Standardized production of hPSC-derived cardiomyocyte aggregates in stirred spinner flasks.
Journal
Nature protocols
ISSN: 1750-2799
Titre abrégé: Nat Protoc
Pays: England
ID NLM: 101284307
Informations de publication
Date de publication:
28 Mar 2024
28 Mar 2024
Historique:
received:
04
07
2023
accepted:
17
01
2024
medline:
29
3
2024
pubmed:
29
3
2024
entrez:
29
3
2024
Statut:
aheadofprint
Résumé
A promising cell-therapy approach for heart failure aims at differentiating human pluripotent stem cells (hPSCs) into functional cardiomyocytes (CMs) in vitro to replace the disease-induced loss of patients' heart muscle cells in vivo. But many challenges remain for the routine clinical application of hPSC-derived CMs (hPSC-CMs), including good manufacturing practice (GMP)-compliant production strategies. This protocol describes the efficient generation of hPSC-CM aggregates in suspension culture, emphasizing process simplicity, robustness and GMP compliance. The strategy promotes clinical translation and other applications that require large numbers of CMs. Using a simple spinner-flask platform, this protocol is applicable to a broad range of users with general experience in handling hPSCs without extensive know-how in biotechnology. hPSCs are expanded in monolayer to generate the required cell numbers for process inoculation in suspension culture, followed by stirring-controlled formation of cell-only aggregates at a 300-ml scale. After 48 h at checkpoint (CP) 0, chemically defined cardiac differentiation is induced by WNT-pathway modulation through use of the glycogen-synthase kinase-3 inhibitor CHIR99021 (WNT agonist), which is replaced 24 h later by the chemical WNT-pathway inhibitor IWP-2. The exact application of the described process parameters is important to ensure process efficiency and robustness. After 10 d of differentiation (CP I), the production of ≥100 × 10
Identifiants
pubmed: 38548938
doi: 10.1038/s41596-024-00976-2
pii: 10.1038/s41596-024-00976-2
doi:
Types de publication
Journal Article
Review
Langues
eng
Sous-ensembles de citation
IM
Subventions
Organisme : Deutsche Forschungsgemeinschaft (German Research Foundation)
ID : REBIRTH EXC 62/2, ZW64/4-2
Organisme : Bundesministerium für Bildung und Forschung (Federal Ministry of Education and Research)
ID : 01EK1601A, 13XP5092B, 031L0249, 01EK2108A
Organisme : Niedersächsische Ministerium für Wissenschaft und Kultur (Lower Saxony Ministry of Science and Culture)
ID : ZN3340, ZN4092
Organisme : European Commission (EC)
ID : 101056712
Informations de copyright
© 2024. Springer Nature Limited.
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