Figure 3

EMD-induced activation of p38 MAPK in MG-63 cells. (A) MG-63 cells were stimulated with 100 μg/ml EMD protein for the indicated times at 37°C. Cells were harvested, and lysates were resolved in 10% SDS-PAGE and were transferred to a PVDF membrane. Membranes were immunoblotted with anti-phospho-p38 MAPK antibody (top), and were then stripped and immunoblotted with anti- p38 MAPK antibody (bottom). Molecular markers (kDa) are shown in the left column. (B) MG-63 cells were treated for 30 min with SB203580 (5 μM) before stimulation with 100 μg/ml EMD protein for 5 min. Cells were harvested, and lysates were resolved in 10% SDS-PAGE and transferred to a PVDF membrane. The membrane was immunoblotted with anti-phospho-p38 MAPK antibody (top), and was then stripped and immunoblotted with anti-p38 MAPK antibody (bottom). Molecular markers (kDa) are shown in the left column. Quantification of p38 MAPK phosphorylation was performed densitometrically and corrected against the amount of total p38 MAPK protein. Peak heights of each density are depicted as percent of maximum value.