The human umbilical vein endothelial cell line EA.hy926 (ATCC; Mannasas, VA, USA) was cultured in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal calf serum (FCS) in an atmosphere of 5% CO2/95% air and 37°C. EA.hy926 cells (105cells/plate) were seeded in transwell inserts (Corning® BiocoatTM Cell Culture Inserts Collagen, Type I Rat Tail, 24-Well, 3 µm; Corning, NY, USA) and were cultured to confluence at 24 h. The cells were starved overnight and then activated with LPS (1 µg/ml) for 4 h. Upper chamber media, containing LPS (1 µg/ml) and soluble endoglin (500 ng/ml) for their respective treatments, were replaced with FITC-Dextran (40 kDa) at 1 mg/ml in DMEM. The bottom chambers were also replaced with DMEM. After 24 h at 37°C the inserts were removed, and the amount of fluorescence in the bottom chambers was measured using a fluorescence plate reader (Fluoroskan Ascent FL; Thermo Electron Corporation, Waltham, MA, USA).