Skip to main content
PMC home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1996 Feb 1;97(3):621–632. doi: 10.1172/JCI118458

Immunity to the G1 globular domain of the cartilage proteoglycan aggrecan can induce inflammatory erosive polyarthritis and spondylitis in BALB/c mice but immunity to G1 is inhibited by covalently bound keratan sulfate in vitro and in vivo.

J Y Leroux 1, A Guerassimov 1, A Cartman 1, N Delaunay 1, C Webber 1, L C Rosenberg 1, S Banerjee 1, A R Poole 1
PMCID: PMC507097  PMID: 8609216

Abstract

Earlier work from this laboratory showed that the human proteoglycan aggrecan from fetal cartilages can induce a CD4+ T cell-dependent inflammatory polyarthritis in BALB/c mice when injected after removal of chondroitin sulfate chains. Adult keratan sulfate (KS)-rich aggrecan does not possess this property. We found that two CD4+ T cell hybridomas (TH5 and TH14) isolated from arthritic mice recognize bovine calf aggrecan and the purified G1 domain of this molecule, which also contains a portion of the interglobular domain to which KS is bound. These hybridoma responses to G1 are enhanced by partial removal of KS by the endoglycosidase keratanase or by cyanogen bromide cleavage of core protein. KS removal results in increased cellular uptake by antigen-present cells in vitro. After removal of KS by keratanase, G1 alone can induce a severe erosive polyarthritis and spondylitis in BALB/c mice identifying it as an arthritogenic domain of aggrecan. The presence of KS prevents induction of arthritis presumably as a result of an impaired immune response as observed in vitro. These observations not only identify the arthritogenic properties of G1 but they also point to the importance of glycosylation and proteolysis in determining the arthritogenicity of aggrecan and fragments thereof.

Full Text

The Full Text of this article is available as a PDF (757.8 KB).

