Abstract
The purpose of the presented work is to examine the response of engineered cartilage to a transient, 2-week application of anabolic growth factors compared to continuous exposure in in vitro culture. Immature bovine chondrocytes were suspended in agarose hydrogel and cultured for 28 days (Study 1) or 42 days (Study 2) in chondrogenic media with TGF-β1, TGF-β3, or IGF-I either added for only the first 14 days in culture or added to the media for the entire study period. In both studies, there were no statistical differences in tissue mechanical or biochemical properties between the growth factors on day 14. In Study 1, growth factor removal led to a significant and drastic increase in Young’s modulus and glycosaminoglycans content compared to continuously exposed controls on day 28. In Study 2, both TGF-β1 and β3 led to significantly higher mechanical properties and collagen content vs. IGF-I on day 42. These results indicate that the rapid rise in tissue properties (previously observed with TGF-β3 only) is not dependent on the type but rather the temporal application of the anabolic growth factor. These findings shed light on possible techniques to rapidly develop engineered cartilage tissue for the future treatment of osteoarthritis.
Similar content being viewed by others
References
Asanbaeva, A., K. Masuda, E. J. Thonar, S. M. Klisch, and R. L. Sah. Regulation of immature cartilage growth by IGF-I, TGF-beta1, BMP-7, and PDGF-AB: role of metabolic balance between fixed charge and collagen network. Biomech. Model. Mechanobiol. 7:263–276, 2008.
Ashcroft, G. S., M. A. Horan, and M. W. Ferguson. The effects of ageing on wound healing: immunolocalisation of growth factors and their receptors in a murine incisional model. J. Anat. 190(Pt 3):351–365, 1997.
Benya, P. D., and J. D. Shaffer. Dedifferentiated chondrocytes reexpress the differentiated collagen phenotype when cultured in agarose gels. Cell 30:215–224, 1982.
Bian, L. M., S. L. Angione, E. G. Lima, K. W. Ng, G. A. Ateshian, and C. T. Hung. Tissue-engineered cartilage constructs using mature bovine chondrocytes: effects of temporal exposure to growth factors and dynamic deformational loading. Trans. Orthop. Res. Soc. 32:1480, 2007
Blunk, T., A. L. Sieminski, K. J. Gooch, D. L. Courter, A. P. Hollander, A. M. Nahir, R. Langer, G. Vunjak-Novakovic, and L. E. Freed. Differential effects of growth factors on tissue-engineered cartilage. Tissue Eng. 8:73–84, 2002.
Bos, P. K., G. J. van Osch, D. A. Frenz, J. A. Verhaar, and H. L. Verwoerd-Verhoef. Growth factor expression in cartilage wound healing: temporal and spatial immunolocalization in a rabbit auricular cartilage wound model. Osteoarthr. Cartil. 9:382–389, 2001.
Brittberg, M., A. Lindahl, A. Nilsson, C. Ohlsson, O. Isaksson, and L. Peterson. Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation. N. Engl. J. Med. 331:889–895, 1994.
Buschmann, M. D., Y. A. Gluzband, A. J. Grodzinsky, J. H. Kimura, and E. B. Hunziker. Chondrocytes in agarose culture synthesize a mechanically functional extracellular matrix. J. Orthop. Res. 10:745–758, 1992.
Byers, B. A., R. L. Mauck, I. E. Chiang, and R. S. Tuan. Transient exposure to transforming growth factor beta 3 under serum-free conditions enhances the biomechanical and biochemical maturation of tissue-engineered cartilage. Tissue Eng. A 14:1821–1834, 2008.
Farndale, R. W., C. A. Sayers, and A. J. Barrett. A direct spectrophotometric microassay for sulfated glycosaminoglycans in cartilage cultures. Connect. Tissue Res. 9:247–248, 1982.
Felson, D. T., R. C. Lawrence, P. A. Dieppe, R. Hirsch, C. G. Helmick, J. M. Jordan, R. S. Kington, N. E. Lane, M. C. Nevitt, Y. Zhang, M. Sowers, T. McAlindon, T. D. Spector, A. R. Poole, S. Z. Yanovski, G. Ateshian, L. Sharma, J. A. Buckwalter, K. D. Brandt, and J. F. Fries. Osteoarthritis: new insights. Part 1. The disease and its risk factors. Ann. Intern. Med. 133:635–646, 2000.
Goldring, M. B., K. Tsuchimochi, and K. Ijiri. The control of chondrogenesis. J. Cell. Biochem. 97:33–44, 2006.
Gooch, K. J., T. Blunk, D. L. Courter, A. L. Sieminski, P. M. Bursac, G. Vunjak-Novakovic, and L. E. Freed. IGF-I and mechanical environment interact to modulate engineered cartilage development. Biochem. Biophys. Res. Commun. 286:909–915, 2001.
Guilak, F., D. L. Butler, and S. A. Goldstein. Functional tissue engineering: the role of biomechanics in articular cartilage repair. Clin. Orthop. S295–S305, 2001.
Hangody, L., and P. Fules. Autologous osteochondral mosaicplasty for the treatment of full-thickness defects of weight-bearing joints: ten years of experimental and clinical experience. J. Bone Joint Surg. Am. 85-A(Suppl. 2):25–32, 2003.
