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. 1978 Mar;75(3):1554–1558. doi: 10.1073/pnas.75.3.1554

Degradation of basic protein in myelin by neutral proteases secreted by stimulated macrophages: A possible mechanism of inflammatory demyelination

Wendy Cammer *,, Barry R Bloom ‡,§, William T Norton *,, Siamon Gordon
PMCID: PMC411512  PMID: 148651

Abstract

In inflammatory demyelinating diseases such as multiple sclerosis and experimental allergic encephalomyelitis, myelin destruction occurs in the vicinity of infiltrating mononuclear cells. The observations that myelin can be altered prior to phagocytosis and in areas not contiguous with inflammatory cells suggests a common mechanism for the initial stages of demyelination. Because stimulated macrophages secrete several neutral proteases, including plasminogen activator, we have investigated the possibility that myelinolysis could be mediated directly or indirectly by these enzymes. Isolated myelin was incubated with conditioned media from cultures of thioglycollate-stimulated mouse peritoneal macrophages in the presence and absence of plasminogen. Myelin appeared to be vulnerable to attack by at least two proteolytic activities secreted by the macrophages, a plasminogen-dependent and a plasminogen-independent activity; of the major proteins in myelin, the basic protein was most susceptible. The direct myelinolytic activity of macrophage-conditioned media was abolished by EDTA, and the plasminogen-dependent hydrolysis was abolished by p-nitrophenylguanidinobenzoate, an inhibitor of plasminogen activator and plasmin. These results suggest that the plasminogen activator released by the stimulated macrophages generated plasmin which hydrolyzed basic protein in intact myelin. This interpretation was confirmed by the observation that urokinase, a plasminogen activator, in the presence of plasminogen brought about marked degradation of basic protein in myelin. We propose that the release of neutral proteases by stimulated macrophages involved in cell-mediated reactions, and its amplification by the plasminogen-plasmin system, may play a significant role in the demyelination observed in several inflammatory demyelinating diseases.

Keywords: plasminogen activator, plasmin, cell-mediated immunity, urokinase, protease inhibitors

