OL-pc expression accelerates cell movement in wound healing. (A) Confluent cell layers were wounded by cell scraping, and the wounded edges were immediately processed for time-lapse recording. Phase-contrast images were taken for 4 h at 3-min intervals. At the end of the recording, individual cells located along the front of cell sheets were marked with colored circles over their nuclear positions, and the same cells were tracked back to earlier time points. At each time point, the circles were connected with lines in a fixed order. Dashed lines represent wound edges. The numbers denote the time elapsed (in minutes). Control and OL-pcΔNBS transfectants roughly maintain their relative positions during migration, whereas OL-pc transfectants irregularly change their spatial relations with neighbors (e.g., the cell marked with the red circle at the right jumped out to the front position from an initially deep site). Videos 3–5 are available. (B) Migration track of individual cells marked with different colors in A, with cell position plotted at 9-min intervals. The arrow indicates the direction of wound healing. Abrupt acceleration of movement is most frequently seen in OL-pc transfectants. (C) Statistical analysis of the instantaneous velocities of cells during wound healing. Comparison of each experimental group with the control cells (n = 38) shows a significant difference for OL-pc (**, P < 0.0005; n = 51) but not for OL-pcΔNBS (P = 0.70; n = 41). (D) Changes in the positions of cells relative to those of their neighbors during migration. At the end of recording (at 240 min after scraping), all cells at the wound edges were marked as in A, and their migration track was drawn as in B. Choosing a pair of cells located next to each other at the 240-min point, the distances between their nuclei at earlier time points were measured at 15-min intervals, and their means as well as the farthest and nearest distances were obtained. The farthest and nearest indices were defined as the ratio of the farthest (Dmax) and nearest (Dmin) distance to the average distance (Davg), respectively, as illustrated in the left panel. The farthest index for OL-pc transfectants is significantly larger (**, P < 0.0005; n = 77) than that for the control cells (n = 58), whereas that for OL-pcΔNBS transfectants (P = 0.39; n = 63) is not. Likewise, the nearest index for OL-pc transfectants is significantly smaller (*, P < 0.005) than that for the controls, whereas that for OL-pcΔNBS (n.s.; P = 0.87) is not. (E) Changes in the mean distance (±SEM [error bars]) of the migrating wound edges from their starting position during a long incubation period up to 11 h. n = 12 for all samples. Bar, 50 μm.