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Effects of Initial Cell Seeding in Self Assembly of Articular Cartilage

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Abstract

Current forays into tissue engineering of articular cartilage in vitro using the self-assembling method have produced constructs possessing significant extracellular matrix and resulting mechanical properties. However, large numbers of native articular chondrocytes are necessary to produce functional engineered cartilage; all previous work with the self-assembling process has used 5.5 × 106 cells/construct. In this study, the effects of initial cell seeding (0.25–11 × 106 cells/construct) on tissue quality were investigated. Results showed that tissue engineered articular cartilage was formed, when using at least 2 million cells/construct, possessing dimensional, compositional, and compressive properties approaching those of native tissue. It was noted that higher seeding contributed to thicker constructs with larger diameters and had a significant effect on resulting biochemical and biomechanical properties. It was further observed that aggregate modulus increased with increased seeding. By combining gross morphological, histological, biochemical, and biomechanical results, an optimal initial seeding for the self-assembling process of 3.75 × 106 cells/construct was identified. This finding enhances the translatability of this tissue engineering process by reducing the number of cells needed for tissue engineering of articular cartilage by 32% while maintaining essential tissue properties.

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References

  1. Almarza A. J., K. A. Athanasiou 2005 Effects of initial cell seeding density for the tissue engineering of the temporomandibular joint disc. Ann. Biomed. Eng. 33, 943–950. doi:10.1007/s10439-005-3311-8

    Article  PubMed  Google Scholar 

  2. Ateshian G. A., C. T. Hung 2003 Functional properties of native articular cartilage. In: F. Guilak, D. L. Butler, S. A. Goldstein (edited by) Functional Tissue Engineering. New York: Springer-Verlag, pp. 46–68

    Chapter  Google Scholar 

  3. Athanasiou, K. A., A. Agarwal, and F. J. Dzida. Comparative study of the intrinsic mechanical properties of the human acetabular and femoral head cartilage. J. Orthop. Res. 12:340–349, 1994. doi:10.1002/jor.1100120306

    Google Scholar 

  4. Athanasiou, K. A., A. Agarwal, A. Muffoletto, F. J. Dzida, G. Constantinides, and M. Clem. Biomechanical properties of hip cartilage in experimental animal models [published erratum appears in Clin. Orthop. 1995 Nov;(320):283]. Clin. Orthop. 254–266, 1995

  5. Athanasiou K. A., D. Korvick, R. C. Schenck 1997 Biodegradable implants for the treatment of osteochondral defects in a goat model. Tissue Eng. 3, 363–373. doi:10.1089/ten.1997.3.363

    Article  Google Scholar 

  6. Athanasiou K. A., G. G. Niederauer, C. M. Agrawal 1996 Sterilization, toxicity, biocompatibility and clinical applications of polylactic acid/polyglycolic acid copolymers. Biomaterials. 17, 93–102. doi:10.1016/0142-9612(96)85754-1

    Article  PubMed  CAS  Google Scholar 

  7. Bernardo M. E., J. A. Emons, M. Karperien, A. J. Nauta, R. Willemze, H. Roelofs, S. Romeo, A. Marchini, G. A. Rappold, S. Vukicevic, F. Locatelli, W. E. Fibbe 2007 Human mesenchymal stem cells derived from bone marrow display a better chondrogenic differentiation compared with other sources. Connect. Tissue Res. 48, 132–140. doi:10.1080/03008200701228464

    Article  PubMed  CAS  Google Scholar 

  8. Brittberg M., A. Lindahl, A. Nilsson, C. Ohlsson, O. Isaksson, L. Peterson 1994 Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation. N. Engl. J. Med. 331, 889–895. doi:10.1056/NEJM199410063311401

    Article  PubMed  CAS  Google Scholar 

  9. Buschmann M. D., Y. A. Gluzband, A. J. Grodzinsky, E. B. Hunziker 1995 Mechanical compression modulates matrix biosynthesis in chondrocyte/agarose culture. J. Cell Sci. 108, 1497–1508