Hollander, A. P., T. F. Heathfield, C. Webber, Y. Iwata, R. Bourne, C. Rorabeck, and A. R. Poole. Increased damage to type II collagen in osteoarthritic articular cartilage detected by a new immunoassay. J. Clin. Invest. 93:1722–1732, 1994.
Honn, K. V., J. A. Singley, and W. Chavin. Fetal bovine serum: a multivariate standard. Proc. Soc. Exp. Biol. Med. 149:344–347, 1975.
Huang, C. Y., M. A. Soltz, M. Kopacz, V. C. Mow, and G. A. Ateshian. Experimental verification of the roles of intrinsic matrix viscoelasticity and tension-compression nonlinearity in the biphasic response of cartilage. J. Biomech. Eng. 125:84–93, 2003.
Kalpakci, K. N., E. J. Kim, and K. A. Athanasiou. Assessment of growth factor treatment on fibrochondrocyte and chondrocyte co-cultures for TMJ fibrocartilage engineering. Acta Biomater. 7:1710–1718, 2011.
Kempson, G. E. Age-related changes in the tensile properties of human articular cartilage: a comparative study between the femoral head of the hip joint and the talus of the ankle joint. Biochim. Biophys. Acta 1075:223–230, 1991.
Kim, M., X. Bi, W. E. Horton, R. G. Spencer, and N. P. Camacho. Fourier transform infrared imaging spectroscopic analysis of tissue engineered cartilage: histologic and biochemical correlations. J. Biomed. Opt. 10:031105, 2005.
Lima, E. G., L. Bian, K. W. Ng, R. L. Mauck, B. A. Byers, R. S. Tuan, G. A. Ateshian, and C. T. Hung. The beneficial effect of delayed compressive loading on tissue-engineered cartilage constructs cultured with TGF-beta3. Osteoarthr. Cartil. 15:1025–1033, 2007.
Lima, E. G., A. R. Tan, T. Tai, L. Bian, A. M. Stoker, G. A. Ateshian, J. L. Cook, and C. T. Hung. Differences in interleukin-1 response between engineered and native cartilage. Tissue Eng. A 14:1721–1730, 2008.
Mankin, H. J., V. C. Mow, J. A. Buckwalter, J. P. Iannotti, and A. Ratcliffe. Articular cartilage structure, composition, and function. In: Orthopaedic Basic Science: Biology and Biomechanics of the Musculoskeletal System, edited by J. A. Buckwalter, T. A. Einhorn, and S. R. Simon. Rosemont, IL: American Academy of Orthopaedic Surgeons, 2000, pp. 443–470.
Mauck, R. L., S. B. Nicoll, S. L. Seyhan, G. A. Ateshian, and C. T. Hung. Synergistic action of growth factors and dynamic loading for articular cartilage tissue engineering. Tissue Eng. 9:597–611, 2003.
Mauck, R. L., M. A. Soltz, C. C. Wang, D. D. Wong, P. H. Chao, W. B. Valhmu, C. T. Hung, and G. A. Ateshian. Functional tissue engineering of articular cartilage through dynamic loading of chondrocyte-seeded agarose gels. J. Biomech. Eng. 122:252–260, 2000.
Mauck, R. L., C. C. Wang, E. S. Oswald, G. A. Ateshian, and C. T. Hung. The role of cell seeding density and nutrient supply for articular cartilage tissue engineering with deformational loading. Osteoarthr. Cartil. 11:879–890, 2003.
Mithoefer, K., T. McAdams, R. J. Williams, P. C. Kreuz, and B. R. Mandelbaum. Clinical efficacy of the microfracture technique for articular cartilage repair in the knee: an evidence-based systematic analysis. Am. J. Sports Med. 37:2053–2063, 2009.
Moseley, J. B., Jr., A. F. Anderson, J. E. Browne, B. Mandelbaum, L. J. Micheli, F. Fu, and C. Erggelet. Long-term durability of autologous chondrocyte implantation: a multicenter, observational study in U.S. patients. Am. J. Sports Med. 38:238–246, 2010.
Mow, V. C., G. A. Ateshian, and A. Ratcliffe. Anatomic form and biomechanical properties of articular cartilage of the knee joint. In: Biology and Biomechanics of the Traumatized Synovial Joint. The Knee as a Model, edited by G. A. M. Finerman, and F. R. Noyes. Rosemont, IL: AAOS, 1992, pp. 55–81.
Ng, K. W., J. G. DeFrancis, L. E. Kugler, T. A. Kelly, M. M. Ho, C. J. O’Conor, G. A. Ateshian, and C. T. Hung. Amino acids supply in culture media is not a limiting factor in the matrix synthesis of engineered cartilage tissue. Amino Acids 35:433–438, 2008.
Ng, K. W., E. G. Lima, L. Bian, C. J. O’Conor, P. S. Jayabalan, A. M. Stoker, K. Kuroki, C. R. Cook, G. A. Ateshian, J. L. Cook, and C. T. Hung. Passaged adult chondrocytes can form engineered cartilage with functional mechanical properties: a canine model. Tissue Eng. A 16:1041–1051, 2010.
