Clin Res Cardiol 108, Suppl 2, October 2019

Influence of substrate stiffness and growth area on structural and functional maturation of induced pluripotent stem cell-derived cardiomyocytes

A. Körner1, J. Balitzki1, M. Hecker1, N. Ullrich2

1Abteilung für Herz- und Kreislaufphysiologie, Universität Heidelberg, Heidelberg; 2Institut für Physiologie und Pathophysiologie, Universitätsklinikum Heidelberg, Heidelberg;

To overcome the inability of the adult heart to regenerate after injury, modern therapeutic approaches are heading for the use of novel cardiomyocytes (CMs) derived from induced pluripotent stem cells (iPSC-CMs) to repair diseased myocardium. Despite their cardiogenic electrophysiological profile and contractile activity, these cells present a highly variable and immature functional phenotype. In order to meet the complex functional features of adult CMs for adequate integration, new strategies are needed to enhance cardiac maturation in iPSC-CMs. In this study, we investigated the influence of defined physical cues during cell culture on structural and functional maturation of iPSC-CMs. We tested the hypothesis that specific cell geometry and growth surface stiffness influence the cardiogenic phenotype of these cells at the level of excitation-contraction (EC) coupling. Murine iPSC-CMs were grown on culture surfaces with defined stiffnesses (glass: 20 gPa; polydimethylsiloxan (PDMS): 255 kPa, 28 kPa, 15 kPa and 1.5 kPa). Cell viability was tested over a period of 24 days. Using the technique of microcontact printing, cell shape was determined to form either cuboid cells based on the shape of adult CMs or hexagonal cells as control. Structural remodeling was assessed by immunocytochemistry and confocal imaging of cardiac proteins. Contractile properties were evaluated by measuring calcium (Ca2+) transients using the Ca2+-sensitive fluorescent indicator fura-2 and cell shortening by edge detection in electrically paced iPSC-CMs. Our data revealed an increased viability of iPSC-CMs grown on soft PDMS surfaces compared with glass, which was significant during prolonged culture times (>14 days). The influence of cell shape on structural remodeling was evaluated considering the deviation of myofibrils from the long axis of the cells grown on microprinted patterns. Our analysis revealed strong parallel alignment of myofibrils in cuboid cells compared to hexagonally shaped cells. Analysis of Ca2+ transients showed faster Ca2+ release and removal dynamics on softer growth surfaces, expressed as decreased time-to-peak and decay rates of Ca2+ transients indicative of functional maturation of Ca2+ handling properties. In addition, time-to-peak of cell shortening increased on softer matrices indicating enhanced contraction efficiency.  Taken together, our results demonstrate that soft surfaces provide a better substrate for iPSC-CMs cell growth and promote maturation of Ca2+ handling mechanisms resulting in enhanced EC coupling. Furthermore, establishment of a cell axis induced strong structural remodeling processes toward a more mature and adult-like phenotype. We conclude that cell shape and surrounding material stiffnesses are critical determinants for structural and functional maturation of iPSC-CMs and need to be considered for future optimization processes of iPSC-CM production and clinical applications.

https://www.abstractserver.com/dgk2019/ht/abstracts//BS289.htm