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Optimizing the structure and contractility of engineered skeletal muscle thin films.
An experimental system was developed to tissue engineer skeletal muscle thin films with well-defined tissue architecture and to quantify the effect on contractility. Using the C2C12 cell line, the authors tested whether tailoring the width and spacing of micropatterned fibronectin lines can be used to increase myoblast differentiation into functional myotubes and maximize uniaxial alignment within a 2-D sheet. Using a combination of image analysis and the muscular thin film contractility assay, it was demonstrated that a fibronectin line width of 100μm and line spacing of 20μm is able to maximize the formation of anisotropic, engineered skeletal muscle with consistent contractile properties at the millimeter length scale. The engineered skeletal muscle exhibited a positive force-frequency relationship, could achieve tetanus and produced a normalized peak twitch stress of 9.4±4.6kPa at 1Hz stimulation. These results establish that micropatterning technologies can be used to control skeletal muscle differentiation and tissue architecture and, in combination with the muscular thin film contractility, assay can be used to probe structure-function relationships. More broadly, an experimental platform is provided with the potential to examine how a range of microenvironmental cues such as extracellular matrix protein composition, micropattern geometries and substrate mechanics affect skeletal muscle myogenesis and contractility.