Abstract
Chondrogenic differentiation of mesenchymal cells is generally thought to be initiated by the inductive action of specific growth factors and depends on intimate cell-cell interactions. The aim of our investigation was to characterize the influences of basic fibroblast growth factor (bFGF) and ferroussulfate (FeSO4) on proliferation and differentiation of human articular chondrocytes (HAC). This is the first report of the effects of FeSO4 on chondrogenesis of HAC. Multiplied chondrocytes of hip and shoulder joints were cultured in chondrocyte growth medium supplemented with bFGF, FeSO4, or both bFGF + FeSO4 for4weeks. A 20 μl aliquot of a cell suspension containing2 × 107 cells ml−1 was delivered onto each well of 24-well tissue culture plates. Cells cultured with the growth medium only was used as a control. Alamar blue and alcian blue staining were done to determine the chondrocyte proliferation and differentiation, respectively, after 4 weeks. The samples exposed to bFGF, FeSO4, and combination of both indicated sufficient cell proliferation similar to the control level. Differentiations of the HAC exposed to bFGF, FeSO4,and bFGF + FeSO4 were 1.2-, 2.0-, and 2.2-fold of the control, respectively. Therefore, chondrocyte differentiation was significantly enhanced by the addition of FeSO4 andbFGF + FeSO4. The combined effects of bFGF and FeSO4 were additive, rather than synergistic. These results suggest that treatment with ferrous sulfate alone or in combination with basic fibroblast growth factor etc, is a powerful tool to promote the differentiation of HAC for the clinical application.
Similar content being viewed by others
References
Brown C.C. and Balian G. 1987. Effect of heparin on synthesis of short chain collagen by chondrocytes and smooth muscle cells. J. Cell. Biol. 105: 1007–1012.
Bohme K., Conscience-Egli M., Tschan T., Winterhalter K.H. and Bruckner P. 1992. Induction of proliferation or hypertrophy of chondrocytes in serum-free culture: the role of insulin-like growth factor-I, insulin, or thyroxine. J. Cell. Biol. 116: 1035–1042.
Bujia J., Sittinger M., Wilmes E. and Hammer C. 1994. Effect of growth factors on cell proliferation by human nasal septal chondrocytes cultured in monolayer. Acta. Otolaryngol. 114: 539–543.
Bujia J., Pitzke P., Kastenbauer E., Wilmes E. and Hammer C. 1996. Effect of growth factors on matrix synthesis by human nasal chondrocytes cultured in monolayer and in agar. Eur. Arch. Otorhinolaryngol. 253: 336–340.
Dreyfus J., Brunet-de Carvalho N., Duprez D., Raulais D. and Vigny M. 1998. HB-GAM/ pleiotrophin but not RIHB/midkine enhances chondrogenesis in micromass culture. Exp. Cell. Res. 241: 171–180.
De Luca F., Uyeda J.A., Mericq V., Mancilla E.E., Yanovski J.A., Barnes K.M. et al. 2000. Retinoic acid is a potent regulator of growth plate chondrogenesis. Endocrinology 141: 346–353.
Fujisato T., Sajiki T., Liu Q. and Ikada Y. 1996. Effect of basic fibroblast growth factor on cartilage regeneration in chondrocyteseeded collagen sponge scaffold. Biomaterials 17: 155–162.
Fukuda K., Dan H., Takayama M., Kumano F., Saitoh M. and Tanaka S. 1996. Hyaluronic acid increases proteoglycan synthesis in bovine articular cartilage in the presence of interleukin-1. J. Pharmacol. Exp. Ther. 277: 1672–1675.
Fujimoto E., Ochi M., Kato Y., Mochizuki Y., Sumen Y. and Ikuta Y. 1999. Beneficial effect of basic fibroblast growth factor on the repair of full-thickness defects in rabbit articular cartilage. Arch. Orthop. Trauma. Surg. 119: 139–145.
Green W.T. Jr. 1977. Articular cartilage repair. Behavior of rabbit chondrocytes during tissue culture and subsequent allografting. Clin. Orthop. 124: 237–250.
Harper J. and Harper E. 1987. Insulin stimulates secretion of a collagenase inhibitor by Swarm rat chondrosarcoma chon-drocytes. Biochem. Biophys. Res. Commun. 147: 550–555.
Ito K., Fujisato T. and Ikada Y. 1992. Implantation of cell seeded biodegradable polymers for tissue reconstruction. Mater. Res. Soc. Symp. Proc. 252: 359–365.
Kato Y., Iwamoto M., Koike T. and Suzuki F. 1987. Effect of vanadate on cartilage-matrix proteoglycan synthesis in rabbit costal chondrocyte cultures. J. Cell. Biol. 104: 311–319.
Kimelman D., Abraham J.A., Haaparanta T., Palisi T.M. and Kirschner M.W. 1988. The presence of fibroblast growth factor in the frog egg: its role as a natural mesoderm inducer. Science 242: 1053–1056.
Kistler A., Tsuchiya T., Tsuchiya M. and Klaus M. 1990. Teratogenicity of arotinoids (retinoids) in vivoand in vitro. Arch. Toxicol. 64: 616–622.
