title: Mouse embryonic stem cell expansion in a microcarrier-based stirred culture system.
abstract: Embryonic stem (ES) cells have the ability to differentiate in vitro into a wide variety of cell types with potential applications for tissue regeneration. However, a large number of cells are required, thus strengthening the need to develop large-scale systems using chemically defined media for ES cell production and/or controlled differentiation. In the present studies, a stirred culture system (i.e. spinner flask) was used to scale-up mouse ES (mES) cell expansion in serum-containing (DMEM/FBS) or serum-free medium, both supplemented with leukemia inhibitory factor (LIF), using either Cytodex 3 or Cultispher S microcarriers. After 8 days, maximal cell densities achieved were (1.9+/-0.1), (2.6+/-0.7) and 3.5x10(6)cells/mL for Cytodex 3 in DMEM/FBS, Cultispher S in DMEM/FBS and Cultispher S in serum-free cultures, respectively, with fold increases of 38+/-2, 50+/-15 and 70. Both microcarriers were suitable to sustain mES cell expansion, though the macroporous Cultispher S seemed to be advantageous in providing a more protective environment against shear stress forces, which harmful effects are exacerbated in serum-free conditions. Importantly, mES cells expanded under stirred conditions using serum-free medium retained their pluripotency and the ability to commit to the neural lineage.
authors.affiliation: Institute for Biotechnology and Bioengineering (IBB), Centre for Biological and Chemical Engineering, Instituto Superior Técnico, Av. Rovisco Pais, 1049-001 Lisboa, Portugal.
authors.forename: A M
authors.initials: AM
authors.lastname: Fernandes
authors.forename: T G
authors.initials: TG
authors.lastname: Fernandes
authors.forename: M M
authors.initials: MM
authors.lastname: Diogo
authors.forename: C Lobato
authors.initials: CL
authors.lastname: da Silva
authors.forename: D
authors.initials: D
authors.lastname: Henrique
authors.forename: J M S
authors.initials: JM
authors.lastname: Cabral
mesh_terms: D000818:Animals; D019149:Bioreactors; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D002460:Cell Line; D016895:Culture Media, Serum-Free; D053595:Embryonic Stem Cells / Q000502:physiology*; D051379:Mice; D008863:Microspheres
chemical_list: D016895:Culture Media, Serum-Free