Fig. 7.

Exposure to intratumoral hypoxia promotes invasion. a Schematic representation of the early and late steps of the metastatic cascade. b Tumor-derived cells were incorporated with 90% non-labeled wild-type MDA-MB-231 cells in a 3D spheroid fully embedded in collagen. After 4 days in culture, spheroids were subjected to time-lapse imaging every 5 min for 16 h. Phase-contrast and fluorescent images taken on days 0 and 4 of the same spheroid. Scale bar = 100 μm. The white dashed ring marks the invasive front of the spheroid. c, d Total diffusivity (mm/min) (c) and persistent time (min) (d) of DsRed+ and GFP+ (mean ± SEM, N = 3, n = 80–92 cells); ****P < 0.0001 GFP versus DsRed (two-tailed t-test). e Projection of DsRed+ (red) or GFP+ (green) cell trajectories. Scale bar = 100 μm. f Number of DsRed+ and GFP+ cells at the invasive front of the spheroid (beyond the dashed white line in (b)) (mean ± SEM, N = 3, n = 44 spheroids); ****P < 0.0001 GFP versus DsRed (two-tailed t-test)