Tissue engineering of skeletal muscle using polymer fiber arrays

The purpose of this study was to assess a new scaffold design for muscle tissue engineering: arrays of parallel-oriented polymer microfibers. First, C2C12 skeletal myoblasts were seeded onto single, laminin-coated polypropylene fibers and their growth and alignment were characterized. With the aim of creating skeletal muscle sheets, it was then investigated whether cell layers of single fibers merged when in close proximity to neighboring fibers. The optimal fiber spacing needed to achieve cell alignment with the lowest possible content of scaffold material was established. Further, it was assessed whether such a cell sheet became contractile and whether it survived in vitro for extended periods of time. C2C12 cells, cultured on fibers 10 to 15 microm in diameter, formed up to 50-microm-thick layers of longitudinally aligned cells. Four different groups based on fiber spacing (30 to 35, 50 to 55, 70 to 75, and 90 to 95 microm) were evaluated. Complete cell sheets formed between fibers that were spaced 55 microm apart or less; larger spacing led to no or incomplete sheets. C2C12 cells, seeded onto a 10 x 20 mm fiber array, formed a contractile cell sheet that was maintained in vitro for 70 days. Larger, three-dimensional structures might be created by arranging fibers in several layers or by stacking cellular sheets.