Expansion of 3D human induced pluripotent stem cell aggregates in bioreactors: Bioprocess intensification and scaling-up approaches

Tiago Aguiar, Rita Costa, Paula M. Alves, Anders Aspegren,, Patrícia Isabel Alves, Bernardo Abecassis, Émilie Arnault, Margarida Serra

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

Human induced pluripotent stem cells (hiPSC) are attractive tools for drug screening and disease modeling and promising candidates for cell therapy applications. However, to achieve the high numbers of cells required for these purposes, scalable and clinical-grade technologies must be established. In this study, we use environmentally controlled stirred-tank bioreactors operating in perfusion as a powerful tool for bioprocess intensification of hiPSC production. We demonstrate the importance of controlling the dissolved oxygen concentration at low levels (4%) and perfusion at 1.3 day−1 dilution rate to improve hiPSC growth as aggregates in a xeno-free medium. This strategy allowed for increased cell specific growth rate, maximum volumetric concentrations (4.7 × 106 cell/mL) and expansion factors (approximately 19 in total cells), resulting in a 2.6-fold overall improvement in cell yields. Extensive cell characterization, including whole proteomic analysis, was performed to confirm that cells’ pluripotent phenotype was maintained during culture. A scalable protocol for continuous expansion of hiPSC aggregates in bioreactors was implemented using mechanical dissociation for aggregate disruption and cell passaging. A total expansion factor of 1100 in viable cells was obtained in 11 days of culture, while cells maintained their proliferation capacity, pluripotent phenotype and potential as well as genomic stability after 3 sequential passages in bioreactors.

Original languageEnglish
Pages (from-to)81-93
Number of pages13
JournalJournal of Biotechnology
Volume246
DOIs
Publication statusPublished - 20 Mar 2017

Keywords

  • Aggregate dissociation
  • Continuous expansion
  • Human induced pluripotent stem cells
  • Scale-up
  • Stirred tank bioreactors
  • Whole proteome

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