Characterisation of chondrocyte activation in response to cytokines synthesised by a synovial cell line

Abstract

The lapine, synovial cell line, HIG-82, secretes 'chondrocyte activating factors' (CAF) which induce the synthesis of collagenase (EC 3.4.24.7), gelatinase, caseinase and prostaglandin E2 (PGE2) by confluent, monolayer cultures of lapine, articular chondrocytes. Partially purified CAF increased the production of PGE2 by chondrocytes within 3 h; in certain cultures this occurred in as little as 1 h. Increased levels of the three neutral metalloproteinases, in contrast, were only measurable in the conditioned medium after a delay of 9-18 h. After removal of the CAF, the synthesis of PGE2 reverted to basal levels within 1-4 h, but synthesis of the three proteinases remained high for an additional 4 days. Indomethacin, at concentrations which completely inhibited PGE2 synthesis, had no effect upon the coordinate induction of collagenase, gelatinase and caseinase. However, cycloheximide, alpha-amanitin and 5,6-dichlororibosylbenzimidazole (DRB) suppressed induction of these proteinases suggesting that CAF derepressed the genes coding for these enzymes. Once the chondrocytes had been activated by CAF, the inhibitors of transcription had a much weaker effect on the production of the neutral proteinases, indicating that their mRNAs may be relatively stable. In the presence of CAF, inhibition under these conditions was weaker still, possibly due to stabilisation of these mRNA molecules. Experiments with a number of compounds which modulate cellular Ca2+, cAMP or cGMP failed to support a straightforward role for these mediators in the induction of neutral metalloproteinases in chondrocytes. High concentrations of phorbol myristate acetate (PMA) provoked only a slight synthesis of these enzymes.