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One of the limitations in embryo implantation research is the lack of an available in vitro model that faithfully replicates embryo-uterine interactions. In previous studies, embryos were cultured on a monolayer of either uterine epithelial cells or extracellular matrix substratum on which embryos could adhere and outgrow. However, these models failed to display embryonic invasion, primarily because of the shortage of critical structural and molecular supports that are available in vivo. In the present study, we used intact mouse uterine endometrium collected on Day 4 of pregnancy and placed in contact with blastocysts to initiate coculture experiments in a defined medium at the air-liquid interface. The culture medium was composed of Ham F-12/Dulbecco modified Eagle medium (1:1), 30% fetal calf serum, 63.5 nmol/L of progesterone, 7.14 nmol/L of estradiol-17beta, 100 mug/ml of insulin, and 20 ng/ml of epidermal growth factor, whereas the incubation condition was mixed air of 50% oxygen, 5% carbon dioxide, and 45% nitrogen with a humidity of greater than 90% at 37 degrees C. Our observations from 24 h of culture clearly demonstrated that embryos were capable of attachment to the uterine endometrium and displayed partial invasion into the endometrial stroma. Interestingly, no outgrowth of trophoblasts on the surface of uterine endometrium was seen, but embryos exhibited a pole-specific attachment. Overall, this model is capable of demonstrating a true invasion of embryo within the endometrial stroma and may be suitable in studies related to early embryo implantation.

A Model for Implantation: Coculture of Blastocysts and Uterine Endometrium in Mice1