    PubMed  Google Scholar 

  10. Buschmann M. D., Y. A. Gluzband, A. J. Grodzinsky, J. H. Kimura, E. B. Hunziker 1992 Chondrocytes in agarose culture synthesize a mechanically functional extracellular matrix. J. Orthop. Res. 10, 745–758. doi:10.1002/jor.1100100602

    Article  PubMed  CAS  Google Scholar 

  11. Darling E. M., K. A. Athanasiou 2003 Biomechanical strategies for articular cartilage regeneration. Ann. Biomed. Eng. 31, 1114–1124. doi:10.1114/1.1603752

    Article  PubMed  Google Scholar 

  12. Elder B. D., K. A. Athanasiou 2008 Effects of confinement on the mechanical properties of self-assembled articular cartilage constructs in the direction orthogonal to the confinement surface. J. Orthop. Res. 26, 238–246. doi:10.1002/jor.20480

    Article  PubMed  Google Scholar 

  13. Foty R. A., C. M. Pfleger, G. Forgacs, M. S. Steinberg 1996 Surface tensions of embryonic tissues predict their mutual envelopment behavior. Development 122, 1611–1620

    PubMed  CAS  Google Scholar 

  14. Foty R. A., M. S. Steinberg 2005 The differential adhesion hypothesis: a direct evaluation. Dev. Biol. 278, 255–263. doi:10.1016/j.ydbio.2004.11.012

    Article  PubMed  Google Scholar 

  15. Freed L. E., A. P. Hollander, I. Martin, J. R. Barry, R. Langer, G. Vunjak-Novakovic 1998 Chondrogenesis in a cell-polymer-bioreactor system. Exp. Cell Res. 240, 58–65. doi:10.1006/excr.1998.4010

    Article  PubMed  CAS  Google Scholar 

  16. Freed L. E., I. Martin, and G. Vunjak-Novakovic. Frontiers in tissue engineering. In vitro modulation of chondrogenesis. Clin. Orthop. S46–S58, 1999. doi:10.1097/00003086-199910001-00006

  17. Freed L. E., G. Vunjak-Novakovic, R. Langer 1993 Cultivation of cell-polymer cartilage implants in bioreactors. J. Cell. Biochem. 51, 257–264. doi:10.1002/jcb.240510304

    Article  PubMed  CAS  Google Scholar 

  18. Glowacki J., K. E. Yates, R. Maclean, S. Mizuno 2005 In vitro engineering of cartilage: effects of serum substitutes, TGF-beta, and IL-1alpha. Orthod. Craniofac. Res. 8, 200–208. doi:10.1111/j.1601-6343.2005.00333.x

    Article  PubMed  CAS  Google Scholar 

  19. Gooch K. J., T. Blunk, D. L. Courter, A. L. Sieminski, P. M. Bursac, G. Vunjak-Novakovic, L. E. Freed 2001 IGF-I and mechanical environment interact to modulate engineered cartilage development. Biochem. Biophys. Res. Commun. 286, 909–915. doi:10.1006/bbrc.2001.5486

    Article  PubMed  CAS  Google Scholar 

  20. Heywood H. K., P. K. Sembi, D. A. Lee, D. L. Bader 2004 Cellular utilization determines viability and matrix distribution profiles in chondrocyte-seeded alginate constructs. Tissue Eng. 10, 1467–1479

    PubMed  Google Scholar 

  21. Hu J. C. Y., K. A. Athanasiou 2003 Chapter 4: structure and function of articular cartilage. In: Y. H. An K. L. Martin (edited by) Handbook of Histology Methods for Bone and Cartilage, Totowa, NJ: Humana Press, pp. 73–95

    Chapter  Google Scholar 

  22. Hu J. C., K. A. Athanasiou 2006 A self-assembling process in articular cartilage tissue engineering. Tissue Eng. 12, 969–979. doi:10.1089/ten.2006.12.969

