Investigation of Hematopoietic Stem Cell and Progenitor Populations: Implication for Cell Fate Determination and Lineage Commitment.

A thorough understanding of the lineage potential of each subset of hematopoietic stem cells (HSC) and progenitor populations is critical to establish an accurate map of cell fate determination during hematopoietic development. A controversy exists whether multipotentiality is conserved until a mutually exclusive segregation of myeloid and lymphoid potentials or whether early progenitor populations sequentially lose lineage potential as they differentiate from the long-term self-renewing HSC (LT-HSC), starting with loss of megakaryocyte/erythrocyte (MegE) potential.

Hematopoietic cells at different developmental stages can be prospectively isolated based on a combination of cell surface phenotypes and functional assays in vitro and in vivo. However, assessment of lineage potential of cells other than LT-HSC is complicated by the progressive loss of self-renewal activity in progenitor populations and the lack of congenic surface markers on mature cells of the MegE lineage. Using sensitive in vitro and in vivo approaches, we quantitatively and kinetically assessed the MegE potential of Lineage⁻/c-Kit⁺/Sca-1⁺ (KLS) subsets of mouse bone marrow, including LT-HSC (Thy1.1^int/Flk-2⁻), sort-term HSC (ST-HSC^F: Thy1.1^int/Flk-2⁺) and multipotent progenitor population (MPP^F: Thy1.1⁻/Flk-2⁺), and compared it with the MegE potential of downstream myeloid progenitors (CMP, GMP and MEP) and with their ability to give rise to mature myelomonocytic and lymphoid cells. In contrast to previous reports, we demonstrate that Flk2-positive ST-HSC^F and MPP^F populations have readily detectable but transient MegE potential in vivo that is more robust than committed myeloid progenitors CMP and MEP. We also show that these cells make clonal colonies in vitro and in vivo in the spleen that contained megakaryocytes and erythrocytes. Moreover, we established the kinetics of mature cell production from each stem and progenitor population, hence providing the timing of these early differentiation events in vivo that is of critical importance when investigating lineage potential.

Our results demonstrate that multipotentiality is retained in the KLS “stem cell” fraction of the bone marrow and support a model of hematopoietic development with mutually exclusive segregation of myeloid and lymphoid lineage potential. Taken together with previous findings, they indicate that transition from LT-HSC to ST-HSC^F and then to MPP^F, is accompanied by progressive lose of self-renewal ability, increased proliferation and change in gene expression programs to prepare multipotent cells to leave the stem cell niche and undergo lineage differentiation. This model is by definition a simplification of a complex biological process but accounts for most, if not all, differentiation events, tolerates plasticity in lineage segregation at early steps of commitment and it accommodates intrinsic lineage preferences during ontogeny and aging.