Fabrication of Multicomponent Tissue-Like Constructs-Toward Spinal Motion Segment Tissue Engineering
Authors: Chik T. K. , Luk K. D. K. , Cheung K. M. C. , Sze K. Y. , Chan B. P. 
Published: June 2012

Introduction

Allogenic spinal motion segment transplantation demonstrated promising results in treating late stage disk degeneration and stimulated immense interests of tissue engineering in the whole spinal motion segment from autologous cell source and safe biomaterials, solving the donor availability problem and potentially alleviating the degeneration and immunogenicity problems. Bioengineering complex tissues such as spinal motion segment is extremely challenging because it involves the integration of multiple tissues with distinct structures and composition such as lamellar annulus fibrosus (AF), gel-like nucleus pulposus (NP), cartilaginous endplate, and bony vertebra. Although attempts have been made to fabricate biphasic structures such as lamellar AF structure wrapping around a NP core, or bony block with cartilaginous structure, good methods integrating different tissue parts to form a combined unit are not available. In this study, a bioreactor-based method integrating multicomponent spinal motion segment-like tissue constructs, together with in vitro mechanical and biological stimulation, was reported, as the first step towards functional spinal motion segment tissue engineering.

Materials and Methods

Rabbit mesenchymal stem cells (rMSC) were used to fabricate cell-collagen constructs. Constructs were cultured separately in chondrogenic and osteogenic differentiation medium for 21 days. A layer of rMSC-collagen solution was added between chondrogenic and osteogenic units for coculture with cyclic compression to form the combined osteochondral constructs with intact interface. Afterward, an acellular cylindrical collagen gel was formed in between two such combined osteochondral constructs before circumferencing the whole construct with rMSC-collagen tubular layers. The multicomponent constructs were subjected to periodic compression and torsion simultaneously. The constructs were evaluated histologically for multicomponent structures. Cell alignment in the tubular layers was quantified using an adhoc MatLab program while collagen alignment was evaluated using scanning electron microscopy (SEM). Moreover, the interfacial strength of the osteochondral units was also evaluated using a tensile test machine.

Results

The combined constructs were able to be dismounted with integrity from the supporting shafts of the bioreactor after 14 days of culture. Every component of the combined constructs remained intact and can be manipulated with forceps. This method proves the feasibility to proceed to bioreactor-based functional remodelling and engineering of spinal motion segment in the future. Alignment analysis of rMSC in the tubular layers of the combined constructs demonstrated that the cell aligned along different preferred direction under torsional stimulation as compared with that from the static loading control (Fig. 1). However, SEM demonstrated no significant collagen alignment after 14 days culture for all groups. Histological results showed good integration among different components while cells were able to migrate into the acellular collagen gel at the center of the combined constructs although the source of the migrated cells was not studied yet. In addition, the osteogenic and chondrogenic phenotype in the osteochondral units were maintained while the interfacial strength of the osteochondral units was significantly higher when both the biological and the compression stimulation were present during their fabrication.

Conclusion

A method fabricating a multicomponent tissue construct with integrity for future functional spinal motion segment tissue engineering is demonstrated. Torsional stimulation can be used to stimulate rMSC re-orientation within a collagen tubular structure. Osteogenic differentiated and chondrogenic differentiated rMSC were able to maintain the phenotype throughout the culture period and their interface stabilized by biological and mechanical stimulation. I confirm having declared any potential conflict of interest for all authors listed on this abstract No Disclosure of Interest None declared Cheng HW, Luk KD, Cheung KM, Chan BP. In vitro generation of an osteochondral interface from mesenchymal stem cell-collagen microspheres. Biomaterials 2011;32(6):1526–1535. Au-Yeung KL, Sze KY, Sham MH, Chan BP. Development of a micromanipulator-based loading device for mechanoregulation study of human mesenchymal stem cells in three-dimensional collagen constructs. Tissue Engineering Part C Methods 2010;16(1):93–107.