    Article  PubMed  Google Scholar 

  23. Hung C. T., R. L. Mauck, C. C. Wang, E. G. Lima, G. A. Ateshian 2004 A paradigm for functional tissue engineering of articular cartilage via applied physiologic deformational loading. Ann. Biomed. Eng. 32, 35–49. doi:10.1023/B:ABME.0000007789.99565.42

    Article  PubMed  Google Scholar 

  24. Hunziker E. B. 1999 Articular cartilage repair: are the intrinsic biological constraints undermining this process insuperable? Osteoarthritis Cartilage 7, 15–28. doi:10.1053/joca.1998.0159

    Article  PubMed  CAS  Google Scholar 

  25. Jadin K. D., B. L. Wong, W. C. Bae, K. W. Li, A. K. Williamson, B. L. Schumacher, J. H. Price, R. L. Sah 2005 Depth-varying density and organization of chondrocytes in immature and mature bovine articular cartilage assessed by 3d imaging and analysis. J. Histochem. Cytochem. 53, 1109–1119. doi:10.1369/jhc.4A6511.2005

    Article  PubMed  Google Scholar 

  26. Klein T. J., B. L. Schumacher, T. A. Schmidt, K. W. Li, M. S. Voegtline, K. Masuda, E. J. Thonar, R. L. Sah 2003 Tissue engineering of stratified articular cartilage from chondrocyte subpopulations. Osteoarthritis Cartilage 11, 595–602. doi:10.1016/S1063-4584(03)00090-6

    Article  PubMed  CAS  Google Scholar 

  27. Koay E. J., G. M. Hoben, K. A. Athanasiou 2007 Tissue engineering with chondrogenically differentiated human embryonic stem cells. Stem Cells 25, 2183–2190. doi:10.1634/stemcells.2007-0105

    Article  PubMed  CAS  Google Scholar 

  28. Litzke L. E., E. Wagner, W. Baumgaertner, U. Hetzel, O. Josimovic-Alasevic, J. Libera 2004 Repair of extensive articular cartilage defects in horses by autologous chondrocyte transplantation. Ann. Biomed. Eng. 32, 57–69. doi:10.1023/B:ABME.0000007791.81433.1a

    Article  PubMed  CAS  Google Scholar 

  29. Mauck R. L., S. B. Nicoll, S. L. Seyhan, G. A. Ateshian, C. T. Hung 2003 Synergistic action of growth factors and dynamic loading for articular cartilage tissue engineering. Tissue Eng. 9, 597–611. doi:10.1089/107632703768247304

    Article  PubMed  Google Scholar 

  30. Mauck R. L., S. L. Seyhan, G. A. Ateshian, C. T. Hung 2002 Influence of seeding density and dynamic deformational loading on the developing structure/function relationships of chondrocyte-seeded agarose hydrogels. Ann. Biomed. Eng. 30, 1046–1056. doi:10.1114/1.1512676

    Article  PubMed  Google Scholar 

  31. Mauck R. L., C. C. Wang, E. S. Oswald, G. A. Ateshian, C. T. Hung 2003 The role of cell seeding density and nutrient supply for articular cartilage tissue engineering with deformational loading. Osteoarthritis Cartilage 11, 879–890. doi:10.1016/j.joca.2003.08.006

    Article  PubMed  CAS  Google Scholar 

  32. Napolitano A. P., P. Chai, D. M. Dean, J. R. Morgan 2007 Dynamics of the self-assembly of complex cellular aggregates on micromolded nonadhesive hydrogels. Tissue Eng. 13, 2087–2094. doi:10.1089/ten.2006.0190

    Article  PubMed  CAS  Google Scholar 

  33. Pietila K., T. Kantomaa, P. Pirttiniemi, A. Poikela 1999 Comparison of amounts and properties of collagen and proteoglycans in condylar, costal and nasal cartilages. Cells Tissues Organs 164, 30–36. doi:10.1159/000016640

