Matrix metalloproteinase 13 loss associated with impaired extracellular matrix remodeling disrupts chondrocyte differentiation by concerted effects on multiple regulatory factors

Rosa Maria Borzí, Eleonora Olivotto, Stefania Pagani, Roberta Vitellozzi, Simona Neri, Michela Battistelli, Elisabetta Falcieri, Annalisa Facchini, Flavio Flamigni, Marianna Penzo, Daniela Platano, Spartaco Santi, Andrea Facchini, Kenneth B. Marcu

Research output: Contribution to journalArticlepeer-review

Abstract

Objective. To link matrix metalloproteinase 13 (MMP-13) activity and extracellular matrix (ECM) remodeling to alterations in regulatory factors leading to a disruption in chondrocyte homeostasis. Methods. MMP-13 expression was ablated in primary human chondrocytes by stable retrotransduction of short hairpin RNA. The effects of MMP-13 knockdown on key regulators of chondrocyte differentiation (SOX9, runt-related transcription factor 2 [RUNX-2], and β-catenin) and angiogenesis (vascular endothelial growth factor [VEGF]) were scored at the protein level (by immunohistochemical or Western blot analysis) and RNA level (by real-time polymerase chain reaction) in high-density monolayer and micromass cultures under mineralizing conditions. Effects on cellular viability in conjunction with chondrocyte progression toward a hypertrophic-like state were assessed in micromass cultures. Alterations in SOX9 subcellular distribution were assessed using confocal microscopy in micromass cultures and also in osteoarthritic cartilage. Results. Differentiation of control chondrocyte micromasses progressed up to a terminal phase, with calcium deposition in conjunction with reduced cell viability and scant ECM. MMP-13 knockdown impaired ECM remodeling and suppressed differentiation in conjunction with reduced levels of RUNX-2, β-catenin, and VEGF. MMP-13 levels in vitro and ECM remodeling in vitro and in vivo were linked to changes in SOX9 subcellular localization. SOX9 was largely excluded from the nuclei of chondrocytes with MMP-13-remodeled or -degraded ECM, and exhibited an intranuclear staining pattern in chondrocytes with impaired MMP-13 activity in vitro or with more intact ECM in vivo. Conclusion. MMP-13 loss leads to a breakdown in primary human articular chondrocyte differentiation by altering the expression of multiple regulatory factors.

Original languageEnglish
Pages (from-to)2370-2381
Number of pages12
JournalArthritis and Rheumatism
Volume62
Issue number8
DOIs
Publication statusPublished - Aug 2010

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy
  • Rheumatology
  • Pharmacology (medical)

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