Embryonic endothelial evolution towards first hematopoietic stem cells revealed by single-cell transcriptomic and functional analyses

摘要

Hematopoietic stem cells (HSCs) in adults are believed to be born from hemogenic endothelial cells (HECs) in mid-gestational embryos.Due to the rare and transient nature,the HSC-competent HECs have never been stringently identified and accurately captured,let alone their genuine vascular precursors.Here,we first used high-precision single-cell transcriptomics to unbiasedly examine the relevant EC populations at continuous developmental stages with intervals of 05 days from embryonic day (E) 9.5 to E11.0.As a consequence,we transcriptomically identified two molecularly different arterial EC populations and putative HSC-pdmed HECs,whose number peaked at E10.0 and sharply decreased thereafter,in the dorsal aorta of the aorta-gonad-mesonephros (AGM) region.Combining computational prediction and in vivo functional validation,we precisely captured HSC-competent HECs by the newly constructed Neud3-EGFP reporter mouse model,and realized the enrichment further by a combination of surface markers (Procr+Kit+CD44+,PK44).Surprisingly,the endothelial-hematopoietic dual potential was rarely but reliably witnessed in the cultures of single HECs.Noteworthy,primitive vascular ECs from E8.0 experienced twc-step fate choices to become HSC-primed HECs,namely an initial arterial fate choice followed by a hemogenic fate conversion.This finding resolves several previously observed contradictions.Taken together,comprehensive understanding of endothelial evolutions and molecular programs underlying HSC-primed HEC specification in vivo will facilitate future investigations directing HSC production in vitro.