Fig. 4

NSC12^sens cells are characterized by stem-like features. A qPCR analysis of NANOG, OCT4, SNAIL, SLUG, TWIST, SOX2, CD44, CD47, and ZEB2 expression in NSC12^res and NSC12^sens subpopulations of 92.1 and Mel270 cells. Data are the mean ± SEM of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 vs NSC12^res, Student’s t-test. B Western blot analysis of Nanog in control and NSC12^res/NSC12^sens Mel270 cells. Right panel: densitometric analysis of immunoreactive bands normalized to GAPDH protein levels. C Nanog^br cells were measured on control and NSC12^res/NSC12^sens 92.1 cells by cytofluorimetric analysis. Data are the mean ± SEM of three independent experiments. *p < 0.05 vs control; ^#p < 0.05 vs NSC12^res, ANOVA. D ALDH^br cells were measured on control and NSC12^res/NSC12^sens 92.1 cells by cytofluorimetric analysis. Representative dot plots are reported in the left panel. Data are the mean ± SEM of three independent experiments. ^#p < 0.05 vs NSC12^res, Student’s t-test. E 92.1 cells were engrafted onto the chick embryo CAM at day 7 post-incubation in the absence or in the presence of 4 pmol/embryo NSC12. Tumor growth was assessed after 7 days and qPCR analysis of NANOG was performed on the explants (right panels). In box and whisker graphs, boxes extend from the 25th to the 75th percentiles, lines indicate the median values, and whiskers indicate the range of values. Data are the mean ± SEM of two independent experiments (n = 20). *p < 0.05, ***p < 0.001 vs control, Student’s t-test. Representative images of tumors are shown in the left panel. Scale bar: 2 mm