Introduction: The feto-maternal interface during bovine implantation was studied in vivo and using
three-dimensional bovine endometrial (BCECph) and trophoblast spheroids (CCS), each with underlying
Methods: The expression of ezrin and cytokeratin 18 (CK18) was analyzed via immunohistochemistry
(IHC), RT-PCR and western blotting in bovine endometrium (GD 18-44) with in vivo (VIVO) and in vitro-
produced embryos (VITRO). BCECph were stimulated with cotyledon-conditioned media (CCM) and
analyzed by TEM/SEM and IHC. CCS were stained (IHC) for TGC markers, to test if spheroidal trophoblast
cells had differentiated into TGC.
Results: At GD 20, caruncular epithelium (CE) and uterine glands (UG) showed a loss of cytosolic ezrin
and CK18 followed by a complete loss of both proteins. At GD 35 both reappeared in CE and UG. The
endometrial expression pattern did not differ between VIVO and VITRO. RT-PCR and western blotting
confirmed the presence of ezrin and CK18. All spheroids had an outer polarized, cytokeratin and ezrin positive epithelium (CE or trophoblast) with apical microvilli. Stimulation of BCECph with CCM induced
similar changes in ezrin expression as observed in endometrial tissue. However, no ultrastructural
alterations were found by transmission electron microscopy. Absence of TGC-specific glycoproteins in
CCS indicated that TGC differentiation was not induced by three-dimensional culture conditions.
Discussion: Ezrin and CK18 are downregulated during implantation in cattle. The expression changes
represent a temporal depolarization, which could be important for an establishment of bovine preg-
nancy. Our in vitro experiments demonstrate that the trophoblast could contribute to this change in vivo.