Bovine oviductal and uterine extracellular vesicles enhance blastocyst development in confined individual embryo culture

Abstract Purpose In vitro -produced embryos exhibit reduced developmental competence compared to their in vivo -derived counterparts due to the inability to accurately replicate the complex milieu of the maternal reproductive tract. A key aspect in vivo is the confinement of embryos in small volumes of oviductal and uterine fluids, where maternal extracellular vesicles (EVs) deliver their cargos into developing embryos, influencing their competence. The main aim of this study was to investigate the effects of the sequential exposure to oviductal and uterine EVs on the developmental competence of bovine embryos cultured in vitro under different embryo densities in groups or individually. Methods Presumptive zygotes were cultured in groups of 25 in 25 μL drops under oil (embryo density: 1/1 µL), or individually in ~ 70 nL in microwell chambers (embryo density: 14.28/1 µL), in medium alone or supplemented with oviductal EVs at days 1–4, and then with uterine EVs at days 5–8 of embryo culture. Results Sequential EVs exposure increases the blastocyst rates, mean cell numbers, and reduces the accumulation of large detrimental lipid droplets only in individually cultured embryos. Blastocyst apoptotic cells and lipid area were reduced by sequential EVs exposure both in individually and group cultured embryos. Conclusions We suggest that confined conditions may either enhance the accumulation of embryo-derived autocrine/paracrine factors acting synergistically with maternal EVs, or allow presumptive EV-induced embryo secretions to reach biologically effective levels only under individual culture. High-density embryo may have a key role in future culture strategies aimed to improve embryo development and competence through supplementation of bioactive maternal factors.