Premium
Scalable stirred suspension culture for the generation of billions of human induced pluripotent stem cells using single‐use bioreactors
Author(s) -
Kwok Chee Keong,
Ueda Yuichiro,
Kadari Asifiqbal,
Günther Katharina,
Ergün Süleyman,
Heron Antoine,
Schnitzler Aletta C.,
Rook Martha,
Edenhofer Frank
Publication year - 2018
Publication title -
journal of tissue engineering and regenerative medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.835
H-Index - 72
eISSN - 1932-7005
pISSN - 1932-6254
DOI - 10.1002/term.2435
Subject(s) - laboratory flask , bioreactor , sox2 , human induced pluripotent stem cells , induced pluripotent stem cell , suspension culture , suspension (topology) , cell culture , chemistry , volume (thermodynamics) , microbiology and biotechnology , biology , embryonic stem cell , biochemistry , mathematics , genetics , physics , organic chemistry , quantum mechanics , homotopy , pure mathematics , gene
The production of human induced pluripotent stem cells (hiPSCs) in quantities that are relevant for cell‐based therapies and cell‐loaded implants through standard adherent culture is hardly achievable and lacks process scalability. A promising approach to overcoming these hurdles is the culture of hiPSCs in suspension. In this study, stirred suspension culture vessels were investigated for their suitability in the expansion of two hiPSC lines inoculated as a single cell suspension, with a free scalability between volumes of 50 and 2400 ml. The simple and robust two‐step process reported here first generates hiPSC aggregates of 324 ± 71 μm diameter in 7 days in 125 ml spinner flasks (100 ml volume). These are subsequently dissociated into a single cell suspension for inoculation in 3000 ml bioreactors (1000 ml volume), finally yielding hiPSC aggregates of 198 ± 58 μm after 7 additional days. In both spinner flasks and bioreactors, hiPSCs can be cultured as aggregates for more than 40 days in suspension, maintain an undifferentiated state as confirmed by the expression of pluripotency markers TRA‐1‐60, TRA‐1‐81, SSEA‐4, OCT4, and SOX2, can differentiate into cells of all three germ layers, and can be directed to differentiate into specific lineages such as cardiomyocytes. Up to a 16‐fold increase in hiPSC quantity at the 100 ml volume was achieved, corresponding to a fold increase per day of 2.28; at the 1000 ml scale, an additional 10‐fold increase was achieved. Taken together, 16 × 10 6 hiPSCs were expanded into 2 × 10 9 hiPSCs in 14 days for a fold increase per day of 8.93. This quantity of hiPSCs readily meets the requirements of cell‐based therapies and brings their clinical potential closer to fruition.