FIG 5.

Pak2 is required for endothelial barrier maintenance. (A) MLECs and HUVECs were plated to confluence on 1- or 0.4-μm Transwell filters. Trypan blue-BSA or FITC-dextran was added to the top well, and the amount of trypan blue-BSA or FITC-dextran in the bottom chamber was measured over 60 min (60′). Exogenous Pak2 was expressed by transducing MLECs with Pak2-expressing adenovirus. OD600, optical density at 600 nm. (B) In vivo permeability assay in VEC-CreER^T2; Pak2^fl/fl, VEC-CreER^T2, and Pak1 KO mice. Western blot shows deletion of Pak2 and Pak1 in corresponding MLECs. (C) Evans blue dye was injected in the tail veins of VEC-CreER^T2; Pak2^fl/fl, VEC-CreER^T2, and Pak1 KO mice that were dosed with 4-OHTx. Evans blue dye was injected into mice, and the dye was extracted with formamide. The amount of extravasated dye was normalized to the tissue mass (in nanograms of Evans blue per milligram of tissue). Error bars represent SD. Experiments were repeated at least three times. **, P < 0.01; NS, not significant. (D) Survival curve for CreER^T2; Pak2^fl/fl or CreER^T2; Pak2^wt/wt mice dosed with 100 mg/kg or 200 mg/kg 4-OHTx.