Cardiac differentiation of human pluripotent stem cells in scalable suspension culture.
Cardiomyocytes (CMs) generated from human pluripotent stem cells (hPSCs) are a potential cell source for regenerative therapies, drug discovery and disease modeling. All these applications require a routine supply of relatively large quantities of in vitro-generated CMs. This protocol describes a suspension culture-based strategy for the generation of hPSC-CMs as cell-only aggregates, which facilitates process development and scale-up. Aggregates are formed for 4 d in hPSC culture medium followed by 10 d of directed differentiation by applying chemical Wnt pathway modulators. The protocol is applicable to static multiwell formats supporting fast adaptation to specific hPSC line requirements. We also demonstrate how to apply the protocol using stirred tank bioreactors at a 100-ml scale, providing a well-controlled upscaling platform for CM production. In bioreactors, the generation of 40-50 million CMs per differentiation batch at >80% purity without further lineage enrichment can been achieved within 24 d.
1] Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany. [2] REBIRTH-Cluster of Excellence, Hannover Medical School, Hannover, Germany.
Henning
H
Kempf
1] Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany. [2] REBIRTH-Cluster of Excellence, Hannover Medical School, Hannover, Germany.
Christina
C
Kropp
1] Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany. [2] REBIRTH-Cluster of Excellence, Hannover Medical School, Hannover, Germany. [3] Member of Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research (DZL), Hannover, Germany.
Ruth
R
Olmer
1] Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany. [2] REBIRTH-Cluster of Excellence, Hannover Medical School, Hannover, Germany. [3] Member of Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research (DZL), Hannover, Germany.
Ulrich
U
Martin
1] Leibniz Research Laboratories for Biotechnology and Artificial Organs (LEBAO), Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany. [2] REBIRTH-Cluster of Excellence, Hannover Medical School, Hannover, Germany.
Robert
0000000246560770
R
Zweigerdt
D018929:Cell Culture Techniques; D006801:Humans; D032383:Myocytes, Cardiac / Q000166:cytology*; D039904:Pluripotent Stem Cells / Q000502:physiology*