Erg is required for EC migration. (A) HUVECs were transfected with either control-GB FITC or Erg-specific GB FITC for 30 hours. Time-lapse confocal microscopy was used to visualize EC directional migration after a scratch wound. (i) Representative images of scratch wounds at the start and 16 hours after scratching show that compared with control cells (left), directional migration in Erg-deficient cells (right) is reduced. Scale bar, 200 μm. (ii) Single-cell tracking was used to analyze the migratory pathway of individual cells adjacent to the wound. Each track represents the migratory routes of a selected FITC-labeled cell during the course of the 16-hour experiment. (iii) Migration plots showing the migration distance and directionality of single-cell tracks. (iv) Quantification of single-cell velocities, Euclidean distance and accumulated distance were determined from single-cell tracking. Three sets of time-lapse images were captured along each scratch, and 25-40 cells were analyzed per image. *P < .05. Values are mean ± SEM, n = 3 separate experiments. (B) HUVECs were transduced with adenovirus (Ad-LacZ or Ad-Erg) for 48 hours. Endothelial cell directional migration was visualized after a scratch wound by time-lapse confocal microscopy. (i) Representative images showing scratch wounds at the start and at 7.5 hours post-scratch show that compared with Ad-LacZ control cells (left), migration of Erg-overexpressing cells (Ad-Erg, right) is increased. Scale bar, 200 μm. (ii) Single-cell tracking was used to map the migratory pathway of individual cells adjacent to the wound, as above. (iii) Migration plots showing the migration distance and directionality of single-cell tracks. (iv) Quantification of single-cell velocities, Euclidean distance, and accumulated distance were determined from single-cell tracking. Three sets of time-lapse images were captured along each scratch, and 25-40 cells were analyzed per image. ***P < .001. Values are mean ± SEM, n = 3 separate experiments.