Role of TIMP and MMP Inhibition in Preventing Connective Tissue Breakdowna

The matrix metalloproteinases (MMPs) are a unique family of enzymes that in concert can degrade all the components of the extracellular matrix.'These potent enzymes are made in a proenzyme form, and activation occurs after leaving the cell.Extracellular activity is also controlled by specific inhibitor called tissue inhibitor of metalloproteinases (TIMPS)~ that bind to the active forms of the enzyme forming 1:l complexes and block their activity.The inflammatory cytokine interleukin-1 (IL-1) can induce resorption of cartilage in vitro3 and in vivo when injected into rabbit joints4 and is also found in joint fluids from patients with rheumatoid a r t h r i t i ~. ~. ~ IL-1 is also known to stimulate the production of the MMPs collagenase and stromelysin from human synovial cells and chondrocyte~.~Raised levels of these enzymes have been localized at both the mRNA and protein level within diseased cartilage from different rheumatic dis-eases8 Consequently, the MMPs are a valid therapeutic target for inhibition in the rheumatic disease^.^A number of studies have shown that IL-1-stimulated cartilage model systems can be used to test the effectiveness of proteinase inhibitors.The release of glycosaminoglycan (GAG) fragments can be prevented by the addition of low molecular weight synthetic but not by the addition of TIMP.lS-l6However, although the release of GAG fragments from cartilage appears to be rapid in vivo in response to IL-14J7 it is also quickly resynthesized by chondrocytes.18In contrast the release of collagen from cartilage is much slower, is less reproducible, and appears to be irreversible as resynthesis is difficult to achieve.19In this study we have examined the release of both collagen and proteoglycan fragments from resorbing cartilage to determine if the naturally occurring MMP inhibitors TIMP and TIMP-2 and low molecular weight synthetic inhibitors can block both proteoglycan and collagen degradation in IL-1 stimulated cartilage. MATERIALS AND METHODS Cartilage Degradation AssayControl culture medium was Dulbecco's modification of Eagle's medium containing 25 mM HEPES (Gibco) supplemented with sodium bicarbonate (0.5 g/L), nThis work was supported by the Arthritis and Rheumatism Council.