Effects of glucocorticoids and interleukin-1 beta on expression and activity of aggrecanases in equine chondrocytes

Evita Busschers, Jeff P Holt, Dean W Richardson

Research output: Contribution to journalArticle (Academic Journal)peer-review

9 Citations (Scopus)

Abstract

OBJECTIVE: To determine effects of interleukin (IL)-1 beta and glucocorticoids on total glycosaminoglycan (GAG) loss and aggrecanase-mediated matrix degradation in equine cartilage.

SAMPLE POPULATION: Cartilage from 24 equine cadavers free of sepsis and musculoskeletal disease.

PROCEDURES: Effects of IL-1 beta, IL-1 beta with glucocorticoids (dexamethasone and triamcinolone, 10(-6) and 10(-7)M), and glucocorticoids alone on degradation of equine articular and nasal cartilage explants were assessed by measuring GAG release in media and GAG content in cartilage. Aggrecanase-mediated cleavage within the interglobular domain at Glu373-Ala374 was evaluated via western blot analysis and ELISAs. Steady-state mRNA concentrations of matrix metalloproteinase (MMP)-3, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)4, and ADAMTS5 were assessed by use of real-time reverse transcriptase PCR assay (cartilage explants) and northern blot analysis (cell culture).

RESULTS: IL-1 beta increased GAG release and aggrecanase activity (11-fold). The MMP-3, MMP-13, and ADAMTS4 mRNA were upregulated with IL-1 beta, whereas ADAMTS5 mRNA was increased (13-fold), but significantly less than ADAMTS4 mRNA (27-fold), suggesting a role for both ADAMTS4 and ADAMTS5 in degradation of cytokine-stimulated cartilage. Despite downregulation of MMP-3 and MMP-13 mRNA, glucocorticoids did not alter GAG degradation. A further increase in aggrecanase activity was detected with ELISAs and western blot analysis, whereas ADAMTS4 mRNA was downregulated and ADAMTS5 mRNA was maintained or upregulated.

CONCLUSIONS AND CLINICAL RELEVANCE: MMP-3, MMP-13, and ADAMTS4 were regulated differently than ADAMTS5. Glucocorticoids increased aggrecanase activity despite down-regulation of ADAMTS4 mRNA, suggesting a major role of ADAMTS5. Effects of glucocorticoids on aggrecanase activity have important implications in terms of treatment.

Original languageEnglish
Pages (from-to)176-85
Number of pages10
JournalAmerican Journal of Veterinary Research
Volume71
Issue number2
DOIs
Publication statusPublished - Feb 2010

Keywords

  • ADAM Proteins
  • Animals
  • Cartilage
  • Cells, Cultured
  • Chondrocytes
  • Dexamethasone
  • Gene Expression Regulation, Enzymologic
  • Glucocorticoids
  • Horses
  • Interleukin-1beta
  • Metalloproteases
  • Tissue Culture Techniques
  • Triamcinolone
  • Journal Article
  • Research Support, Non-U.S. Gov't

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