An ultrastructural study of mouse and rat embryo implantation sites was undertaken to determine whether the uterine luminal epithelial cells surrounding the blastocyst exhibited the morphologic characteristics of apoptotic or necrotic cell death. In both species the epithelial cells exhibited all of the characteristics of apoptosis, including surface blebbing, shrinkage and fragmentation of the cells, condensation of chromatin, and indentation and fragmentation of nuclei. Cytoplasmic organelles remained morphologically intact, and the cytoplasm maintained normal or increased staining density. Also, the epithelial cells and cell fragments were phagocytosed by the adjacent trophoblast cells. The epithelial cells did not exhibit the characteristics of necrotic cell death, such as swollen cells and mitochondria, damaged surface membranes, and disintegrated cytoplasmic organelles. We conclude that uterine epithelial cells surrounding mouse and rat embryos during implantation undergo apoptotic cell death leading to their phagocytosis by trophoblast cells.
Ham's F-10 medium (Gibco, Grand Island, NY) and medium T6 with and without 15% fetal calf serum (FCS) were compared for their ability to support development of murine blastocysts with the capacity to implant and produce normal fetuses when transferred to pseudopregnant females. All media supported equal rates of blastocyst development from 2-cell embryos. In addition, there were no differences in the rates of blastocyst implantation. However, once implanted, blastocysts grown in T6 produced a significantly higher proportion of normal fetuses (58.5% to 65.9%) than blastocysts grown in either Ham's F-10 (2.4%) or T6 with FCS (27.6%). These results demonstrate that the rate of murine blastocyst development from 2-cell embryos in vitro is not a good criterion of healthy embryos. Murine blastocysts transferred in medium with 0% versus 50% FCS implanted and developed into normal fetuses at equal rates.
Blastocysts were recovered from intact mice at various times on the fourth and fifth days of pregnancy and incubated in vitro with 35S-methionine. Labeled proteins synthesized by the embryos and secreted into the medium were separated in two-dimensions on polyacrylamide gels by electrophoresis and localized by fluorography. The array of proteins synthesized and secreted by late stage blastocysts was found to be qualitatively and quantitatively different from those released by embryos at earlier stages of development. Similar changes were also observed in secreted proteins when delayed-implanting embryos were reactivated after an injection of estrogen. Furthermore, there was a temporal correlation between the appearance of certain proteins secreted by the embryos and changes in specific proteins synthesized and released by the uterus. It is suggested that these various secreted proteins constitute a signal-response mechanism that is important for the process of embryo implantation in mice.