Abstract
The culture of chondrocytes is one of the most powerful tool for exploring the intracellular and molecular features of chondrocyte differentiation and activation. However, chondrocytes tend to dedifferentiate to fibroblasts when they are subcultured, which is a major problem. This chapter describes several protocols for culturing chondrocytes of different anatomical origins (articular and costal chondrocytes) from various species (humans, mice, rabbits, and cattle). All these protocols involve primary cultures in order to limit dedifferentiation. This chapter also describes a new protocol for culturing mouse articular chondrocytes.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Aydelotte, M. B., and Kuettner, K. E. (1988) Differences between sub-populations of cultured bovine articular chondrocytes. I. Morphology and cartilage matrix production. Connect. Tissue Res. 18, 205–222
von der Mark, K. and Conrad, G. (1979) Cartilage cell differentiation. Clin. Orthop. 139, 185–205
Stokes, D. G., Liu, G., Coimbra, I. B., Piera-Velazquez, S., Crowl, R. M., and Jimenez, S. A. (2002) Assessment of the gene expression profile of differentiated and dedifferentiated human fetal chondrocytes by microarray analysis. Arthritis Rheum. 46, 404–419.
Kergosien, N., Sautier, J., and Forest, N. (1998) Gene and protein expression during differentiation and matrix mineralization in a chondrocyte cell culture system. Calcif. Tissue Int. 62, 114–121.
Hauselmann, H. J., Aydelotte, M. B., Schumacher, B. L., Kuettner, K. E., Gitelis, S. H., and Thonar, E. J. (1992) Synthesis and turnover of proteoglycans by human and bovine adult articular chondrocytes cultured in alginate beads. Matrix 12, 116–129.
Karsenty, G. (2001) Chondrogenesis just ain’t what it used to be. J. Clin. Invest. 107, 405–407.
Corvol, M. T., Dumontier, M. F., and Rappaport, R. (1975) Culture of chondrocytes from the proliferative zone of epiphyseal growth plate cartilage from prepubertal rabbits. Biomedicine 23, 103–107.
Mwale, F., Billinghurst, C., Wu, W., et al. (2000) Selective assembly and remodelling of collagens II and IX associated with expression of the chondrocyte hypertrophic phenotype. Dev. Dyn. 218, 648–662.
Goldring, M. B. (2000) The role of the chondrocyte in osteoarthritis. Arthritis Rheum. 43, 1916–1926.
Goldring, M. B. and Berenbaum, F. (1999) Human chondrocyte culture models for studying cyclooxygenase expression and prostaglandin regulation of collagen gene expression. Osteoarthritis Cartilage 7, 386–388.
Watt, F. M. (1988) Effect of seeding density on stability of the differentiated phenotype of pig articular chondrocytes in culture. J. Cell. Sci. 89, 373–378.
Stokes, D. G., Liu, G., Dharmavaram, R., Hawkins, D., Piera-Velazquez, S., and Jimenez, S. A. (2001) Regulation of type-II collagen gene expression during human chondrocyte de-differentiation and recovery of chondrocyte-specific phenotype in culture involves Sry-type high-mobility-group box (SOX) transcription factors. Biochem. J. 360, 461–470.
Goldring, M. B., Birkhead, J., Sandell, L. J., Kimura, T., and Krane, S. M. (1988) Interleukin 1 suppresses expression of cartilage-specific types II and IX collagens and increases types I and III collagens in human chondrocytes. J. Clin. Invest. 82, 2026–2037.
Goldring, M. B., Birkhead, J. R., Suen, L. F., et al. (1994) Interleukin-1 beta-modulated gene expression in immortalized human chondrocytes. J. Clin. Invest. 94, 2307–2316.
Demoor-Fossard, M., Redini, F., Boittin, M., and Pujol, J. P. (1998) Expression of decorin and biglycan by rabbit articular chondrocytes. Effects of cytokines and phenotypic modulation. Biochim. Biophys. Acta 1398, 179–191.
Gibson, G. J., Schor, S. L., and Grant, M. E. (1982) Effects of matrix macromolecules on chondrocyte gene expression: synthesis of a low molecular weight collagen species by cells cultured within collagen gels. J. Cell Biol. 93, 767–774.
Takigawa, M., Pan, H. O., Kinoshita, A., Tajima, K., and Takano, Y. (1991) Establishment from a human chondrosarcoma of a new immortal cell line with high tumorigenicity in vivo, which is able to form proteoglycan-rich cartilage-like nodules and to respond to insulin in vitro. Int. J. Cancer 48, 717–725.
Mallein-Gerin, F., and Olsen, B. R. (1993) Expression of simian virus 40 large T (tumor) oncogene in mouse chondrocytes induces cell proliferation without loss of the differentiated phenotype. Proc. Natl. Acad. Sci. USA 90, 3289–3293.
Robbins, J. R., Thomas, B., Tan, L., et al. (2000) Immortalized human adult articular chondrocytes maintain cartilage-specific phenotype and responses to interleukin-1β. Arthritis Rheum. 43, 2189–2201.
