Induction of stromelysin-1 and collagenase synthesis in fibrochondrocytes by tumor necrosis factor-α

Stromelysin-1 and collagenase mRNA levels were assayed in fibrochondrocytes by Northern blot analysis at 0, 2, 4 8 and 24 h after stimulation with tissue necrosis for-α (TNF-α). Peak collagenase mRNA levels occurred 24 after stimulation and were increased nine-fold over the level at time 0. Stromelysin-1 mRNA levels peaked 8 h after stimulation, with a five-fold increase over the level at time 0. A TNF-α dose-related response to both collagenase and stromelysin-1 mRNA accumulation was also demonstrated. Confirmation of the presence of secreted metalloproteinases in the conditioned media was established by immunoprecipitation of stromelsyin-1 and Western blotting of collagenase. Both enzymes were secreted in latent forms. Consistent with stromelysin-1 activity, substrate gels demonstrated a doublet of caseinase activity with molecular masses at 57 kDa and 59 kDa in TNF-α stimulated samples. Collagenase assays of conditioned media also demonstrated a significant increase in collagenase activity after stimulation by TNF-α. While epidermal growth factor had a minimal effect on stromelysin-1 and collagenase expression, transforming growth factor-β, and insulin-like growth factor-1 did not induce either enzyme activity.