Ng, K. W., C. J. O’Conor, E. G. Lima, S. B. Lo, G. A. Ateshian, J. L. Cook, and C. T. Hung. Primed mature canine chondrocytes can develop an engineered cartilage tissue with physiologic properties. Trans. Orthop. Res. Soc. 31:599, 2008.
Ng, K. W., C. C. Wang, R. L. Mauck, T. A. Kelly, N. O. Chahine, K. D. Costa, G. A. Ateshian, and C. T. Hung. A layered agarose approach to fabricate depth-dependent inhomogeneity in chondrocyte-seeded constructs. J. Orthop. Res. 23:134–141, 2005.
Pei, M., J. Seidel, G. Vunjak-Novakovic, and L. E. Freed. Growth factors for sequential cellular de- and re-differentiation in tissue engineering. Biochem. Biophys. Res. Commun. 294:149–154, 2002.
Price, P. J., and E. A. Gregory. Relationship between in vitro growth promotion and biophysical and biochemical properties of the serum supplement. In Vitro 18:576–584, 1982.
Riesle, J., A. P. Hollander, R. Langer, L. E. Freed, and G. Vunjak-Novakovic. Collagen in tissue-engineered cartilage: types, structure, and crosslinks. J. Cell. Biochem. 71:313–327, 1998.
Scuderi, G. R., and J. N. Insall. Total knee arthroplasty. Current clinical perspectives. Clin. Orthop. Relat. Res. 276:26–32, 1992.
Selmi, T. A., P. Verdonk, P. Chambat, F. Dubrana, J. F. Potel, L. Barnouin, and P. Neyret. Autologous chondrocyte implantation in a novel alginate-agarose hydrogel: outcome at two years. J. Bone Joint Surg. Br. 90:597–604, 2008.
Shah, M., D. M. Foreman, and M. W. Ferguson. Neutralisation of TGF-beta 1 and TGF-beta 2 or exogenous addition of TGF-beta 3 to cutaneous rat wounds reduces scarring. J. Cell Sci. 108(Pt 3):985–1002, 1995.
Slover, J. D., and H. E. Rubash. Hip resurfacing arthroplasty: time to consider it again? Instr. Course Lect. 57:267–271, 2008.
Solheim, E., J. Hegna, J. Oyen, O. K. Austgulen, T. Harlem, and T. Strand. Osteochondral autografting (mosaicplasty) in articular cartilage defects in the knee: results at 5 to 9 years. Knee 17:84–87, 2010.
Steadman, J. R., W. G. Rodkey, and J. J. Rodrigo. Microfracture: surgical technique and rehabilitation to treat chondral defects. Clin. Orthop. Relat. Res. S362–S369, 2001.
Stegemann, H., and K. Stalder. Determination of hydroxyproline. Clin. Chim. Acta 18:267–273, 1967.
Theoret, C. L., S. M. Barber, T. N. Moyana, and J. R. Gordon. Expression of transforming growth factor beta(1), beta(3), and basic fibroblast growth factor in full-thickness skin wounds of equine limbs and thorax. Vet. Surg. 30:269–277, 2001.
van Osch, G. J., W. B. van den Berg, E. B. Hunziker, and H. J. Hauselmann. Differential effects of IGF-1 and TGF beta-2 on the assembly of proteoglycans in pericellular and territorial matrix by cultured bovine articular chondrocytes. Osteoarthr. Cartil. 6:187–195, 1998.
van Osch, G. J., S. W. van der Veen, P. Buma, and H. L. Verwoerd-Verhoef. Effect of transforming growth factor-beta on proteoglycan synthesis by chondrocytes in relation to differentiation stage and the presence of pericellular matrix. Matrix Biol. 17:413–424, 1998.
Weisser, J., B. Rahfoth, A. Timmermann, T. Aigner, R. Brauer, and K. von der Mark. Role of growth factors in rabbit articular cartilage repair by chondrocytes in agarose. Osteoarthr. Cartil. 9(Suppl. A):S48–S54, 2001.
Williams, G. M., K. J. Dills, C. R. Flores, M. E. Stender, K. M. Stewart, L. M. Nelson, A. C. Chen, K. Masuda, S. J. Hazelwood, S. M. Klisch, and R. L. Sah. Differential regulation of immature articular cartilage compressive moduli and Poisson’s ratios by in vitro stimulation with IGF-1 and TGF-beta1. J. Biomech. 43:2501–2507, 2010.
Acknowledgments
Research funding was provided by the National Institutes of Health (AR46568, AR52871, and AR53530).
Author information
Authors and Affiliations
Corresponding author
Additional information
Associate Editor Kent Leach oversaw the review of this article.
Rights and permissions
About this article
Cite this article
Ng, K.W., O’Conor, C.J., Kugler, L.E. et al. Transient Supplementation of Anabolic Growth Factors Rapidly Stimulates Matrix Synthesis in Engineered Cartilage. Ann Biomed Eng 39, 2491–2500 (2011). https://doi.org/10.1007/s10439-011-0356-8
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10439-011-0356-8