Endometrial cells and extracellular vesicles response to high body energy reserves in bovine: Insights into miRNA and mRNA regulation before embryo arrival

Body energy reserves influence reproductive performance in cattle. Previous findings from our laboratory showed that cows with high body energy reserves (HBER) have lower ovulation and embryo recovery rates compared to cows with moderate reserves (MBER). To investigate whether these reproductive differences are associated with changes in the uterine environment, Nelore cows from the same herd were assigned to MBER or HBER groups through nutritional management. Following estrous synchronization and artificial insemination, animals were slaughtered ∼120 h after ovulation induction. Samples from the uterotubal junction (UTJ) and anterior uterine horn (ANT) were collected. Extracellular vesicles (EVs) were isolated from uterine fluid by flushing, and endometrial tissue was sampled for molecular analysis. Nanoparticle tracking analysis revealed no differences in EV concentration or size between groups. However, when comparing MBER and HBER groups, miRNA profiling identified 8 and 9 differentially expressed miRNAs between MBER and HBER in EVs from the UTJ and ANT, respectively, and 2 differentially expressed miRNAs in endometrial cells from the UTJ, suggesting potential differences in molecular profiles. Transcriptomic analysis of endometrial cells revealed 430 and 35 differentially expressed genes (DEGs) in the UTJ and ANT, respectively, between MBER and HBER groups. The higher number of DEGs in the UTJ may suggest a greater molecular response, which is reflected by more extensive pathway enrichment compared to the ANT. miRNA–mRNA integration, performed by intersecting predicted miRNA targets with the differentially expressed mRNAs from our RNA-seq data, suggests that differentially expressed genes may be regulated by miRNAs altered between groups, indicating miRNA-mediated effects of metabolic condition on the uterine transcriptome. These findings suggest that high body energy reserves are associated with enrichment of immune and metabolism related pathways in the uterine environment, especially in the UTJ, which may reflect a pro-inflammatory, metabolically altered state potentially impairing early embryo development and maternal-embryonic communication.