Isogenic monocytes improve the responsiveness of hiPSC cardiac spheroids to cardiac stressors

AimsHeart failure remains a leading cause of morbidity and mortality worldwide. Suitable in vitro models to accurately replicate the pathological environment in heart failure with reduced and preserved ejection fraction (HFrEF/HFpEF) are limited, hampering mechanistic studies and drug screening. In particular, these models rarely incorporate immune cells, which play a critical role in heart failure. To address these limitations, we developed an isogenic 3D induced pluripotent stem cell (iPSC)-derived cardiac spheroid model incorporating monocytes. Methods and resultsCardiac spheroids were assembled from three healthy female iPSC lines: three-cell-type (3CT) spheroids consisting of iPSC-derived cardiomyocytes, cardiac fibroblasts, and endothelial cells, and four-cell-type (4CT) spheroids additionally containing monocytes. After six days of culture, established spheroids were treated for 24 h with different known heart failure-associated triggers (glucose & tumour necrosis factor alpha (TNF) or ischaemia with/without reoxygenation). Differences between treated and control 3CT and 4CT spheroids were investigated at the cellular, molecular, and functional levels using confocal microscopy, RNA expression (qPCR and RNA sequencing), protein secretion using proximity extension assay technology (Olink), and functional analyses of beating rate, contraction, and relaxation. The results confirmed successful monocyte integration in 4CT spheroids, and only spheroids with monocytes (4CTs) exhibited changes in beating rate and relaxation duration upon stimulation, highlighting the necessity of incorporating immune cells to successfully mimic heart failure-associated functional changes. Along with a more pronounced global transcriptomic treatment response and inflammatory changes, additional transcriptomic alterations previously linked to heart failure in patients, as well as changes in metabolism, ion channels, and extracellular matrix pathways, were observed in 4CT compared with 3CT spheroids. ConclusionWe showed that immune cell incorporation enhances the functional and transcriptional responses of engineered cardiac tissue to relevant heart failure triggers in vitro and is essential for future studies to elucidate the cellular crosstalk and pathomechanisms. Translational perspectiveHeart failure continues to be a predominant cause of morbidity and mortality, necessitating the development of innovative therapeutic strategies, particularly in light of the rising prevalence of obesity and diabetes mellitus. We introduced an isogenic in vitro spheroid model comprising iPSC-derived cardiomyocytes, cardiac fibroblasts, endothelial cells, and monocytes to examine the effects of heart failure-associated triggers on cardiac tissue. Our findings indicate that spheroids incorporating monocytes exhibit a more pronounced response to heart failure-associated triggers and demonstrate greater differential transcriptional and functional responses than spheroids lacking immune cells. This model