Design, development, and benchmarking of a bioreactor integrated with 3D bioprinting: Application to skeletal muscle regeneration

In recent years, great efforts have been spent to create engineered muscle constructs recapitulating the 3D architecture and applying external stimulations. In this regard, tissue engineering approaches could be very promising in regenerating skeletal muscle, in which bioprinting techniques have produced encouraging results especially regarding 3D architecture. Tensile stimuli showed a fundamental role in regulating the behavior of muscle cells both in terms of 3D organizations and protein expression. Despite this promising premise, the combination of 3D bioprinting and mechanical stimulation has been poorly investigated, calling for novel approaches dealing with the mechanical stimulation of the 3D bioprinted construct and the integration of the bioprinting phase into the stimulation device. To this aim, the present work proposes the design, manufacturing, and benchmarking of a bioprinting-integrated mechanical platform conceived for mechanically stimulating a 3D muscle model directly printed into the bioreactor to foster the integration of printing and stimulation. The study consists of three main steps: 1) the design, fabrication, and mechanical characterization of stretchable supports suitable for bioprinting and long-term cell culture; 2) the design, assisted by computational tools, and the fabrication of the smart Petri dish containing the stimulation mechanism and of the final cyclic mechanical platform; 3) the in-vitro validation of the proposed platform in terms of transmission of the mechanical stimulation to the 3D construct and the biological effect of dynamic culture on 3D bioprinted muscle cells. The results highlighted excellent viability and demonstrated that the external stimulus influences the murine myoblasts behavior already after 7 days of culture. In conclusion, prototypes are now available of a mechanical platform that integrates the 3D bioprinting and is capable of stimulating 3D biological constructs for applications in the field of muscle tissue engineering.