Characterization and generation of human definitive multipotent hematopoietic stem/progenitor cells

Definitive hematopoiesis generates hematopoietic stem/progenitor cells (HSPCs) that give rise to all mature blood and immune cells, but remains poorly defined in human. Here, we resolve human hematopoietic populations at the earliest hematopoiesis stage by single-cell RNA-seq. We characterize the distinct molecular profiling between early primitive and definitive hematopoiesis in both human embryonic stem cell (hESC) differentiation and early embryonic development. We generate definitive HSPCs from hESCs that hold the multipotency to differentiate various blood and immune cells, as validated by single-cell clonal assay. Strikingly, the generated HSPCs from hESCs give rise to various blood and lymphoid lineages in vivo. Lastly, we characterize gene-expression dynamics in definitive and primitive hematopoiesis and reveal an unreported role of ROCK-inhibition in enhancing human definitive hematopoiesis. Our study provides a prospect for understanding human early hematopoiesis and a firm basis to generate blood and immune cells for clinical purposes.