title: Characterization of Notch Signaling During Osteogenic Differentiation in Human Osteosarcoma Cell Line MG63.
abstract: Osteogenic differentiation is a multi-step process controlled by a complex molecular framework. Notch is an evolutionarily conserved intercellular signaling pathway playing a prominent role in cell fate and differentiation, although the mechanisms by which this pathway regulates osteogenesis remain controversial. This study aimed to investigate, in vitro, the involvement of Notch pathway during all the developmental stages of osteogenic differentiation in human osteosarcoma cell line MG63. Cells were cultured in basal condition (control) and in osteoinductive medium (OM). Notch inhibitors were also added in OM to block Notch pathway. During osteogenic differentiation, early (alkaline phosphatase activity and collagen type I) and late osteogenic markers (osteocalcin levels and matrix mineralization), as well as the gene expression of the main osteogenic transcription factors (Runx2, Osterix, and Dlx5) increased. Time dependent changes in the expression of specific Notch receptors were identified in OM versus control with a significant reduction in the expression of Notch1 and Notch3 receptors in the early phase of differentiation, and an increase of Notch2 and Notch4 receptors in the late phase. Among Notch nuclear target genes, Hey1 expression was significantly higher in OM than control, while Hes5 expression decreased. Osteogenic markers were reduced and Hey1 was significantly inhibited by Notch inhibitors, suggesting a role for Notch through the canonical pathway. In conclusion, Notch pathway might be involved with a dual role in osteogenesis of MG63, through the activation of Notch2, Notch4, and Hey1, inducing osteoblast differentiation and the depression of Notch1, Notch3, and Hes5, maintaining an undifferentiated status. J. Cell. Physiol. 231: 2652-2663, 2016. © 2016 Wiley Periodicals, Inc.
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.
authors.forename: Alessia
authors.initials: A
authors.lastname: Ongaro
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.
authors.forename: Agnese
authors.initials: A
authors.lastname: Pellati
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.
authors.forename: Leila
authors.initials: L
authors.lastname: Bagheri
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.
authors.forename: Paola
authors.initials: P
authors.lastname: Rizzo
authors.affiliation: Department of Chemistry "Giacomo Ciamician," Alma Mater Studiorum, University of Bologna, Bologna, Italy.
authors.forename: Cristiana
authors.initials: C
authors.lastname: Caliceti
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.
authors.forename: Leo
authors.initials: L
authors.lastname: Massari
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Ferrara, Italy.
authors.forename: Monica
authors.initials: M
authors.lastname: De Mattei
mesh_terms: D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D004151:Dipeptides / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D008024:Ligands; D010012:Osteogenesis / Q000187:drug effects / Q000235:genetics; D012516:Osteosarcoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D051880:Receptors, Notch / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism
chemical_list: D014408:Biomarkers, Tumor; D050659:Core Binding Factor Alpha 1 Subunit; D004151:Dipeptides; D008024:Ligands; C419410:N-(N-(3,5-difluorophenacetyl)alanyl)phenylglycine tert-butyl ester; D051880:Receptors, Notch; D014157:Transcription Factors