title: Inhibition of receptor activator of nuclear factor-κB ligand- or lipopolysaccharide-induced osteoclast formation by conophylline through downregulation of CREB.
abstract: The effect of conophylline (CNP) on the receptor activator of nuclear factor-κB ligand (RANKL) or lipopolysaccharide (LPS)-induced osteoclast formation was studied in vitro using bone marrow-derived macrophages (BMMs) or the mouse macrophage-like cell line RAW 264.7. CNP inhibited RANKL-induced formation of osteoclasts identified as tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells in a culture of BMMs. It also inhibited RANKL- or LPS-induced osteoclast formation in RAW 264.7 cells. CNP lowered the osteoclast maturation markers such as calcitonin receptor, MMP9 and cathepsin K in BMMs, suggesting that CNP would inhibit the process of osteoclast differentiation. CNP inhibited the RANKL-induced expressions of c-Fos and nuclear factor of activated T cells (NFATc1), key transcription factors for osteoclastogenesis. On the other hand, CNP did not inhibit the signaling pathway of NF-κB and mitogen-activated protein kinases (MAPKs) in RANKL-stimulated BMMs. Interestingly, CNP inhibited RANKL-induced CREB activation that can mediate c-Fos and NFATc1. CNP also inhibited RANKL- or LPS-induced CREB, c-Fos and NFATc1 activation in RAW 264.7 cells. We have previously found that CNP directly binds to ADP-ribosylation-like factor-6 interacting protein (ARL6ip), although its role in osteoclastogenesis is not clear. Gene knockdown of ARL6ip by siRNA inhibited RANKL-induced c-Fos expression, suggesting that inactivation of ARL6ip may be involved in an inhibitory effect of CNP. Taken together, CNP was shown to inhibit osteoclast formation possibly via CREB inactivation following a decrease in c-Fos and NFATc1 expression.
authors.affiliation: Department of Microbiology and Immunology, Aichi Medical University School of Medicine, Nagakute, Japan. Electronic address: koide@aichi-med-u.ac.jp.
authors.forename: Naoki
authors.initials: N
authors.lastname: Koide
authors.affiliation: Department of Applied Chemistry, Faculty of Science and Technology, Keio University, Yokohama, Japan.
authors.forename: Yuichiro
authors.initials: Y
authors.lastname: Kondo
authors.affiliation: Department of Microbiology and Immunology, Aichi Medical University School of Medicine, Nagakute, Japan.
authors.forename: Erdenezaya
authors.initials: E
authors.lastname: Odkhuu
authors.affiliation: Department of Microbiology and Immunology, Aichi Medical University School of Medicine, Nagakute, Japan.
authors.forename: Jambalganiin
authors.initials: J
authors.lastname: Ulziisaikhan
authors.affiliation: Department of Molecular Target Medicine Screening, Aichi Medical University School of Medicine, Nagakute, Japan.
authors.forename: Tamami
authors.initials: T
authors.lastname: Ukaji
authors.affiliation: Department of Microbiology and Immunology, Aichi Medical University School of Medicine, Nagakute, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Yokochi
authors.affiliation: Department of Molecular Target Medicine Screening, Aichi Medical University School of Medicine, Nagakute, Japan.
authors.forename: Kazuo
authors.initials: K
authors.lastname: Umezawa
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects / Q000378:metabolism; D002460:Cell Line; D017362:Cyclic AMP Response Element-Binding Protein / Q000235:genetics* / Q000378:metabolism; D015536:Down-Regulation; D005786:Gene Expression Regulation / Q000187:drug effects*; D008070:Lipopolysaccharides / Q000494:pharmacology*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D050778:NFATC Transcription Factors / Q000235:genetics; D010010:Osteoclasts / Q000187:drug effects* / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics; D053245:RANK Ligand / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D014748:Vinca Alkaloids / Q000494:pharmacology*
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D008070:Lipopolysaccharides; D016328:NF-kappa B; D050778:NFATC Transcription Factors; D016760:Proto-Oncogene Proteins c-fos; D053245:RANK Ligand; D014748:Vinca Alkaloids; C099167:conophylline; D020928:Mitogen-Activated Protein Kinases
keywords: ARL6ip; CREB; Conophylline; Lipopolysaccharide; Osteoclast; Receptor activator of nuclear factor-κB ligand