The high levels of pro-inflammatory cytokines in rheumatoid arthritis elicit increased synthesis of matrix metalloproteinases in the rheumatoid joint leading to degradation of articular cartilage and loss of joint function. Transforming growth factor beta 1 (TGF-beta1) acts antagonistically to the actions of TNF-alpha and IL-1beta decreasing synthesis of collagenase-1, -2 and -2 in normal human chrondrocytes stimulated with these cytokines. We propose this growth factor may be effective in reducing collagenase synthesis in RA and will study the mechanism of its effects. Specifically we will (a) Determine the levels of different collagenases produced by RA synovial tissue and cartilage explants and evaluate the effects of incubation with TGF-beta1. (2) Characterize the mechanism of TGFbeta1 inhibition of collagenase-3 expression in human chrondrocytes. Inhibition of collagenase-3 by TGF- beta1 will be examined with regard to transcriptional or translational control using a model of normal and immortalized human articular chrondrocytes stimulates with TNF-alpha or IL-1beta to induce up- regulation of collagenase synthesis. The region/s of the collagenase-3 promoter which are necessary for the inhibition will be identified with transient transfections of progressive deletions of the promoter constructs. We will further define these regions with DNase footprinting and electrophoretic shift mobility assays. (3) Characterize the mechanism of TGF-beta1 induction of collagenase-3 expression. We will determine if low levels of inflammatory cytokines are permissive for TGF-beta1 induction of collagenase-3 in human articular chrondrocytes. We will define the transcriptional factors which relate to this phenomena. We postulate that TGF-beta1 would be an effective modulator of all three collagenases in RA tissues and may be useful chondroprotective agent to regulate collagenase synthesis and influence the rate of cartilage degradation.
