Transcriptional dynamics of pluripotent stem cell-derived endothelial cell differentiation revealed by single-cell RNA sequencing.
Ian R McCracken, Richard S Taylor, Fatma O Kok, Fernando de la Cuesta, Ross Dobie, Beth E P Henderson, Joanne C Mountford, Axelle Caudrillier, Neil C Henderson, Chris P Ponting, Andrew H Baker
Author Information
Ian R McCracken: Centre for Cardiovascular Science, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, UK.
Richard S Taylor: Centre for Cardiovascular Science, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, UK.
Fatma O Kok: Centre for Cardiovascular Science, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, UK.
Fernando de la Cuesta: Centre for Cardiovascular Science, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, UK.
Ross Dobie: Centre for Inflammation Research, University of Edinburgh, Edinburgh EH16 4TJ, UK.
Beth E P Henderson: Centre for Inflammation Research, University of Edinburgh, Edinburgh EH16 4TJ, UK.
Joanne C Mountford: Scottish National Blood Transfusion Service, Edinburgh EH14 4BE, UK.
Axelle Caudrillier: Centre for Cardiovascular Science, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, UK.
Neil C Henderson: Centre for Inflammation Research, University of Edinburgh, Edinburgh EH16 4TJ, UK.
Chris P Ponting: MRC Human Genetics Unit, IGMM, University of Edinburgh, Edinburgh EH4 2XU, UK.
Andrew H Baker: Centre for Cardiovascular Science, University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, UK.
AIMS: Pluripotent stem cell-derived endothelial cell products possess therapeutic potential in ischaemic vascular disease. However, the factors that drive endothelial differentiation from pluripotency and cellular specification are largely unknown. The aims of this study were to use single-cell RNA sequencing (scRNA-seq) to map the transcriptional landscape and cellular dynamics of directed differentiation of human embryonic stem cell-derived endothelial cells (hESC-EC) and to compare these cells to mature endothelial cells from diverse vascular beds. METHODS AND RESULTS: A highly efficient directed 8-day differentiation protocol was used to generate a hESC-derived endothelial cell product (hESC-ECP), in which 66% of cells co-expressed CD31 and CD144. We observed largely homogeneous hESC and mesodermal populations at Days 0 and 4, respectively, followed by a rapid emergence of distinct endothelial and mesenchymal populations. Pseudotime trajectory identified transcriptional signatures of endothelial commitment and maturation during the differentiation process. Concordance in transcriptional signatures was verified by scRNA-seq analysis using both a second hESC line RC11, and an alternative hESC-EC differentiation protocol. In total, 105 727 cells were subjected to scRNA-seq analysis. Global transcriptional comparison revealed a transcriptional architecture of hESC-EC that differs from freshly isolated and cultured human endothelial cells and from organ-specific endothelial cells. CONCLUSION: A transcriptional bifurcation into endothelial and mesenchymal lineages was identified, as well as novel transcriptional signatures underpinning commitment and maturation. The transcriptional architecture of hESC-ECP was distinct from mature and foetal human EC.
