Figure 5. Periostin regulates TGF-β-induced U-87 MG cell invasion, cell migration and Smad2, Akt and Fak signaling pathways.

U-87 MG cells were (A) treated with different concentrations of TGF-β for 48 h, or (B-E) transfected with 2 nM periostin siRNA for 24 h then treated with 10 ng/mL TGF-β for 48 h before (B) adhesion onto CIM-Plates coated (or not) with Matrigel or (C, D) adhesion onto CIM-Plates coated with 0.15% gelatin or (E) cell lysis and immunoblotting. Data showing (A, B) TGF-β-induced invasion of U-87 MG cells at 10 h, (C) Real-Time migration of TGF-β-induced U-87 MG cells and (D) TGF-β-induced migration of U-87 MG cells at 10 h. Values are means ± SEM of three independent experiments performed in quadruplicate. Cell invasion represents the percentage of the ratio of cell index of Matrigel-coated wells to cell index of uncoated wells. Cell migration represents the percentage of the normalized cell index of coated wells. (E) Western blot analysis shows levels of phosphorylated proteins. The immunoreactive band intensities were analyzed by densitometry using ImageJ software and expressed in arbitrary units as a ratio of levels of phosphorylated protein to those of the total protein to correct for variation in the amount of loaded proteins. The relative levels of phosphorylated proteins were also normalized to that of TGF-β stimulated controls. Statistically significant differences were calculated by one-way ANOVA followed by Bonferroni’s test (^*P < 0.05, ^**P < 0.01, and ^***P < 0.001 versus stimulated control).