Extracellular Vesicles, Lipid Droplets and AhR Ligands in Early Implantation: The Dynamics of Embryo-Maternal Crosstalk

Intercellular communication is essential for healthy embryo development, yet the role and dynamics of extracellular space in the maternal-embryonic dialogue remain unclear. Furthermore, little is known of maternal and embryonic metabolic states during early endometrial preparation and after the embryo enters the uterine cavity. Using a human in vitro co-culture model and extracellular vesicle (EV)-specific tools, we dynamically tracked EV secretion, uptake and processing between embryonic and endometrial cells at early stages of cell-to-cell communication. Hormonal stimulation altered endometrial secretory output, producing distinct EV populations. Stimulated EVs (St-EVs) differed from non-stimulated EVs (NSt-EVs) in size, secretion dynamics, uptake efficiency and metabolic cargo, selectively packaging energy-related metabolites and aryl hydrocarbon receptor (AhR) ligands. AhR inhibition increased spheroid attachment, suggesting that AhR signalling regulates implantation by modulating the endometrial environment. Additionally, lipid droplets (LDs) affected by endometrial- and embryo-derived EVs were actively secreted and taken up by embryonic cells, highlighting their role in implantation. EVs were not only exchanged between the embryo and endometrium but were also rapidly internalized, influencing mitochondrial activity, lipid metabolism and extracellular matrix remodelling. Translation of EV-derived mRNA occurred within 1 h of uptake, driving cellular changes and enhancing embryo attachment. These findings suggest EVs, extracellular metabolites and LDs mobilized between the endometrium and embryo coordinate to promote embryo attachment and implantation. This study advances our understanding of embryo-maternal EV-mediated communication and provides a valuable model for investigating EV-mediated simultaneous intercellular bidirectional crosstalk in other biological contexts.