Speaker
Description
Myotendinous junction disfunctions due to degenerative musculoskeletal diseases or injuries resulting from strenuous physical activities are still considered an ongoing issue in the field of musculoskeletal tissue engineering. Indeed, the main challenge of biofabrication strategies relies on the development of methods enabling the generation of artificial bio-constructs that can replicate the complexity of the muscle-tendon tissue interface. In this work, a core-shell Y-shaped microfluidic chip has been developed to alternatively deliver two different hydrogel-based bioinks that enable the mimicking of the tendon and the muscle tissue, respectively. As a result, core-shell microfibers are extruded and collected on a rotating drum to form heterogenous hydrogel yarns. In order to fabricate a myotendinous-like construct, the time-switching between the two bioinks, the drum rotational speed, and the core and shell flow rate, have been optimized. Once flow rates have been selected, different values of drum rotational speeds (i.e., 20, 30, 40, 50 and 60 rpm) have been tested to evaluate the overall effect on both the microfiber diameter and the core dimension. Moreover, NIH 3T3 fibroblasts and C2C12 myoblasts, used to mimic the tendon and muscle side respectively, were encapsulated in the core bioink and spun at the selected rotational speed values. As a result, the effect of the increasing velocity induced a decreasing of the microfiber and core diameter along with the shell thickness. Furthermore, rotational speeds up to 40 rpm showed high viability and cell alignment along the fiber direction for both NIH 3T3 fibroblasts and C2C12 myoblasts. On the other hand, rotational speeds of 20 rpm and 30 rpm induced low cell alignment and spreading along with a low cell viability, due to the increasing of shell diameter that prevents the oxygen and nutrient exchange. In addition, it was observed that rotational speeds up to 40 rpm were related with higher switching-time that create flow rate turbulences and a reduced ratio of muscle/tendon tissue-only section compared to the tissue interface one. Hence, 40 rpm has been selected as optimal rotational speed velocity for the fabrication of myotendinous-like constructs. Cell-laden heterogenous scaffolds showed high degree of compartmentalization and enabled the recreation of the tissue-specific biological heterogeneity. Moreover, C2C12/NIH 3T3-laden constructs showed high cell proliferation up to 14 days of culture. Finally, immunochemistry analysis will be performed in order to investigate myosin heavy chain as well as collagen I and III expression at the muscle and tendon side, respectively. Thus, such biofabrication method could be validated for the generation of a biomimetic heterogenous scaffold that can recapitulate the biological complexity of the muscle tendon unit.
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