title: Cellular and molecular characterization of a novel primary osteoblast culture from the vertebrate model organism Xenopus tropicalis.
abstract: Osteogenesis is the fundamental process by which bones are formed, maintained and regenerated. The osteoblasts deposit the bone mineralized matrix by secreting large amounts of extracellular proteins and by allowing the biochemical conditions for the nucleation of hydroxyapatite crystals. Normal bone formation requires a tight control of osteoblastic activity, and therefore, osteoblasts represent a major focus of interest in biomedical research. Several crucial features of osteogenesis can be readily recapitulated using murine, avian and fish primary and immortalized osteoblastic cultures. Here, we describe a novel and straightforward in vitro culture of primary osteoblasts from the amphibian Xenopus tropicalis, a major vertebrate model organism. X. tropicalis osteoblasts can readily be extracted from the frontoparietal bone of pre-metamorphosing tadpole skulls by series of gentle protease treatments. Such primary cultures efficiently proliferate and can conveniently be grown at room temperature, in the absence of CO2, on a variety of substrates. X. tropicalis primary osteoblasts express well-characterized genes known to be active during osteogenesis of teleost fish, chick, mouse and human. Upon differentiation, such cultures mineralize and activate DMP1, an osteocyte-specific gene. Importantly, X. tropicalis primary osteoblasts can be efficiently transfected and respond to the forced activation of the bone morphogenetic protein pathway by increasing their nuclear levels of phospho-Smad. Therefore, this novel primary culture is amenable to experimental manipulations and represents a valuable tool for improving our understanding of the complex network of molecular interactions that govern vertebrate bone formation.
authors.affiliation: Department of Cell Biology, Faculty of Biological Sciences, Universidad de Concepción, Casilla, 160-C, Concepción, Chile.
authors.forename: Ariana
authors.initials: A
authors.lastname: Bertin
authors.forename: Patricia
authors.initials: P
authors.lastname: Hanna
authors.forename: Gaston
authors.initials: G
authors.lastname: Otarola
authors.forename: Alan
authors.initials: A
authors.lastname: Fritz
authors.forename: Juan Pablo
authors.initials: JP
authors.lastname: Henriquez
authors.forename: Sylvain
authors.initials: S
authors.lastname: Marcellini
mesh_terms: D000818:Animals; D052005:Bone Morphogenetic Protein Receptors, Type I / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D002469:Cell Separation; D002478:Cells, Cultured; D018507:Gene Expression Regulation, Developmental; D007814:Larva / Q000166:cytology / Q000502:physiology; D010006:Osteoblasts / Q000502:physiology*; D010012:Osteogenesis / Q000235:genetics; D010294:Parietal Bone / Q000196:embryology / Q000502:physiology*; D010766:Phosphorylation; D061251:Primary Cell Culture; D051785:Smad Proteins / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D014981:Xenopus / Q000196:embryology / Q000235:genetics / Q000502:physiology*; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D051785:Smad Proteins; D029867:Xenopus Proteins; D052005:Bone Morphogenetic Protein Receptors, Type I