title: Hedgehog signalling does not stimulate cartilage catabolism and is inhibited by Interleukin-1β.
abstract: BACKGROUND
  In osteoarthritis, chondrocytes adopt an abnormal hypertrophic morphology and upregulate the expression of the extracellular matrix-degrading enzymes, MMP-13 and ADAMTS-5. The activation of the hedgehog signalling pathway has been established in osteoarthritis and is thought to influence both of these processes. However, the role of this pathway in the initiation and progression of osteoarthritis is unclear as previous studies have been unable to isolate the effects of hedgehog pathway activation from other pathological processes. In this study we test the hypothesis that hedgehog pathway activation causes cartilage degradation in healthy cartilage and in an in vitro model of inflammatory arthritis.
  
  
  METHODS
  Isolated articular chondrocytes from the bovine metacarpal-phalangeal joint were stimulated for up to 24 hours with the agonist, recombinant Indian hedgehog (r-Ihh). ADAMTS-5 and MMP-13 gene expression was quantified by real-time PCR. In addition, healthy bovine cartilage explants were treated with r-Ihh or the hedgehog antagonist, cyclopamine, and sGAG release into the media was measured over 72 hours. Studies were repeated using chondrocytes and cartilage explants from human knee joint. Finally, studies were conducted to determine the effect of hedgehog pathway activation on matrix catabolism in the presence of the pro-inflammatory cytokine, IL-1β.
  
  
  RESULTS
  Addition of r-Ihh activated hedgehog signalling, confirmed by upregulation of Gli1 and Ptch1 expression, but did not increase ADAMTS-5 or MMP-13 expression in bovine or human chondrocytes. Furthermore, r-Ihh did not induce sGAG release in healthy bovine or human cartilage explants. IL-1β treatment induced sGAG release, but this response was not altered by the stimulation or inhibition of hedgehog signalling. Hedgehog pathway activation was downregulated by IL-1β. Conversely, r-Ihh weakly suppressed IL-1β-induced ADAMTS-5 expression.
  
  
  CONCLUSIONS
  Our results show for the first time that Indian hedgehog does not cause extracellular matrix degradation in healthy ex vivo cartilage or in the presence of IL-1β and that IL-1β downregulates Indian hedgehog induced signalling. Thus, we suggest reported hedgehog induced matrix catabolism in osteoarthritis must be due to its interaction with pathological factors other than IL-1β. Hence, hedgehog signalling and its downstream effects are highly context-dependent.
authors.affiliation: Institute of Bioengineering and School of Engineering and Material Sciences, Queen Mary University of London, Mile End Road, London, E1 4NS, UK. clare.l.thompson@qmul.ac.uk.
authors.forename: Clare L
authors.initials: CL
authors.lastname: Thompson
authors.affiliation: Institute of Bioengineering and School of Engineering and Material Sciences, Queen Mary University of London, Mile End Road, London, E1 4NS, UK. r.patel@smd11.qmul.ac.uk.
authors.forename: Riana
authors.initials: R
authors.lastname: Patel
authors.affiliation: Institute of Bioengineering and School of Engineering and Material Sciences, Queen Mary University of London, Mile End Road, London, E1 4NS, UK. t.a.kelly@qmul.ac.uk.|Department of Biomedical Engineering, Columbia University, New York, NY, USA. t.a.kelly@qmul.ac.uk.
authors.forename: Terri-Ann N
authors.initials: TA
authors.lastname: Kelly
authors.affiliation: Institute of Bioengineering and School of Engineering and Material Sciences, Queen Mary University of London, Mile End Road, London, E1 4NS, UK. angus.wann@kennedy.ox.ac.uk.|Kennedy Institute of Rheumatology, University of Oxford, Oxford, UK. angus.wann@kennedy.ox.ac.uk.
authors.forename: Angus K T
authors.initials: AK
authors.lastname: Wann
authors.affiliation: Department of Biomedical Engineering, Columbia University, New York, NY, USA. cth6@columbia.edu.
authors.forename: Clark T
authors.initials: CT
authors.lastname: Hung
authors.affiliation: Center for Endocrinology, William Harvey Research Institute, School of Medicine and Dentistry, Queen Mary University of London, London, UK. j.p.chapple@qmul.ac.uk.
authors.forename: J Paul
authors.initials: JP
authors.lastname: Chapple
authors.affiliation: Institute of Bioengineering and School of Engineering and Material Sciences, Queen Mary University of London, Mile End Road, London, E1 4NS, UK. m.m.knight@qmul.ac.uk.
authors.forename: Martin M
authors.initials: MM
authors.lastname: Knight
mesh_terms: D000328:Adult; D000818:Animals; D002358:Cartilage, Articular / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism; D005260:Female; D053823:Hedgehog Proteins / Q000378:metabolism*; D006801:Humans; D053583:Interleukin-1beta / Q000378:metabolism*; D008297:Male; D008660:Metabolism; D008875:Middle Aged; D009924:Organ Culture Techniques; D010003:Osteoarthritis / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D053823:Hedgehog Proteins; D053583:Interleukin-1beta