Histological and histochemical analysis of embryoid bodies

We examined the histological structure of embryoid bodies arising from aggregation of mouse embryonic stem (ES D3) cells after 7, 12, 18 and 26 days of in vitro culture. Morphology of originally solid embryoid bodies was affected by the process of cavitation that resulted in formation of cystic embryoid bodies and by spontaneous differentiation of the ES D3 cells. We applied in situ immunophenotyping to characterise cell populations that spontaneously differentiated inside the embryoid bodies in the various stages. The most distinct cell populations that were found inside embryoid bodies were alpha-fetoprotein-positive endodermal cells and myogenic cells that expressed desmin, myogenin or smooth muscle actin. ES D3-derived endothelial cells generated during vasculogenesis inside the embryoid bodies differed from mature endothelial cells because they did not stain for von Willebrand factor. These cells also differed from endothelial cell that were generated during angiogenesis since they did not stain for the intermediate filament nestin. Our results demonstrate the usefulness of this in vitro model for studying early embryogenesis.