Myosin expression and contractile function are altered by replating stem cell-derived cardiomyocytes

Myosin heavy chain (MyHC) is the main determinant of contractile function. Human ventricular cardiomyocytes (CMs) predominantly express the β-isoform. We previously demonstrated that ∼80% of human embryonic stem cell-derived cardiomyocytes (hESC-CMs) express exclusively β-MyHC after long-term cultur...

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Published in:	The Journal of general physiology Vol. 155; no. 11; p. 1
Main Authors:	Osten, Felix, Weber, Natalie, Wendland, Meike, Holler, Tim, Piep, Birgit, Kröhn, Simon, Teske, Jana, Bodenschatz, Alea K, Devadas, Santoshi Biswanath, Menge, Kaja S, Chatterjee, Shambhabi, Schwanke, Kristin, Kosanke, Maike, Montag, Judith, Thum, Thomas, Zweigerdt, Robert, Kraft, Theresia, Iorga, Bogdan, Meissner, Joachim D
Format:	Journal Article
Language:	English
Published:	United States Rockefeller University Press 06-11-2023
Subjects:	Cardiomyocytes Cell Biology Cell culture Cellular Physiology Contractility Embryo cells Focal adhesion kinase Gene expression Humans Immunofluorescence Induced Pluripotent Stem Cells Integrins Kinases Laminin Mechanotransduction Myocytes, Cardiac Myofibrils Myosin Myosin Heavy Chains - genetics Myosins Pluripotency Signal Transduction Stem cells Ventricle
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Summary:	Myosin heavy chain (MyHC) is the main determinant of contractile function. Human ventricular cardiomyocytes (CMs) predominantly express the β-isoform. We previously demonstrated that ∼80% of human embryonic stem cell-derived cardiomyocytes (hESC-CMs) express exclusively β-MyHC after long-term culture on laminin-coated glass coverslips. Here, we investigated the impact of enzymatically detaching hESC-CMs after long-term culture and subsequently replating them for characterization of cellular function. We observed that force-related kinetic parameters, as measured in a micromechanical setup, resembled α- rather than β-MyHC-expressing myofibrils, as well as changes in calcium transients. Single-cell immunofluorescence analysis revealed that replating hESC-CMs led to rapid upregulation of α-MyHC, as indicated by increases in exclusively α-MyHC- and in mixed α/β-MyHC-expressing hESC-CMs. A comparable increase in heterogeneity of MyHC isoform expression was also found among individual human induced pluripotent stem cell (hiPSC)-derived CMs after replating. Changes in MyHC isoform expression and cardiomyocyte function induced by replating were reversible in the course of the second week after replating. Gene enrichment analysis based on RNA-sequencing data revealed changes in the expression profile of mechanosensation/-transduction-related genes and pathways, especially integrin-associated signaling. Accordingly, the integrin downstream mediator focal adhesion kinase (FAK) promoted β-MyHC expression on a stiff matrix, further validating gene enrichment analysis. To conclude, detachment and replating induced substantial changes in gene expression, MyHC isoform composition, and function of long-term cultivated human stem cell-derived CMs, thus inducing alterations in mechanosensation/-transduction, that need to be considered, particularly for downstream in vitro assays.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Disclosures: T. Thum reported other from Cardior Pharmaceuticals outside the submitted work. No other disclosures were reported. S. Kröhn’s current address is Clinic for Nuclear Medicine, Hannover Medical School, Hannover, Germany. B. Iorga and J.D. Meissner contributed equally to this paper. F. Osten and N. Weber contributed equally to this paper.
ISSN:	0022-1295 1540-7748
DOI:	10.1085/jgp.202313377