Much evidence implicates collagenolysis by specific enzymes in joint destruction in rheumatoid arthritis and other chronic states of synovial inflammation. Monolayer culture of synovial cells has facilitated determination of mechanisms involved during induction, biosynthesis, secretion and activation of collagenase. In the rabbit system (and preliminary data indicate that the same is true for human rheumatoid synovial cells), procollagenase (proC) is secreted along with a latent procollagenase activator (proA). ProA is essential for activation of proC. Like proC, proA is a metalloproteinase. ProA must itself be activated before it can activate proC. In order to study in more detail the induction, activation and inhibition of collagenolysis in disease states, pure probes that can measure the synthetic machinery for proC and proA must be constructed. Rabbit and human cDNA for proC and proA will be generated in expressing plasmid vectors introduced into E.coli. The probes for proC and proA will be isolated, purified, and characterized by several techniques (i.e., direct immunologic screening, use of immunoprecipitated polysomes, differential hybridization, and/or hybridization with synthetic oligonucleotides). The cDNA probes will be used to: (1) characterize genomic DNA for proC and proA; (2) determine amino acid sequences for proC and proA in the rabbit and human; (3) help determine how proC is activated by proA so that the value of generating an inhibitor of this process may be assessed; (4) study the kinetics of induction of mRNA for proC and proA and the kinetics of inhibition of synthesis of proC and proA by glucocorticoids and retinoids; (5) examine by in situ hybridization the localization of mRNA for proC and proA (compared with mRNA for procollagen) in cartilage/pannus junction and surrounding tissue in joints removed at arthroplasty in RA; (6) sort out the apparent abnormalities in the sequence of biosynthesis and/or activation of proC in """"""""Milwaukee Shoulder"""""""" and (7) examine the hypothesis that complexes of alpha 2 macroglobulin collagenase phagocytosed by synovial cells results in alterations of gene expression by these cells. It is hoped with precise knowledge about the intricacies of biosynthesis and activation of proC, useful and specific ways of intervening to retard the collagenolytic cascade might be suggested.
| Buttice, G; Quinones, S; Kurkinen, M (1991) The AP-1 site is required for basal expression but is not necessary for TPA-response of the human stromelysin gene. Nucleic Acids Res 19:3723-31 |
| Otani, Y; Quinones, S; Saus, J et al. (1990) Cycloheximide induces stromelysin mRNA in cultured human fibroblasts. Eur J Biochem 192:75-9 |
| Quinones, S; Saus, J; Otani, Y et al. (1989) Transcriptional regulation of human stromelysin. J Biol Chem 264:8339-44 |
| Saus, J; Quinones, S; Otani, Y et al. (1988) The complete primary structure of human matrix metalloproteinase-3. Identity with stromelysin. J Biol Chem 263:6742-5 |
