Morpho-functional comparison of differentiation protocols to create iPSC-derived cardiomyocytes
Aleksandra Nijak,
Eline Simons,
Bert Vandendriessche,
Dieter Van de Sande,
Erik Fransen,
Ewa Sieliwończyk,
Ilse Van Gucht,
Emeline Van Craenenbroeck,
Johan Saenen,
Hein Heidbuchel,
Peter Ponsaerts,
Alain J. Labro,
Dirk Snyders,
Winnok De Vos,
Dorien Schepers,
Maaike Alaerts,
Bart L. Loeys
Affiliations
Aleksandra Nijak
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Eline Simons
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Bert Vandendriessche
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Dieter Van de Sande
Laboratory of Molecular Biophysics, Cellular and Network Excitability, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium
Erik Fransen
StatUa Center of Statistics, University of Antwerp 2650, Antwerp, Belgium
Ewa Sieliwończyk
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Ilse Van Gucht
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Emeline Van Craenenbroeck
Department of Cardiology, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, Antwerp 2650, Belgium
Johan Saenen
Department of Cardiology, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, Antwerp 2650, Belgium
Hein Heidbuchel
Department of Cardiology, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, Antwerp 2650, Belgium
Peter Ponsaerts
Laboratory of Experimental Hematology, Vaccine & Infectious Disease Institute, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium
Alain J. Labro
Laboratory of Molecular Biophysics, Cellular and Network Excitability, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium
Dirk Snyders
Laboratory of Molecular Biophysics, Cellular and Network Excitability, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium
Winnok De Vos
Laboratory of Cell Biology and Histology, Faculty of Veterinary Sciences, University of Antwerp, Antwerp 2610, Belgium
Dorien Schepers
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Maaike Alaerts
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Bart L. Loeys
Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium
Cardiomyocytes derived from induced pluripotent stem cells (iPSC-CMs) offer an attractive platform for cardiovascular research. Patient-specific iPSC-CMs are very useful for studying disease development, and bear potential for disease diagnostics, prognosis evaluation and development of personalized treatment. Several monolayer-based serum-free protocols have been described for the differentiation of iPSCs into cardiomyocytes, but data on their performance are scarce. In this study, we evaluated two protocols that are based on temporal modulation of the Wnt/β-catenin pathway for iPSC-CM differentiation from four iPSC lines, including two control individuals and two patients carrying an SCN5A mutation. The SCN5A gene encodes the cardiac voltage-gated sodium channel (Nav1.5) and loss-of-function mutations can cause the cardiac arrhythmia Brugada syndrome. We performed molecular characterization of the obtained iPSC-CMs by immunostaining for cardiac specific markers and by expression analysis of selected cardiac structural and ionic channel protein-encoding genes with qPCR. We also investigated cell growth morphology, contractility and survival of the iPSC-CMs after dissociation. Finally, we performed electrophysiological characterization of the cells, focusing on the action potential (AP) and calcium transient (CT) characteristics using patch-clamping and optical imaging, respectively. Based on our comprehensive morpho-functional analysis, we concluded that both tested protocols result in a high percentage of contracting CMs. Moreover, they showed acceptable survival and cell quality after dissociation (>50% of cells with a smooth cell membrane, possible to seal during patch-clamping). Both protocols generated cells presenting with typical iPSC-CM AP and CT characteristics, although one protocol (that involves sequential addition of CHIR99021 and Wnt-C59) rendered iPSC-CMs, which were more accessible for patch-clamp and calcium transient experiments and showed an expression pattern of cardiac-specific markers more similar to this observed in human heart left ventricle samples.