Collagen Sponge Scaffold for Tissue-Engineered Skeletal Muscle

It is still a big challenge to construct a matured muscle tissue in vitro having applicable mass for tissue transplantation. It is well known that nutrient and oxygen supplies to seeded cells limit the scale of tissue-engineered constructs. In this study, a collagen sponge scaffold having uniaxial porous structure was prepared for the large-scaled tissue-engineered skeletal muscle. Unidirectional collagen sponge (US) was prepared from Type I collagen gel solution filled in PTFE tube by placing vertically and freezing with gradual immersion into liquid nitrogen from its bottom to top. Also, homogenous collagen sponge (HS) was prepared by freezing in deep freezer statically. The both of frozen collagen gel were freeze-dried and then crosslinked in a vacuum oven. The collagen scaffolds were observed by SEM and analyzed their pore orientation. C2C12 myoblasts were seeded in both scaffolds and cultured up to 5 weeks and then isometric contractile force of the constructs was determined. In addition, the constructs were studied histologically by HE staining. The isometric contractile force of tissue-engineered skeletal muscle of US was much higher than that of HS. Multinucleated myotubes were observed in both scaffolds, however many of cells located inside of the scaffold were dead probably because of a shortage of nutrients and oxygen supply resulting from shrinkage of the constructs. It may be important to keep the porous structure of sponge scaffold for long-term culture of tissue-engineered skeletal muscle.