Speaker
Description
"Introduction
The engineering of functional macrotissues, such as muscle tissue, is challenging due to the limited degree of diffusion, resulting in poor cell survival rates. To overcome this problem, the incorporation of a (mature) vascularized network is important. Therefore, a possible strategy is conducting bottom-up tissue engineering by generating vascularized tissue-specific spheroids that can serve as tissue building blocks. The use of these microtissue building blocks is advantageous compared to single cells because of their high cell density, improved cell survival, microarchitecture and accelerated ECM production. When spheroids are combined with a suitable hydrogel, a bioink can be obtained. This bioink can be used for the creation of more complex macrotissues via 3D bioprinting.
Methodology
Myogenic vascularized spheroids were generated by seeding different ratios of human myoblasts, endothelial cells and mesenchymal stem cells (1,0x106 cells) in an in-house developed microchip system with 400 µm pores. After 4 days of cultivation in medium specialized for endothelial cell growth (EGM-2), spheroids were exposed to myogenic differentiation medium consisting of DMEM:glutamax supplemented with 50 µg/ml gentamicin, 50 µg/ml bovine serum albumin, 10 µg/ml human insulin and 10-10 g/ml human epidermal growth factor. Analysis was performed after 4, 8 and 12 days of chip culture. Immunohistochemical staining for tropomyosin and CD31, as well as, histological and live/dead analysis was conducted. With this, the morphology, ECM production, differentiation and spheroid viability was evaluated.
Results
Vascularized myogenic spheroids could be obtained containing several ratios of cells. The spheroids remained viable over time. (Immuno)histochemical analysis showed the presence of tissue-specific ECM components and the formation of microvascular networks.
Conclusion
During this study it was demonstrated that vascularized myogenic spheroids can be obtained to use as a part of a printable bioink. Future work will investigate the fusion capacities of the spheroids, as well as, the alignment of fibers and optimal spheroid maturation stage for encapsulation in several types of hydrogels."
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