Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1981 Jul 1;90(1):78–83. doi: 10.1083/jcb.90.1.78

Extracellular matrix formation by chondrocytes in monolayer culture

PMCID: PMC2111838  PMID: 6788783

Abstract

In previous studies were have reported on the secretion and extracellular deposition of type II collagen and fibronectin (Dessau et al., 1978, J. Cell Biol., 79:342-355) and chondroitin sulfate proteoglycan (CSPG) (Vertel and Dorfman, 1979, Proc. Natl. Acad. Sci. U. S. A. 76:1261-1264) in chondrocyte cultures. This study describes a combined effort to compare sequence and pattern of secretion and deposition of all three macromolecules in the same chondrocyte culture experiment. By immunofluorescence labeling experiments, we demonstrate that type II collagen, fibronectin, and CSPG reappear on the cell surface after enzymatic release of chondrocytes from embryonic chick cartilage but develop different patterns in the pericellular matrix. When chondrocytes spread on the culture dish, CSPG is deposited in the extracellular space as an amorphous mass and fibronectin forms fine, intercellular strands, whereas type II collagen disappears from the chondrocyte surface and remains absent from the extracellular space in early cultures. Only after cells in the center of chondrocyte colonies shape reassume spherical shape does the immunofluorescence reveal type II collagen in the refractile matrix characteristic of differentiated cartilage. By immunofluorescence double staining of the newly formed cartilage matrix, we demonstrate that CSPG spreads farther out into the extracellular space that type II collagen. Fibronectin finally disappears from the cartilage matrix.

Full Text

The Full Text of this article is available as a PDF (803.4 KB).

