Fig. 3: p38 MAPK regulates laminin-332 expression in RT3 cells.

See also Supplementary Fig. S3. A RT3 cells were treated in 2D condition with p38 signaling inhibitors SB203580 (10 μM) or BIRB796 (10 μM) for 24 h, followed by 3D spheroid formation with skin primary fibroblasts. The spheroids were allowed to grow for 3 days before harvesting for western blotting. Laminin-332 level was analyzed by western blotting and β-actin was used as a loading control. Representative images from three independent biological replicates are shown. B RT3 cells were treated with 10 μM PLX8394 for 24 h in 2D condition, followed by spheroid formation with skin primary fibroblasts. The spheroids were grown for 3 days. The level of phosphorylated p38 (p-p38) was analyzed by western blotting and β-actin was used as a loading control. Representative images from three independent biological replicates are shown. C RT3 cells were treated with 10 μM PLX8394 for 24 h in 2D condition, followed by spheroid formation with skin primary fibroblasts. The spheroids were grown for 1–5 days. The level of p-CREB was analyzed by western blotting. The graph shows relative p-CREB expression to β-actin from three independent biological replicates ± S.E.M. *<0.05; paired t-test. D RT3 cells were treated in 2D condition with 10 μM PLX8394 for 24 h, followed by treatments with p38 signaling activators TNF-α (10 ng/ml, 30 min), IL-1β (10 ng/ml, 30 min) and sorbitol (400 mM, 2 h). The level of p-CREB (arrow) was analyzed by western blotting and β-actin was used as a loading control. Densitometric quantitation of p-CREB levels corrected for β-actin is shown below the blots. Values are relative to the levels (1.0) of control samples (without PLX8394) of each activator.