title: Harmine promotes osteoblast differentiation through bone morphogenetic protein signaling.
abstract: Bone mass is regulated by osteoblast-mediated bone formation and osteoclast-mediated bone resorption. We previously reported that harmine, a β-carboline alkaloid, inhibits osteoclast differentiation and bone resorption in vitro and in vivo. In this study, we investigated the effects of harmine on osteoblast proliferation, differentiation and mineralization. Harmine promoted alkaline phosphatase (ALP) activity in MC3T3-E1 cells without affecting their proliferation. Harmine also increased the mRNA expressions of the osteoblast marker genes ALP and Osteocalcin. Furthermore, the mineralization of MC3T3-E1 cells was enhanced by treatment with harmine. Harmine also induced osteoblast differentiation in primary calvarial osteoblasts and mesenchymal stem cell line C3H10T1/2 cells. Structure-activity relationship studies using harmine-related β-carboline alkaloids revealed that the C3-C4 double bond and 7-hydroxy or 7-methoxy group of harmine were important for its osteogenic activity. The bone morphogenetic protein (BMP) antagonist noggin and its receptor kinase inhibitors dorsomorphin and LDN-193189 attenuated harmine-promoted ALP activity. In addition, harmine increased the mRNA expressions of Bmp-2, Bmp-4, Bmp-6, Bmp-7 and its target gene Id1. Harmine also enhanced the mRNA expressions of Runx2 and Osterix, which are key transcription factors in osteoblast differentiation. Furthermore, BMP-responsive and Runx2-responsive reporters were activated by harmine treatment. Taken together, these results indicate that harmine enhances osteoblast differentiation probably by inducing the expressions of BMPs and activating BMP and Runx2 pathways. Our findings suggest that harmine has bone anabolic effects and may be useful for the treatment of bone-decreasing diseases and bone regeneration as a lead compound.
authors.affiliation: Department of Nutriproteomics, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Yonezawa
authors.forename: Ji-Won
authors.initials: JW
authors.lastname: Lee
authors.forename: Ayaka
authors.initials: A
authors.lastname: Hibino
authors.forename: Midori
authors.initials: M
authors.lastname: Asai
authors.forename: Hironori
authors.initials: H
authors.lastname: Hojo
authors.forename: Byung-Yoon
authors.initials: BY
authors.lastname: Cha
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Teruya
authors.forename: Kazuo
authors.initials: K
authors.lastname: Nagai
authors.forename: Ung-Il
authors.initials: UI
authors.lastname: Chung
authors.forename: Kazumi
authors.initials: K
authors.lastname: Yagasaki
authors.forename: Je-Tae
authors.initials: JT
authors.lastname: Woo
mesh_terms: D000818:Animals; D019485:Bone Morphogenetic Proteins / Q000378:metabolism* / Q000494:pharmacology; D002243:Carbolines / Q000737:chemistry / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects* / Q000235:genetics; D002460:Cell Line; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects; D006247:Harmine / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D051798:Inhibitor of Differentiation Protein 1 / Q000235:genetics; D051379:Mice; D008815:Mice, Inbred Strains; D010006:Osteoblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects* / Q000235:genetics; D015398:Signal Transduction; D012886:Skull / Q000166:cytology; D000076182:Sp7 Transcription Factor; D013329:Structure-Activity Relationship; D014157:Transcription Factors / Q000235:genetics
chemical_list: D019485:Bone Morphogenetic Proteins; D002243:Carbolines; D050659:Core Binding Factor Alpha 1 Subunit; C497455:Idb1 protein, mouse; D051798:Inhibitor of Differentiation Protein 1; C493716:Runx2 protein, mouse; D000076182:Sp7 Transcription Factor; C479466:Sp7 protein, mouse; D014157:Transcription Factors; D006247:Harmine