Selected References

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

  1. Abou-Zeid C., Filley E., Steele J., Rook G. A. A simple new method for using antigens separated by polyacrylamide gel electrophoresis to stimulate lymphocytes in vitro after converting bands cut from Western blots into antigen-bearing particles. J Immunol Methods. 1987 Apr 2;98(1):5–10. doi: 10.1016/0022-1759(87)90429-7. [DOI] [PubMed] [Google Scholar]
  2. Allen P. M., Unanue E. R. Differential requirements for antigen processing by macrophages for lysozyme-specific T cell hybridomas. J Immunol. 1984 Mar;132(3):1077–1079. [PubMed] [Google Scholar]
  3. Banerjee S., Webber C., Poole A. R. The induction of arthritis in mice by the cartilage proteoglycan aggrecan: roles of CD4+ and CD8+ T cells. Cell Immunol. 1992 Oct 15;144(2):347–357. doi: 10.1016/0008-8749(92)90250-s. [DOI] [PubMed] [Google Scholar]
  4. Barry F. P., Gaw J. U., Young C. N., Neame P. J. Hyaluronan-binding region of aggrecan from pig laryngeal cartilage. Amino acid sequence, analysis of N-linked oligosaccharides and location of the keratan sulphate. Biochem J. 1992 Sep 15;286(Pt 3):761–769. doi: 10.1042/bj2860761. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Barry F. P., Rosenberg L. C., Gaw J. U., Gaw J. U., Koob T. J., Neame P. J. N- and O-linked keratan sulfate on the hyaluronan binding region of aggrecan from mature and immature bovine cartilage. J Biol Chem. 1995 Sep 1;270(35):20516–20524. doi: 10.1074/jbc.270.35.20516. [DOI] [PubMed] [Google Scholar]
  6. Bonnet F., Dunham D. G., Hardingham T. E. Structure and interactions of cartilage proteoglycan binding region and link protein. Biochem J. 1985 May 15;228(1):77–85. doi: 10.1042/bj2280077. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Campbell I. K., Golds E. E., Mort J. S., Roughley P. J. Human articular cartilage secretes characteristic metal dependent proteinases upon stimulation by mononuclear cell factor. J Rheumatol. 1986 Feb;13(1):20–27. [PubMed] [Google Scholar]
  8. Caterson B., Christner J. E., Baker J. R. Identification of a monoclonal antibody that specifically recognizes corneal and skeletal keratan sulfate. Monoclonal antibodies to cartilage proteoglycan. J Biol Chem. 1983 Jul 25;258(14):8848–8854. [PubMed] [Google Scholar]
  9. Cawston T. E., Barrett A. J. A rapid and reproducible assay for collagenase using [1-14C]acetylated collagen. Anal Biochem. 1979 Nov 1;99(2):340–345. doi: 10.1016/s0003-2697(79)80017-2. [DOI] [PubMed] [Google Scholar]
  10. Fosang A. J., Hardingham T. E. 1-C-6 epitope in cartilage proteoglycan G2 domain is masked by keratan sulphate. Biochem J. 1991 Jan 15;273(Pt 2):369–373. doi: 10.1042/bj2730369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Glant T. T., Mikecz K., Arzoumanian A., Poole A. R. Proteoglycan-induced arthritis in BALB/c mice. Clinical features and histopathology. Arthritis Rheum. 1987 Feb;30(2):201–212. doi: 10.1002/art.1780300211. [DOI] [PubMed] [Google Scholar]
  12. Glant T. T., Mikecz K., Roughley P. J., Buzás E., Poole A. R. Age-related changes in protein-related epitopes of human articular-cartilage proteoglycans. Biochem J. 1986 May 15;236(1):71–75. doi: 10.1042/bj2360071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Golds E. E., Stephen I. B., Esdaile J. M., Strawczynski H., Poole A. R. Lymphocyte transformation to connective tissue antigens in adult and juvenile rheumatoid arthritis, osteoarthritis, ankylosing spondylitis, systemic lupus erythematosus, and a nonarthritic control population. Cell Immunol. 1983 Nov;82(1):196–209. doi: 10.1016/0008-8749(83)90153-3. [DOI] [PubMed] [Google Scholar]
  14. Heinegård D., Sommarin Y. Isolation and characterization of proteoglycans. Methods Enzymol. 1987;144:319–372. doi: 10.1016/0076-6879(87)44186-4. [DOI] [PubMed] [Google Scholar]
  15. Ishioka G. Y., Lamont A. G., Thomson D., Bulbow N., Gaeta F. C., Sette A., Grey H. M. MHC interaction and T cell recognition of carbohydrates and glycopeptides. J Immunol. 1992 Apr 15;148(8):2446–2451. [PubMed] [Google Scholar]
  16. Kappler J. W., Skidmore B., White J., Marrack P. Antigen-inducible, H-2-restricted, interleukin-2-producing T cell hybridomas. Lack of independent antigen and H-2 recognition. J Exp Med. 1981 May 1;153(5):1198–1214. doi: 10.1084/jem.153.5.1198. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kozarsky K., Kingsley D., Krieger M. Use of a mutant cell line to study the kinetics and function of O-linked glycosylation of low density lipoprotein receptors. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4335–4339. doi: 10.1073/pnas.85.12.4335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. 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]
  19. 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]
  20. Laurent T. C., Fraser J. R., Pertoft H., Smedsrød B. Binding of hyaluronate and chondroitin sulphate to liver endothelial cells. Biochem J. 1986 Mar 15;234(3):653–658. doi: 10.1042/bj2340653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Leroux J. Y., Poole A. R., Webber C., Vipparti V., Choi H. U., Rosenberg L. C., Banerjee S. Characterization of proteoglycan-reactive T cell lines and hybridomas from mice with proteoglycan-induced arthritis. J Immunol. 1992 Apr 1;148(7):2090–2096. [PubMed] [Google Scholar]
