HiPS-cardiomyocyte derived engineered heart tissues (EHT) as a tool for cardiomyocytes maturation improvement

Abstract

Advances in human induced pluripotent stem cell (hiPSC) technology and the ability to create patient-derived hiPSC lines has raised the possibility for studying pathology-associated specific mutations consequences. However, studies focusing on cardiac pathologies suffer from the lack of maturity of hiPSC-derived cardiomyocytes (CM) cultured in 2D conditions, the difficulty for measuring physiologic parameters such as contractility because of their impaired cellular structure, and the lack of organized interfaces between CM. EHT, which were shown to favor hiPSC maturation both at the intra- and intercellular levels, represent a promising tool to overcome these hurdles. We describe the establishment of the EHT technology together with readouts to assess CM maturation degree. hiPSC were differentiated into CM through a small molecule-based protocol. 3D strip-format fibrin-based EHT were then generated from freshly isolated CM and kept in culture for > 3 weeks under auxotonic stretch conditions to allow progressive CM maturation. Over the culture period, the force developed by the EHT, contraction-relaxation parameters and beating frequency–reflecting together CM maturation–were assessed by video-optical recording. MUSCLEMOTION® was used for data extraction. At the end of the culture period, EHT were fixed for intracellular structures evaluation via immunofluorescence. We report the successful establishment of a standardized protocol combining hiPSC differentiation into CM and their subsequent use for EHT production. We show that CM casted in 3D EHT submitted to auxotonic stretch conditions exhibit signs of advanced maturation. These results raise the possibility of the EHT technology transposition to patient-derived iPSC lines allowing for comparative studies between mutation carrying and isogenic control cell lines to define the impact of precise genetic mutations on CM contractile function and structure.