Servicenavigation

Project Details

Back

Projekt Print View

Structural remodeling and functional maturation of stem cell-derived cardiomyocytes in novel 3D microprinted scaffolds.

Subject Area Anatomy and Physiology
Cell Biology
Term from 2019 to 2023
Project identifier Deutsche Forschungsgemeinschaft (DFG) - Project number 417543243
 
Final Report Year 2023

Final Report Abstract

Human cardiomyocytes derived from induced pluripotent stem cells (hiPSC-CM) form a promising basis for the development of new cell-based therapeutic approaches for heart diseases. However, despite successful cardiogenic differentiation, these cells retain characteristics of immature cardiomyocytes, particularly in terms of structural cell organization, their microarchitecture, the arrangement and orientation of myofibrils, and spontaneous contractility. In order to achieve successful clinical application of hiPSC-CM in the future, it is necessary that hiPSC-CM have the same physiological complexity as mature, adult cardiomyocytes. For this reason, we need a better understanding of the various external triggers as well as the molecular mechanisms that drive the maturation of cardiomyocytes towards the adult stage. These new findings will serve to develop new strategies to improve the cardiac phenotype in hiPSC-CM. In this project, we tested the hypothesis that structural and functional characteristics of hiPSC-CM are influenced by biomechanical properties of the immediate cell environment in culture. Thus, we postulated that maturation of cardiomyocytes can be achieved by optimizing culture conditions. Using direct laser writing technology in collaboration with Prof. Martin Bastmeyer's team at KIT, we developed flexible 3D scaffolds made of light-sensitive polyethylene glycol-based polymers in defined rectangular and, as a control, polygonal shapes in order to match the cuboid adult cell shape as closely as possible. hiPSC-CM were seeded into these precoated 3D molds and cultured in the predefined geometry. In addition, we forced the expression of a membrane protein (BIN1) that is suspected of being involved in the formation of transverse tubules. We investigated the effect of different growth forms of single cells on cell morphology and orientation of the contractile myofibrils, as well as the resulting functional changes at the level of Ca2+ regulation during excitation-contraction (EC) coupling. Structural changes in the cell membrane caused by reshaping and BIN1 expression, as well as the expression patterns of relevant proteins for Ca2+ regulation were evaluated by immunocytochemistry. Functional analysis of intracellular Ca2+ signals was performed by live imaging using a Ca2+-sensitive fluorescent dye (Fluo-4) via confocal laser scanning microscopy. Compared to cells cultured without peripheral constraints, rectangular shapes mimicking the characteristic shape of adult cardiomyocytes improved myofibril alignment in hiPSC-CM, formed transverse tubule precursors, and synchronized Ca2+ transients, resulting in improved temporal properties cytosolic Ca2+ signals. These results suggest a functional maturation of intracellular Ca2+ regulation during EC coupling. In addition, cuboid cardiomyocytes showed improved rhythmicity in their spontaneous activity. In summary, we showed that anisotropic cell shape critically influences the morphological and functional development of hiPSC-CM at the level of Ca2+ signaling during EC coupling. These special requirements of hiPSC-CM must be taken into account in the future when developing cardiac cell constructs for regenerative therapeutic approaches so that functional integration becomes compatible with adult cardiac tissue.

Publications

 
 

Additional Information

© 2025 DFG Disclaimer / Copyright / Privacy Policy

Textvergrößerung und Kontrastanpassung