Differentiation and lineage selection of mouse embryonic stem cells in a stirred bench scale bioreactor with automated process control

Magnus Schroeder, Sylvia Niebruegge, Andreas Werner, Elmar Willbold, Monika Burg, Manfred Ruediger, Loren Field, Juergen Lehmann, Robert Zweigerdt

Research output: Contribution to journalArticle

125 Citations (Scopus)

Abstract

It is well established that embryonic stem (ES) cells can differentiate into functional cardiomyocytes in vitro. ES-derived cardiomyocytes could be used for pharmaceutical and therapeutic applications, provided that they can be generated in sufficient quantity and with sufficient purity. To enable large-scale culture of ES-derived cells, we have developed a robust and scalable bioprocess that allows direct embryoid body (EB) formation in a fully controlled, stirred 2 L bioreactor following inoculation with a single cell suspension of mouse ES cells. Utilizing a pitched-blade-turbine, parameters for optimal cell expansion as well as efficient ES cell differentiation were established. Optimization of stirring conditions resulted in the generation of high-density suspension cultures containing 12.5 × 106 cells/mL after 9 days of differentiation. Approximately 30%-40% of the EBs formed in this process vigorously contracted, indicating robust cardiomyogenic induction. An ES cell clone carrying a recombinant DNA molecule comprised of the cardiomyocyte-restricted alpha myosin heavy chain (αMHC) promoter and a neomycin resistance gene was used to establish the utility of this bioprocess to efficiently generate ES-derived cardiomyocytes. The genetically engineered ES cells were cultured directly in the stirred bioreactor for 9 days, followed by antibiotic treatment for another 9 days. The protocol resulted in the generation of essentially pure cardiomyocyte cultures, with a total yield of 1.28 × 109 cells in a single 2 L bioreactor run. This study thus provides an important step towards the large-scale generation of ES-derived cells for therapeutic and industrial applications.

Original languageEnglish
Pages (from-to)920-933
Number of pages14
JournalBiotechnology and Bioengineering
Volume92
Issue number7
DOIs
StatePublished - Dec 30 2005

Fingerprint

Bioreactors
Embryonic Stem Cells
Stem cells
Process control
Cardiac Myocytes
Suspensions
Ventricular Myosins
Neomycin
Recombinant DNA
Myosin Heavy Chains
Embryoid Bodies
Antibiotics
Drug products
Turbomachine blades
Industrial applications
Turbines
Genes
Mouse Embryonic Stem Cells
Cell Differentiation
Anti-Bacterial Agents

Keywords

  • Bioreactor agitation
  • Cardiomyocytes
  • Embryoid bodies
  • Embryonic stem cells
  • Process control
  • Scale-up

ASJC Scopus subject areas

  • Biotechnology
  • Microbiology

Cite this

Differentiation and lineage selection of mouse embryonic stem cells in a stirred bench scale bioreactor with automated process control. / Schroeder, Magnus; Niebruegge, Sylvia; Werner, Andreas; Willbold, Elmar; Burg, Monika; Ruediger, Manfred; Field, Loren; Lehmann, Juergen; Zweigerdt, Robert.

In: Biotechnology and Bioengineering, Vol. 92, No. 7, 30.12.2005, p. 920-933.

Research output: Contribution to journalArticle

Schroeder, M, Niebruegge, S, Werner, A, Willbold, E, Burg, M, Ruediger, M, Field, L, Lehmann, J & Zweigerdt, R 2005, 'Differentiation and lineage selection of mouse embryonic stem cells in a stirred bench scale bioreactor with automated process control', Biotechnology and Bioengineering, vol. 92, no. 7, pp. 920-933. https://doi.org/10.1002/bit.20668
Schroeder, Magnus ; Niebruegge, Sylvia ; Werner, Andreas ; Willbold, Elmar ; Burg, Monika ; Ruediger, Manfred ; Field, Loren ; Lehmann, Juergen ; Zweigerdt, Robert. / Differentiation and lineage selection of mouse embryonic stem cells in a stirred bench scale bioreactor with automated process control. In: Biotechnology and Bioengineering. 2005 ; Vol. 92, No. 7. pp. 920-933.
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