FIGURE 1.

Erk activation is involved in BMP-2-stimulated osteoblast differentiation. A, BMP-2 induced Erk activation. C2C12 cells were serum-starved for 16 h and treated with BMP-2 for the indicated times. Whole cell lysates were prepared and subjected to immunoblot analyses. B–D, Erk inhibition by U0126 suppressed BMP-2-induced ALP and OC expression. C2C12 cells were treated with BMP-2 and/or U0126 (40 μm) for 2 days and then ALP histochemical staining (B) and semiquantitative RT-PCR (C and D, upper panels) or real-time PCR (C and D, lower panel) were performed. Data represent the mean ± S.E. (n = 4). *, p < 0.01. E and F, Erk signaling increased Runx2 transcriptional activity. C2C12 cells were transfected with pGL3–6XOSE2-Luc and FLAG-Runx2 or a constitutively active MEK1 expression vector. After overnight recovery from transfection, cells were incubated in the presence or absence of U0126 for an additional 24 h. Data are shown as activity relative to Renilla luciferase activity and represent the mean ± S.E. (n = 6). * p < 0.01, compared with control; #, p < 0.01, compared with FLAG-Runx2 overexpressed group.