Introduction The epicardium is a heterogeneous cell layer covering the mammalian heart. During embryogenesis, the epicardial lineage is essential to heart and vascular development, yielding cardiac fibroblasts and coronary vascular smooth muscle cells. The epicardium also has a trophic effect on developing cardiomyocytes. It is quiescent in adulthood but reactivates post-injury to a limited degree to yield cardiac fibroblasts, which allows fast healing, yet also causes fibrosis. Epicardial functional heterogeneity remains incompletely characterised.

Methods The Sinha group has derived a robust model of human epicardium (hpsc-epi) from human pluripotent stem cells; this was used for single-cell RNA sequencing (scRNA-seq). Immunohistochemistry and immunocytochemistry were used to validate scRNA-seq results in primary human foetal tissue. The candidate gene BNC1 has been investigated by siRNA-mediated knockdown studies in vitro.

Results Single-cell RNA-seq identified two main epicardial subpopulations in hpsc-epi: WT1^high/BNC1^high/TCF21^low and WT1^low/BNC1^low/TCF21^high. Here we show validation of our scRNA-seq data in human foetal epicardium by immunohistochemistry in cryosections and human foetal epicardial explants, confirming our hpsc-epi model is representative of the in vivo situation. We show preliminary data from siRNA-mediated knockdown of BNC1, which indicate this gene may play a role in epicardial function, possibly in regulating cell migration in a model of epithelial-to-mesenchymal transition.

Implications Improved understanding of developmental epicardial regulation could pave the way towards harnessing epicardial potential in prospective strategies to aid revascularisation and regeneration of the injured heart.