DOI RECORD
Oviduct epithelial spheroids as a model to study maternal communication with gametes and embryos in sheep
Abstract
The influence of sperm quality on embryo development and the challenges of in vitro embryo production in sheep highlight the need for models that reproduce oviductal functions and physiological sperm-selection mechanisms. Therefore, this study aimed to establish and characterize an in vitro model of ovine oviduct epithelial spheroids from the ampulla (AES) and isthmus (IES), evaluate their maintenance under different culture conditions, and investigate sperm–oviduct interactions. Pairs of oviducts were collected from cycling ewes, and mucosal fragments were cultured for 5 days in DMEM/F-12 to generate AES and IES. Spheroids were then cultured for 7 days in 500 μL DMEM/F-12 (DM/500), 25-μL DMEM/F-12 (DM/25), or 25-μL synthetic oviductal fluid (SOF/25). Spheroid morphology, cell viability, number and ciliation were evaluated, as well as cytokeratin, vimentin and OVGP1 immunodetection. Spheroid motion velocity was used to evaluate ciliary activity. Finally, the ability of AES and IES to bind frozen-thawed ram sperm was assessed. Overall, 88% of AES and IES remained in suspension with more than 80% cell viability during culture, although cavity maintenance, diameter and cell number decreased over time in all conditions. The expression of cytokeratin, vimentin and OVGP1 was maintained in AES and IES in all conditions. Cell numbers per spheroid and ciliary activity were better maintained in DM compared to SOF. The density of bound sperm per surface unit was higher on IES than AES. Collectively, AES and IES preserved the morphofunctional properties of the oviduct epithelium and provide a valuable tool to investigate gamete/embryo interactions.