 in this process. P-Rex1 knockout mice were also refractory to TNF-alpha-induced lung vascular hyper-permeability and edema. Conclusions: These results demonstrate for the first time that P-Rex1 expressed in endothelial cells, is activated downstream of TNF-alpha, which is not a GPCR agonist. We conclude that P-Rex1 is a critical mediator of endothelial barrier disruption and therefore could be a novel therapeutic target in the control of vascular permeability and neutrophil infiltration to inflammatory tissues.