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Human Embryonic Stem Cell-Derived Cardiomyocytes Self-Arrange with Areas of Different Subtypes During Differentiation.

Stem cells and development (2017-08-11)
Maj Linea Vestergaard, Søren Grubb, Karen Koefoed, Zoe Anderson-Jenkins, Kristina Grunnet-Lauridsen, Kirstine Calloe, Christian Clausen, Søren Tvorup Christensen, Kjeld Møllgård, Claus Yding Andersen
ABSTRACT

The derivation of functional cardiomyocytes (CMs) from human embryonic stem cells (hESCs) represents a unique way of studying human cardiogenesis, including the development of CM subtypes. In this study, we investigated the development and organization of hESC-derived cardiomyocytes (hESC-CMs) and examined how the expression levels of CM subtypes correspond to human in vivo cardiogenesis. Beating clusters were used to determine cardiac differentiation, which was evaluated by the expression of cardiac genes GATA4 and TNNT2 and subcellular localization of GATA4 and NKX2.5. Sharp electrode recordings to determine action potentials (APs) further revealed spatial organization of intracluster CM subtypes (ie, complex clusters). Nodal-, atrial-, and ventricular-like AP morphologies were detected within distinct regions of complex clusters. The ability of different CM subtypes to self-organize was documented by immunohistochemical analyses and a differential spatial expression of β-III tubulin, myosin light chain 2v (MLC-2V), and α-smooth muscle actin (α-SMA). Furthermore, all hESC-CM subtypes formed expressed primary cilia, which are known to coordinate cellular signaling pathways during cardiomyogenesis and heart development. This study expands the foundation for studying regulatory pathways for spatial and temporal CM differentiation during human cardiogenesis.

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Casein from bovine milk, technical grade