Metabolomic Profile of Oviductal Extracellular Vesicles across the Estrous Cycle in Cattle

Abstract

Oviductal extracellular vesicles (oEVs) have been proposed as key modulators of gamete/embryo maternal interactions. The aim of this study was to examine the metabolite content of oEVs and its regulation across the estrous cycle in cattle. Oviductal EVs were isolated from bovine oviducts ipsilateral and contralateral to ovulation at four stages of the estrous cycle (post-ovulatory stage, early and late luteal phases, and pre-ovulatory stage). The metabolomic profiling of EVs was performed by proton nuclear magnetic resonance spectroscopy (NMR). NMR identified 22 metabolites in oEVs, among which 15 were quantified. Lactate, myoinositol, and glycine were the most abundant metabolites throughout the estrous cycle. The side relative to ovulation had no effect on the oEVs' metabolite concentrations. However, levels of glucose-1-phosphate and maltose were greatly affected by the cycle stage, showing up to 100-fold higher levels at the luteal phase than at the peri-ovulatory phases. In contrast, levels of methionine were significantly higher at peri-ovulatory phases than at the late-luteal phase. Quantitative enrichment analyses of oEV-metabolites across the cycle evidenced several significantly regulated metabolic pathways related to sucrose, glucose, and lactose metabolism. This study provides the first metabolomic characterization of oEVs, increasing our understanding of the potential role of oEVs in promoting fertilization and early embryo development.

Oviduct Fluid Extracellular Vesicles Change the Phospholipid Composition of Bovine Embryos Developed In Vitro

“…In the study from Alminana et al [10] and ours, frozen-thawed oEVs collected from oviductal fluids at the postovulatory stage were used. Furthermore, as previously reported [7], the proportions of exosomes (30-100 nm: 76%) and of microvesicles (100-500 nm: 24%), as measured by transmission electron microscopy in the oEV samples used, were very similar in the two studies. However, the differences between the two studies may be due to differences in oEV content in the culture media: based on preliminary (unpublished) data comparing different doses of oEVs proteins (0.005, 0.05, 0.5 mg/mL) in the culture medium, the 0.05 dose tended to increase the rates of blastocysts at days 7-8 compared to controls without oEVs and was chosen for the present study, whereas Alminana et al [10] used final concentrations of 0.2-0.4 mg of proteins/mL.…”

“…The oEVs used were obtained from a pool of bovine oviductal fluids collected at a local commercial slaughterhouse and isolated by centrifugation (12,000× g, 15 min) and two successive ultracentrifugations (100,000× g, 90 min), then characterized by transmission electron microscopy and western blotting as detailed in our previous study [7]. Only oviducts at the postovulatory stage of cycle (recently formed corpus luteum, absence of follicles >10 mm; estimated Days 1-5 postovulation) and ipsilateral to ovulation, i.e., at the expected time and site of embryo development, were used.…”

“…EVs are lipid bilayer-enclosed nanosized vesicles, including exosomes (40-100 nm) and microvesicles (100-1000 nm) [6]. As EVs in other biological fluids, oEVs contain a range of biomolecules such as proteins, small RNAs, metabolites, and genomic DNA [4,[7][8][9]. Using fluorescent-labelled oEVs and confocal microscopy, it was shown that bovine blastocysts were able to take up oEVs added to the culture medium and internalize them inside their blastomeres [10].…”

The Oviductal Extracellular Vesicles’ RNA Cargo Regulates the Bovine Embryonic Transcriptome

“…Given the wide range of oEV components recently identified in our laboratory (mRNAs, proteins, ncRNAs including miRNAs, snoRNAs, snRNAs, and metabolites) [19,20], it is difficult to select potential candidates as modulators of embryonic development. To date, only a few miRNAs and proteins have been proven to be responsible for oEVs' functional effects on spermatozoa [11,16], while the functional impact of the oEV cargo on embryos and the extent of those effects is not yet fully understood.…”

Embryo–Maternal Interactions Underlying Reproduction in Mammals

“…Bridi et al summarize the current knowledge about the potential roles of both maternal (including oviduct and uterus) and embryonic EVs in the crucial events leading to successful pregnancy [ 7 ]. To increase the current knowledge of the oviductal EVs molecular cargo, Gatien et al focused on a less known cargo, characterizing metabolomic profile of bovine oviductal EVs during the estrous cycle [ 28 ]. Their study revealed up to 100-fold higher levels of glucose-1-phosphate and maltose at the luteal phase compared to the peri-ovulatory phases.…”