    Article  PubMed  CAS  Google Scholar 

  34. Saini S., T. M. Wick 2003 Concentric cylinder bioreactor for production of tissue engineered cartilage: effect of seeding density and hydrodynamic loading on construct development. Biotechnol. Prog. 19, 510–521. doi:10.1021/bp0256519

    Article  PubMed  Google Scholar 

  35. Scott C. C., K. A. Athanasiou 2006 Design, validation, and utilization of an articular cartilage impact instrument. Proc. Inst. Mech. Eng. [H] 220, 845–855. doi:10.1243/09544119JEIM97

    CAS  Google Scholar 

  36. Steinberg M. S. 1962 Mechanism of tissue reconstruction by dissociated cells. II. Time-course of events. Science. 137, 762–763. doi:10.1126/science.137.3532.762

    Article  PubMed  CAS  Google Scholar 

  37. Varghese S., N. S. Hwang, A. C. Canver, P. Theprungsirikul, D. W. Lin, J. Elisseeff 2008 Chondroitin sulfate based niches for chondrogenic differentiation of mesenchymal stem cells. Matrix Biol. 27, 12–21. doi:10.1016/j.matbio.2007.07.002

    Article  PubMed  CAS  Google Scholar 

  38. Vunjak-Novakovic G., I. Martin, B. Obradovic, S. Treppo, A. J. Grodzinsky, R. Langer, L. E. Freed 1999 Bioreactor cultivation conditions modulate the composition and mechanical properties of tissue-engineered cartilage. J. Orthop. Res. 17, 130–138. doi:10.1002/jor.1100170119

    Article  PubMed  CAS  Google Scholar 

  39. Vunjak-Novakovic G., B. Obradovic, I. Martin, P. M. Bursac, R. Langer, L. E. Freed 1998 Dynamic cell seeding of polymer scaffolds for cartilage tissue engineering. Biotechnol. Prog. 14, 193–202. doi:10.1021/bp970120j

    Article  PubMed  CAS  Google Scholar 

  40. Wang Y., D. J. Blasioli, H. J. Kim, H. S. Kim, D. L. Kaplan 2006 Cartilage tissue engineering with silk scaffolds and human articular chondrocytes. Biomaterials 27, 4434–4442. doi:10.1016/j.biomaterials.2006.03.050

    Article  PubMed  Google Scholar 

  41. Williams C. G., T. K. Kim, A. Taboas, A. Malik, P. Manson, J. Elisseeff 2003 In vitro chondrogenesis of bone marrow-derived mesenchymal stem cells in a photopolymerizing hydrogel. Tissue Eng. 9, 679–688. doi:10.1089/107632703768247377

    Article  PubMed  Google Scholar 

  42. Woessner J. F. Jr. 1961 The determination of hydroxyproline in tissue and protein samples containing small proportions of this imino acid. Arch. Biochem. Biophys. 93, 440–447. doi:10.1016/0003-9861(61)90291-0

    Article  PubMed  CAS  Google Scholar 

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Acknowledgment

The authors would like to gratefully acknowledge funding support from NIAMS RO1 AR053286.

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Authors and Affiliations

  1. Department of Bioengineering, Rice University, 6100 Main Street, Keck Hall, Suite 116, Houston, TX, 77005, USA

    Christopher M. Revell & Kyriacos A. Athanasiou

  2. St. John’s School, 2401 Claremont Lane, Houston, TX, 77019, USA

    Catherine E. Reynolds

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Correspondence to Kyriacos A. Athanasiou.

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Revell, C.M., Reynolds, C.E. & Athanasiou, K.A. Effects of Initial Cell Seeding in Self Assembly of Articular Cartilage. Ann Biomed Eng 36, 1441–1448 (2008). https://doi.org/10.1007/s10439-008-9524-x

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