Skip to main content
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1978 Nov;62(5):987–992. doi: 10.1172/JCI109228

Activation in vitro of rheumatoid synovial collagenase from cell cultures.

C A Vater, C L Mainardi, E D Harris Jr
PMCID: PMC371857  PMID: 213448

Abstract

Rheumatoid synovial cells dissociated from matrix and adherent to culture dishes released a latent form of collagenase into culture medium. Previous studies have shown that the latent enzyme does not complex with alpha2-macroglobulin and binds to fibrillar substrate. We now show that serum-free culture medium of the synovial cells contains an inhibitor of collagenase as well as latent enzyme; the two were separated on a column of acrylamide/agarose. Latent collagenase (estimated mol wt 45,000-49,000) was transformed by trypsin to active collagenase of approximately equal to mol wt 33,000. When mixed with inhibitor the active enzyme formed an inactive complex again with approximately equal to mol wt 45,000-49,000. The inhibitor(s) itself was found in one major peak of mol wt 33,000-35,000 and several minor peaks eluting with lower apparent molecular weight. Mersalyl, an organic mercurial compound, effectively activated latent collagenase producing an active enzyme with approximately equal to mol wt 33,000. Bacterial collagenase did not activate latent enzyme. We suggest that latent rheumatoid synovial collagenase, as it is harvested from synovial cells in culture, is an enzyme-inhibitor complex.

Full text

PDF
987

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bauer E. A., Eisen A. Z., Jeffrey J. J. Studies on purified rheumatoid synovial collagenase in vitro and in vivo. J Clin Invest. 1971 Oct;50(10):2056–2064. doi: 10.1172/JCI106699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Chibber B. A., Deutsch D. G., Mertz E. T. Affinity chromatography of plasminogen. Methods Enzymol. 1974;34:424–432. doi: 10.1016/s0076-6879(74)34051-7. [DOI] [PubMed] [Google Scholar]
  3. Dayer J. M., Krane S. M., Russell R. G., Robinson D. R. Production of collagenase and prostaglandins by isolated adherent rheumatoid synovial cells. Proc Natl Acad Sci U S A. 1976 Mar;73(3):945–949. doi: 10.1073/pnas.73.3.945. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Eeckhout Y., Vaes G. Further studies on the activation of procollagenase, the latent precursor of bone collagenase. Effects of lysosomal cathepsin B, plasmin and kallikrein, and spontaneous activation. Biochem J. 1977 Jul 15;166(1):21–31. doi: 10.1042/bj1660021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Evanson J. M., Jeffrey J. J., Krane S. M. Studies on collagenase from rheumatoid synovium in tissue culture. J Clin Invest. 1968 Dec;47(12):2639–2651. doi: 10.1172/JCI105947. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fiedler-Nagy C., Coffey J. W., Salvador R. A. Factors influencing the apparent molecular weight of collagenase produced by human-skin explants. Eur J Biochem. 1977 Jun 1;76(1):291–297. doi: 10.1111/j.1432-1033.1977.tb11595.x. [DOI] [PubMed] [Google Scholar]
  7. GLIMCHER M. J., FRANCOIS C. J., RICHARDS L., KRANE S. M. THE PRESENCE OF ORGANIC PHOSPHORUS IN COLLAGENS AND GELATINS. Biochim Biophys Acta. 1964 Dec 9;93:585–602. doi: 10.1016/0304-4165(64)90342-3. [DOI] [PubMed] [Google Scholar]
  8. GROSS J., KIRK D. The heat precipitation of collagen from neutral salt solutions: some rate-regulating factors. J Biol Chem. 1958 Aug;233(2):355–360. [PubMed] [Google Scholar]
  9. Harris E. D., Jr, DiBona D. R., Krane S. M. Collagenases in human synovial fluid. J Clin Invest. 1969 Nov;48(11):2104–2113. doi: 10.1172/JCI106177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Harris E. D., Jr Recent insights into the pathogenesis of the proliferative lesion in rheumatoid arthritis. Arthritis Rheum. 1976 Jan-Feb;19(1):68–72. doi: 10.1002/art.1780190111. [DOI] [PubMed] [Google Scholar]
  11. Krane S. M. Collagenase production by human synovial tissues. Ann N Y Acad Sci. 1975 Jun 13;256:289–303. doi: 10.1111/j.1749-6632.1975.tb36056.x. [DOI] [PubMed] [Google Scholar]
  12. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  13. McCroskery P. A., Richards J. F., Harris E. D., Jr Purification and characterization of a collagenase extracted from rabbit tumours. Biochem J. 1975 Oct;152(1):131–142. doi: 10.1042/bj1520131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Reynolds J. J., Murphy G., Sellers A., Cartwright E. A new factor that may control collagen resorption. Lancet. 1977 Aug 13;2(8033):333–335. doi: 10.1016/s0140-6736(77)91490-8. [DOI] [PubMed] [Google Scholar]
  15. Sellers A., Cartwright E., Murphy G., Reynolds J. J. An inhibitor of mammalian collagenase from foetal rabbit bone in culture. Biochem Soc Trans. 1977;5(1):227–229. doi: 10.1042/bst0050227. [DOI] [PubMed] [Google Scholar]
  16. Sellers A., Cartwright E., Murphy G., Reynolds J. J. Evidence that latent collagenases are enzyme-inhibitor complexes. Biochem J. 1977 May 1;163(2):303–307. doi: 10.1042/bj1630303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Stricklin G. P., Bauer E. A., Jeffrey J. J., Eisen A. Z. Human skin collagenase: isolation of precursor and active forms from both fibroblast and organ cultures. Biochemistry. 1977 Apr 19;16(8):1607–1615. doi: 10.1021/bi00627a013. [DOI] [PubMed] [Google Scholar]
  18. Vater C. A., Mainardi C. L., Harris E. D., Jr Binding of latent rheumatoid synovial collagenase to collagen fibrils. Biochim Biophys Acta. 1978 Mar 1;539(2):238–247. doi: 10.1016/0304-4165(78)90010-7. [DOI] [PubMed] [Google Scholar]
  19. Werb Z., Burleigh M. C., Barrett A. J., Starkey P. M. The interaction of alpha2-macroglobulin with proteinases. Binding and inhibition of mammalian collagenases and other metal proteinases. Biochem J. 1974 May;139(2):359–368. doi: 10.1042/bj1390359. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Werb Z., Mainardi C. L., Vater C. A., Harris E. D., Jr Endogenous activiation of latent collagenase by rheumatoid synovial cells. Evidence for a role of plasminogen activator. N Engl J Med. 1977 May 5;296(18):1017–1023. doi: 10.1056/NEJM197705052961801. [DOI] [PubMed] [Google Scholar]
  21. Werb Z., Reynolds J. J. Stimulation by endocytosis of the secretion of collagenase and neutral proteinase from rabbit synovial fibroblasts. J Exp Med. 1974 Dec 1;140(6):1482–1497. doi: 10.1084/jem.140.6.1482. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Woolley D. E., Glanville R. W., Crossley M. J., Evanson J. M. Purification of rheumatoid synovial collagenase and its action on soluble and insoluble collagen. Eur J Biochem. 1975 Jun;54(2):611–622. doi: 10.1111/j.1432-1033.1975.tb04173.x. [DOI] [PubMed] [Google Scholar]
  23. Woolley D. E., Roberts D. R., Evanson J. M. Small molecular weight beta 1 serum protein which specifically inhibits human collagenases. Nature. 1976 May 27;261(5558):325–327. doi: 10.1038/261325a0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES