Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1989 Sep 1;109(3):1363–1369. doi: 10.1083/jcb.109.3.1363

Cellular heterogeneity in cultured human chondrocytes identified by antibodies specific for alpha 2(XI) collagen chains

PMCID: PMC2115754  PMID: 2670958

Abstract

Collagen type XI is a component of hyaline cartilage consisting of alpha 1(XI), alpha 2(XI), and alpha 3(XI) chains; with 5-10% of the total collagen content, it is a minor but significant component next to type II collagen, but its function and precise localization in cartilaginous tissues is still unclear. Owing to the homology of the alpha 3(XI) and alpha 1(II) collagen chains, attempts to prepare specific antibodies to native type XI collagen have been unsuccessful in the past. In this study, we report on the preparation and use for immunohistochemistry of a polyclonal antibody specific for alpha 2(XI) denatured collagen chains. The antibody was prepared by immunization with the isolated alpha 2(XI) chain and reacts neither with native type XI collagen nor type I, II, V, or IX by ELISA or immunoblotting, nor with alpha 1(XI) or alpha 3(XI), but with alpha 2(XI) chains. Using this antibody, it was possible to specifically localize alpha 2(XI) in cartilage by pretreating tissue sections with 6 M urea. In double immunofluorescence staining experiments, the distribution of alpha 2(XI) as indicative for type XI collagen in fetal bovine and human cartilage was compared with that of type II collagen, using a monoclonal antibody to alpha 1(II). Type XI collagen was found throughout the matrix of hyaline cartilage. However, owing to cross- reactivity of the monoclonal anti-alpha 1(II) with alpha 3(XI), both antibodies produced the same staining pattern. Cellular heterogeneity was, however, detected in monolayer cultures of human chondrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Full Text

The Full Text of this article is available as a PDF (2.5 MB).

