Robust cardiomyocyte differentiation from human pluripotent stem cells via temporal modulation of canonical Wnt signaling

Proc Natl Acad Sci U S A. 2012 Jul 3;109(27):E1848-57. doi: 10.1073/pnas.1200250109. Epub 2012 May 29.

Abstract

Human pluripotent stem cells (hPSCs) offer the potential to generate large numbers of functional cardiomyocytes from clonal and patient-specific cell sources. Here we show that temporal modulation of Wnt signaling is both essential and sufficient for efficient cardiac induction in hPSCs under defined, growth factor-free conditions. shRNA knockdown of β-catenin during the initial stage of hPSC differentiation fully blocked cardiomyocyte specification, whereas glycogen synthase kinase 3 inhibition at this point enhanced cardiomyocyte generation. Furthermore, sequential treatment of hPSCs with glycogen synthase kinase 3 inhibitors followed by inducible expression of β-catenin shRNA or chemical inhibitors of Wnt signaling produced a high yield of virtually (up to 98%) pure functional human cardiomyocytes from multiple hPSC lines. The robust ability to generate functional cardiomyocytes under defined, growth factor-free conditions solely by genetic or chemically mediated manipulation of a single developmental pathway should facilitate scalable production of cardiac cells suitable for research and regenerative applications.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Cell Culture Techniques / methods*
  • Cell Differentiation / physiology
  • Culture Media / pharmacology
  • Enzyme Inhibitors / pharmacology
  • Glycogen Synthase Kinase 3 / antagonists & inhibitors
  • Glycogen Synthase Kinase 3 / metabolism
  • Humans
  • Myocytes, Cardiac / cytology*
  • Pluripotent Stem Cells / cytology*
  • Pluripotent Stem Cells / metabolism*
  • RNA, Small Interfering / genetics
  • Transforming Growth Factor beta / metabolism
  • Wnt Signaling Pathway / drug effects
  • Wnt Signaling Pathway / physiology*
  • beta Catenin / genetics

Substances

  • CTNNB1 protein, human
  • Culture Media
  • Enzyme Inhibitors
  • RNA, Small Interfering
  • Transforming Growth Factor beta
  • beta Catenin
  • Glycogen Synthase Kinase 3