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3R-Project 112-08
A novel in-vitro model for the holistic assessment and optimisation of engineered tissue for functional cartilage repair
Zhijie Luo and Bahaa Seedhom
Leeds Dental Institute, Clarendon Way, Leeds LS9 9LU, UK
medzjl@leeds.ac.uk
Keywords: human; cell cultures: bioreactor; reduction; replacement
Duration: 1.5 years End of the Project: 2011
Background and Aim
Tissue engineering offers potential solutions to address strategies for cartilage repair and regeneration. Animal models are still widely used in evaluation of tissue engineering techniques for the repair of damaged human articular cartilage. However, there is no consensus on the most appropriate animal model and none of the species used replicate the anatomical, cellular and biomechanical properties of human articular cartilage. A further major problem is the poor integration of newly implanted engineered cartilage with the adjacent healthy tissue(1, 2). Without full integration into native cartilage, the implants will experience abnormal mechanical stress during their post-surgical maturation and remodeling, leading to further damage to both new and adjacent tissue and eventually to failure of the repair. Application of mechanical stimulation to engineered tissue has been reported to enhance cartilaginous matrix formation and improve tissue growth(3-5). However, despite reports of improvements in the quality of in-vitro engineered cartilage tissue, there is a lack of information about whether these ‘better quality’ engineered tissue constructs ever achieve successful lateral integration between repair tissue and adjacent cartilage after implantation.
Our goal is to replace the use of animals in cartilage research, which will be achieved by the development and testing of a screening model to undertake in-vitro simulative studies of cartilage repair, with special focus on (a) the sideway integration of the repair tissue and (b) its mechanical properties.
Method and Results
in progress (present status)
We will use a novel implant co-culture model designed to permit a holistic in-vitro evaluation of tissue-engineering techniques intended to provide constructs for cartilage repair. These constructs will be developed in our specially designed bioreactors, and exposed to cyclic compressive loads (CCL) over prolonged periods in culture. The resulting engineered tissue will then be evaluated in a 3D co-culture implant model in simulation of proposed in-vivo repair. The optimal loading regimes will be applied for 30 min. per day, 5 days per week for 12 weeks. Identical samples will be cultured in the bioreactor chamber without application of CCL and used as controls. The maturation of implant tissue will be evaluated by measuring DNA, GAG and collagen content, together with RT-PCR for relevant markers of chondrocyte gene expression. Mechanical properties of the constructs will be determined by measuring the compressive modulus using an unconfined uniaxial compression test. To test the degree of integration of the implant construct with the surrounding cartilage, a ‘push out’ test will be undertaken to determine the strength of adhesion of the implant to the adjacent cartilage. The integration between engineered tissue and surrounding cartilage will be also evaluated histologically.
Conclusions and Relevance for 3R
This in-vitro model could be used for developing new techniques and will hopefully be used as an in-vitro screening strategy for newly developed technologies intended for cartilage repair and so would provide an alternative experimental model to replace current animal experimentation.
References
1. Archer C.W, Redman S, Khan I, Bishop J and Kirsty R (2006) Enhancing tissue integration in cartilage repair procedures. J. Anat. 209, 481-493.
2. Chang C.H, Kuo TF, Lin C. C, Chou C.H, Chen K.H, Lin F.H, Liu H.C. (2006) Tissue engineering-based cartilage repair with allogenous chondrocytes and gelatin-chondroitin-hyaluronan tri-copolymer scaffold: A procine model assessed at 18, 24, and 36 weeks. Biomaterials 27, 1876-1888.
3. Mauck R.L, Soltz M.A., Wang C.C, Wong D.D, Chao P.H, Valhmu W.B., Hung C.T, and Ateshian G.A. (2000) Functional tissue engineering of articular cartilage through dynamic loading of chondrocyte-seeded agarose gels J Biomech Eng 122, 252-60
4. Mauck R.L, Nicoll S. B, Seyhan S. L, Ateshian G. A and Hung C. T. (2003) Synergistic action of growth factors and dynamic loading for articular cartilage tissue engineering Tissue Eng 9(4):597-611.
5. Miyanishi K, Trindade M.C, Lindsey D.P, Beaupre G.S, Carter D.R, Goodman S. B, Schurman D.J, Smith R.L. (2006). Dose- and time-dependent effects of cyclic hydrostatic pressure on transforming growth factor-beta3-induced chondrogenesis by adult human mesenchymal stem cells in vitro. Tissue Eng 12(8): 2253-2262
| Dernières modifications: 29.08.2010 |  |