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Glucuronic acid supplementation during maturation improves the zona reaction during in vitro fertilization of pig oocytes
An incomplete zona reaction during in vitro fertilization (IVF) could be responsible for high incidences of polyspermic penetration in pig oocytes. The objective of this study was to evaluate the effects of supplementing perivitelline space components during oocyte maturation on the incidence of polyspermic penetration, cortical granule exocytosis, and completion of the zona reaction. Oocytes (n=800) were supplemented during the last 24 h of maturation with either 0.01 mM glucuronic acid (GA), 0.01 mM N-acteylglucosamine (GlcNAc), 0.01 mM GA and GlcNAc, or 0.005 mM GA and GlcNAc and then subject to IVF and subsequent embryonic development. Oocytes were either evaluated for the presence of cortical granules using the intensity of fluorescence or zona pellucida hardness using a zona dissolution assay. Fertilization characteristics were evaluated 12 h after IVF and rates of embryonic cleavage and blastocyst development were observed at 48 h and 144 h after IVF, respectively. There were no significant differences in the percent of oocytes at metaphase II by then end of maturation, penetration rates, and male pronuclear formation. Oocytes supplemented with GA had a significantly less (P < 0.05) incidence of polyspermic penetration compared to unsupplemented (32.61 ± 4.80%) and GlcNAc only supplemented oocytes (39.78 ± 7.93%). Zona pellucida digestion times were significantly longer (P < 0.05) in oocytes that were supplemented with GA compared to those that were not. Fluorescent intensity of cortical granules was significantly decreased (P < 0.05) after IVF compared to the end of maturation for each treatment. Supplementation with GA had significantly lower (P < 0.05) fluorescent intensities of cortical granules after IVF compared to no supplementation and 0.01 mM GlcNAc supplementation. Supplementation of 0.005 mM GA and 0.005 mM GlcNAc to the maturation media produced a significantly higher (P < 0.05) percentage of embryos cleaved by 48 h after IVF (75.33 ± 5.14%) compared to the other treatment groups. Supplementation of GA during oocyte maturation produced a significant higher (P < 0.05) percentage of embryos reaching the blastocyst stage of development compared to no supplementation or 0.01 mM GlcNAc supplemented oocytes. The results of this study suggest that supplementing GA during oocyte maturation improves IVF success and subsequent embryo development in pigs.
Keywords:
zona reaction, polyspermy, pig