Fig. 5

Effects of MS-275 on FN-1 expression and extracellular secretion (A) RT-qPCR experiment showing FN-1 expression from total RNA of mesenchymal-like MeT5A cells treated with MS-275 (250 nM) for 72 hours. Bars represent means±SEM of 5 experiments (B-C) Adhesion assay of GFP-SKOV3/MeT5A cells (B) and GFP-OVCAR-3/MeT5A cells (C) treated with an anti-FN-1 blocking antibody. Results are shown as relative number of adherent SKOV3 cells. 3 fields for each sample were analyzed. This experiment was performed 3 times. D Representative Western blot showing expression FN-1 from cell lysates of mesenchymal-like MeT5A cells treated as above or from cell supernatant. HSP90 and anti-anti trypsin were used as a loading control. One experiment is shown of 3 performed. Quantifications are shown in the right. E Immunofluorescence of mesenchymal-like MeT5A cells treated with MS-275. Cells were fixed, permeabilized and stained with an anti-FN-1 antibody. Nuclei are stained with DAPI (blue). Confocal images are shown from one representative experiment of four performed. Scale bar: 10 μm, Quantifications of FN-1 perinuclear proportion and fiberness are shown on the right of the figure. Differences were considered significant at P < 0.05 (*p < 0.05; **p < 0.01; ***p < 0.001; **** p < 0.0001). F, G FN-1 staining of MeT5A cells fixed and permeabilized (F) or decellularized matrices (G) after treatment with MS-275 (250 nM) for 72 h. Nuclei are stained with DAPI. Decellularized matrices are shown on the right. Representative images are shown from one of three experiments performed