ISOLATION AND PROPAGATION OF YOLK-SAC-DERIVED ENDOTHELIAL-CELLS FROM A HYPERVASCULAR TRANSGENIC MOUSE EXPRESSING A GAIN-OF-FUNCTION FPS FESPROTOONCOGENE/

We report on the isolation and propagation of endothelial cells from t he mouse embryonic yolk sac, the earliest site of blood vessel develop ment, and on the advantages of a hypervascular transgenic mouse source of these cells. These transgenic mice express multiple copies of an a ctivated allele of the human fps/fes proto-oncogene and display hyperv ascularity progressing to multifocal hemangiomas. This phenotype sugge sted a role of the fps/fes proto-oncogene in vasculogenesis and angiog enesis and led us to investigate the growth characteristics of polk-sa c-derived endothelial cells from transgenic fps/fes embryos. We have e stablished eight independent cell clones from a mixture of transgenic and control yolk sacs from Day 12 embryos. Southern blot hybridization analysis showed all eight clones to be derived from transgenic cells, suggesting a growth advantage of cells carrying the activated fps/fes gene. A cell line, Clone 166 (C166), established from one of these cl ones, was more fully characterized. C166 exhibits normal endothelial c haracteristics, such as rearrangement into tubelike structures when pl aced on Matrigel, expression of angiotensin converting enzyme, retenti on of cobblestone morphology at confluence, and the presence of cell s urface receptors for acetylated low density lipoprotein. The cells con stitutively express murine endothelial cell adhesion molecule VCAM-1 a nd the vascular addressin identified by antibody MECA-99. As expected, the cell line expresses high levels of the cytoplasmic protein-tyrosi ne kinase encoded by the fps/fes protooncogene. The clone we have desc ribed as well as other endothelial cell lines that we have established from the mouse embryonic yolk sac should prove useful for the study o f endothelial cell differentiation and for the determination of the me chanisms underlying the establishment of organ-specific endothelial ce ll heterogeneity.