A potential role for reactive oxygen species and the HIF-1α–VEGF pathway in hypoxia-induced pulmonary vascular leak

Acute hypoxia causes pulmonary vascular leak and is involved in the pathogenesis of pulmonary edema associated with inflammation, acute altitude exposure, and other critical illnesses. Reactive oxygen species, HIF-1, and VEGF have all been implicated in various hypoxic pathologies, yet the ROS–HIF-1–VEGF pathway in pulmonary vascular leak has not been defined. We hypothesized that the ROS–HIF-1–VEGF pathway has an important role in producing hypoxia-induced pulmonary vascular leak. Human pulmonary artery endothelial cell (HPAEC) monolayers were exposed to either normoxia (21% O2) or acute hypoxia (3% O2) for 24 h and monolayer permeability and H2O2, nuclear HIF-1α, and cytosolic VEGF levels were determined. HPAEC were treated with antioxidant cocktail (AO; ascorbate, glutathione, and α-tocopherol), HIF-1 siRNA, or the VEGF soluble binding protein fms-like tyrosine kinase-1 (sFlt-1) to delineate the role of the ROS–HIF-1–VEGF pathway in hypoxia-induced HPAEC leak. Additionally, mice exposed to hypobaric hypoxia (18,000 ft, 10% O2) were treated with the same antioxidant to determine if in vitro responses corresponded to in vivo hypoxia stress. Hypoxia increased albumin permeativity, H2O2 production, and nuclear HIF-1α and cytosolic VEGF concentration. Treatment with an AO lowered the hypoxia-induced HPAEC monolayer permeability as well as the elevation of HIF-1α and VEGF. Treatment of hypoxia-induced HPAEC with either an siRNA designed against HIF-1α or the VEGF antagonist sFlt-1 decreased monolayer permeability. Mice treated with AO and exposed to hypobaric hypoxia (18,000 ft, 10% O2) had less pulmonary vascular leak than those that were untreated. Our data suggest that hypoxia-induced permeability is due, in part, to the ROS–HIF-1α–VEGF pathway.