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Engineering of aligned skeletal muscle by micropatterning.

Tissue engineered skeletal muscle has tremendous potential for the treatment of muscular injury or muscular dysfunction. However, in vitro methods to generate skeletal muscle with physiologically aligned myofiber structure remains limited. To develop a robust in vitro model that resembles the physiologically aligned structure of muscle fibers, we fabricated micropatterned polymer membranes of poly(dimethylsiloxane) (PDMS) with parallel microgrooves, and then examined the effect of micropatterning on myoblast cellular organization and the cell fusion process. In comparison to the myoblasts on non-patterned PDMS films, myoblasts on micropatterned PDMS films had well-organized F-actin assembly in close proximity to the direction of microgrooves, along with enhanced levels of myotube formation at early time points. The increase of cell cycle regulator p21(WAF/Cip1) and the organized interactions of N-cadherin in myoblasts on micropatterned surfaces may contribute to the enhanced formation of myotubes. Similar results of cellular alignment was observed when myoblasts were cultured on microfluidically patterned poly(2-hydroxyethyl methacrylate) (pHEMA) microgrooves, and the micropatterns were found to detach from the Petri dish over time. To apply this technology for generating aligned tissue-like muscle constructs, we developed a methodology to transfer the aligned myotubes to biodegradable collagen gels. Histological analysis revealed the persistence of aligned cellular organization in the collagen gels. Together, these results demonstrate that micropatterned PDMS or pHEMA can promote cell alignment and fusion along the direction of the microgrooves, and this platform can be utilized to transfer aligned myotubes on biodegradable hydrogels. This study highlights the importance of spatial cues in creating aligned skeletal muscle for tissue engineering and muscular regeneration applications.