  22. Matsudaira P. Sequence from picomole quantities of proteins electroblotted onto polyvinylidene difluoride membranes. J Biol Chem. 1987 Jul 25;262(21):10035–10038. [PubMed] [Google Scholar]
  23. Mikecz K., Glant T. T., Baron M., Poole A. R. Isolation of proteoglycan-specific T lymphocytes from patients with ankylosing spondylitis. Cell Immunol. 1988 Mar;112(1):55–63. doi: 10.1016/0008-8749(88)90275-4. [DOI] [PubMed] [Google Scholar]
  24. Mikecz K., Glant T. T., Buzás E., Poole A. R. Proteoglycan-induced polyarthritis and spondylitis adoptively transferred to naive (nonimmunized) BALB/c mice. Arthritis Rheum. 1990 Jun;33(6):866–876. doi: 10.1002/art.1780330614. [DOI] [PubMed] [Google Scholar]
  25. Mikecz K., Glant T. T., Poole A. R. Immunity to cartilage proteoglycans in BALB/c mice with progressive polyarthritis and ankylosing spondylitis induced by injection of human cartilage proteoglycan. Arthritis Rheum. 1987 Mar;30(3):306–318. doi: 10.1002/art.1780300310. [DOI] [PubMed] [Google Scholar]
  26. Mosmann T. R., Cherwinski H., Bond M. W., Giedlin M. A., Coffman R. L. Two types of murine helper T cell clone. I. Definition according to profiles of lymphokine activities and secreted proteins. J Immunol. 1986 Apr 1;136(7):2348–2357. [PubMed] [Google Scholar]
  27. Nakazawa K., Suzuki S. Purification of Keratan Sulfate-endogalactosidase and its action on keratan sulfates of different origin. J Biol Chem. 1975 Feb 10;250(3):912–917. [PubMed] [Google Scholar]
  28. Neame P. J., Christner J. E., Baker J. R. Cartilage proteoglycan aggregates. The link protein and proteoglycan amino-terminal globular domains have similar structures. J Biol Chem. 1987 Dec 25;262(36):17768–17778. [PubMed] [Google Scholar]
  29. Oike Y., Kimata K., Shinomura T., Nakazawa K., Suzuki S. Structural analysis of chick-embryo cartilage proteoglycan by selective degradation with chondroitin lyases (chondroitinases) and endo-beta-D-galactosidase (keratanase). Biochem J. 1980 Oct 1;191(1):193–207. doi: 10.1042/bj1910193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Olden K., Pratt R. M., Yamada K. M. Role of carbohydrate in biological function of the adhesive glycoprotein fibronectin. Proc Natl Acad Sci U S A. 1979 Jul;76(7):3343–3347. doi: 10.1073/pnas.76.7.3343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Poole A. R., Webber C., Reiner A., Roughley P. J. Studies of a monoclonal antibody to skeletal keratan sulphate. Importance of antibody valency. Biochem J. 1989 Jun 15;260(3):849–856. doi: 10.1042/bj2600849. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Roughley P. J., White R. J. Age-related changes in the structure of the proteoglycan subunits from human articular cartilage. J Biol Chem. 1980 Jan 10;255(1):217–224. [PubMed] [Google Scholar]
  33. Roughley P. J., White R. J., Poole A. R. Identification of a hyaluronic acid-binding protein that interferes with the preparation of high-buoyant-density proteoglycan aggregates from adult human articular cartilage. Biochem J. 1985 Oct 1;231(1):129–138. doi: 10.1042/bj2310129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sandy J. D., Flannery C. R., Boynton R. E., Neame P. J. Isolation and characterization of disulfide-bonded peptides from the three globular domains of aggregating cartilage proteoglycan. J Biol Chem. 1990 Dec 5;265(34):21108–21113. [PubMed] [Google Scholar]
  35. Sandy J. D., Flannery C. R., Neame P. J., Lohmander L. S. The structure of aggrecan fragments in human synovial fluid. Evidence for the involvement in osteoarthritis of a novel proteinase which cleaves the Glu 373-Ala 374 bond of the interglobular domain. J Clin Invest. 1992 May;89(5):1512–1516. doi: 10.1172/JCI115742. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sandy J. D., Neame P. J., Boynton R. E., Flannery C. R. Catabolism of aggrecan in cartilage explants. Identification of a major cleavage site within the interglobular domain. J Biol Chem. 1991 May 15;266(14):8683–8685. [PubMed] [Google Scholar]
  37. Saxne T., Heinegård D. Synovial fluid analysis of two groups of proteoglycan epitopes distinguishes early and late cartilage lesions. Arthritis Rheum. 1992 Apr;35(4):385–390. doi: 10.1002/art.1780350404. [DOI] [PubMed] [Google Scholar]
  38. Stevens J. W., Oike Y., Handley C., Hascall V. C., Hampton A., Caterson B. Characteristics of the core protein of the aggregating proteoglycan from the Swarm rat chondrosarcoma. J Cell Biochem. 1984;26(4):247–259. doi: 10.1002/jcb.240260405. [DOI] [PubMed] [Google Scholar]
  39. Tang L. H., Rosenberg L., Reiner A., Poole A. R. Proteoglycans from bovine nasal cartilage. Properties of a soluble form of link protein. J Biol Chem. 1979 Oct 25;254(20):10523–10531. [PubMed] [Google Scholar]
  40. Unanue E. R. Antigen-presenting function of the macrophage. Annu Rev Immunol. 1984;2:395–428. doi: 10.1146/annurev.iy.02.040184.002143. [DOI] [PubMed] [Google Scholar]
  41. Witter J., Roughley P. J., Webber C., Roberts N., Keystone E., Poole A. R. The immunologic detection and characterization of cartilage proteoglycan degradation products in synovial fluids of patients with arthritis. Arthritis Rheum. 1987 May;30(5):519–529. doi: 10.1002/art.1780300506. [DOI] [PubMed] [Google Scholar]

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

RESOURCES