A SERINE PROTEINASE INACTIVATOR INHIBITS CHONDROCYTE-MEDIATED CARTILAGE PROTEOGLYCAN BREAKDOWN OCCURRING IN RESPONSE TO PROINFLAMMATORY CYTOKINES

The role played by serine proteinases with trypsinlike specificity in chondrocyte-mediated cartilage proteoglycan breakdown was investigated by use of a selective proteinase inactivator, no-4-chloro-3-(-3-isoth iureidopropoxy)isocoumarin, in explant culture systems. This compound was a rapid inactivator of urokinase-type plasminogen activator. It po tently inhibited interleukin 1- and tumor necrosis factor-stimulated p roteoglycan release from both nasal and articular cartilage. Its less potent inhibition of basal and retinoic acid-stimulated release appear ed to be due to cytotoxic effects. The functional half-life of the ina ctivator in culture medium was 95 min, and its concentration in cartil age was 2.5-fold higher than in the surrounding medium. Following spon taneous hydrolysis the breakdown products of the inactivator were unab le to inhibit proteoglycan release. Trypsin-like activity was demonstr ated by enzyme histochemistry to be chondrocyte-associated and inhibit ed by the serine proteinase inactivator. Cell-associated and secreted plasminogen activator activity was detected by zymography. These resul ts suggest the involvement of a serine proteinase(s) with trypsin-like specificity, possibly urokinase-type plasminogen activator, in chondr ocyte-mediated cartilage proteoglycan breakdown occurring as a result of stimulation with proinflammatory cytokines. Basal proteoglycan brea kdown may occur via a different pathway. Our findings point to a patho logical role for serine proteinase(s) in the development of cartilage diseases such as arthritis, possibly in a cascade which results in the activation of the enzyme(s) directly responsible for proteoglycan bre akdown. It remains to be shown whether the target serine proteinase is urokinase-type plasminogen activator. (C) 1998 Academic Press.