Functional characterization of human pluripotent stem cell-derived arterial endothelial cells

Significance Generating fully functional arterial endothelial cells is a critical problem for vascular development and disease research. Currently, the arterial endothelial cells derived from human pluripotent stem cells lack the range of arterial-specific functions in vitro and the protective function for ischemic tissues in vivo. Here, we combine single-cell RNA sequencing and CRISPR-Cas9 technology to identify pathways for regulating arterial endothelial cell differentiation. We then manipulate these pathways and generate arterial endothelial cells that demonstrate unprecedented arterial-specific functions as well as improve survival of myocardial infarction. These findings facilitate the understanding of vascular development and disease and provide a source of cells that have broad applications for vascular disease modeling and regenerative medicine. Here, we report the derivation of arterial endothelial cells from human pluripotent stem cells that exhibit arterial-specific functions in vitro and in vivo. We combine single-cell RNA sequencing of embryonic mouse endothelial cells with an EFNB2-tdTomato/EPHB4-EGFP dual reporter human embryonic stem cell line to identify factors that regulate arterial endothelial cell specification. The resulting xeno-free protocol produces cells with gene expression profiles, oxygen consumption rates, nitric oxide production levels, shear stress responses, and TNFα-induced leukocyte adhesion rates characteristic of arterial endothelial cells. Arterial endothelial cells were robustly generated from multiple human embryonic and induced pluripotent stem cell lines and have potential applications for both disease modeling and regenerative medicine.

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