The overall goal of our work is to elucidate the roles and molecular basis of endothelial and hematopoietic cell interactions in the establishment of the hematopoietic system in the mouse. Examination of the mechanisms of yolk sac blood island organogenesis is important to understand how the first organ in the mouse is formed and as a model to investigate how the hematopoietic and endothelial lineages arise from a common precursor. However, reagents to identify the earliest events in yolk sac hematopoiesis have been lacking. We present novel preliminary data that CD41 expression marks the onset of primitive and definitive hematopoiesis in the murine embryo. We have utilized confocal microscopy to identify the site of emergence of the progenitor cells of both primitive and definitive progenitors. We have also developed a strategy that permits isolation of pure populations of endothelial cells from embryonic or adult mouse organs. We now propose to address the following areas where significant information gaps exist in our field: 1. The mechanisms of yolk sac blood island development remain undefined. We hypothesize that blood cell formation occurs concomitant with vasculogenesis but that primitive erythroblast progenitor cells are formed outside of any endothelial lined lumen. We further hypothesize that definitive hematopoietic progenitor cells emerge as clusters of cells from the endothelial lining of the blood islands. Thus, primitive and definitive progenitors arise via different mechanisms, raising the question of whether they share a common precursor. 2. Direct evidence of hematopoietic progenitor cell seeding of the liver is lacking. We hypothesize that the day 10 fetal liver is seeded with circulating yolk sac CD41bright definitive progenitor cells in vivo and that yolk sac CD41 bright progenitor cells will preferentially adhere to fetal liver-derived endothelial cells in vitro. 3. Cells from vascular grafts protect mice from radiation-induced death but do not possess hematopoietic stem cell activity (HSC). We present preliminary evidence that yolk sac endothelial cells preferentially maintain HSC repopulating ability ex vivo compared to adult kidney or liver endothelial cells. We hypothesize that yolk sac and embryo proper endothelial cells will facilitate HSC engraftment upon transplantation in vivo ? ?
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