title: Biomechanical modulation of collagen fragment-induced anabolic and catabolic activities in chondrocyte/agarose constructs.
abstract: BACKGROUND
  The present study examined the effect of collagen fragments on anabolic and catabolic activities by chondrocyte/agarose constructs subjected to dynamic compression.
  
  
  METHODS
  Constructs were cultured under free-swelling conditions or subjected to continuous and intermittent compression regimes, in the presence of the N-terminal (NT) and C-terminal (CT) telopeptides derived from collagen type II and/or 1400 W (inhibits inducible nitric oxide synthase (iNOS)). The anabolic and catabolic activities were compared to the amino-terminal fibronectin fragment (NH2-FN-f) and assessed as follows: nitric oxide (NO) release and sulphated glycosaminoglycan (sGAG) content were quantified using biochemical assays. Tumour necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta) release were measured by ELISA. Gene expression of matrix metalloproteinase-3 (MMP-3), matrix metalloproteinase-13 (MMP-13), collagen type II and fibronectin were assessed by real-time quantitative polymerase chain reaction (qPCR). Two-way ANOVA and the post hoc Bonferroni-corrected t-test was used to examine data.
  
  
  RESULTS
  The presence of the NT or CT peptides caused a moderate to strong dose-dependent stimulation of NO, TNFalpha and IL-1beta production and inhibition of sGAG content. In some instances, high concentrations of telopeptides were just as potent in stimulating catabolic activities when compared to NH2-FN-f. Depending on the concentration and type of fragment, the increased levels of NO and cytokines were inhibited with 1400 W, resulting in the restoration of sGAG content. Depending on the duration and type of compression regime employed, stimulation with compression or incubation with 1400 W or a combination of both, inhibited telopeptide or NH2-FN-f induced NO release and cytokine production and enhanced sGAG content. All fragments induced MMP-3 and MMP-13 expression in a time-dependent manner. This effect was reversed with compression and/or 1400 W resulting in the restoration of sGAG content and induction of collagen type II and fibronectin expression.
  
  
  CONCLUSIONS
  Collagen fragments containing the N- and C-terminal telopeptides have dose-dependent catabolic activities similar to fibronectin fragments and increase the production of NO, cytokines and MMPs. Catabolic activities were downregulated by dynamic compression or by the presence of the iNOS inhibitor, linking reparative activities by both types of stimuli. Future investigations which examine the signalling cascades of chondrocytes in response to matrix fragments with mechanical influences may provide useful information for early osteoarthritis treatments.
authors.affiliation: School of Engineering and Materials Science, Queen Mary University of London, Mile End Road, London, E1 4NS, UK. t.t.chowdhury@qmul.ac.uk
authors.forename: Tina T
authors.initials: TT
authors.lastname: Chowdhury
authors.forename: Ronny M
authors.initials: RM
authors.lastname: Schulz
authors.forename: Sonpreet S
authors.initials: SS
authors.lastname: Rai
authors.forename: Christian B
authors.initials: CB
authors.lastname: Thuemmler
authors.forename: Nico
authors.initials: N
authors.lastname: Wuestneck
authors.forename: Augustinus
authors.initials: A
authors.lastname: Bader
authors.forename: Gene A
authors.initials: GA
authors.lastname: Homandberg
mesh_terms: D000818:Animals; D001696:Biomechanical Phenomena; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D024042:Collagen Type I / Q000494:pharmacology*; D024043:Collagen Type II / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005353:Fibronectins / Q000378:metabolism; D006025:Glycosaminoglycans / Q000378:metabolism; D053583:Interleukin-1beta / Q000378:metabolism; D053509:Matrix Metalloproteinase 13 / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000378:metabolism; D008660:Metabolism / Q000187:drug effects; D009569:Nitric Oxide / Q000378:metabolism; D010446:Peptide Fragments / Q000494:pharmacology*; D010455:Peptides / Q000494:pharmacology*; D012685:Sepharose; D013552:Swine; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D024042:Collagen Type I; D024043:Collagen Type II; D005353:Fibronectins; D006025:Glycosaminoglycans; D053583:Interleukin-1beta; D010446:Peptide Fragments; D010455:Peptides; D014409:Tumor Necrosis Factor-alpha; C052929:collagen type I trimeric cross-linked peptide; D009569:Nitric Oxide; D012685:Sepharose; D053509:Matrix Metalloproteinase 13; D019278:Matrix Metalloproteinase 3
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