Implication of prostaglandin E2 in TNF-α-induced release of m-calpain from HCS-2/8 chondrocytes. Inhibition of m-calpain release by NSAIDs

K. Fushimi, S. Nakashima, Y. Banno, A. Akaike, M. Takigawa, Katsuji Shimizu

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

Objectives: Calpains are known as Ca2+-dependent intracellular neutral cysteine proteases. However, m-calpain is detected in synovial fluid of arthritic joints and is shown to possess the proteoglycanase activity in vitro. The mechanism of m-calpain release into the extracellular spaces during arthritis has not yet been well characterized. In the present study, we have analyzed m-calpain release from cultured chondrocytes stimulated by a proinflammatory cytokine, tumor necrosis factor-α (TNF-α). The effects of non-steroidal anti-inflammatory drugs (NSAIDs) on m-calpain release were also examined. Methods: Human chondrocytic HCS-2/8 cells were stimulated by TNF-α in the presence or absence of an NSAID. m-Calpain in the cells and culture medium was quantified by Western blot analysis using an anti-m-calpain antibody. Western blots were subjected to densitometric analysis and band intensities were determined. Results: TNF-α (10 ng/ml) stimulated m-calpain release with transient increase in cellular m-calpain in HCS-2/8 cells. NSAIDs examined (aspirin, loxoprofen-SRS, diclofenac sodium, indomethacin and NS398) inhibited m-calpain release and production of prostaglandin E2 (PGE2) induced by 10 ng/ml TNF-α. Exogenously added PGE2 accelerated the release of m-calpain in response to a lower concentration of TNF-α (1 ng/ml). AH6809, an EP1/2 antagonist, but not SC19220 (an EP1 antagonist), effectively inhibited TNF-α-induced m-calpain release. In contrast, butaprost, an EP2 agonist, accelerated release of m-calpain by 1 ng/ml TNF-α. Conclusions: These results suggest that TNF-α stimulates upregulation and release of m-calpain in chondrocytic HCS-2/8 cells, and that stimulation of EP2-PGE2 receptor by produced PGE2 is deeply involved in this process.

Original languageEnglish
Pages (from-to)895-903
Number of pages9
JournalOsteoarthritis and Cartilage
Volume12
Issue number11
DOIs
Publication statusPublished - Nov 1 2004

Keywords

  • Human chondrocytes
  • PGE
  • TNF-α
  • m-Calpain

ASJC Scopus subject areas

  • Rheumatology
  • Biomedical Engineering
  • Orthopedics and Sports Medicine

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