Figure 5.

PC3 cells expressing CD44v8-10 display a high invasive potential in vitro (A) Transwell migration/invasion assays of PC3 subpopulations. Data are expressed as fold increase of migration and invasion relative to CD44v8-10^neg cells (set arbitrarily at 1). (B) Representative images of migration/invasion rate of DAPI-labeled PC3 different populations. (C) Representative zymography of three performed, of PA secreted by CD44v8-10^pos and CD44v8-10^neg PC3 cells. At the indicated times, conditioned media were collected and analyzed by casein-agar underlay. The lytic zones were plasminogen dependent and were inhibited by 1 mM amiloride, an uPA-specific inhibitor. (D) The same conditioned media were assayed for uPA activity by chromogenic substrate assay. Results represent the mean ± SEM of three independently sorted cell subpopulations. (^***p < 0.001; ^**p< 0.01). (E) Transwell migration/invasion assays of CD44v8-10^pos PC3 after specific CD44v8-10 down-regulation by siRNA. Data are expressed as fold decrease of migration and invasion relative to CD44v8-10^pos cells treated with scramble RNA (set arbitrarily at 1). Three sample of each subpopulation were examined and the cells counted in seven fields/sample. (^***p < 0.001). (F) Extent of CD44v8-10 knockdown protein after specific CD44v8-10 siRNA or control scramble RNA assayed by flow cytometry. All the data were analyzed by Student's paired t-test. n=3 mean ± S.E.M. (G) Representative FACS plots of CD44v8-10 membrane protein expression in CD44v8-10^pos PC3 cells four days after specific CD44v8-10 siRNA or control scramble RNA treatments.