Figure 3. Isothiocyanates inhibit phosphorylation of ERK, Akt, and FAK in U2OS cells.

(A and C) U2OS cells were incubated with isothiocyanates and TPA for 24 h, and expression of the indicated proteins was determined by western blotting. (B) U2OS cells were treated with PD98059 or wortmannin for 24 h. Expression of MMP-9 and total and phosphorylated forms of ERK1/2 and Akt were measured by western blotting. β-actin is shown as a control. (D) Immunofluorescence microscopy analysis of changes in FAK phosphorylation after isothiocyanate treatment. U2OS cells were co-stained with an anti-p-FAK antibody and Hoechst 33342 and observed under a fluorescence microscope. Data are representative of three independent experiments.