Differences in the metabolomic signatures of porcine follicular fluid collected from environments associated with good and poor oocyte quality

Abstract

The microenvironment of the developing follicle is critical to the acquisition of oocyte developmental competence, which is influenced by several factors including follicle size and season. The aim of this study was to characterise the metabolomic signatures of porcine follicular fluid (FF) collected from good and poor follicular environments, using high-resolution proton nuclear magnetic resonance ( 1 H-NMR) spectroscopy. Sow ovaries were collected at slaughter, 4 days after weaning, in summer and winter. The contents of small (3-4 mm) and large (5-8 mm) diameter follicles were aspirated and pooled separately for each ovary pair. Groups classified as summer-small (nZ8), summer-large (nZ15), winter-small (nZ9) and winter-large (nZ15) were analysed by 1 H-NMR spectroscopy.The concentrations of 11 metabolites differed due to follicle size alone (P!0.05), including glucose, lactate, hypoxanthine and five amino acids. The concentrations of all these metabolites, except for glucose, were lower in large FF compared with small FF. Significant interaction effects of follicle size and season were found for the concentrations of glutamate, glycine, N-acetyl groups and uridine. Succinate was the only metabolite that differed in concentration due to season alone (P!0.05). The FF levels of progesterone, androstenedione and oestradiol were correlated with the concentrations of most of the metabolites examined. The results indicate that there is a distinct shift in follicular glucose metabolism as follicles increase in diameter and suggest that follicular cells may be more vulnerable to oxidative stress during the summer months. Our findings demonstrate the power of 1 H-NMR spectroscopy to expand our understanding of the dynamic and complex microenvironment of the developing follicle.

Reproductive Physiology and Ovarian Folliculogenesis Examined via 1H-NMR Metabolomics Signatures: A Comparative Study of Large and Small Follicles in Three Mammalian Species (Bos taurus, Sus scrofa domesticus and Equus ferus caballus)

“…Lactate dehydrogenase catalyses the conversion of pyruvate to lactate, which in cows was at a reduced level in large FF compared with small FF. The finding that glucose levels increased and lactate levels decreased as follicle size increased is consistent with previous findings in cattle (Landau et al, 2000;Leroy et al, 2004), sheep (Ying et al, 2011), buffalo (Nandi et al, 2008), and pigs (Bertoldo et al, 2013). These results support the proposal that there is a distinct shift in carbohydrate metabolism as follicle development progresses and dominance is attained.…”

“…As previously described (Bertoldo et al, 2013), a linear mixed model was fitted to the data using the statistical software package GenStat 14 th edition (VSN International, Ltd., Hemel Hempstead, UK). For each metabolite, the treatment effects ''follicle type'' and ''species'' were assessed for significance ( p < 0.05), as was treatment interaction.…”

“…Therefore, one would expect the FF changes taking place as the follicles of sows increase in size to be more subtle than those occurring as the follicles of cows and mares attain dominance. Recently, Bertoldo et al (2013) reported an effect of follicle size on the metabolic profiles of sow FF and found differences in the concentrations of numerous metabolites that were also identified using 1 H-NMR spectroscopy. Despite the fact no significant differences were detected in the present study between the small FF and large FF samples in sows, the concentrations and relative variations of seven metabolites (acetate, alanine, glucose, lactate, phenylalanine, N-acetyl groups, and succinate) corresponded very closely with those for the same seven metabolites reported by Bertoldo et al (2013).…”

“…Recently, Bertoldo et al (2013) reported an effect of follicle size on the metabolic profiles of sow FF and found differences in the concentrations of numerous metabolites that were also identified using 1 H-NMR spectroscopy. Despite the fact no significant differences were detected in the present study between the small FF and large FF samples in sows, the concentrations and relative variations of seven metabolites (acetate, alanine, glucose, lactate, phenylalanine, N-acetyl groups, and succinate) corresponded very closely with those for the same seven metabolites reported by Bertoldo et al (2013). A possible explanation for the disparate findings in sows is that the diameters of the follicles aspirated differed slightly between the studies (small: 1-3 mm vs. 3-4 mm; large: 3-5 mm vs. 5-8 mm).…”

“…As previously described (Bertoldo et al, 2013), the 1 H-NMR spectra were performed on a Bruker DRX-500 spectrometer (Bruker SADIS, Wissembourg, France), operating at 11.7 T, with a Broad Band Inverse (BBI) probe head equipped with a Z gradient coil. Conventional 1 H-NMR spectra were recorded at 298 K with 90°pulse with residual water presaturation (single-frequency irradiation during the relaxation delay).…”