Phosphate stimulates matrix Gla protein expression in chondrocytes through the extracellular signal regulated kinase signaling pathway

Whereas increasing evidences suggest that inorganic phosphate (Pi) may act as a signaling molecule in mineralization-competent cells, its mechanisms of action remain largely unknown. The aims of the present work were to determine whether Pi regulates expression of matrix Gla protein (MGP), a mineralization inhibitor, in growth plate chondrocytes and to identify the involved signaling pathways. Chondrogenic ATDC5 cells and primary growth plate chondrocytes were used. Messenger RNA analysis was performed by RT-PCR and real-time quantitative PCR. Activation and role of mitogen-activated protein kinases (MAPK) were respectively determined by Western blotting and the use of specific inhibitors. Immunohistological detection of extracellular signal-regulated kinase 1 and 2 (ERK1/2) was performed in rib organ cultures from newborn mice. Results indicated that Pi markedly stimulated expression of MGP in ATDC5 cells and primary growth plate chondrocytes. Investigation of the involved intracellular signaling pathways revealed that Pi activated ERK1/2. The activation of ERK1/2 appeared cell-specific. Indeed, although Pi stimulated ERK1/2 in MC3T3-E1 osteoblasts and ST2 stromal cells, ERK1/2 phosphorylation could not be detected in L929 fibroblasts or C2C12 myogenic cells. Accordingly, immunohistological detection of ERK1/2 phosphorylation in rib growth plates revealed a marked signal in chondrocytes. Finally, a specific ERK1/2 inhibitor, UO126, blocked Pi-stimulated MGP expression in ATDC5 cells, indicating that ERK1/2 mediates, at least in part, the effects of Pi. These data demonstrate for the first time that Pi regulates MGP expression in growth plate chondrocytes, thereby suggesting a key role for Pi and ERK1/2 in the regulation of bone formation.