The fine line between efficient maturation and metabolic stress in human engineered heart tissues

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) represent an unlimited source for human cardiomyocytes to study the physiology and pathophysiology of the heart. The application of hiPSC-CMs in engineered heart muscles (EHMs) together with human ventricular fibroblasts improves their maturation state and allows direct force measurements to assess physiologically relevant characteristics of human myocardium. 

In this study, we aimed to enhance the maturation of EHMs through cultivation in a recently developed lipid-enriched maturation medium (MM) and by the supplementation of growth factors VEGF, bFGF and IGF-1 (MM+GFs), which are commonly used in EHM media. 

EHMs in MM and MM+GFs were similar with respect to general characteristics such as morphology, cross-sectional area, and tissue weight. Interestingly, we observed only slightly increased contraction force of EHMs cultured in MM+GFs compared to MM, and no differences in their response to isoprenaline stimulation. However, EHMs cultured in MM+GFs had strongly increased spontaneous beat rate and showed an arrhythmic behaviour, compared to EHMs in MM. Increased expression levels of NPPA, NPPB and IL-1β supported our hypothesis of an enhanced stress state of EHMs in MM+GFs compared to MM. Furthermore, we observed an increased stiffness of EHMs and enhanced expression of COL1A1, COL3A1 and TGF-β in EHMs cultured in MM+GFs in comparison to MM, indicating that GF supplementation induced fibrosis. Measurement of lipid staining intensities in hiPSC-CMs isolated from EHMs revealed an increased lipid accumulation in CMs from EHMs cultured in MM+GFs vs. MM, suggesting that GF-supplementation may cause lipotoxicity. Further experiments using standard 2D hiPSC-CM culture confirmed an increase in lipid accumulation, reactive oxygen species (ROS) levels and mitochondrial superoxide levels in CMs cultured in MM+GFs compared to MM. Importantly, these characteristics represent hallmarks of myocardial tissue from patients with metabolic diseases such as obesity or type 2 diabetes^1,2. 

Taken together, the results of our study highlight the fine line between the efficient maturation and metabolic stress of EHMs and iPSC-CMs, especially when combining lipid-enriched media with growth factors. Understanding these processes may pave the way towards more adult-like human heart models, and eventually provide insights about the development of heart disease.