title: Improved tissue culture conditions for engineered skeletal muscle sheets.
abstract: The potential clinical utility of engineered muscle is currently restricted by limited in vitro capacity of expanded muscle precursor cells to fuse and form mature myofibers. The purpose of this study was to use isotropic skeletal muscle sheets to explore the impact of (1) fibroblast coculture and (2) fibroblast-conditioned media (fCM) on in vitro myogenesis. Muscle sheets were prepared by seeding varying ratios of skeletal myoblasts and fibroblasts on a biomimetic substrate and culturing the resulting tissue in either control media or fCM. Muscle sheets were prepared from two cell subpopulations, (1) C2C12 and NOR-10 and (2) primary neonatal rat skeletal muscle cells (nSKM). In C2C12/Nor-10 muscle sheets fCM conferred a myogenic advantage early in culture; at D1 a statistically significant 3.12 ± 0.8-fold increase in myofiber density was observed with fCM. A high purity satellite cell population was collected from an initially mixed population of nSKMs via cell sorting for positive α 7-integrin expression. On D6, tissue sheets with low fibroblast concentrations (0 & 10%) cultured in fCM had increased average myofiber density (4.8 ± 0.2 myofibers/field) compared to tissue sheets with high fibroblast concentrations (50%) cultured in control media (1.0 ± 0.1 myofibers/field). Additionally, fCM promoted longer, thicker myofibers with a mature phenotype.
authors.affiliation: Department of Basic Sciences, New York University, 345 East 24th Street, New York, NY 10010, USA. sh2620@nyu.edu
authors.forename: Sara
authors.initials: S
authors.lastname: Hinds
authors.forename: Natalia
authors.initials: N
authors.lastname: Tyhovych
authors.forename: Clint
authors.initials: C
authors.lastname: Sistrunk
authors.forename: Louis
authors.initials: L
authors.lastname: Terracio
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D015703:Antigens, CD / Q000378:metabolism; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D039001:Integrin alpha Chains / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D024510:Muscle Development; D018482:Muscle, Skeletal / Q000254:growth & development* / Q000378:metabolism; D032448:Myoblasts, Skeletal / Q000378:metabolism*; D051381:Rats; D046509:Tissue Culture Techniques / Q000379:methods*; D023822:Tissue Engineering
chemical_list: D015703:Antigens, CD; D017077:Culture Media, Conditioned; D039001:Integrin alpha Chains; C085725:integrin alpha7