title: Identification of novel osteogenic compounds by an ex-vivo sp7:luciferase zebrafish scale assay.
abstract: Tight interactions among different cell types contributing to bone formation are of key importance in the maintenance of bone homeostasis. Based on the high similarity in responses to (anti)osteogenic signals between zebrafish scales and mammalian bone, we developed and validated a model to screen large numbers of compounds using ex-vivo cultured scales of a sp7:luciferase transgenic zebrafish. This model combines the high predictive value of explant cultures with quick, sensitive, and quantifiable readout converging the effects via various pathways including WNT-signaling, to SP7/osterix promoter activity. Sp7 is pivotal in osteoblast differentiation and activity and its promoter activity provides an excellent surrogate for sp7 expression. Bmp-2a was shown to dose-dependently increase sp7-driven luciferase activity ex vivo. Next, we identified novel effects on bone for 51.7% of the compounds from a small library of WNT-signaling modulators, including a strong osteogenic effect for niclosamide. From all previously characterized compounds, the effect on bone was correctly predicted for 70% of compounds, resulting in a 7% false positive- and 21% false negative rate. The proposed sp7:luciferase zebrafish scale model is unique, powerful and efficient new tool to assess compounds with osteogenic effects, prior to further testing in rodents.
authors.affiliation: Department of Organismal Animal Physiology, Institute for Water and Wetland Research, Faculty of Science, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ, Nijmegen, The Netherlands. Electronic address: e.devrieze@science.ru.nl.
authors.forename: Erik
authors.initials: E
authors.lastname: de Vrieze
authors.affiliation: Department of Organismal Animal Physiology, Institute for Water and Wetland Research, Faculty of Science, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ, Nijmegen, The Netherlands.
authors.forename: Jan
authors.initials: J
authors.lastname: Zethof
authors.affiliation: Hubrecht Institute-KNAW & UMC Utrecht, Utrecht 3584 CT, The Netherlands; Institute for Cardiovascular Organogenesis and Regeneration, Faculty of Medicine, WWU Münster, 48149 Münster, Germany; Cells-in-Motion Cluster of Excellence (EXC 1003 - CiM), University of Münster, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Schulte-Merker
authors.affiliation: Department of Organismal Animal Physiology, Institute for Water and Wetland Research, Faculty of Science, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ, Nijmegen, The Netherlands.
authors.forename: Gert
authors.initials: G
authors.lastname: Flik
authors.affiliation: Department of Organismal Animal Physiology, Institute for Water and Wetland Research, Faculty of Science, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ, Nijmegen, The Netherlands.
authors.forename: Juriaan R
authors.initials: JR
authors.lastname: Metz
mesh_terms: D000825:Animal Structures / Q000378:metabolism*; D000818:Animals; D030801:Animals, Genetically Modified; D001681:Biological Assay / Q000379:methods*; D055396:Bone Morphogenetic Protein 2; D008156:Luciferases / Q000378:metabolism*; D010006:Osteoblasts / Q000378:metabolism; D010012:Osteogenesis; D011401:Promoter Regions, Genetic / Q000235:genetics; D012038:Regeneration; D015398:Signal Transduction; D000076182:Sp7 Transcription Factor; D014157:Transcription Factors / Q000235:genetics; D051153:Wnt Proteins / Q000378:metabolism; D015027:Zebrafish / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000235:genetics
chemical_list: D055396:Bone Morphogenetic Protein 2; D000076182:Sp7 Transcription Factor; D014157:Transcription Factors; D051153:Wnt Proteins; D029961:Zebrafish Proteins; C541742:sp7 protein, zebrafish; D008156:Luciferases
keywords: Bone; Drug design; Elasmoid scales; Osteoblast; Sp7; Zebrafish