, Volume 52, Issue 2, pp 99–106 | Cite as

Development of porous collagen beads for chondrocyte culture

  • Tracy A. Tebb
  • Shiao-Wen Tsai
  • Veronica Glattauer
  • Jacinta F. White
  • John A. M. Ramshaw
  • Jerome A. WerkmeisterEmail author
Original Paper


A method for the preparation of bioresorbable collagen beads with an open porous structure is presented. These beads were prepared from collagen-alginate composite beads by removal of the alginate component. These collagen beads were suitable for rapid proliferation of chondrocytes in a dynamic, spinner culture system. Histology and immuno-histology showed that biochemical markers of chondrocytes are present during this cell proliferation, indicating that there was control of phenotype and that cell de-differentiation had not occurred. Unlike other 3-D scaffolds that have been used, the cells were amplified from very low cell densities and were able to proliferate freely without loss of phenotype.


Collagen beads Alginate Cell culture Chondrocytes Tissue engineering 


  1. Bouchet BY, Colon M, Polotsky A, Shikani AH, Hungerford DS, Frondoza CG (2000) β1-integrin expression by human nasal chondrocytes in microcarrier spinner culture. J Biomed Mater Res 52:716–724CrossRefGoogle Scholar
  2. Brittberg M (1999) Autologous chondrocyte transplantation. Clin Orthopaed Rel Res 367S:147–155CrossRefGoogle Scholar
  3. Brittberg M, Lindahl A, Nilsson A, Ohlsson C, Isaksson O, Peterson L (1994) Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation. N Eng J Med 331:879–895CrossRefGoogle Scholar
  4. Buckwalter JA, Mankin HJ (1997) Articular cartilage 2: degeneration and osteoarthrosis, repair, regeneration, and transplantation. J Bone Joint Surg Am 79A:612–632Google Scholar
  5. Caterson B, Christner JE, Baker JR (1983) Identification of a monoclonal antibody that specifically recognizes corneal and skeletal keratan sulfate. J Biol Chem 258:8848–8854Google Scholar
  6. Chapman JA, Hulmes DJS (1984) Electron microscopy of the collagen fibril. In: Ruggeri A, Motta PM (eds) Ultrastructure of the connective tissue matrix. Martinus Nijhoff Pub., Boston, pp 1–33Google Scholar
  7. Chiang H, Kuo T-F, Tsai C-C, Lin M-C, She B-R, Huang Y-Y, Lee H-S, Shieh C-S, Chen M-H, Ramshaw JAM, Werkmeister JA, Tuan RS, Jiang C-C (2005) Repair of porcine articular cartilage defect with autologous chondrocyte transplantation. J Orth Res 23:584–593CrossRefGoogle Scholar
  8. Dean RC, Silver FH, Berg RA (1989) Weighted collagen microsponge for immobilising bioactive materials. US Patent, 4,863,856Google Scholar
  9. Frondoza C, Sohrabi A, Hungerford D (1996) Human chondrocytes proliferate and produce matrix components in microcarrier suspension culture. Biomaterials 17:879–888CrossRefGoogle Scholar
  10. Gilbert J (1998) Current treatment options for the restoration of articular cartilage. Am J Knee Surg 11:42–46Google Scholar
  11. Glattauer V, White JF, Tsai W-B, Tsai C-C, Tebb TA, Werkmeister JA, Ramshaw JAM (2004) Preparation and properties of extra-cellular matrix-based beads for spinner culture. In: Proceedings 28th Annual Scientific Conference of the Matrix Biology Society of Australia & New Zealand, Rottnest Island, Western Australia 2004; s22. ISBN 0 9585892 5 9Google Scholar
  12. Grohn P, Kloch G, Zimmermann U (1997) Collagen-coated Ba2+-alginate microcarriers for the culture of anchorage-dependent mammalian cells. BioTechniques 22:970–975Google Scholar
  13. Henderson I, Francisco R, Oakes B, Cameron J (2005) Autologous chondrocyte implantation for treatment of focal chondral defects of the knee—a clinical, arthroscopic, MRI and histologic evaluation at 2 years. Knee 12:209–216Google Scholar
  14. Hsu FY, Tsai S-W, Wang FF, Wang YJ (2000) The collagen-containing alginate/poly(lysine)/alginate microcapsules. Art Cells Blood Subs Immob Biotech 28:147–154Google Scholar
  15. Hunziker EB (2001) Articular cartilage repair: basic science and clinical progress. A review of the current status and prospects. Osteoarthritis Cartilage 10:432–463CrossRefGoogle Scholar
