Early megakaryocyte lineage-committed progenitors in adult mouse bone marrow

Abstract Hematopoietic stem cells (HSCs) have been considered to progressively lose their self-renewal and differentiation potentials prior to the commitment to each blood lineage. However, recent studies have suggested that megakaryocyte progenitors are generated at the level of HSCs. In this study, we newly identified early megakaryocyte lineage-committed progenitors (MgPs) in CD201-CD48- cells and CD48+ cells separated from the CD150+CD34-Kit+Sca-1+Lin- HSC population of the bone marrow in C57BL/6 mice. Single-cell transplantation and single-cell colony assay showed that MgPs, unlike platelet-biased HSCs, had little repopulating potential in vivo, but formed larger megakaryocyte colonies in vitro (on average eight megakaryocytes per colony) than did previously reported megakaryocyte progenitors (MkPs). Single-cell RNA-sequencing supported that these MgPs lie between HSCs and MkPs along the megakaryocyte differentiation pathway. Single-cell colony assay and single-cell RT-PCR analysis suggested the coexpression of CD41 and Pf4 is associated with megakaryocyte colony-forming activity. Single-cell colony assay of a small number of cells generated from single HSCs in culture suggested that MgPs are not direct progeny of HSCs. In this study, we propose a differentiation model in which HSCs give rise to MkPs through MgPs.