Comprehensive human stem cell differentiation in a 2D and 3D mode to cardiomyocytes for long-term cultivation and multiparametric monitoring on a multimodal microelectrode array setup.

Human pluripotent stem cell derived cardiomyocytes are a promising cell source for research and clinical applications like investigation of cardiomyopathies and therefore, identification and testing of novel therapeutics as well as for cell based therapy approaches. However, actually it´s a challenge to generate matured adult cardiomyocyte-like phenotype in a reasonable time. Moreover, there is a lack of applicable non-invasive label-free monitoring techniques providing quantitative parameters for analysing the culture stability and maturation status. In this context, we established an efficient protocol based on a combined differentiation of hiPSC in 2D cultures followed by a forced reaggregation step that leads to highly enriched (>90% cardiomyocytes) cardiomyocyte clusters. Interestingly, 3D cultures revealed an accelerated maturation as well as phenotype switch from atrial to ventricular cardiomyocytes. More strikingly using combined impedimetric and electrophysiological monitoring the high functionality and long-term stability of 3D cardiomyocyte cultures, especially in comparison to 2D cultures could be demonstrated. Additionally, chronotropic as well as QT-prolongation causing reference compounds were used for validating the cardio specific and sensitive reaction over the monitored time range of more than 100 days. Thus, the approach of multiparametric bioelectronic monitoring offers capabilities for the long-term quantitative analysis of hiPS derived cardiomyocyte culture functionality and long-term stability. Moreover, the same multiparametric bioelectronic platform can be used in combination with validated long-term stable cardiomyocyte cultures for the quantitative detection of compound induced effects. This could pave the way for more predictive in vitro chronic/repeated dose cardiotoxicity testing assays.