Adolphe, M., Froger, B., Ronot, X., Corvol, M. T., and Forest, N. (1984) Cell multiplication and type II collagen production by rabbit articular chondrocytes cultivated in a defined medium. Exp. Cell Res. 155, 527–536.
Martin, I., Vunjak-Novakovic, G., Yang, J., Langer, R., and Freed, L. E. (1999) Mammalian chondrocytes expanded in the presence of fibroblast growth factor 2 maintain the ability to differentiate and regenerate three-dimensional cartilaginous tissue. Exp. Cell Res. 253, 681–688.
Kuettner, K. E., Memoli, V. A., Pauli, B. U., Wrobel, N. C., Thonar, E. J., and Daniel, J. C. (1982) Synthesis of cartilage matrix by mammalian chondrocytes in vitro. II. Maintenance of collagen and proteoglycan phenotype. J. Cell Biol. 93, 751–757.
Domm, C., Schunke, M., Christesen, K., and Kurz, B. (2002) Redifferentiation of dedifferentiated bovine articular chondrocytes in alginate culture under low oxygen tension. Osteoarthritis Cartilage 10, 13–22.
Zaucke, F., Dinser, R., Maurer, P., and Paulsson, M. (2001) Cartilage oligomeric matrix protein (COMP) and collagen IX are sensitive markers for the differentiation state of articular primary chondrocytes. Biochem. J. 358, 17–24.
Rahfoth, B., Weisser, J., Sternkopf, F., Aigner, T., von der Mark, K., and Brauer, R. (1998) Transplantation of allograft chondrocytes embedded in agarose gel into cartilage defects of rabbits. Osteoarthritis Cartilage 6, 50–65.
Robbins, J. R. and Goldring, M. B. (1998) Preparation of immortalized human chondrocyte cell lines, in Tissue Engineering, Vol. 18 (Morgan, J. R., and Yarmush, M. L., eds.), Humana, Totowa, NJ, pp. 173–192.
Goldring, M. B. (1996) Human chondrocyte cultures as models of cartilage-specific gene regulation, in Human Cell Culture Protocols, Vol. 2 (Gareth, E. J., ed.), Humana, Totowa, NJ, pp. 217–232.
Aulthouse, A. L., Beck, M., Griffey, E., et al. (1989) Expression of the human chondrocyte phenotype in vitro. In Vitro Cell Dev. Biol. 25, 659–668.
Carrascosa, A., Audi, L., and Ballabriga, A. (1985) Morphologic and metabolic development of human fetal epiphyseal chondrocytes in primary culture. Pediatr. Res. 19, 720–727.
Reginato, A. M., Iozzo, R. V., and Jimenez, S. A. (1994) Formation of nodular structures resembling mature articular cartilage in long-term primary cultures of human fetal epiphyseal chondrocytes on a hydrogel substrate. Arthritis Rheum. 37, 1338–1349.
Stove, J., Gerlach, C., Huch, K., et al. (2001) Gene expression of stromelysin and aggrecan in osteoarthritic cartilage. Pathobiology 69, 333–338.
Kawiak, J., Moskalewski, S., and Darzynkiewicz, Z. (1965) Isolation of chondrocytes from calf cartilage. Exp. Cell Res. 39, 59–68.
Nedelec, E., Abid, A., Cipolletta, C., et al. (2001) Stimulation of cyclooxygenase-2-activity by nitric oxide-derived species in rat chondrocyte: lack of contribution to loss of cartilage anabolism. Biochem. Pharmacol. 61, 965–978.
Okazaki, M., Higuchi, Y., and Kitamura, H. (2003) AG-041R stimulates cartilage matrix synthesis without promoting terminal differentiation in rat articular chondrocytes. Osteoarthritis Cartilage 11, 122–132.
Gouze, J. N., Bordji, K., Gulberti, S., et al. (2001) Interleukin-1beta down-regulates the expression of glucuronosyltransferase I, a key enzyme priming glycosaminoglycan biosynthesis: influence of glucosamine on interleukin-1β-mediated effects in rat chondrocytes. Arthritis Rheum. 44, 351–360.
Gouze, J. N., Bianchi, A., Becuwe, P., et al. (2002) Glucosamine modulates IL-1-induced activation of rat chondrocytes at a receptor level, and by inhibiting the NF-k B pathway. FEBS Lett. 510, 166–170.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2004 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Thirion, S., Berenbaum, F. (2004). Culture and Phenotyping of Chondrocytes in Primary Culture. In: Sabatini, M., Pastoureau, P., De Ceuninck, F. (eds) Cartilage and Osteoarthritis. Methods in Molecular Medicine™, vol 100. Humana Press. https://doi.org/10.1385/1-59259-810-2:001
Download citation
DOI: https://doi.org/10.1385/1-59259-810-2:001
Publisher Name: Humana Press
Print ISBN: 978-1-58829-247-6
Online ISBN: 978-1-59259-810-6
eBook Packages: Springer Protocols