Abstract
In stem cell-based chondrogenesis for articular cartilage regeneration, TGF-β3 is dosed to the stem cells to drive differentiation into chondrocytic cells. Meanwhile, type I collagen, which is endogenously expressed in some stem cells (e.g., synovium-derived mesenchymal stem cells) and upregulated by TGF-β3, poses a threat to chondrogenesis, as type I collagen may alter the components and stiffness of articular cartilage. Therefore, a wiser strategy would be to feed the cells with TGF-β3 while at the same time silencing the expression of type I collagen. In this chapter, methods for construction of adenoviral vectors and lentiviral vectors having both of the above functions are given. Their transduction into synovium-derived mesenchymal stem cells for articular cartilage engineering and following characterizations are also described.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Flugge LA, Miller-Deist LA, Petillo PA (1999) Towards a molecular understanding of arthritis. Chem Biol 6:R157–R166
Bader RMSA (2007) Cartilage tissue engineering and bioreactor systems for the cultivation and stimulation of chondrocytes. Eur Biophys J 36:539–568
Chancellor JV, Hunsche E, de Cruz E, Sarasin FP (2001) Economic evaluation of celecoxib, a new cyclo-oxygenase 2 specific inhibitor, in Switzerland. Pharmacoeconomics 19:59–75
Darling EM, Hu JC, Athanasiou KA (2004) Zonal and topographical differences in articular cartilage gene expression. J Orthop Res 22:1182–1187
Hidaka C, Cheng C, Alexandre D, Bhargava M, Torzilli PA (2006) Maturational differences in superficial and deep zone articular chondrocytes. Cell Tissue Res 323:127–135
Goessler UR, Bugert P, Bieback K, Baisch A, Sadick H, Verse T, Klüter H, Hörmann K, Riedel F (2004) Expression of collagen and fiber-associated proteins in human septal cartilage during in vitro dedifferentiation. Int J Mol Med 14:1015–1022
Goessler UR, Bugert P, Bieback K, Sadick H, Baisch A, Hormann K, Riedel F (2006) In vitro analysis of differential expression of collagens, integrins, and growth factors in cultured human chondrocytes. Otolaryngol Head Neck Surg 134:510–515
Yoshimura H, Muneta T, Nimura A, Yokoyama A, Koga H, Sekiya I (2007) Comparison of rat mesenchymal stem cells derived from bone marrow, synovium, periosteum, adipose tissue, and muscle. Cell Tissue Res 327:449–642
Na K, Kim S, Woo DG, Sun BK, Yang HN, Chung HM, Park KH (2007) Synergistic effect of TGFbeta-3 on chondrogenic differentiation of rabbit chondrocytes in thermo-reversible hydrogel constructs blended with hyaluronic acid by in vivo test. J Biotechnol 128:412–422
Yun K, Moon HT (2008) Inducing chondrogenic differentiation in injectable hydrogels embedded with rabbit chondrocytes and growth factor for neocartilage formation. J Biosci Bioeng 105:122–126
Awad HA, Wickham MQ, Leddy HA, Gimble JM, Guilak F (2004) Chondrogenic differentiation of adipose-derived adult stem cells in agarose, alginate, and gelatin scaffolds. Biomaterials 25:3211–3222
Steinert A, Weber M, Dimmler A, Julius C, Schütze N, Nöth U, Cramer H, Eulert J, Zimmermann U, Hendrich C (2003) Chondrogenic differentiation of mesenchymal progenitor cells encapsulated in ultrahigh-viscosity alginate. J Orthop Res 21:1090–1097
Park H, Kang SW, Kim BS, Mooney DJ, Lee KY (2009) Shear-reversibly crosslinked alginate hydrogels for tissue engineering. Macromol Biosci 9:895–901
Yao Y, Zhang F, Zhou RJ, Li M, Wang DA (2010) Continuous supply of TGFbeta3 via adenoviral vector promotes type I collagen and viability of fibroblasts in alginate hydrogel. J Tissue Eng Regen Med 4:497–504
Zhang F, Yao Y, Hao J, Zhou R, Liu C, Gong Y, Wang DA (2010) A dual-functioning adenoviral vector encoding both transforming growth factor-beta3 and shRNA silencing type I collagen: construction and controlled release for chondrogenesis. J Control Release 142:70–77
Oberholzer A, John T, Kohl B, Gust T, Müller RD, La Face D, Hutchins B, Zreiqat H, Ertel W, Schulze-Tanzil G (2007) Adenoviral transduction is more efficient in alginate-derived chondrocytes than in monolayer chondrocytes. Cell Tissue Res 328:383–390
Zhang F, Yao YC, Zhou RJ, Su K, Citra F, Wang DA (2011) Optimal construction and delivery of dual-functioning lentiviral vectors for type I collagen-suppressed chondrogenesis in synovium-derived mesenchymal stem cells. Pharm Res 28:1338–1348
Wang CM, Hao JH, Zhang F, Su K, Wang DA (2008) RNA extraction from polysaccharide-based cell-laden hydrogel scaffolds. Anal Biochem 380:333–334
Wang CM, Li X, Yao YC, Wang DA (2009) Concurrent extraction of proteins and RNA from cell-laden hydrogel scaffold free of polysaccharide interference. J Chromatogr B 877:3762–3766
Acknowledgements
This work was supported by AcRF Tier 1 RG 36/12 and AcRF Tier 2 ARC 1/13, Ministry of Education, Singapore.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Zhang, F., Wang, DA. (2015). Gene Transfer and Gene Silencing in Stem Cells to Promote Chondrogenesis. In: Doran, P. (eds) Cartilage Tissue Engineering. Methods in Molecular Biology, vol 1340. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2938-2_7
Download citation
DOI: https://doi.org/10.1007/978-1-4939-2938-2_7
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2937-5
Online ISBN: 978-1-4939-2938-2
eBook Packages: Springer Protocols