title: Stiff matrix induces switch to pure β-cardiac myosin heavy chain expression in human ESC-derived cardiomyocytes.
abstract: Human pluripotent stem cell (hPSC)-derived cardiomyocytes hold great potential for in vitro modeling of diseases like cardiomyopathies. Yet, knowledge about expression and functional impact of sarcomeric protein isoforms like the myosin heavy chain (MyHC) in hPSC-cardiomyocytes is scarce. We hypothesized that ventricular β-MyHC expression alters contraction and calcium kinetics and drives morphological and electrophysiological differentiation towards ventricular-like cardiomyocytes. To address this, we (1) generated human embryonic stem cell-derived cardiomyocytes (hESC-CMs) that switched towards exclusive β-MyHC, and (2) functionally and morphologically characterized these hESC-CMs at the single-cell level. MyHC-isoforms and functional properties were investigated during prolonged in vitro culture of cardiomyocytes in floating cardiac bodies (soft conditions) vs. culture on a stiff matrix. Using a specific anti-β-MyHC and a newly generated anti-α-MyHC-antibody, we found individual cardiomyocytes grown in cardiac bodies to mostly express both α- and β-MyHC-protein isoforms. Yet, 35 and 75 days of cultivation on laminin-coated glass switched 66 and 87 % of all cardiomyocytes to exclusively express β-MyHC, respectively. Twitch contraction and calcium transients were faster for CMs on laminin-glass. Surprisingly, both parameters were only little affected by the MyHC-isoform, although hESC-CMs with only β-MyHC had much lower ATP-turnover and tension cost, just as in human ventricular cardiomyocytes. Spontaneous contractions and no strict coupling of β-MyHC to ventricular-like action potentials suggest that MyHC-isoform expression does not fully determine the hESC-CM differentiation status. Stiff substrate-induced pure β-MyHC-protein expression in hESC-CMs, with several contractile parameters close to ventricular cardiomyocytes, provides a well-defined in vitro system for modeling of cardiomyopathies and drug screening approaches.
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Natalie
authors.initials: N
authors.lastname: Weber
authors.affiliation: Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany.|Leibniz Research Laboratories for Biotechnology and Artificial Organs, Hannover Medical School, Hannover, Germany.|HTTG, LEBAO and REBIRTH-Center for Regenerative Medicine, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Kristin
authors.initials: K
authors.lastname: Schwanke
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Stephan
authors.initials: S
authors.lastname: Greten
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Meike
authors.initials: M
authors.lastname: Wendland
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.|Department of Physical Chemistry, Faculty of Chemistry, University of Bucharest, Bucharest, Romania.
authors.forename: Bogdan
authors.initials: B
authors.lastname: Iorga
authors.affiliation: Institute of Neurophysiology, Hannover Medical School, Hannover, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Fischer
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Cornelia
authors.initials: C
authors.lastname: Geers-Knörr
authors.affiliation: Institute of Functional and Applied Anatomy, Hannover Medical School, Hannover, Germany.|REBIRTH Excellence Cluster, Hannover, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Hegermann
authors.affiliation: Institute of Functional and Applied Anatomy, Hannover Medical School, Hannover, Germany.|REBIRTH Excellence Cluster, Hannover, Germany.
authors.forename: Christoph
authors.initials: C
authors.lastname: Wrede
authors.affiliation: Institute of Molecular and Translational Therapeutic Strategies (IMTTS), Hannover Medical School, Hannover, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Fiedler
authors.affiliation: Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany.|Leibniz Research Laboratories for Biotechnology and Artificial Organs, Hannover Medical School, Hannover, Germany.|HTTG, LEBAO and REBIRTH-Center for Regenerative Medicine, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Henning
authors.initials: H
authors.lastname: Kempf
authors.affiliation: Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany.|Leibniz Research Laboratories for Biotechnology and Artificial Organs, Hannover Medical School, Hannover, Germany.|HTTG, LEBAO and REBIRTH-Center for Regenerative Medicine, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Annika
authors.initials: A
authors.lastname: Franke
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Birgit
authors.initials: B
authors.lastname: Piep
authors.affiliation: Institute of Molecular and Translational Therapeutic Strategies (IMTTS), Hannover Medical School, Hannover, Germany.
authors.forename: Angelika
authors.initials: A
authors.lastname: Pfanne
authors.affiliation: Institute of Molecular and Translational Therapeutic Strategies (IMTTS), Hannover Medical School, Hannover, Germany.|Integrated Research and Treatment Center Transplantation (IFB-Tx) and REBIRTH Excellence Cluster, Hannover Medical School, Hannover, Germany.|National Heart and Lung Institute, Imperial College London, London, UK.
authors.forename: Thomas
authors.initials: T
authors.lastname: Thum
authors.affiliation: Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany.|Leibniz Research Laboratories for Biotechnology and Artificial Organs, Hannover Medical School, Hannover, Germany.|HTTG, LEBAO and REBIRTH-Center for Regenerative Medicine, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Martin
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Brenner
authors.affiliation: Department of Cardiac, Thoracic, Transplantation and Vascular Surgery, Hannover Medical School, Hannover, Germany. Zweigerdt.Robert@mh-hannover.de.|Leibniz Research Laboratories for Biotechnology and Artificial Organs, Hannover Medical School, Hannover, Germany. Zweigerdt.Robert@mh-hannover.de.|HTTG, LEBAO and REBIRTH-Center for Regenerative Medicine, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany. Zweigerdt.Robert@mh-hannover.de.
authors.forename: Robert
authors.initials: R
authors.lastname: Zweigerdt
authors.affiliation: Institute of Molecular and Cell Physiology, Hannover Medical School, Carl Neuberg Str. 1, 30625, Hannover, Germany. Kraft.Theresia@mh-hannover.de.
authors.forename: Theresia
authors.identifier: 0000-0002-8386-8088
authors.initials: T
authors.lastname: Kraft
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000502:physiology; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D046529:Microscopy, Electron, Transmission; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000096:biosynthesis*; D016133:Polymerase Chain Reaction; D020033:Protein Isoforms; D060888:Real-Time Polymerase Chain Reaction; D024723:Ventricular Myosins / Q000096:biosynthesis*
chemical_list: D020033:Protein Isoforms; D024723:Ventricular Myosins; D018995:Myosin Heavy Chains
keywords: Calcium transients; Cardiac myosin heavy chain isoforms; Cardiomyocytes; Pluripotent stem cells; Tension cost; Twitch kinetics