Myeloid Lineage Restriction Is Prior to and Independent of Lymphoid Lineage Restriction.

Abstract

It has been widely accepted that hematopoietic stem cells (HSCs) exclusively give rise to either myeloid or lymphoid lineage along their differentiation. Phenotypically defined as FcRloCD34+IL-7R−Sca-1−Kit+Lin− cells, common myeloid progenitors (CMPs) are supposedly at the first branched point for myeloid lineage commitment. Phenotypically defined as Thy1.1−IL-7R+KitloSca-1loLin− cells, common lymphoid progenitors are supposedly at the first branched point for both B- and T-lymphoid lineage commitment. Contradicting this model, we previously made observations of the myeloid lineage restriction in very early stages of HSC differentiation. We therefore decided to compare the myeloid and lymphoid differentiation potentials in long-term HSCs, short-term HSCs, multipotent progenitors (MPPs), CMPs, granulocyte/macrophage progenitors (GMPs), megakaryocyte/erythrocyte progenitors (MEPs), and CLPs at the single cell level. Because any available assays are not so sensitive enough for detection of lymphoid lineage differentiation potential, we primarily focused on myeloid differentiation potentials in this study. Here we provide data indicative of the absence of CMPs in the adult mouse bone marrow. Long-term HSCs (CD34−Kit+Sca-1+Lin− cells), short-term HSCs (Flt-3−CD34+Kit+Sca-1+Lin− cells), MPPs (Flt-3+CD34+Kit+Sca-1+Lin− cells), CMPs (FcRloCD34+IL-7R−Sca-1−Kit+Lin− cells), GMPs (FcRhiCD34+IL-7R−Sca-1−Kit+Lin− cells), MEPs (FcRloCD34−IL-7R−Sca-1−Kit+Lin− cells), and CLPs (IL-7R+KitloSca-1loLin− cells) were purified from adult B6 mice by flow cytometry. Single cell cultures were performed in the presence of SCF+TPO+IL-3+EPO. All colonies made by single cells were subjected to Cytospin preparations, followed by May-Gruenwald-Giemsa staining, for morphological classifications of colony cells: neutrophil (n), macrophage (m), erythroblast (E), or megakaryocyte (M). Cells with all n, m, E, M differentiation potentials (nmEM cells) were detected in on average 31%, 5%, and fewer than 1 % of long-term HSCs, short-term HSCs, and MPPs, repectively. Notably, none of a total of over 800 CMPs showed the full nmEM differentiation potential. These CMPs instead showed potentials belonging to members of GMPs and MEPs, suggesting CMPs are mostly an overlaping population of GMPs and MEPs rather than a distinct population. Single cell transplantation experiments revealed the coexistence of cells with the myleoid and B-lymphoid potentials or cells with the myeloid and T-lymphoid potentials among CD34−Kit+Sca-1+Lin− long-term HSC population. These progentiors are likely to be immediate progeny of HSCs. Together, these data support our view that myeloid lineage restriction takes place prior to and independent of lymphoid lineage restriction.