Figure 1

AT-MSCs induced epithelial–to-mesenchymal transition and increased mammosphere formation. A) SKBR3 cells were cultured in DMEM or MSC-CM for 6 days. Phase-contrast images revealed shift from epithelial to mesenchymal-like morphology in the majority of tumor cells (magnification 40x). B) AT-MSCs and EGFP-SKBR3 were directly cocultured for 6 days. Fluorescence microscopy confirmed morphological signs of an EMT in the tumor cells (magnification 100x). C) Markers of the EMT and pluripotency were up-regulated in the tumor cells cultured in MSC-CM. Quantitative RT-PCR confirmed significant increase in the expression of Nanog, Oct, Twist, Snail1, Snail2, αSMA and FAP in EGFP-SKBR3 cells exposed to MSC-CM in comparison to EGFP-SKBR maintained under the standard culture conditions. The data are expressed as means ± SD, *p < 0.05, ^#p < 0.001. D) Non-adherent culture conditions increased EGFP-SKBR3 mammosphere formation in the presence of MSC-CM, which could be abrogated by specific inhibitors of PI3K (1.63 μM LY294002) and p38 MAP kinases (0.5 μM SB203580) (magnification: light microscopy 40x, fluorescent microscopy 100x). Relative viability of EGFP-SKBR3 cells exposed to MSC-CM in the presence of inhibitors did not significantly differ from the viability in DMEM as evaluated by luminescent viability assay (right panel).