Kireeva M.L., MO F.E., Yang G.P. and Lau L.F. 1996. Cyr61, a product of a growth factor-inducible immediate-early gene, promotes cell proliferation, migration, and adhesion. Mol. Cell. Biol. 16: 1326–1334.
Kawasaki K., Ochi M., Uchio Y., Adachi N. and Matsusaki M. 1999. Hyaluronic acid enhances proliferation and chondroitin sulfate synthesis in cultured chondrocytes embedded in collagen gels. J. Cell. Physiol. 179: 142–148.
Lee J.D., Hwang O., Kim S.W. and Han S. 1977. Primary cultured chondrocytes of different origins respond differently to bFGF and TGF-beta. Life Sci. 61: 293–299.
Larsen N.E., Lombard K.M., Parent E.G. and Balazs E.A. 1992. Effect of hylan on cartilage and chondrocyte cultures. J. Orthop. Res. 10: 23–32.
Mancilla E.E., De Luca F., Uyeda J.A., Czerwiec F.S. and Baron J. 1998. Effects of fibroblast growth factor-2 on longitudinal bone growth. Endocrinology 139: 2900–2904.
Matsusaki M., Ochi M., Uchio Y., Shu N., Kurioka H., Kawasaki K. et al. 1998. Effects of basic fibroblast growth factor on proliferation and phenotype expression of chondrocytes embedded in collagen gel. Gen. Pharmacol. 31: 759–764.
Panagakos F.S. 1994. Transforming growth factor- alpha stimulates chemotaxis of osteoblasts and osteoblast-like cells in vitro. Biochem. Mol. Biol. Int. 33: 643–650.
Pizette S. and Niswander L. 2000. BMPs are required at two steps of limb chondrogenesis: formation of prechondrogenic condensations and their differentiation into chondrocytes. Dev. Biol. 219: 237–249.
Quarto R., Campanile G., Cancedda R. and Dozin B. 1992. Thyroid hormone, insulin, and glucocorticoids are sufficient to support chondrocyte differentiation to hypertrophy: a serum-free analysis. J. Cell. Biol. 119: 989–995.
Quatela V.C., Sherris D.A. and Rosier R.N. 1993. The human auricular chondrocyte. Responses to growth factors. Arch. Otolaryngol. Head Neck Surg. 119: 32–37.
Redini F., Daireaux M., Mauviel A., Galera P., Loyau G. and Pujol J.P. 1991. Characterization of proteoglycans synthesized by rabbit articular chondrocytes in response to transforming growth factor-beta (TGF-beta). Biochim. Biophys. Acta. 1093: 196–206.
Stevens R.L. and Hascall V.C. 1981. Characterization of proteoglycans synthesized by rat chondrosarcoma chondrocytes treated with multiplication-stimulating activity and insulin. J. Biol. Chem. 256: 2053–2058.
Slack J.M.W., Darlington B.G., Heath J.K. and Godsave S.F. 1987. Mesoderm induction in early Xenopus embryos by heparin binding growth factors. Nature 326: 197–200.
Shimazu A., Jikko A., Iwamoto M., Koike T., Yan W., Okada Y. et al. 1993. Effects of hyaluronic acid on the release of proteoglycan from the cell matrix in rabbit chondrocyte cultures in the presence and absence of cytokines. Arthritis Rheum. 36: 247–253.
Sugiura N., Sakurai K., Hori Y., Karasawa K., Suzuki S. and Kimata K. 1993. Preparation of lipid-derivatized glycosaminoglycans to probe a regulatory function of the carbohydrate moieties of proteoglycans in cell-matrix interaction. J. Biol. Chem. 268: 15779–15787.
Seko Y., Tanaka Y. and Tokoro T. 1995. Influence of bFGF as a potent growth stimulator and TGF-beta as a growth regulator on scleral chondrocytes and scleral fibroblasts in vitro. Ophthalmic Res. 27: 144–152.
Toolan B.C., Frenkel S.R., Pachence J.M., Yalowitz L. and Alexander H. 1996. Effects of growth-factor-enhanced culture on a chondrocyte-collagen implant for cartilage repair. J. Biomed Mater Res. 31: 273–280.
Wiebkin O.W. and Muir H. 1975. Influence of the cells on the pericellular environment. The effect of hyaluronic acid on proteoglycan synthesis and secretion by chondrocytes of adult cartilage. Philos. Trans. R. Soc. Lond. B. Biol. Sci. 271: 283–291.
Wataha J.C., Lockwood P.E. and Schedle A. 2000. Effect of silver, copper, mercury, and nickel ions on cellular proliferation during extended, low-dose exposures. J. Biomed. Mater. Res. 52: 360–364.
Yamagata M., Suzuki S., Akiyama S.K., Yamada K.M. and Kimata K. 1989. Regulation of cell-substrate adhesion by proteoglycans immobilized on extracellular substrates. J. Biol. Chem. 264: 8012–8018.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Rahman, M.S., Tsuchiya, T. In vitro culture of human chondrocytes (1): A novel enhancement action of ferrous sulfate on the differentiation of human chondrocytes. Cytotechnology 37, 163–169 (2001). https://doi.org/10.1023/A:1020506821201
Issue Date:
DOI: https://doi.org/10.1023/A:1020506821201