Selected References

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

  1. Abbott J., Holtzer H. The loss of phenotypic traits by differentiated cells. 3. The reversible behavior of chondrocytes in primary cultures. J Cell Biol. 1966 Mar;28(3):473–487. doi: 10.1083/jcb.28.3.473. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ali I. U., Mautner V., Lanza R., Hynes R. O. Restoration of normal morphology, adhesion and cytoskeleton in transformed cells by addition of a transformation-sensitive surface protein. Cell. 1977 May;11(1):115–126. doi: 10.1016/0092-8674(77)90322-1. [DOI] [PubMed] [Google Scholar]
  3. 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]
  4. Dehm P., Prockop D. J. Biosynthesis of cartilage procollagen. Eur J Biochem. 1973 May;35(1):159–166. doi: 10.1111/j.1432-1033.1973.tb02821.x. [DOI] [PubMed] [Google Scholar]
  5. 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]
  6. Dessau W., von der Mark H., von der Mark K., Fischer S. Changes in the patterns of collagens and fibronectin during limb-bud chondrogenesis. J Embryol Exp Morphol. 1980 Jun;57:51–60. [PubMed] [Google Scholar]
  7. Dorfman A., Vertel B. M., Schwartz N. B. Immunological methods in the study of chondroitin sulfate proteoglycans. Curr Top Dev Biol. 1980;14(Pt 2):169–198. doi: 10.1016/s0070-2153(08)60194-5. [DOI] [PubMed] [Google Scholar]
  8. Hajek A. S., Solursh M. The effect of ascorbic acid on growth and synthesis of matrix components by cultured chick embryo chondrocytes. J Exp Zool. 1977 Jun;200(3):377–388. doi: 10.1002/jez.1402000308. [DOI] [PubMed] [Google Scholar]
  9. Hascall V. C. Interaction of cartilage proteoglycans with hyaluronic acid. J Supramol Struct. 1977;7(1):101–120. doi: 10.1002/jss.400070110. [DOI] [PubMed] [Google Scholar]
  10. Hascall V. C., Oegema T. R., Brown M., Caplan A. I. Isolation and characterization of proteoglycans from chick limb bud chondrocytes grown in vitro. J Biol Chem. 1976 Jun 10;251(11):3511–3519. [PubMed] [Google Scholar]
  11. Hascall V. C., Sajdera S. W. Physical properties and polydispersity of proteoglycan from bovine nasal cartilage. J Biol Chem. 1970 Oct 10;245(19):4920–4930. [PubMed] [Google Scholar]
  12. Hynes R. O., Destree A. T., Mautner V. M., Ali I. U. Synthesis, secretion, and attachment of LETS glycoprotein in normal and transformed cells. J Supramol Struct. 1977;7(3-4):397–408. doi: 10.1002/jss.400070311. [DOI] [PubMed] [Google Scholar]
  13. Levitt D., Dorfman A. Concepts and mechanisms of cartilage differentiation. Curr Top Dev Biol. 1974;8:103–149. doi: 10.1016/s0070-2153(08)60607-9. [DOI] [PubMed] [Google Scholar]
  14. Linder E., Vaheri A., Ruoslahti E., Wartiovaara J. Distribution of fibroblast surface antigen in the developing chick embryo. J Exp Med. 1975 Jul 1;142(1):41–49. doi: 10.1084/jem.142.1.41. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Mayne R., Vail M. S., Mayne P. M., Miller E. J. Changes in type of collagen synthesized as clones of chick chondrocytes grow and eventually lose division capacity. Proc Natl Acad Sci U S A. 1976 May;73(5):1674–1678. doi: 10.1073/pnas.73.5.1674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Miller E. J. Biochemical characteristics and biological significance of the genetically-distinct collagens. Mol Cell Biochem. 1976 Dec 10;13(3):165–192. doi: 10.1007/BF01731779. [DOI] [PubMed] [Google Scholar]
  17. Miller E. J., Matukas V. J. Chick cartilage collagen: a new type of alpha 1 chain not present in bone or skin of the species. Proc Natl Acad Sci U S A. 1969 Dec;64(4):1264–1268. doi: 10.1073/pnas.64.4.1264. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Müller P. K., Lemmen C., Gay S., Gauss V., Kühn K. Immunochemical and biochemical study of collagen synthesis by chondrocytes in culture. Exp Cell Res. 1977 Aug;108(1):47–55. [PubMed] [Google Scholar]
  19. Oakes B. W., Handley C. J., Lisner F., Lowther D. A. An ultrastructural and biochemical study of high density primary cultures of embryonic chick chondrocytes. J Embryol Exp Morphol. 1977 Apr;38:239–263. [PubMed] [Google Scholar]
  20. Okayama M., Pacifici M., Holtzer H. Differences among sulfated proteoglycans synthesized in nonchondrogenic cells, presumptive chondroblasts, and chondroblasts. Proc Natl Acad Sci U S A. 1976 Sep;73(9):3224–3228. doi: 10.1073/pnas.73.9.3224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Pennypacker J. P., Hassell J. R., Yamada K. M., Pratt R. M. The influence of an adhesive cell surface protein on chondrogenic expression in vitro. Exp Cell Res. 1979 Jul;121(2):411–415. doi: 10.1016/0014-4827(79)90022-3. [DOI] [PubMed] [Google Scholar]
  22. Perkins M. E., Ji T. H., Hynes R. O. Cross-linking of fibronectin to sulfated proteoglycans at the cell surface. Cell. 1979 Apr;16(4):941–952. doi: 10.1016/0092-8674(79)90109-0. [DOI] [PubMed] [Google Scholar]
  23. Peterkofsky B. The effect of ascorbic acid on collagen polypeptide synthesis and proline hydroxylation during the growth of cultured fibroblasts. Arch Biochem Biophys. 1972 Sep;152(1):318–328. doi: 10.1016/0003-9861(72)90221-4. [DOI] [PubMed] [Google Scholar]
  24. Solursh M., Meier S. Effects of cell density on the expression of differentiation by chick embryo chondrocytes. J Exp Zool. 1974 Mar;187(3):311–322. doi: 10.1002/jez.1401870302. [DOI] [PubMed] [Google Scholar]
  25. Upholt W. B., Vertel B. M., Dorfman A. Translation and characterization of messenger RNAs in differentiating chicken cartilage. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4847–4851. doi: 10.1073/pnas.76.10.4847. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Vaheri A., Mosher D. F. High molecular weight, cell surface-associated glycoprotein (fibronectin) lost in malignant transformation. Biochim Biophys Acta. 1978 Sep 18;516(1):1–25. doi: 10.1016/0304-419x(78)90002-1. [DOI] [PubMed] [Google Scholar]
  27. Vertel B. M., Dorfman A. An immunohistochemical study of extracellular matrix formation during chondrogenesis. Dev Biol. 1978 Jan;62(1):1–12. doi: 10.1016/0012-1606(78)90088-x. [DOI] [PubMed] [Google Scholar]
  28. Vertel B. M., Dorfman A. Simultaneous localization of type II collagen and core protein of chondroitin sulfate proteoglycan in individual chondrocytes. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1261–1264. doi: 10.1073/pnas.76.3.1261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. West C. M., Lanza R., Rosenbloom J., Lowe M., Holtzer H., Avdalovic N. Fibronectin alters the phenotypic properties of cultured chick embryo chondroblasts. Cell. 1979 Jul;17(3):491–501. doi: 10.1016/0092-8674(79)90257-5. [DOI] [PubMed] [Google Scholar]
  30. Yamada K. M., Olden K. Fibronectins--adhesive glycoproteins of cell surface and blood. Nature. 1978 Sep 21;275(5677):179–184. doi: 10.1038/275179a0. [DOI] [PubMed] [Google Scholar]
  31. 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]
  32. von der Mark K., Conrad G. Cartilage cell differentiation: review. Clin Orthop Relat Res. 1979 Mar-Apr;(139):185–205. [PubMed] [Google Scholar]
  33. von der Mark K., Gauss V., von der Mark H., Müller P. Relationship between cell shape and type of collagen synthesised as chondrocytes lose their cartilage phenotype in culture. Nature. 1977 Jun 9;267(5611):531–532. doi: 10.1038/267531a0. [DOI] [PubMed] [Google Scholar]

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

RESOURCES