TY - JOUR

T1 - Impact of Feeding Strategies on the Scalable Expansion of Human Pluripotent Stem Cells in Single‐Use Stirred Tank Bioreactors

AU - Kropp, Christina

AU - Kempf, Henning

AU - Halloin, Caroline

AU - Robles-Diaz, Diana

AU - Franke, Annika

AU - Scheper, Thomas

AU - Kinast, Katharina

AU - Knorpp, Thomas

AU - Joos, Thomas O.

AU - Haverich, Axel

AU - Martin, Ulrich

AU - Zweigerdt, Robert

AU - Olmera, Ruth

N1 - © 2016 AlphaMed Press

PY - 2016/7/1

Y1 - 2016/7/1

N2 - Theroutine applicationofhumanpluripotent stemcells (hPSCs)andtheirderivatives inbiomedicineanddrug discoverywill require the constant supply of high-quality cells by defined processes. Culturing hPSCs as cellonly 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 permilliliter 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 supportsmaintenance 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 efficientGMP-compliant cell production and underscores the enormous process development potential of hPSCs in suspension culture.

AB - Theroutine applicationofhumanpluripotent stemcells (hPSCs)andtheirderivatives inbiomedicineanddrug discoverywill require the constant supply of high-quality cells by defined processes. Culturing hPSCs as cellonly 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 permilliliter 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 supportsmaintenance 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 efficientGMP-compliant cell production and underscores the enormous process development potential of hPSCs in suspension culture.

KW - Human pluripotent stem cell expansion

KW - Induced pluripotent stem cells

KW - Metabolism

KW - Perfusion

KW - Single-use stirred tank bioreactors

KW - Three-dimensional suspension culture

UR - http://www.scopus.com/inward/record.url?scp=84988596815&partnerID=8YFLogxK

U2 - 10.5966/sctm.2015-0253

DO - 10.5966/sctm.2015-0253

M3 - Article

C2 - 27369897

AN - SCOPUS:84988596815

VL - 5

SP - 1289

EP - 1301

JO - Stem cells translational medicine

JF - Stem cells translational medicine

SN - 2157-6564

IS - 10

ER -