title: Aberrant hypertrophy in Smad3-deficient murine chondrocytes is rescued by restoring transforming growth factor beta-activated kinase 1/activating transcription factor 2 signaling: a potential clinical implication for osteoarthritis.
abstract: OBJECTIVE
  To investigate the biologic significance of Smad3 in the progression of osteoarthritis (OA), the crosstalk between Smad3 and activating transcription factor 2 (ATF-2) in the transforming growth factor beta (TGFbeta) signaling pathway, and the effects of ATF-2 overexpression and p38 activation in chondrocyte differentiation.
  
  
  METHODS
  Joint disease in Smad3-knockout (Smad3(-/-)) mice was examined by microfocal computed tomography and histologic analysis. Numerous in vitro methods including immunostaining, real-time polymerase chain reaction, Western blotting, an ATF-2 DNA-binding assay, and a p38 kinase activity assay were used to study the various signaling responses and protein interactions underlying the altered chondrocyte phenotype in Smad3(-/-) mice.
  
  
  RESULTS
  In Smad3(-/-) mice, an end-stage OA phenotype gradually developed. TGFbeta-activated kinase 1 (TAK1)/ATF-2 signaling was disrupted in Smad3(-/-) mouse chondrocytes at the level of p38 MAP kinase (MAPK) activation, resulting in reduced ATF-2 phosphorylation and transcriptional activity. Reintroduction of Smad3 into Smad3(-/-) cells restored the normal p38 response to TGFbeta. Phosphorylated p38 formed a complex with Smad3 by binding to a portion of Smad3 containing both the MAD homology 1 and linker domains. Additionally, Smad3 inhibited the dephosphorylation of p38 by MAPK phosphatase 1 (MKP-1). Both ATF-2 overexpression and p38 activation repressed type X collagen expression in wild-type and Smad3(-/-) chondrocytes. P38 was detected in articular cartilage and perichondrium; articular and sternal chondrocytes expressed p38 isoforms alpha, beta, and gamma, but not delta.
  
  
  CONCLUSIONS
  Smad3 is involved in both the onset and progression of OA. Loss of Smad3 abrogates TAK1/ATF-2 signaling, most likely by disrupting the Smad3-phosphorylated p38 complex, thereby promoting p38 dephosphorylation and inactivation by MKP-1. ATF-2 and p38 activation inhibit chondrocyte hypertrophy. Modulation of p38 isoform activity may provide a new therapeutic approach for OA.
authors.affiliation: University of Rochester, Rochester, New York 14642, USA.
authors.forename: Tian-Fang
authors.initials: TF
authors.lastname: Li
authors.forename: Lin
authors.initials: L
authors.lastname: Gao
authors.forename: Tzong-Jen
authors.initials: TJ
authors.lastname: Sheu
authors.forename: Erik R
authors.initials: ER
authors.lastname: Sampson
authors.forename: Lisa M
authors.initials: LM
authors.lastname: Flick
authors.forename: Yrjö T
authors.initials: YT
authors.lastname: Konttinen
authors.forename: Di
authors.initials: D
authors.lastname: Chen
authors.forename: Edward M
authors.initials: EM
authors.lastname: Schwarz
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Zuscik
authors.forename: Jennifer H
authors.initials: JH
authors.lastname: Jonason
authors.forename: Regis J
authors.initials: RJ
authors.lastname: O'Keefe
mesh_terms: D051698:Activating Transcription Factor 2 / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D018450:Disease Progression; D007150:Immunohistochemistry; D051379:Mice; D018345:Mice, Knockout; D010003:Osteoarthritis / Q000378:metabolism* / Q000473:pathology; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism*; D053773:Transforming Growth Factor beta1 / Q000378:metabolism* / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D051698:Activating Transcription Factor 2; D051900:Smad3 Protein; D053773:Transforming Growth Factor beta1; D048051:p38 Mitogen-Activated Protein Kinases