Feeder-Free Generation and Characterization of Endocardial and Cardiac Valve Cells from Human Pluripotent Stem Cells
Valvular heart disease presents a significant health burden, yet advancements in valve biology and novel therapeutics have been hindered by the lack of accessibility to human valve cells. In this study, we have developed a scalable and feeder-free method to differentiate human induced pluripotent stem cells (iPSCs) into endocardial cells. Importantly, we show that these endocardial cells are transcriptionally and phenotypically distinct from vascular endothelial cells and can be directed to undergo endothelial-to-mesenchymal transition (EndMT) to generate cardiac valve cell populations. Following this, we identified two distinct populations—one population undergoes EndMT to become valvular interstitial cells (VICs), while the other population reinforces their endothelial identity to become valvular endothelial cells (VECs). We then characterized our iPSC-derived cell populations and identified putative markers for these populations through bulk RNAseq transcriptome analyses. Lastly, we validated our VIC and VEC populations by comparing our transcriptomic data to pseudobulk data generated from single-cell RNAseq of normal valve tissue from a 15-week-old human fetus. By increasing the accessibility to these cell populations, we aim to accelerate discoveries for cardiac valve biology and disease. Overall design: Two wild-type iPSC lines (MSN02-4 and WTC11) were used. Each of these lines were differentiated to endocardial, endothelial, valvular interstitial cells (VICs), and valvular endothelial cells (VECs). These differentiated cell populations were sorted and total RNA was extracted for bulk RNAseq. Paired-end 150bp reads were performed at a depth of ~20 million reads per sample. A total of 3 replicates for each cell type, from each iPSC line was sequenced.