Muscle-derived extracellular matrix on sinusoidal wavy surfaces synergistically promotes myogenic differentiation and maturation.

The generation of physiologically aligned multinucleated myotubes is critical in the fabrication of functional engineered skeletal muscle. Although micro-/nano-topographical contact guidance, such as groove/ridge structures, has induced the alignment of muscle fibers by providing cells with extracellular matrix (ECM) topography, the complex biochemical microenvironment of the ECM cannot be recapitulated. Here, we report the enhancement of myogenic differentiation and maturation using muscle decellularized ECM (mdECM) and sinusoidal wavy surfaces, which provided a biochemical microenvironment and microscale contact guidance, respectively. Sinusoidal wavy polystyrene surfaces with wavelengths of 20, 40, and 80 μm were fabricated by a deep X-ray lithography-based process. The mdECM was prepared by decellularization of porcine tibialis anterior skeletal muscle. An mdECM coating significantly improved the surface wettability of polystyrene substrates and exhibited higher seeding efficiency, cell viability, and proliferation compared with collagen- and non-coating cases. The sinusoidal wavy surfaces induced well-aligned myotubes and showed significantly enhanced formation of myotubes and myogenic differentiation when the surface was coated with mdECM. Particularly, there was an approximately 1.5-2 fold improvement in morphological analysis and gene expression for mdECM-compared to non-coated sinusoidal wavy surfaces. These results suggest that the consideration of both topographical and biochemical environmental cues can generate a highly mimicked ECM environment, thereby providing cells with a synergistic effect on myogenic differentiation and maturation. The outcome of this study will be useful in developing of functional engineered muscle for application in tissue regeneration or a high-throughput in vitro model for drug screening.