Figure 6
Suppression of p38 signaling in androgen-independent cells results in reversal of EMT, significant decrease in FOXC2-dependent stem-like properties and restoration of sensitivity to Enzalutamide and Docetaxel. (a) Morphology of DU145 cells on SB203580 (specific p38 signaling inhibitor) treatment for 7 days. (b) Representative images of wound healing assay performed with DU145 cells treated with SB203580. (c) Quantitation of cell migration in b (n=5; ***P<0.001; error bars indicate s.e.m.). (d) qRT–PCR analyses for various markers in SB203580-treated DU145 cells. Y-axis represents fold change in HPRT-normalized mRNA expression in SB203580-treated cells compared with vehicle-treated cells, relative to day 0 (n=3; error bars indicate s.e.m.). (e) Immunoblotting for FOXC2, key EMT markers and p38 signaling components in DU145 cells on SB203580 treatment for 7 days. (f) Representative FACS plots for CD44 (APC) and CD24 (PE) surface marker expression in DU145 cells treated with vehicle or SB203580 for 7 days. (g). Quantification of FACS analysis shown in f (n=3; error bars indicate s.e.m.; ***P<0.001). (h) Quantitation of tumorspheres formed per 1000 DU145 cells (treated either with vehicle or SB203580 for 7 days; n=5, error bars indicate s.e.m., ***P<0.001). (i) IF staining for AR and PSA in DU145 cells treated with vehicle or SB203580 for 7 days (scale bar=100 μm, DAPI pseudo-colored to red). (j and k) Quantification of cell survival using MTS Assay in DU145 cells treated as indicated, with Enzalutamide (j) or Docetaxel (k). Data are represented as absorbance (OD) at 490nm (n=3); *P<0.05, **P<0.01, ***P<0.001.
