Impact of Feeding Strategies on the Scalable Expansion of Human Pluripotent Stem Cells in Single-Use Stirred Tank Bioreactors

作者：Christina Kropp, Henning Kempf, Caroline Halloin, Diana Robles-Diaz, Annika Franke, Thomas H. Scheper, Katharina Kinast, Thomas Knorpp, Thomas O Joos, Axel Haverich, Ulrich Martin, Robert Zweigerdt, Ruth Olmer · 发表于：Stem Cells Translational Medicine · 年份：2016 · DOI：10.5966/sctm.2015-0253 · 被引用次数：165 · 研究领域：Pluripotent Stem Cells Research、Biomedical Ethics and Regulation、Mesenchymal stem cell research

Abstract The routine application of human pluripotent stem cells (hPSCs) and their derivatives in biomedicine and drug discovery will require the constant supply of high-quality cells by defined processes. Culturing hPSCs as cell-only aggregates in (three-dimensional [3D]) suspension has the potential to overcome numerous limitations of conventional surface-adherent (two-dimensional [2D]) cultivation. Utilizing single-use instrumented stirred-tank bioreactors, we showed that perfusion resulted in a more homogeneous culture environment and enabled superior cell densities of 2.85 × 106 cells per milliliter and 47% higher cell yields compared with conventional repeated batch cultures. Flow cytometry, quantitative reverse-transcriptase polymerase chain reaction, and global gene expression analysis revealed a high similarity across 3D suspension and 2D precultures, underscoring that matrix-free hPSC culture efficiently supports maintenance of pluripotency. Interestingly, physiological data and gene expression assessment indicated distinct changes of the cells' energy metabolism, suggesting a culture-induced switch from glycolysis to oxidative phosphorylation in the absence of hPSC differentiation. Our data highlight the plasticity of hPSCs' energy metabolism and provide clear physiological and molecular targets for process monitoring and further development. This study paves the way toward more efficient GMP-compliant cell production and underscores the enormous process developmen...