Fig. 2

ADAM12 knockdown reduces cell migration, invasion, resistance to anoikis, and sphere formation. SUM159PT cells expressing inducible ADAM12 shRNA (shADAM12) or control shRNA (shControl) were treated with doxycycline (Dox; 1 μg/ml) for 4 days. a Cell migration was analyzed using Transwell assays. The data are shown as means ± SEM from 3 determinations. n.s., non-significant, P > 0.05. b Representative phase-contrast images of SUM159PT_shADAM12 and SUM159PT_shControl cells embedded in 3D Matrigel. Scale bar, 100 μm. c Anoikis (apoptotic cell death upon detachment) was assayed using a DNA fragmentation ELISA kit. A time course of cell death was measured in triplicates (raw data, left), and the mean values of DNA fragmentation, ±SEM, after 24 h of detachment obtained in three independent experiments were calculated (right). *, P < 0.05. d Sphere formation was evaluated after seeding single-cell suspensions in serum-free sphere medium containing 1% methylcellulose into ultra-low attachment plates. After 7 days, spheres with diameters >50 μm were photographed and counted using ImageJ. The results are shown as mean fold change in sphere numbers after doxycycline treatment, normalized to shControl cells without doxycycline, ±SEM, obtained in 3 independent experiments