Fibronectin fragment-induced expression of matrix metalloproteinases is mediated by MyD88-dependent TLR-2 signaling pathway in human chondrocytes

Arthritis Res Ther. 2015 Nov 12:17:320. doi: 10.1186/s13075-015-0833-9.

Abstract

Introduction: Fibronectin fragments (FN-fs) are increased in the cartilage of patients with osteoarthritis (OA) and have a potent chondrolytic effect. However, little is known about the cellular receptors and signaling mechanisms that are mediated by FN-fs. We investigated whether the 29-kDa amino-terminal fibronectin fragment (29-kDa FN-f) regulates cartilage catabolism via the Toll-like receptor (TLR)-2 signaling pathway in human chondrocytes.

Methods: Small interfering RNA was used to knock down TLR-2 and myeloid differentiation factor 88 (MyD88). TLR-2 was overexpressed in chondrocytes transfected with a TLR-2 expression plasmid. The expression levels of matrix metalloproteinase (MMP)-1, MMP-3, and MMP-13 were analyzed using quantitative real-time reverse transcription polymerase chain reactions, immunoblotting, or enzyme-linked immunosorbent assay. The effect of TLR-2 on 29-kDa FN-f-mediated signaling pathways was investigated by immunoblotting.

Results: TLR-2, TLR-3, TLR-4, and TLR-5 mRNA were significantly overexpressed in OA cartilage compared with normal cartilage, whereas no significant difference of TLR-1 mRNA expression was found. 29-kDa FN-f significantly increased TLR-2 expression in human chondrocytes in a dose- and time-dependent manner. Knockdown of TLR-2 or MyD88, the latter a downstream adaptor of TLR-2, significantly inhibited 29-kDa FN-f-induced MMP production at the mRNA and protein levels. Conversely, TLR-2 overexpression led to enhanced MMP production by 29-kDa FN-f. In addition, TLR-2 knockdown apparently inhibited 29-kDa FN-f-mediated activation of phosphorylated nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha, and p38, but not of c-Jun N-terminal kinase or extracellular signal-regulated kinase. Exposure to synovial fluid (SF) from affected joints of patients with OA elevated MMP-1, MMP-3, and MMP-13 expression markedly in primary chondrocytes without reducing cell viability. However, TLR-2 knockdown in chondrocytes significantly suppressed SF-induced MMP induction.

Conclusions: Our data demonstrate that the MyD88-dependent TLR-2 signaling pathway may be responsible for 29-kDa FN-f-mediated cartilage catabolic responses. Our results will enhance understanding of cartilage catabolic mechanisms driven by cartilage degradation products, including FN-f. The modulation of TLR-2 signaling activated by damage-associated molecular patterns, including 29-kDa FN-f, is a potential therapeutic strategy for the prevention of cartilage degradation in OA.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aged
  • Aged, 80 and over
  • Cells, Cultured
  • Chondrocytes / drug effects
  • Chondrocytes / metabolism*
  • Female
  • Fibronectins / pharmacology*
  • Gene Expression Regulation, Enzymologic
  • Humans
  • Male
  • Matrix Metalloproteinases / biosynthesis*
  • Middle Aged
  • Myeloid Differentiation Factor 88 / biosynthesis*
  • Peptide Fragments / pharmacology
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Toll-Like Receptor 2 / biosynthesis*

Substances

  • Fibronectins
  • MYD88 protein, human
  • Myeloid Differentiation Factor 88
  • Peptide Fragments
  • TLR2 protein, human
  • Toll-Like Receptor 2
  • Matrix Metalloproteinases