Induction of Mitogen‐Activated Protein Kinase Phosphatase‐1 and Down‐Regulation of p38 and ERK1/2 Phosphorylation in PTHrP Stimulated Differentiated Osteoblasts

Parathyroid hormone (PTH) and PTH related protein (PTHrP) have been implicated in the control of bone cell turnover through complex signaling mechanisms. Mitogen‐activated protein (MAP) kinases play a critical role in this process. In order to understand the signaling pathways, this study examined the role of MAP kinase phosphatase‐1 (MKP‐1) in regulating MAPKs, cyclin D1 and osteoblast cell growth arrest using mouse MC3T3‐E1 osteoblastic cells. Analysis of MAPK activity revealed down‐regulation of cyclin D1 (5–10 fold) and p38 and ERK1/2 phosphorylation after PTHrP stimulation. Interestingly, MKP‐1 was induced concurrently with the attenuation of p38 and ERK1/2 phosphorylation. Real‐time PCR and western blot analysis revealed 4‐ 8 fold increase in MKP‐1 expression following 5 min to 2h PTHrP stimulation compared to vehicle controls in differentiated osteoblasts. Forskolin, a cAMP agonist, mimicked PTHrP action, and the PKC inhibitor, GF109203, partially blocked induction of MKP‐1, implying involvement of both cAMP and PKC. MKP‐1 expression was also induced in differentiated primary bone marrow stromal cells (BMSCs). Microarray analysis of in vivo ectopic ossicles (implanted BMSCs) generated in mice, demonstrated down‐regulation of MKP‐1 in less differentiated ossicles following acute injection of PTH. These findings highlight MKP‐1 as a critical regulator of the PTH and PTHrP response in osteoblasts and may be an important target gene in the anabolic action of PTH in bone. This work was supported by NIH grants DE016865, DE14073, and DK53904.