title: Analyses of bone modeling and remodeling using in vitro reconstitution system with two-photon microscopy.
abstract: Bone modeling and remodeling are cellular events during which osteoblast lineage cells and osteoclasts interact. During these events, cells undergo drastic changes with time as they become differentiated. Their morphology, topology, and activity are affected by other cells and the extracellular matrices. Since the mechanisms underlying the cellular events of bone metabolism have not been elucidated, there is a need for systems to analyze these cellular networks and their microenvironments spatiotemporally at the cellular level. Here we report a novel in vitro system for reconstituting the bone cell network of osteoclasts, osteoblasts, and osteocytes in the mineralized nodule, allowing for observation of bone modeling and remodeling phenomena by 2-photon microscopy. Using this system, the change in morphology of osteoblasts from cuboidal to flat cells was observed and measured during the formation of mineralized nodules. Furthermore, the recruitment of osteoblasts to resorption pits and their replenishment by newly formed matrices were successfully observed, providing strong evidence for the coupling of bone resorption and bone formation at cellular level. During such remodeling cycle, flat osteoblasts that survived more than 7 weeks were recruited to resorption pits, where they became cuboidal osteoblasts that express osteocalcin. This novel system permitted the elucidation of cellular behavior during bone modeling and remodeling, and can be used to analyze cellular events involved in bone metabolism.
authors.affiliation: Department of Molecular Medicine for Pathogenesis, Ehime University Graduate School of Medicine, Ehime, Japan; Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Ehime, Japan; Division of Bio-imaging, Proteo-Science Center, Ehime University, Ehime, Japan; Department of Cartilage & Bone Regeneration (Fujisoft), Graduate School of Medicine, The University of Tokyo, Tokyo, Japan. Electronic address: ahikita-tky@umin.ac.jp.
authors.forename: Atsuhiko
authors.initials: A
authors.lastname: Hikita
authors.affiliation: Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Ehime, Japan; Division of Bio-imaging, Proteo-Science Center, Ehime University, Ehime, Japan.
authors.forename: Tadahiro
authors.initials: T
authors.lastname: Iimura
authors.affiliation: Department of Molecular Medicine for Pathogenesis, Ehime University Graduate School of Medicine, Ehime, Japan; Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Ehime, Japan; Division of Bio-imaging, Proteo-Science Center, Ehime University, Ehime, Japan; Translational Research Center, Ehime University Hospital, Ehime, Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Oshima
authors.affiliation: Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Ehime, Japan; Translational Research Center, Ehime University Hospital, Ehime, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Saitou
authors.affiliation: Department of Molecular Medicine for Pathogenesis, Ehime University Graduate School of Medicine, Ehime, Japan; Department of Gastroenterology and Metabiology, Ehime University, Ehime, Japan.
authors.forename: Shin
authors.initials: S
authors.lastname: Yamamoto
authors.affiliation: Department of Molecular Medicine for Pathogenesis, Ehime University Graduate School of Medicine, Ehime, Japan; Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Ehime, Japan; Division of Bio-imaging, Proteo-Science Center, Ehime University, Ehime, Japan; Translational Research Center, Ehime University Hospital, Ehime, Japan. Electronic address: timamura-ind@umin.ac.jp.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Imamura
mesh_terms: D000818:Animals; D015415:Biomarkers; D016723:Bone Remodeling; D002454:Cell Differentiation; D002478:Cells, Cultured; D066298:In Vitro Techniques; D051379:Mice; D008810:Mice, Inbred C57BL; D008853:Microscopy / Q000379:methods*; D010006:Osteoblasts / Q000166:cytology; D010011:Osteocytes / Q000166:cytology
chemical_list: D015415:Biomarkers
keywords: Bone modeling; Bone remodeling; Osteoblasts; Osteoclasts; Osteocytes; Two-photon microscopy