Fabrication of multi-well platform for efficient myogenic commitment of C2C12 cells via electrical stimulation

Abstract

To engineer tissue-like structures, cells are required to organize themselves into three-dimensional networks that mimic the native tissue microarchitecture. Here, we present agarose-based multi-well platform incorporated with electrical stimulation (ES) to build skeletal muscle-like structures in a facile and highly reproducible fashion. Our observations showed that ES improved myogenic differentiation of C2C12 cells in two-dimensional monolayer culture and increased the diameter of the resulting myotubes. Furthermore, C2C12 skeletal muscle cells were encapsulated in collagen/matrigel scaffolds in 3D manner, and electrical stimulation was applied for efficient myogenic commitment. ES also facilitated the formation of free-standing 3D muscle tissues with or without mechanical stretch at both ends of the gel structure. Consequently, we confirmed the transcriptional upregulations of myogenic related genes in the ES group compared to non-stimulated control group in our multi-well 3D culture platform. Interestingly, the cell-encapsulated gel consisted solely of matrigel did not give admirable myotube formation

however, collagen showed upregulated myosin heavy chain, muscle regulatory factor expression and prominent effect in building myotubular structures. Given the robust fabrication, engineered muscle tissues in multi-well platform may find their use in high-throughput biological studies drug screenings.