The emergence and development of the hematopoietic and endothelial lineages in the developing murine embryo are intimately linked. Both mesoderm-derived lineages appear to descend from a common precursor cell, the hemangioblast, and require yolk sac visceral endoderm-derived factors for normal progenitor cell proliferation and differentiation and for normal blood island formation.
Vascular endothelial growth factor A (VEGF-A) represents one of the critical molecules that is required for yolk sac blood island development. VEGF-A interaction with Flk1 (kinase domain receptor [KDR] or VEGF receptor-2) on extraembryonic mesoderm cells appears to be required for proper mesoderm migration to the proximal yolk sac where the blood islands develop, but not for the initial emergence and differentiation of hematopoietic and endothelial cells from mesoderm cells. Thus, Flk1 receptor null embryos fail to develop blood islands, though Flk1 null embryonic stem (ES) cell–derived embryoid bodies produce some hematopoietic and endothelial cells. Little is known of the intracellular signaling molecules that act down-stream of VEGF-A/Flk1 interactions in the yolk sac mesoderm cells or hemangioblasts leading to blood island formation.
The cytoplasmic tyrosine kinase Fps/Fes has been implicated in several receptor signaling pathways regulating endothelial cell migration and capillary tube formation. Activation of Fps/Fes via myristoylation leads to plasma membrane localization and increased kinase activity in endothelial cells, and expression of myristoylated human fps (MFps/Fes) in transgenic mice results in the development of widespread hypervascularity and multifocal hemangiomas in the MFps/Fes–expressing animals.
Similarities in vascular transformation in endothelial cells expressing activated MFps/Fes and excessive VEGF-A/Flk1 signaling led Haigh and colleagues (page ) to investigate whether Fps/Fes plays a role in VEGF-A/Flk1 signaling. VEGF-A stimulation of endothelial cell lines in vitro recruited Fps/Fes from cytoplasmic vesicles to the plasma membrane and was associated with increased Fps/Fes tyrosine phosphorylation, suggesting that Fps/Fes might be a downstream mediator of VEGF-A/Flk1 signaling. Flk1 null ES cells overexpressing MFps/Fes were generated, and it was demonstrated that embryoid bodies derived from these clones contain migratory endothelial cells and vascular channel formation, in contrast with control Flk1 null ES cell–derived embryoid bodies that lacked such elements. Chimeric embryo studies confirmed that Flk1 null ES-derived cells failed to contribute to vessel formation in vivo. In contrast, embryos chimeric for Flk1 null ES-derived cells expressing MFps/Fes demonstrated a 30-fold increase in contribution to the endothelial cell lineage. However, the embryos containing a high percentage of Flk1 null ES-derived cells expressing MFps/Fes displayed numerous aberrant vascular structures in the yolk sac and defects in cardiovascular and craniofacial development. These and other results suggest that constitutive MFps/Fes activity in the absence of normal Flk1 signaling leads to exuberant vasculogenesis/angiogenesis. Finally, Haigh and colleagues demonstrate that MFps/Fes expression in Flk1 null or heterozygous ES cells results in an increase in migratory behavior of mesodermal cells in vivo and a 5- to 10-fold increase in hemangioblast development in vitro. Though no direct interaction between Flk1 and Fps/Fes was demonstrated, the results of the study suggest that Fps/Fes may play a role downstream of VEGF-A/Flk1 signaling and/or complement a parallel signaling pathway.
One surprising result was that MFps/Fes expression failed to rescue hematopoietic development in the Flk1 null ES cells. The authors suggest that a more detailed evaluation may indicate some level of rescue that was missed in the current study. As vascular smooth muscle cells are also derived from Flk1-expressing cells, future studies may also wish to address whether MFps/Fes rescues this lineage in Flk1 null ES cells.
