Combined Effects of Electrical Stimulation and Protein Coatings on Myotube Formation in a Soft Porous Scaffold

Compared to two-dimensional cell cultures, three-dimensional ones potentially allow recreating natural tissue environments with higher accuracy. The three-dimensional approach is being investigated in the field of tissue engineering targeting the reconstruction of various tissues, among which skeletal muscle. Skeletal muscle is an electroactive tissue which strongly relies upon interactions with the extracellular matrix for internal organization and mechanical function. Studying the optimization of myogenesis in vitro implies focusing on appropriate biomimetic stimuli, as biochemical and electrical ones. Here we present a three-dimensional polyurethane-based soft porous scaffold (porosity ~ 86%) with a Young's modulus in wet conditions close to the one of natural skeletal muscle tissue (~ 9 kPa). To study the effect of external stimuli on muscle cells, we functionalized the scaffold with extracellular matrix components (laminin and fibronectin) and observed an increase in myoblast proliferation over three days. Furthermore, the combination between laminin coating and electrical stimulation resulted in more spread and thicker myotubes compared to non-stimulated samples and samples receiving the single (non-combined) inputs. These results pave the way to the development of mature muscle tissue within three-dimensional soft scaffolds, through the combination of biochemical and electrical stimuli.