Advertisement

High density continuous production of murine pluripotent cells in an acoustic perfused bioreactor at different oxygen concentrations

Authors

  • Ricardo P. Baptista,

    1. Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada
    2. Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, Ontario, Canada
    Search for more papers by this author
  • David A. Fluri,

    1. Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada
    2. Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, Ontario, Canada
    Search for more papers by this author
  • Peter W. Zandstra

    Corresponding author
    1. Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada
    2. Department of Chemical Engineering and Applied Chemistry, University of Toronto, Toronto, Ontario, Canada
    3. McEwen Centre for Regenerative Medicine, University Health Network and Heart & Stroke Richard Lewar Centre of Excellence, University of Toronto, Toronto, Ontario, Canada
    4. Terrence Donnelly Centre for Cellular and Biomolecular Research, 160 College Street, Rm 1116, Toronto, ON, Canada M5S 3E1; telephone: 416-978-8888; fax: 416-978-4317
    • Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada.
    Search for more papers by this author

  • The authors declare no competing financial interests.

Abstract

Strategies for the production of pluripotent stem cells (PSCs) rely on serially dissociated adherent or aggregate-based culture, consequently limiting robust scale-up of cell production, on-line control and optimization of culture conditions. We recently developed a method that enables continuous (non-serially dissociated) suspension culture-mediated reprogramming to pluripotency. Herein, we use this method to demonstrate the scalable production of PSCs and early derivatives using acoustic filter technology to enable continuous oxygen-controlled perfusion culture. Cell densities of greater than 1 × 107 cells/mL were achieved after 7 days of expansion at a specific growth rate (µ) of 0.61 ± 0.1 day−1 with a perfusion rate (D) of 5.0 day−1. A twofold increase in maximum cell density (to greater than 2.5 × 107 cells/mL) was achieved when the medium dissolved oxygen was reduced (5% DO). Cell densities and viabilities >80% were maintained for extended production periods during which maintenance of pluripotency was confirmed by stable expression of pluripotency factors (SSEA-1 and Nanog), as well as the capacity to differentiate into all three germ layers. This work establishes a versatile biotechnological platform for the production of pluripotent cells and derivatives in an integrated, scalable and intensified stirred suspension culture. Biotechnol. Bioeng. 2013; 110: 648–655. © 2012 Wiley Periodicals, Inc.

Ancillary