Ectopic expression of WIF1 in PC3 cells results in a reduction of cell migratory and invasive capacity and a decrease in MMP-2 and-9 expression and activities. A, representative photomicrographs of scratch wounds at 0 and 30 h after wounds were made. Quantitative measurement of wound gaps by Photoshop software showed a reduced cellular motility in the WIF1-transfected PC3 cells compared with vector control cells. Columns, mean fold changes of wound width in WIF1 versus vector control-transfected PC3 cells; bars, SD. Experiments were replicated thrice. B, cells were applied to the upper surface of a Matrigel-coated membrane. After incubation for 48 hours, the upper surface of the membrane was scrubbed free of cells; the membrane was fixed, stained, and photographed. Representative pictures were taken from the lower surface of four independent membranes at ×100 magnification. Invasion index was calculated by adjusting cellular motility as described in detail in Materials and Methods. Columns, mean of invasion index calculated from four independent membranes; bars, SE. C, MMP-2 and MMP-9 activities in the conditioned medium from the transfectants were assayed by zymography as described in Materials and Methods. Semiquantitative comparison of MMP activities between PCDNA3.1 vector control and WIF1-transfected cells was done by densitometry. Columns, mean from three independent experiments; bars, SE. D, Western blot analysis of MMP-2 and MMP-9 expression. Representative blots were from three independent experiments. β-Actin serves as loading control.