Proteoglycan degradation by a chondrocyte metalloprotease. Effects of synthetic protease inhibitors.
Synthetic inhibitors of a chondrocyte metalloprotease (CMP) were assessed for potency. Proteoglycan core protein was used as substrate. The IC50 values were between 2 X 10(-6) and 7 X 10(-6) M for two types of inhibitors, thiol tripeptides and N-carboxyalkyl peptides. Hydroxamic acid peptides were more potent, with IC50 values of 3.2 X 10(-8) to 6.0 X 10(-8) M. These results confirm inhibitory concentrations reported using a proteoglycan-polyacrylamide bead assay. The slopes of the dose-response curves for the thiol compounds were steeper than the slopes for the other two types of compounds. All of the culture media tested inhibited CMP to some extent. Some media also interfered with inhibitor activity. In Ham's F10 nutrient medium, minimum CMP inhibition occurred, and all four hydroxamic acid peptides retained their activity for 1-2 days at 37 degrees. One thiol peptide compound assayed lost activity in 1 hr in thiocyanate-treated serum. All four hydroxamic acid peptides assayed retained activity in thiocyanate-treated serum after 3 days at 37 degrees. The hydroxamic acid peptides may provide a way to block endogenous CMP activity in vivo and to assess the role of CMP in normal and experimentally altered cartilage. They are more potent than other known CMP inhibitors. They retain activity in culture media and serum conditions used for in vivo and in vitro tests of CMP activity and toxicity.
D000818:Animals; D002358:Cartilage, Articular / Q000201:enzymology*; D002478:Cells, Cultured; D003470:Culture Media; D004355:Drug Stability; D010450:Endopeptidases; D007700:Kinetics; D008666:Metalloendopeptidases; D011480:Protease Inhibitors / Q000494:pharmacology*; D011509:Proteoglycans / Q000378:metabolism*; D011817:Rabbits
D003470:Culture Media; D011480:Protease Inhibitors; D011509:Proteoglycans; D010450:Endopeptidases; D008666:Metalloendopeptidases