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Selected References

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  1. Althaus H. H., Pilz H., Müller D. The protein composition of myelin in multiple sclerosis (MS) and orthochromatic leukodystrophy (OLD). Z Neurol. 1973 Nov 5;205(3):229–241. doi: 10.1007/BF00316087. [DOI] [PubMed] [Google Scholar]
  2. Arstila A. U., Riekkinen P., Rinne U. K., Laitinen L. Studies on the pathogenesis of multiple sclerosis. Participation of lysosomes on demyelination in the central nervous system white matter outside plaques. Eur Neurol. 1973;9(1):1–20. doi: 10.1159/000114197. [DOI] [PubMed] [Google Scholar]
  3. Banik N. L., Davison A. N. Lipid and basic protein interaction of myelin. Biochem J. 1974 Oct;143(1):39–45. doi: 10.1042/bj1430039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Boxer P. A., Leibovich S. J. Production of collagenase by mouse peritoneal macrophages in vitro. Characterization of sites of cleavage of tropocollagen. Biochim Biophys Acta. 1976 Sep 24;444(2):626–632. doi: 10.1016/0304-4165(76)90408-6. [DOI] [PubMed] [Google Scholar]
  5. Buletza G. F., Jr, Smith M. E. Enzymic hydrolysis of myelin basic protein and other proteins in central nervous system and lymphoid tissues from normal and demyelinating rats. Biochem J. 1976 Jun 15;156(3):627–633. doi: 10.1042/bj1560627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cammer W., Norton W. T. Disc gel electrophoresis of myelin proteins: new observations on development of the intermediate proteins (DM-20). Brain Res. 1976 Jun 18;109(3):643–648. doi: 10.1016/0006-8993(76)90046-9. [DOI] [PubMed] [Google Scholar]
  7. Cohen S. R., Herndon R. M., McKhann G. M. Radioimmunoassay of myelin basic protein in spinal fluid. An index of active demyelination. N Engl J Med. 1976 Dec 23;295(26):1455–1457. doi: 10.1056/NEJM197612232952604. [DOI] [PubMed] [Google Scholar]
  8. Cuatrecasas P., Wilchek M., Anfinsen C. B. Selective enzyme purification by affinity chromatography. Proc Natl Acad Sci U S A. 1968 Oct;61(2):636–643. doi: 10.1073/pnas.61.2.636. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cuzner M. L., Barnard R. O., MacGregor B. J., Borshell N. J., Davison A. N. Myelin composition in acute and chronic multiple sclerosis in relation to cerebral lysosomal activity. J Neurol Sci. 1976 Oct;29(2-4):323–334. doi: 10.1016/0022-510x(76)90181-7. [DOI] [PubMed] [Google Scholar]
  10. Dalcanto M. C., Wiśniewski H. M., Johnson A. B., Brostoff S. W., Raine C. S. Vesicular disruption of myelin in autoimmune demyelination. J Neurol Sci. 1975 Mar;24(3):313–319. doi: 10.1016/0022-510x(75)90251-8. [DOI] [PubMed] [Google Scholar]
  11. Einstein E. R., Csejtey J., Dalal K. B., Adams C. W., Bayliss O. B., Hallpike J. F. Proteolytic activity and basic protein loss in and around multiple sclerosis plaques: combined biochemical and histochemical observations. J Neurochem. 1972 Mar;19(3):653–662. doi: 10.1111/j.1471-4159.1972.tb01382.x. [DOI] [PubMed] [Google Scholar]
  12. Einstein E. R., Dalal K. B., Csejtey J. Increased protease activity and changes in basic proteins and lipids in multiple sclerosis plaques. J Neurol Sci. 1970 Aug;11(2):109–121. doi: 10.1016/0022-510x(70)90121-8. [DOI] [PubMed] [Google Scholar]
  13. Gordon S., Unkeless J. C., Cohn Z. A. Induction of macrophage plasminogen activator by endotoxin stimulation and phagocytosis: evidence for a two-stage process. J Exp Med. 1974 Oct 1;140(4):995–1010. doi: 10.1084/jem.140.4.995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Govindarajan K. R., Rauch H. C., Clausen J., Einstein E. R. Changes in cathepsins B-1 and D, neutral proteinase and 2',3'-cyclic nucleotide-3'-phosphohydrolase activities in monkey brain with experimental allergic encephalomyelitis. J Neurol Sci. 1974 Oct;23(2):295–306. doi: 10.1016/0022-510x(74)90232-9. [DOI] [PubMed] [Google Scholar]
  15. Greenfield S., Norton W. T., Morell P. Quaking mouse: isolation and characterization of myelin protein. J Neurochem. 1971 Nov;18(11):2119–2128. doi: 10.1111/j.1471-4159.1971.tb05070.x. [DOI] [PubMed] [Google Scholar]
  16. Hallpike J. F., Adams C. W., Bayliss O. B. Histochemistry of myelin. 8. Proteolytic activity around multiple sclerosis plaques. Histochem J. 1970 May;2(3):199–208. doi: 10.1007/BF01003469. [DOI] [PubMed] [Google Scholar]
  17. Hirsch H. E., Duquette P., Parks M. E. The quantitative histochemistry of multiple sclerosis plaques: acid proteinase and other acid hydrolases. J Neurochem. 1976 Mar;26(3):505–512. doi: 10.1111/j.1471-4159.1976.tb01503.x. [DOI] [PubMed] [Google Scholar]
  18. Hirsch H. E., Parks M. E. Acid proteinases and other acid hydrolases in experimental allergic encephalomyelitis: pinpointing the source. J Neurochem. 1975 May;24(5):853–858. doi: 10.1111/j.1471-4159.1975.tb03647.x. [DOI] [PubMed] [Google Scholar]
  19. Horwitz A. L., Kelman J. A., Crystal R. G. Activation of alveolar macrophage collagenase by a neutral protease secreted by the same cell. Nature. 1976 Dec 23;264(5588):772–774. doi: 10.1038/264772a0. [DOI] [PubMed] [Google Scholar]
  20. Klimetzek V., Sorg C. Lymphokine-induced secretion of plasminogen activator by murine macrophages. Eur J Immunol. 1977 Mar;7(3):185–187. doi: 10.1002/eji.1830070314. [DOI] [PubMed] [Google Scholar]
