Subsequent to implantation of the mouse blastocyst there is rapid proliferation of the egg cylinder, which consists of three cell types: the more proximal extra-embryonic ectoderm, the more distal embryonic ectoderm or epiblast, and an outer layer of visceral endoderm [29]. The visceral endoderm originates from the primitive endoderm, a layer of cells organized at E4.5, which is composed of cells from the ICM of the E3.5 blastocyst expressing Gata6 but not Nanog [30]. At ∼E5.5, a specialized cluster of endoderm cells, the DVE, arises at the distal tip of the embryo. DVE cells migrate toward the prospective anterior, to form the anterior visceral endoderm (AVE). DVE/AVE cells secrete molecules such as cerberus (Cer1) and Lefty1, antagonists of the Transforming Growth Factor β (TGFβ)-related protein Nodal [29]. These antagonists restrict the activity of Nodal to the posterior pole of the embryo at E6.0 [31]. The primitive streak forms at E6.5, indicating that gastrulation has begun, and gives rise to the mesoderm and definitive endoderm germ layers [29].
