Osthole-mediated cell differentiation through bone morphogenetic protein-2/p38 and extracellular signal-regulated kinase 1/2 pathway in human osteoblast cells.
The survival of osteoblast cells is one of the determinants of the development of osteoporosis in patients. Osthole (7-methoxy-8-isopentenoxycoumarin) is a coumarin derivative present in many medicinal plants. By means of alkaline phosphatase (ALP) activity, osteocalcin, osteopontin, and type I collagen, enzyme-linked immunosorbent assay, we have shown that osthole exhibits a significant induction of differentiation in two human osteoblast-like cell lines, MG-63 and hFOB. Induction of differentiation by osthole was associated with increased bone morphogenetic protein (BMP)-2 production and the activations of SMAD1/5/8 and p38 and extracellular signal-regulated kinase (ERK) 1/2 kinases. Addition of purified BMP-2 protein did not increase the up-regulation of ALP activity and osteocalcin by osthole, whereas the BMP-2 antagonist noggin blocked both osthole and BMP-2-mediated ALP activity enhancement, indicating that BMP-2 production is required in osthole-mediated osteoblast maturation. Pretreatment of osteoblast cells with noggin abrogated p38 activation but only partially decreased ERK1/2 activation, suggesting that BMP-2 signaling is required in p38 activation and is partially involved in ERK1/2 activation in osthole-treated osteoblast cells. Cotreatment of p38 inhibitor SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole] or p38 small interfering RNA (siRNA) expression inhibited osthole-mediated activation of ALP but only slightly affected osteocalcin production. In contrast, the production of osteocalcin induced by osthole was inhibited by the mitogen-activated protein kinase kinase inhibitor PD98059 (2'-amino-3'-methoxyflavone) or by expression of an ERK2 siRNA. These data suggest that BMP-2/p38 pathway links to the early phase, whereas ERK1/2 pathway is associated with the later phase in osthole-mediated differentiation of osteoblast cells. In this study, we demonstrate that osthole is a promising agent for treating osteoporosis.
Kuo
Po-Lin
PL
Department of Biotechnology, Chia-Nan University of Pharmacy and Science, Tainan, Taiwan. kuopolin@seed.net.tw
Hsu
Ya-Ling
YL
Chang
Cheng-Hsiung
CH
Chang
Jiunn-Kae
JK
D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000502:physiology*; D002454:Cell Differentiation / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003374:Coumarins / Q000494:pharmacology*; D004268:DNA-Binding Proteins / Q000502:physiology; D006801:Humans; D020935:MAP Kinase Signaling System / Q000502:physiology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000502:physiology*; D048052:Mitogen-Activated Protein Kinase 3 / Q000502:physiology*; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects*; D010024:Osteoporosis / Q000188:drug therapy / Q000209:etiology; D034741:RNA, Small Interfering / Q000494:pharmacology; D051785:Smad Proteins; D051898:Smad1 Protein; D015534:Trans-Activators / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000502:physiology*; D048051:p38 Mitogen-Activated Protein Kinases / Q000502:physiology*
C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D003374:Coumarins; D004268:DNA-Binding Proteins; D034741:RNA, Small Interfering; C497399:SMAD1 protein, human; D051785:Smad Proteins; D051898:Smad1 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases; C046627:osthol