Selected References

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

  1. Adachi E., Hayashi T. In vitro formation of fine fibrils with a D-periodic banding pattern from type V collagen. Coll Relat Res. 1985 Jun;5(3):225–232. doi: 10.1016/s0174-173x(85)80012-1. [DOI] [PubMed] [Google Scholar]
  2. Ayad S., Evans H., Weiss J. B., Holt L. Type VI collagen but not type V collagen is present in cartilage. Coll Relat Res. 1984 Mar;4(2):165–168. doi: 10.1016/s0174-173x(84)80023-0. [DOI] [PubMed] [Google Scholar]
  3. Bentz H., Bächinger H. P., Glanville R., Kühn K. Physical evidence for the assembly of A and B chains of human placental collagen in a single triple helix. Eur J Biochem. 1978 Dec;92(2):563–567. doi: 10.1111/j.1432-1033.1978.tb12778.x. [DOI] [PubMed] [Google Scholar]
  4. Birk D. E., Fitch J. M., Babiarz J. P., Linsenmayer T. F. Collagen type I and type V are present in the same fibril in the avian corneal stroma. J Cell Biol. 1988 Mar;106(3):999–1008. doi: 10.1083/jcb.106.3.999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Burgeson R. E., Hollister D. W. Collagen heterogeneity in human cartilage: identification of several new collagen chains. Biochem Biophys Res Commun. 1979 Apr 27;87(4):1124–1131. doi: 10.1016/s0006-291x(79)80024-8. [DOI] [PubMed] [Google Scholar]
  6. Capasso O., Tajana G., Cancedda R. Location of 64K collagen producer chondrocytes in developing chicken embryo tibiae. Mol Cell Biol. 1984 Jun;4(6):1163–1168. doi: 10.1128/mcb.4.6.1163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dessau W., Sasse J., Timpl R., Jilek F., von der Mark K. Synthesis and extracellular deposition of fibronectin in chondrocyte cultures. Response to the removal of extracellular cartilage matrix. J Cell Biol. 1978 Nov;79(2 Pt 1):342–355. doi: 10.1083/jcb.79.2.342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dessau W., Vertel B. M., von der Mark H., von der Mark K. Extracellular matrix formation by chondrocytes in monolayer culture. J Cell Biol. 1981 Jul;90(1):78–83. doi: 10.1083/jcb.90.1.78. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Dodge G. R., Poole A. R. Immunohistochemical detection and immunochemical analysis of type II collagen degradation in human normal, rheumatoid, and osteoarthritic articular cartilages and in explants of bovine articular cartilage cultured with interleukin 1. J Clin Invest. 1989 Feb;83(2):647–661. doi: 10.1172/JCI113929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Engvall E., Perlmann P. Enzyme-linked immunosorbent assay (ELISA). Quantitative assay of immunoglobulin G. Immunochemistry. 1971 Sep;8(9):871–874. doi: 10.1016/0019-2791(71)90454-x. [DOI] [PubMed] [Google Scholar]
  11. Eyre D. R., Apon S., Wu J. J., Ericsson L. H., Walsh K. A. Collagen type IX: evidence for covalent linkages to type II collagen in cartilage. FEBS Lett. 1987 Aug 17;220(2):337–341. doi: 10.1016/0014-5793(87)80842-6. [DOI] [PubMed] [Google Scholar]
  12. Furuto D. K., Miller E. J. Different levels of glycosylation contribute to the heterogeneity of alpha 1(II) collagen chains derived from a transplantable rat chondrosarcoma. Arch Biochem Biophys. 1983 Oct 15;226(2):604–611. doi: 10.1016/0003-9861(83)90329-6. [DOI] [PubMed] [Google Scholar]
  13. Gibson G. J., Flint M. H. Type X collagen synthesis by chick sternal cartilage and its relationship to endochondral development. J Cell Biol. 1985 Jul;101(1):277–284. doi: 10.1083/jcb.101.1.277. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Haralson M. A., Mitchell W. M., Rhodes R. K., Kresina T. F., Gay R., Miller E. J. Chinese hamster lung cells synthesize and confine to the cellular domain a collagen composed solely of B chains. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5206–5210. doi: 10.1073/pnas.77.9.5206. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Herrmann H., Dessau W., Fessler L. I., von der Mark K. Synthesis of types I, III and AB2 collagen by chick tendon fibroblasts in vitro. Eur J Biochem. 1980 Mar;105(1):63–74. doi: 10.1111/j.1432-1033.1980.tb04474.x. [DOI] [PubMed] [Google Scholar]
  16. Holmdahl R., Rubin K., Klareskog L., Larsson E., Wigzell H. Characterization of the antibody response in mice with type II collagen-induced arthritis, using monoclonal anti-type II collagen antibodies. Arthritis Rheum. 1986 Mar;29(3):400–410. doi: 10.1002/art.1780290314. [DOI] [PubMed] [Google Scholar]
  17. Keene D. R., Engvall E., Glanville R. W. Ultrastructure of type VI collagen in human skin and cartilage suggests an anchoring function for this filamentous network. J Cell Biol. 1988 Nov;107(5):1995–2006. doi: 10.1083/jcb.107.5.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. 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]
