Abstract
Background
Aiming to construct an osteochondral-like structure, the combination of a β-tricalcium phosphate (βTCP) block with a scaffold-free sheet formed using mesenchymal stem cells (MSCs) was investigated.
Methods
Human bone marrow MSCs in a cell culture insert that was set in a 24-well plate were cultivated using a chondrogenic medium containing dexamethasone, IGF-1, and TGFβ3 for 3 weeks during which a cylindrical βTCP block was put on the sheet at day 1, and the cell sheet construct was harvested. In other experiments, at day 14, the construct was put on a cell sheet that was prepared the day before and cultivated for 3 weeks.
Results
The addition of a βTCP block resulted in a combined osteochondral-like construct and comparable staining intensity by Alcian blue, while the expression levels of the aggrecan and type II collagen genes decreased a little. During the culture with the βTCP block, the expression levels of the aggrecan gene increased monotonically. The increase in the inoculum cell number from 1.86 to 3.72 × 106 cells resulted in marked increases in the thickness of cell sheet parts in the βTCP block and expression levels of the aggrecan and type II collagen genes, while the thickness of cell sheet parts on the βTCP block scarcely changed. On the other hand, the addition of a cell sheet that was prepared a day before to the construct at day 14 resulted in the marked increase in thickness of the cell sheet part on the βTCP block, while the thickness of that in the βTCP block did not increase.
Conclusion
A combined osteochondral-like structure was produced by putting a βTCP block on the sheet of MSC. The thickness of the cell sheet parts in and on the βTCP block could be increased by the increase in inoculum cell number and by providing an additional cell sheet, respectively.
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References
Warren L, Grayson P-H, Grace C, Darja M, David LK, Gordana V-N. Engineering custom-designed osteochondral tissue grafts. Trends Biotechnol. 2008;26:181–9.
Mano JF, Reis RL. Osteochondral defects: present situation and tissue engineering approaches. J Tissue Eng Regen Med. 2007;1:261–73.
Ivan M, Sylvie M, Andrea B, Marcel J, David W. Osteochondral tissue engineering. J Biomech. 2007;40:750–65.
Schaefer D, Martin I, Shastri P, Padera RF, Langer R, Freed LE. In vitro generation of osteochondral composites. Biomaterials. 2000;1:2599–606.
Niyama K, Ide N, Onoue K, Kawaguchi A, Okabe T, Wakitani S, Takagi M. Construction of osteochondral-like tissue graft combining β-tricalcium phosphate block and scaffold-free centrifuged chondrocyte cell sheet. J Orthop Sci. 2011;16:613–21.
Maeda S, Fujitomo T, Okabe T, Wakitani S, Takagi M. Shrinkage-free preparation of scaffold-free cartilage-like disc-shaped cell sheet using human bone marrow mesenchymal stem cells. J Biosci Bioeng. 2011;111:489–92.
Takagi M, Nakamura T, Matsuda C, Hattori T, Wakitani S, Yoshida T. In vitro proliferation of human bone marrow mesenchymal stem cells employing donor serum and basic fibroblast growth factor. Cytotechnology. 2003;43:89–96.
Marino G, Rosso F, Cafiero G, Tortora C, Moraci M, Barbarisi M, Barbarisi A. β-Tricalcium phosphate 3D scaffold promote alone osteogenic differentiation of human adipose stem cells: in vitro study. J Mater Sci. 2010;21:353–63.
Barry F, Boynton RE, Liu B, Mary Murphy J. Chondrogenic differentiation of mesenchymal stem cells from bone marrow: differentiation-dependent gene expression of matrix components. Exp Cell Res. 2001;268:189–200.
Markway BD, Tan GK, Brooke G, Hudson JE, Cooper-White JJ, Doran MR. Enhanced chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells in low oxygen environment micropellet cultures. Cell Transplant. 2010;19(1):29–42.
Khan WS, Adesida AB, Tew SR, Lowe ET, Hardingham TE. Bone marrow-derived mesenchymal stem cells express the pericyte marker 3G5 in culture and show enhanced chondrogenesis in hypoxic conditions. J Orthop Res. 2010;28:834–40.
Verzijl N, Degroot J, Bank RA, Bayliss MT, Bijlsma JW, Lafeber FP, Maroudas A, TeKoppele JM. Age-related accumulation of the advanced glycation endproduct pentosidine levels in human articular cartilage aggrecan: the use of pentosidine levels as a quantitative measure of protein turnover. Matrix Biol. 2001;20:409–17.
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Miyagi, S., Tensho, K., Wakitani, S. et al. Construction of an osteochondral-like tissue graft combining β-tricalcium phosphate block and scaffold-free mesenchymal stem cell sheet. J Orthop Sci 18, 471–477 (2013). https://doi.org/10.1007/s00776-013-0368-3
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DOI: https://doi.org/10.1007/s00776-013-0368-3