The continuous and life-long production of all mature blood cells in the circulation and hematopoietic tissues is contingent upon hematopoietic stem cells (HSCs). These rare cells, at the base of the hematopoietic differentiation hierarchy, reside in the bone marrow of adult mammals. In human medicine, HSCs are clinically important cells in transplantation therapies for blood-related diseases and leukemias. The ability to manipulate adult HSCs, particularly to expand the limiting numbers of these cells in vitro, has been difficult. Despite the isolation and characterization of many hematopoietic growth factors, HSC numbers can only (at best) be maintained in culture. Taking a developmental approach, we have been investigating the origins and growth of HSCs in the mouse embryo. We conjectured that at some early stage the embryo must induce the production of HSCs, generate a supportive microenvironment, and expand HSC numbers. With insight into these processes, novel factors to promote HSC induction/expansion may be discovered. We have shown that the aorta-gonads-mesonephros (AGM) region autonomously generates the first fully potent adult HSCs and that HSC activity is localized to the major vascular regions of the embryo body; the dorsal aorta, vitelline and umbilical arteries. Presently, the direct precursor cells to HSCs are unknown but are speculated to be hemangioblasts, hemogenic endothelial cells or mesenchymal cells in these vascular regions. Additionally, the lineage relationships of the first intraembryonic HSCs to those in the adult have not yet been established. Thus, we propose to examine cellular source of HSCs in the mouse embryo and to follow the migration of HSCs and the colonization of fetal and adult hematopoietic tissues by HSCs. Cultured whole mouse embryos will be injected with dye to fate map putative HSC precursors. Also, sorted putative HSC precursors will be injected into cultured whole mouse embryos to follow their migration and functional potential. Finally, cre-lox recombination will be used to specifically mark putative pre-HSCs so as to follow their functional potential and fate in fetal and adult hematopoietic tissues. These results should give insight into the source and complexity of adult HSC production.
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