  16. Iwamoto S, Nakagawa K, Sugiura S, Nakajima M (2002) Preparation of gelatin microbeads with a narrow size distribution using microchannel emulsification. AAPS Pharma Sci Tech 3:E25CrossRefGoogle Scholar
  17. Izuta Y, Ochi M, Adachi N, Deie M, Yamasaki T, Shinomiya R (2005) Meniscal repair using bone marrow-derived mesenchymal stem cells: experimental study using green fluorescent protein transgenic rats. Knee 12:217–223Google Scholar
  18. King D (1936) The healing of semilunar cartilages. J Bone Joint Surg 18:333Google Scholar
  19. Kleinman HR, Klebe R, Martin GR (1981) Role of collagenous matrices in the adhesion and growth of cells. J Cell Biol 88:473–485CrossRefGoogle Scholar
  20. Kwon YJ, Peng CA (2002) Calcium-alginate gel bead cross-linked with gelatin as microcarrier for anchorage-dependent cell culture. BioTechniques 33:212–214Google Scholar
  21. Majmudar G, Bole D, Goldstein SA, Bonadio J (1991) Bone cell culture in a three-dimensional polymer bead stabilizes the differentiated phenotype and provides evidence that osteoblastic cells synthesize type III collagen and fibronectin. J Bone Miner Res 6:869–881CrossRefGoogle Scholar
  22. Malda J, van Blitterswijk CA, Grojec M, Martens DE, Tramper J, Riesle J (2003) Expansion of bovine chondrocytes on microcarriers enhances redifferentiation. Tissue Eng 9:939–948CrossRefGoogle Scholar
  23. Miller EJ, Rhodes RK (1982) Preparation and characterisation of the different types of collagens. Method Enzymol 82:33–64CrossRefGoogle Scholar
  24. Nakata K, Shino K, Hamada M, Mae T, Miyama T, Shinjo H, Horibe S, Tada K, Ochi T, Yoshikawa H (2001) Human meniscus cell. Characterisation of the primary culture and use for tissue engineering. Clin Orthopaed Rel Res 391S:s208–s218CrossRefGoogle Scholar
  25. Overstreet M, Sohrabi A, Polotsky A, Hungerford D, Frondoza CG (2003) Collagen microcarrier spinner culture promotes osteoblast proliferation and synthesis of matrix proteins. In Vitro Cell Dev Biol - Animal 39:228–234CrossRefGoogle Scholar
  26. Poole CA, Ayad S, Gilbert RT (1992) Chondrons from articular cartilage—V. Imunohistochemical evaluation of type VI collagen organization in isolated chondrons by light, confocal and electron microscopy. J Cell Sci 103:1101–1110Google Scholar
  27. Ramshaw JAM, Werkmeister JA, Glattauer V (1995) Collagen-based biomaterials. Biotechnol Gen Eng Rev 13:335–382Google Scholar
  28. Sorrell JM, Caterson B (1989) Detection of age-related changes in the distributions of keratan sulfates and chondroitin sulfates in developing chick limbs; an immunocytochemical study. Development 106:657–663Google Scholar
  29. Temenoff JS, Mikos AG (2000) Review: tissue engineering for regeneration of articular cartilage. Biomaterials 21:431–440CrossRefGoogle Scholar
  30. Thissen H, Chang K-Y, Tebb TA, Tsai W-B, Glattauer V, Ramshaw JAM, Werkmeister JA (2005) Synthetic biodegradable microparticles for articular cartilage tissue engineering. J Biomed Mater Res 77A:590–598CrossRefGoogle Scholar
  31. von der Mark K, Gauss V, von der Mark H, Muller P (1977) Relationship between cell shape and type of collagen synthesised as chondrocytes lose their cartilage phenotype in culture. Nature 267:531–532CrossRefGoogle Scholar
  32. Werkmeister JA, Tebb TA, White JF, Ramshaw JAM (1993) Monoclonal antibodies to type VI collagen demonstrate new tissue augmentation of a collagen-based biomaterial implant. J Histochem Cytochem 41:1701–1706Google Scholar
  33. Werkmeister JA, Tsai W-B, Ramshaw JAM, Thissen HW, Chang K-Y (2002) Methods and devices for tissue repair. Patent WO 02/062357-A1Google Scholar

Copyright information

© Springer Science+Business Media, Inc. 2006

Authors and Affiliations

  • Tracy A. Tebb
    • 1
  • Shiao-Wen Tsai
    • 2
    • 3
  • Veronica Glattauer
    • 1
  • Jacinta F. White
    • 1
  • John A. M. Ramshaw
    • 1
  • Jerome A. Werkmeister
    • 1
    Email author
  1. 1.CSIRO, Molecular and Health TechnologiesClayton SouthAustralia
  2. 2.ITRI, Biomedical Engineering CenterHsinchuTaiwan, R.O.C.
  3. 3.Institute of Biochemical and Biomedical EngineeringChang-Gung UniversityTaoyuanTaiwan, R.O.C.

Personalised recommendations