Morpho-functional comparison of differentiation protocols to create iPSC-derived cardiomyocytes.

Autor:	Nijak A; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium., Simons E; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium., Vandendriessche B; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium., Van de Sande D; Laboratory of Molecular Biophysics, Cellular and Network Excitability, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium., Fransen E; StatUa Center of Statistics, University of Antwerp 2650, Antwerp, Belgium., Sieliwończyk E; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium., Van Gucht I; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium., Van Craenenbroeck E; Department of Cardiology, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, Antwerp 2650, Belgium., Saenen J; Department of Cardiology, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, Antwerp 2650, Belgium., Heidbuchel H; Department of Cardiology, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, Antwerp 2650, Belgium., Ponsaerts P; Laboratory of Experimental Hematology, Vaccine & Infectious Disease Institute, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium., Labro AJ; Laboratory of Molecular Biophysics, Cellular and Network Excitability, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium.; Department of Basic and Applied Medical Sciences, Faculty of Medicine and Health Sciences, Ghent University, Ghent 9000, Belgium., Snyders D; Laboratory of Molecular Biophysics, Cellular and Network Excitability, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium., De Vos W; Laboratory of Cell Biology and Histology, Faculty of Veterinary Sciences, University of Antwerp, Antwerp 2610, Belgium., Schepers D; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium.; Laboratory of Molecular Biophysics, Cellular and Network Excitability, Department of Biomedical Sciences, University of Antwerp, Antwerp 2610, Belgium., Alaerts M; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium., Loeys BL; Center of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp & Antwerp University Hospital, Antwerp 2650, Belgium.; Department of Human Genetics, Radboud University Medical Centre, Nijmegen 6525, The Netherlands.
Jazyk:	angličtina
Zdroj:	Biology open [Biol Open] 2022 Feb 15; Vol. 11 (2). Date of Electronic Publication: 2022 Feb 23.
DOI:	10.1242/bio.059016
Abstrakt:	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. Competing Interests: Competing interests The authors declare no competing or financial interests. (© 2022. Published by The Company of Biologists Ltd.)
Databáze:	MEDLINE
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