Adhesion and fusion of the extraembryonic epiblast

A tissue culture model system has been devised to examine the attachment, expansion, and fusion of epithelial cell sheets. A normal embryonic epithelial tissue, the extraembryonic epiblast of the chick, is isolated mechanically and cultured on its natural substratum, the vitelline membrane. This persistently migratory tissue has distinct adhesive and non-adhesive regions. A serum-free chemically-defined culture medium has been formulated that permits determination of the effects of individual growth and trophic factors. Attachment of transferred epiblasts is dependent upon the presence of mineralocorticoids in the medium. This suggests that fluid transport is required for the cell sheet to make its initial attachment to the culture substratum. Expansion of the cell sheet following attachment, and the fusion of epiblasts advancing toward each other, does not require the presence of mineralocorticoid. No exogenous adhesive glycoproteins are required for attachment, expansion, or fusion. Antibody localization shows that endogenous laminin is present on the attachment surface of the specialized adhesive edge region of the extraembryonic epiblast. Following fusion of confronted epiblasts into one coherent cell sheet, the laminin disappears. Throughout these studies the adhesive and non-adhesive regions of the epiblast are identified by their characteristic distributions of actin microfilaments, localized using rhodamine-phalloidin staining.