title: Notch signaling through Jagged-1 is necessary to initiate chondrogenesis in human bone marrow stromal cells but must be switched off to complete chondrogenesis.
abstract: We investigated Notch signaling during chondrogenesis in human bone marrow stromal cells (hMSC) in three-dimensional cell aggregate culture. Expression analysis of Notch pathway genes in 14-day chondrogenic cultures showed that the Notch ligand Jagged-1 (Jag-1) sharply increased in expression, peaking at day 2, and then declined. A Notch target gene, HEY-1, was also expressed, with a temporal profile that closely followed the expression of Jag-1, and this preceded the rise in type II collagen expression that characterized chondrogenesis. We demonstrated that the shut-down in Notch signaling was critical for full chondrogenesis, as adenoviral human Jag-1 transduction of hMSC, which caused continuous elevated expression of Jag-1 and sustained Notch signaling over 14 days, completely blocked chondrogenesis. In these cultures, there was inhibited production of extracellular matrix, and the gene expression of aggrecan and type II collagen were strongly suppressed; this may reflect the retention of a prechondrogenic state. The JAG-1-mediated Notch signaling was also shown to be necessary for chondrogenesis, as N-[N-(3,5-difluorophenacetyl-L-alanyl)]-(S)-phenylglycine t-butyl ester (DAPT) added to cultures on days 0-14 or just days 0-5 inhibited chondrogenesis, but DAPT added from day 5 did not. The results thus showed that Jag-1-mediated Notch signaling in hMSC was necessary to initiate chondrogenesis, but it must be switched off for chondrogenesis to proceed.
authors.affiliation: UK Centre for Tissue Engineering, Faculty of Life Sciences, University of Manchester, Oxford Road, Manchester, M13 9PT, United Kingdom.
authors.forename: Rachel A
authors.initials: RA
authors.lastname: Oldershaw
authors.forename: Simon R
authors.initials: SR
authors.lastname: Tew
authors.forename: Amanda M
authors.initials: AM
authors.lastname: Russell
authors.forename: Kate
authors.initials: K
authors.lastname: Meade
authors.forename: Robert
authors.initials: R
authors.lastname: Hawkins
authors.forename: Tristan R
authors.initials: TR
authors.lastname: McKay
authors.forename: Keith R
authors.initials: KR
authors.lastname: Brennan
authors.forename: Timothy E
authors.initials: TE
authors.lastname: Hardingham
mesh_terms: D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002449:Cell Aggregation / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000187:drug effects; D004151:Dipeptides / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D000072100:Jagged-1 Protein; D008024:Ligands; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D051880:Receptors, Notch / Q000235:genetics / Q000378:metabolism*; D000072099:Serrate-Jagged Proteins; D015398:Signal Transduction / Q000187:drug effects; D017154:Stromal Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014161:Transduction, Genetic
chemical_list: D002135:Calcium-Binding Proteins; D004151:Dipeptides; D036341:Intercellular Signaling Peptides and Proteins; C000606273:JAG1 protein, human; D000072100:Jagged-1 Protein; D008024:Ligands; D008565:Membrane Proteins; C419410:N-(N-(3,5-difluorophenacetyl)alanyl)phenylglycine tert-butyl ester; D051880:Receptors, Notch; D000072099:Serrate-Jagged Proteins