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Expansion and patterning of cardiovascular progenitors derived from human pluripotent stem cells

Nature Biotechnology volume 33, pages 970–979 (2015)Cite this article

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Abstract

The inability of multipotent cardiovascular progenitor cells (CPCs) to undergo multiple divisions in culture has precluded stable expansion of precursors of cardiomyocytes and vascular cells. This contrasts with neural progenitors, which can be expanded robustly and are a renewable source of their derivatives. Here we use human pluripotent stem cells bearing a cardiac lineage reporter to show that regulated MYC expression enables robust expansion of CPCs with insulin-like growth factor-1 (IGF-1) and a hedgehog pathway agonist. The CPCs can be patterned with morphogens, recreating features of heart field assignment, and controllably differentiated to relatively pure populations of pacemaker-like or ventricular-like cardiomyocytes. The cells are clonogenic and can be expanded for >40 population doublings while retaining the ability to differentiate into cardiomyocytes and vascular cells. Access to CPCs will allow precise recreation of elements of heart development in vitro and facilitate investigation of the molecular basis of cardiac fate determination. This technology is applicable for cardiac disease modeling, toxicology studies and tissue engineering.

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Figure 1: The expansion of early hPSC-derived cardiac cells.
Figure 2: NKX2-5–eGFP expression is activated in spheres by concerted FGF and BMP signaling.
Figure 3: Characterization of sphere heterogeneity and differentiation potential.
Figure 4: Recapitulation of ventricular-like and pacemaker-like cardiomyocyte functions.
Figure 5: Long-term expansion and differentiation analysis of CPCs.
Figure 6: Working model of hPSC-derived cardiovascular progenitor self-renewal and differentiation in the context of heart development.

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Acknowledgements

We acknowledge the contributions of I. Marcuccio and S. Casini. Work in C.L.M.'s lab is supported by Cardiovascular Research Netherlands (CVON HUSTCARE), The Netherlands Institute of Regenerative Medicine (NIRM), the European Research Council (ERCAdG 323182 STEMCARDIOVASC) and The Netherlands Organization for Scientific Research (NWO-FOM FOM 09MMC02). Work in R.P.'s lab is supported by the Netherlands Organization for Scientific Research (ZonMW-TOP 40-00812-98-12086 and ZonMW-MKMD 40-42600-98-036) and the Rembrandt Institute of Cardiovascular Science. V.V.O. was supported by the European Community's Seventh Framework Programme (FP7/2007-2013) grant agreement 602423.

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Authors and Affiliations

  1. Leiden University Medical Center, Leiden, the Netherlands

    Matthew J Birket, Marcelo C Ribeiro, Dorien Ward, Ana Rita Leitoguinho, Sabine C den Hartogh, Valeria V Orlova, Harsha D Devalla, Verena Schwach, Milena Bellin, Robert Passier & Christine L Mummery

  2. Amsterdam Medical Center, Amsterdam, the Netherlands

    Arie O Verkerk

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Contributions

M.J.B. and C.L.M. designed the study. M.J.B., M.C.R., A.O.V., A.R.L. and V.S. performed experiments and analyzed data. H.D.D. analyzed data. M.J.B., C.L.M. and A.O.V. wrote the manuscript. D.W. maintained and differentiated the PSCs. S.C.d.H. generated and provided the MESP1+ RNA. M.B. generated and provided the original hPSC lines. V.V.O. helped with experimental design. R.P. and C.L.M. supervised the study. All authors edited the manuscript.

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Correspondence to Christine L Mummery.

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Birket, M., Ribeiro, M., Verkerk, A. et al. Expansion and patterning of cardiovascular progenitors derived from human pluripotent stem cells. Nat Biotechnol 33, 970–979 (2015). https://doi.org/10.1038/nbt.3271

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