Inhibition of Rap1 reduced endothelial adhesion. (A) HUVECs were transfected with Rap1GAP1-FLAG or empty vector, and adhesion assays were performed with transfected endothelial cells. After 24 hours, cells were allowed to adhere for one hour on fibronectin (n = 6) or collagen (n = 9; *P < .05 vs empty vector). (B) Adhesion assay with endothelial cells transfected with siRNAs targeted against Rap1a, Rap1b, Rap1a/Rap1b, or scrambled siRNA. After 48 hours, cells were allowed to adhere for 1 hour on fibronectin-coated wells (n = 5). Data are presented as percentage of adhering cells plus or minus SEM (*P < .05 vs scrambled). (C) Twelve hours after transfection, HUVEC cells were grown on 4-well chamber slides. At 48 hours after transfection, serum-starved cells were left untreated or stimulated with 100 μM 8-pCPT-2′-O-Me-cAMP for 10 minutes. Immunofluorescence was performed using HUTS21 antibodies. Representative pictures from 3 different experiments are depicted. Bar represents 10 μm. Micrographs were acquired with an LSM 510 confocal microscope fitted with a Plan-Neofluar 40×/1.3 NA oil objective and LSM5 image acquisition software (all from Carl Zeiss).