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.

Maturation of human induced pluripotent stem cell-derived cardiomyocytes for modeling hypertrophic cardiomyopathy

“…The activation kinetics we measured in hiPSC-CMs myofibrils (0.95-1.04) are lower than previously reported values in human atrial tissue myofibrils (3.73) 33 or in eSC-CM myofibrils expressing either mostly MYH7 or mixed MYH6/7 (1.70 or 2.44 respectively). 34 Together, these findings are indicative of sarcomeric maturity and potentially a more ventricular-like identity in our MPAT cells. As forced expression of MLC-2V results in increased contractility in isolated cardiomyocytes 35 , this could be a mechanism underlying the increased myofibril force generation we observe in MPAT cells.…”

A microwell cell capture device reveals variable response to dobutamine in isolated cardiomyocytes

“…These arrays allow a variety of functional measurements to be performed on many individual cells. Others have succeeded in relating functional measurements of individual iPSC-derived cardiomyocytes with cell-specific myosin heavy chain isoform expression (6). The microwell capture device described here is a technical advance that provides new and useful ways to link isolated adult cardiomyocyte function with gene expression.…”

Engineered Maturation Approaches of Human Pluripotent Stem Cell-Derived Ventricular Cardiomyocytes

“…Long-term cultivation and stiff matrix have been shown to enhance human pluripotent stem cell-derived cardiomyocyte sarcomere formation, calcium handling, and ion channel protein expression [26,27]. Collagen-coated polyacrylamide gels with an elastic moduli of 10 kPa have been shown to lead to aligned sarcomeres in comparison with a stiffer substrate [28].…”