Mouse embryonic stem cell expansion in a microcarrier-based stirred culture system

Fernandes, A. M.; Diogo, M. M.; da Silva, C. Lobato; Henrique, D.; Cabral, J. M. S.; Fernandes, T.G.; Domingos Manuel Pinto

Key information

Authors:

Fernandes, A. M.; Diogo, M. M. (Maria Margarida Fonseca Rodrigues Diogo); da Silva, C. Lobato (Cláudia Lobato da Silva); Henrique, D.; Cabral, J. M. S. (Joaquim Manuel Sampaio Cabral); Fernandes, T.G. (Tiago Paulo Gonçalves Fernandes); Domingos Manuel Pinto (Domingos Manuel Pinto Henrique)

Published in

October 2007

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 3.5 x 10(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.