Controlling Expansion and Cardiomyogenic Differentiation of Human Pluripotent Stem Cells in Scalable Suspension Culture

Summary

To harness the potential of human pluripotent stem cells (hPSCs), an abundant supply of their progenies is required. Here, hPSC expansion as matrix-independent aggregates in suspension culture was combined with cardiomyogenic differentiation using chemical Wnt pathway modulators. A multiwell screen was scaled up to stirred Erlenmeyer flasks and subsequently to tank bioreactors, applying controlled feeding strategies (batch and cyclic perfusion). Cardiomyogenesis was sensitive to the GSK3 inhibitor CHIR99021 concentration, whereas the aggregate size was no prevailing factor across culture platforms. However, in bioreactors, the pattern of aggregate formation in the expansion phase dominated subsequent differentiation. Global profiling revealed a culture-dependent expression of BMP agonists/antagonists, suggesting their decisive role in cell-fate determination. Furthermore, metallothionein was discovered as a potentially stress-related marker in hPSCs. In 100 ml bioreactors, the production of 40 million predominantly ventricular-like cardiomyocytes (up to 85% purity) was enabled that were directly applicable to bioartificial cardiac tissue formation.

Graphical Abstract

Highlights

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  Efficient cardiac differentiation protocol in suspension by chemical Wnt modulators

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  Differentiation is CHIR concentration dependent, but aggregate size independent

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  Bioreactor-controlled hPSC expansion dictates subsequent lineage differentiation

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  Metallothionein is a potentially stress-induced marker of hPSC culture

Abstract

In this article, Zweigerdt and colleagues show hPSC expansion as matrix-independent aggregates in suspension culture combined with efficient and scalable cardiac differentiation in stirred tank bioreactors. The strategy enables the generation of 40 million cardiomyocytes (up to 85% purity) of predominantly ventricular-like phenotype per run that were directly applicable to bioartificial cardiac tissue formation.