  21. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  22. Macrophages and cathepsin A activity in multiple sclerosis brain. J Neurol Sci. 1974 Feb;21(2):227–231. doi: 10.1016/0022-510x(74)90074-4. [DOI] [PubMed] [Google Scholar]
  23. Marks N., Grynbaum A., Levine S. Proteolytic enzymes in ordinary, hyperacute, monocytic and passive transfer forms of experimental allergic encephalomyelitis. Brain Res. 1977 Mar 4;123(1):147–157. doi: 10.1016/0006-8993(77)90649-7. [DOI] [PubMed] [Google Scholar]
  24. Nogueira N., Gordon S., Cohn Z. Trypanosoma cruzi: the immunological induction of macrophage plasminogen activator requires thymus-derived lymphocytes. J Exp Med. 1977 Jul 1;146(1):172–183. doi: 10.1084/jem.146.1.172. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Norton W. T., Poduslo S. E. Myelination in rat brain: method of myelin isolation. J Neurochem. 1973 Oct;21(4):749–757. doi: 10.1111/j.1471-4159.1973.tb07519.x. [DOI] [PubMed] [Google Scholar]
  26. Prineas J. Pathology of the early lesion in multiple sclerosis. Hum Pathol. 1975 Sep;6(5):531–554. doi: 10.1016/s0046-8177(75)80040-2. [DOI] [PubMed] [Google Scholar]
  27. Raghavan S. S., Rhoads D. B., Kanfer J. N. The effects of trypsin on purified myelin. Biochim Biophys Acta. 1973 Nov 11;328(1):205–212. doi: 10.1016/0005-2795(73)90346-2. [DOI] [PubMed] [Google Scholar]
  28. Raine C. S., Snyder D. H., Valsamis M. P., Stone S. H. Chronic experimental allergic encephalomyelitis in inbred guinea pigs. An ultrastructural study. Lab Invest. 1974 Oct;31(4):369–380. [PubMed] [Google Scholar]
  29. Rauch H. C., Einstein E. R., Csejtey J. Enzymatic degradation of myelin basic protein in central nervous system lesions of monkeys with experimental allergic encephalomyelitis. Neurobiology. 1973;3(3):195–205. [PubMed] [Google Scholar]
  30. Riekkinen P. J., Clausen J., Frey H. J., Fog T., Rinne U. K. Acid proteinase activity of white matter and plaques in multiple sclerosis. Acta Neurol Scand. 1970;46(3):349–353. doi: 10.1111/j.1600-0404.1970.tb05799.x. [DOI] [PubMed] [Google Scholar]
  31. Riekkinen P., Rinne U. K., Savolainen H., Palo J., Kivalo E., Arstila A. Studies on the pathogenesis of multiple sclerosis. Basic proteins in the myelin and white matter of multiple sclerosis, subacute sclerosing panencephalitis and postvaccinal leucoencephalitis. Eur Neurol. 1971;5(4):229–244. doi: 10.1159/000114075. [DOI] [PubMed] [Google Scholar]
  32. Smith M. E. A lymph node neutral proteinase acting on myelin basic protein. J Neurochem. 1976 Nov;27(5):1077–1082. doi: 10.1111/j.1471-4159.1976.tb00311.x. [DOI] [PubMed] [Google Scholar]
  33. Smith M. E., Sedgewick L. M., Tagg J. S. Proteolytic enzymes and experimental demyelination in the rat and monkey. J Neurochem. 1974 Nov;23(5):965–971. doi: 10.1111/j.1471-4159.1974.tb10748.x. [DOI] [PubMed] [Google Scholar]
  34. Unkeless J. C., Gordon S., Reich E. Secretion of plasminogen activator by stimulated macrophages. J Exp Med. 1974 Apr 1;139(4):834–850. doi: 10.1084/jem.139.4.834. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Vassalli J. D., Hamilton J., Reich E. Macrophage plasminogen activator: modulation of enzyme production by anti-inflammatory steroids, mitotic inhibitors, and cyclic nucleotides. Cell. 1976 Jun;8(2):271–281. doi: 10.1016/0092-8674(76)90011-8. [DOI] [PubMed] [Google Scholar]
  36. Vassalli J. D., Reich E. Macrophage plasminogen activator: induction by products of activated lymphoid cells. J Exp Med. 1977 Feb 1;145(2):429–437. doi: 10.1084/jem.145.2.429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Wahl L. M., Wahl S. M., Mergenhagen S. E., Martin G. R. Collagenase production by lymphokine-activated macrophages. Science. 1975 Jan 24;187(4173):261–263. doi: 10.1126/science.163038. [DOI] [PubMed] [Google Scholar]
  38. Werb Z., Gordon S. Elastase secretion by stimulated macrophages. Characterization and regulation. J Exp Med. 1975 Aug 1;142(2):361–377. doi: 10.1084/jem.142.2.361. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Werb Z., Gordon S. Secretion of a specific collagenase by stimulated macrophages. J Exp Med. 1975 Aug 1;142(2):346–360. doi: 10.1084/jem.142.2.346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Whitaker J. N. Myelin encephalitogenic protein fragments in cerebrospinal fluid of persons with multiple sclerosis. Neurology. 1977 Oct;27(10):911–920. doi: 10.1212/wnl.27.10.911. [DOI] [PubMed] [Google Scholar]
  41. Wisniewski H. M., Bloom B. R. Primary demyelination as a nonspecific consequence of a cell-mediated immune reaction. J Exp Med. 1975 Feb 1;141(2):346–359. doi: 10.1084/jem.141.2.346. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Wiśniewski H. M., Bloom B. R. Experimental allergic optic neuritis (EAON) in the rabbit. A new model to study primary demyelinating diseases. J Neurol Sci. 1975 Feb;24(2):257–263. doi: 10.1016/0022-510x(75)90237-3. [DOI] [PubMed] [Google Scholar]
  43. Wiśniewski H., Prineas J., Raine C. S. An ultrastructural study of experimental demyelination and remyelination. I. Acute experimental allergic encephalomyelitis in the peripheral nervous system. Lab Invest. 1969 Aug;21(2):105–118. [PubMed] [Google Scholar]
  44. Wood J. G., Dawson R. M., Hauser H. Effect of proteolytic attack on the structure of CNS myelin membrane. J Neurochem. 1974 May;22(5):637–643. doi: 10.1111/j.1471-4159.1974.tb04275.x. [DOI] [PubMed] [Google Scholar]

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