Salinomycin Promotes Anoikis and Decreases the CD44+/CD24- Stem-Like Population via Inhibition of STAT3 Activation in MDA-MB-231 Cells
Fig 3
Salinomycin suppresses STAT3 activation.
(A) Basal levels of STAT3 and phospho-STAT3 (Tyr705) in seven breast cancer cell lines: luminal-type (MCF7 and T47D), HER2-amplified (BT474, MDA-MB-453, and SKBR3), and TNBC (Hs578T and MDA-MB-231). (B) After exposure of MDA-MB-231 cells to salinomycin (1–10 μM) or DMSO for 48 h, STAT3 and phospho-STAT3 protein content were determined by Western blot analysis. A quantitative graph of the phospho-STAT/STAT3 ratio is shown (bottom panel, * p<0.05). (C) Cells were treated with salinomycin (2 μM, 48 h) and immunostained for STAT3 (1:100, green) or phospho-STAT3 (1:100, green), with DAPI nuclear staining (blue). F-actin (1:100, red) was used as a cytosolic marker. Nuclear phospho-STAT3 intensity (y-axis) is represented in arbitrary units as defined by the software. (D) Cells were pretreated with IL-6 (0–10 ng/ml) for 1 h before salinomycin treatment (2 μM, 48 h). STAT3 and phospho-STAT3 protein levels were determined by Western blot analysis. Quantification of the phospho-STAT/STAT3 ratio is shown (bottom panel, *** p<0.001 and ### p<0.001). The results are expressed as mean ± SEM (n = 3, independent experiments) and were analyzed by one-way ANOVA followed by Bonferroni’s post hoc test (* p<0.05, ** p<0.01 and *** p<0.001, versus DMSO control; ### p<0.001, versus each treatment).