  19. Linsenmayer T. F., Fitch J. M., Schmid T. M., Zak N. B., Gibney E., Sanderson R. D., Mayne R. Monoclonal antibodies against chicken type V collagen: production, specificity, and use for immunocytochemical localization in embryonic cornea and other organs. J Cell Biol. 1983 Jan;96(1):124–132. doi: 10.1083/jcb.96.1.124. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Linsenmayer T. F., Smith G. N., Jr, Hay E. D. Synthesis of two collagen types by embryonic chick corneal epithelium in vitro. Proc Natl Acad Sci U S A. 1977 Jan;74(1):39–43. doi: 10.1073/pnas.74.1.39. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Mayne R., Elrod B. W., Mayne P. M., Sanderson R. D., Linsenmayer T. F. Changes in the synthesis of minor cartilage collagens after growth of chick chondrocytes in 5-bromo-2'-deoxyuridine or to senescence. Exp Cell Res. 1984 Mar;151(1):171–182. doi: 10.1016/0014-4827(84)90366-5. [DOI] [PubMed] [Google Scholar]
  22. Mendler M., Eich-Bender S. G., Vaughan L., Winterhalter K. H., Bruckner P. Cartilage contains mixed fibrils of collagen types II, IX, and XI. J Cell Biol. 1989 Jan;108(1):191–197. doi: 10.1083/jcb.108.1.191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Morris N. P., Bächinger H. P. Type XI collagen is a heterotrimer with the composition (1 alpha, 2 alpha, 3 alpha) retaining non-triple-helical domains. J Biol Chem. 1987 Aug 15;262(23):11345–11350. [PubMed] [Google Scholar]
  24. Reese C. A., Mayne R. Minor collagens of chicken hyaline cartilage. Biochemistry. 1981 Sep 15;20(19):5443–5448. doi: 10.1021/bi00522a014. [DOI] [PubMed] [Google Scholar]
  25. Sangiorgi F. O., Benson-Chanda V., de Wet W. J., Sobel M. E., Ramirez F. Analysis of cDNA and genomic clones coding for the pro alpha 1 chain of calf type II collagen. Nucleic Acids Res. 1985 Apr 25;13(8):2815–2826. doi: 10.1093/nar/13.8.2815. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Schmid T. M., Linsenmayer T. F. Immunohistochemical localization of short chain cartilage collagen (type X) in avian tissues. J Cell Biol. 1985 Feb;100(2):598–605. doi: 10.1083/jcb.100.2.598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Shimokomaki M., Duance V. C., Bailey A. J. Identification of a new disulphide bonded collagen from cartilage. FEBS Lett. 1980 Nov 17;121(1):51–54. doi: 10.1016/0014-5793(80)81265-8. [DOI] [PubMed] [Google Scholar]
  28. Swann D. A., Sotman S. S. The chemical composition of bovine vitreous-humour collagen fibres. Biochem J. 1980 Mar 1;185(3):545–554. doi: 10.1042/bj1850545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Thorogood P., Bee J., von der Mark K. Transient expression of collagen type II at epitheliomesenchymal interfaces during morphogenesis of the cartilaginous neurocranium. Dev Biol. 1986 Aug;116(2):497–509. doi: 10.1016/0012-1606(86)90150-8. [DOI] [PubMed] [Google Scholar]
  30. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Vaughan L., Mendler M., Huber S., Bruckner P., Winterhalter K. H., Irwin M. I., Mayne R. D-periodic distribution of collagen type IX along cartilage fibrils. J Cell Biol. 1988 Mar;106(3):991–997. doi: 10.1083/jcb.106.3.991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. von Der Mark K., Ocalan M. Immunofluorescent localization of type V collagen in the chick embryo with monoclonal antibodies. Coll Relat Res. 1982 Nov;2(6):541–555. doi: 10.1016/s0174-173x(82)80008-3. [DOI] [PubMed] [Google Scholar]
  33. von der Mark H., von der Mark K., Gay S. Study of differential collagen synthesis during development of the chick embryo by immunofluorescence. I. Preparation of collagen type I and type II specific antibodies and their application to early stages of the chick embryo. Dev Biol. 1976 Feb;48(2):237–249. doi: 10.1016/0012-1606(76)90088-9. [DOI] [PubMed] [Google Scholar]
  34. von der Mark K., van Menxel M., Wiedemann H. Isolation and characterization of new collagens from chick cartilage. Eur J Biochem. 1982 May;124(1):57–62. doi: 10.1111/j.1432-1033.1982.tb05905.x. [DOI] [PubMed] [Google Scholar]
  35. von der Mark K., von der Mark H. Immunological and biochemical studies of collagen type transition during in vitro chrondrogenesis of chick limb mesodermal cells. J Cell Biol. 1977 Jun;73(3):736–747. doi: 10.1083/jcb.73.3.736. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. von der Mark K., von der Mark H., Timpl R., Trelstad R. L. Immunofluorescent localization of collagen types I, II, and III in the embryonic chick eye. Dev Biol. 1977 Aug;59(1):75–85. doi: 10.1016/0012-1606(77)90241-x. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES