Encapsulation of Human Articular Chondrocytes into 3D Hydrogel: Phenotype and Genotype Characterization
This chapter is intended to provide a summary of the current materials used in cell encapsulation technology as well as methods for evaluating the performance of cells encapsulated in a polymeric matrix. In particular, it describes the experimental procedure to prepare a hydrogel matrix based on natural polymers for encapsulating and culturing human articular chondrocytes with the interest in cartilage regeneration. Protocols to evaluate the viability, proliferation, differentiation, and matrix production of embedded cells are also described and include standard protocols such as the MTT and [3H] Thymidine assays, reverse transcription polymerase chain reaction (RT-PCR) technique, histology, and immunohistochemistry analysis. The assessment of cell distribution within the 3D hydrogel construct is also described using APoTome analysis.
Key wordsNatural polymers Hydrogels Gelation Chondrocyte encapsulation Cell culture and survival within 3D matrix Cell-based technology Cartilage regeneration
The authors would like to thank Recco Orthopedic staff members for helpful discussions, particularly to Dr. Michele Grandizio; patients for biopsy material donation, as well as to Mrs Daniela Marubbi and Monica Scaranari for technical assistance with histological sample processing. This work was supported by funds from the Italian MUR (FIRB-Tissuenet project), the European Union funded STREP Project HIPPOCRATES (NMP3-CT-2003-505758), and the European NoE EXPERTISSUES (NMP3-CT-2004-500283).
- 14.Pereira, R.C., Scaranari, M., Castagnola, P., Grandizio, M., Azevedo, H.S., Reis, R.L., Cancedda, R., and Gentili, C. (2009) Novel injectable gel (system) as a vehicle for human articular chondrocytes in cartilage tissue regeneration. J. Tissue Eng. Regen. Med. 3, 97–106.PubMedCrossRefGoogle Scholar