title: Continuous inhibition of MAPK signaling promotes the early osteoblastic differentiation and mineralization of the extracellular matrix.
abstract: We screened the small molecule compounds that stimulate osteogenesis by themselves or promote bone morphogenetic protein (BMP)-induced bone formation. We found that a specific inhibitor for MAPK/extracellular signal-regulated kinase kinase (MEK)-1, promoted the early osteoblastic differentiation and mineralization of extracellular matrix (ECM) in C2Cl2 pluripotent mesenchymal cells treated with recombinant human BMP-2 (rhBMP-2) and MC3T3-E1 preosteoblastic cells. ALP activity was synergistically increased by the treatment with a specific MEK-1 inhibitor PD98059 and rhBMP-2 in both cell lines. Twenty-five micromolar PD98059 promoted mineralization of ECM in rhBMP-2-treated C2Cl2 cells and MC3T3-E1 cells. In contrast, PD98059 reduced osteocalcin (OCN) secretion and its transcriptional level in rhBMP-2-treated C2Cl2 cells but increased its secretion and mRNA level in MC3T3-E1 cells. Stable expression of a dominant-negative MEK-1 mutant in C2Cl2 cells represented high ALP activity and low osteocalcin production in the presence of rhBMP-2, while a constitutively active mutant of MEK-1 attenuated both of them. Together, our results indicated that BMP-2-induced mineralization of ECM of pluripotent mesenchymal stem cells and preosteoblastic cells could be controlled by a fine tuning of the MAPK signaling pathway. Further, MEK-1 inhibitors would be useful for the promotion of bone formation, for instance, the treatments for delayed fracture healing or advance of localized osteoporotic change after fracture healing.
mesh_terms: D000818:Animals; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line / Q000187:drug effects / Q000378:metabolism; D004790:Enzyme Induction / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology*; D005799:Genes, Dominant; D006801:Humans; D048369:MAP Kinase Kinase 1; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D008648:Mesoderm / Q000166:cytology; D051379:Mice; D008903:Minerals / Q000378:metabolism*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors* / Q000502:physiology; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors* / Q000502:physiology; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015675:Osteocalcin / Q000096:biosynthesis / Q000187:drug effects / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000187:drug effects*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000502:physiology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000494:pharmacology / Q000502:physiology; D011994:Recombinant Proteins; D014158:Transcription, Genetic / Q000187:drug effects; D016212:Transforming Growth Factor beta; D048051:p38 Mitogen-Activated Protein Kinases
chemical_list: C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; C525720:Bmp2 protein, rat; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D004791:Enzyme Inhibitors; D005419:Flavonoids; D008903:Minerals; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; C499469:recombinant human bone morphogenetic protein-2; D015675:Osteocalcin; D017346:Protein Serine-Threonine Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; C482127:Map2k1 protein, mouse; D020929:Mitogen-Activated Protein Kinase Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one