256 LEPTIN AND GLUCOSE INFLUENCE PORCINE NUCLEAR MATURATION

Abstract

Leptin (LEP), a product of the obese gene, regulates food intake and may be associated with the metabolic syndrome. In addition, leptin receptor has been identified in the luteal and granulosa cells of the pig, suggesting that this protein may play a role in fertility. Research indicates that LEP is associated with insulin sensitivity; thus, the addition of different concentrations of LEP and glucose (GLUC) in maturation media may have an impact on oocyte maturation. The objective of the current experiments was to determine the effect of LEP (Experiment 1), and LEP in combination with GLUC (Experiment 2) on the nuclear maturation of porcine oocytes. This could potentially be a good model system in which to study the effects of obesity and diabetes on oocyte quality. Cumulus–oocyte complexes were cultured in a chemically defined medium, Purdue porcine medium (PPM) for 42 h, in 7% CO₂ in air and at 38.7°C. In Experiment 1, medium was supplemented with 5 different concentrations (0, 1, 10, 20, and 100 ng mL^–1) of LEP, in the presence of 2 mm GLUC. Experiment 2 was designed as a 3 × 3 factorial, with LEP (0, 1, and 10 ng mL^–1) and GLUC (0, 5, and 50 mm). Oocytes were fixed and stained after IVM (136 to 155/treatment, 7 replicates; and 39 to 90/treatment, 5 replicates for Experiment 1 and 2, respectively). Nuclear maturation was scored as 1 (mature; T or MII) or 0 (not mature). Data were analyzed by GLM ANOVA and chi-square. There was no difference in nuclear maturation between 0, 1, 10, 20, and 100 ng mL^–1 LEP (79.4, 82.3, 74.5, 77.2, and 83.7% mature, respectively). According to these data, LEP alone did not have an effect on in vitro maturation of porcine oocytes when using a chemically defined maturation medium. Absence of GLUC in the medium had a negative effect on maturation (39.5%; P < 0.01) compared with treatment with 5 mm (88.2%) and 50 mm (74.7%) GLUC, independent of LEP. In the presence of 5 mm GLUC, there was no difference between 0, 1, and 10 ng mL^–1 LEP (83.9, 94.9, and 88.3%, respectively). However, LEP did affect nuclear maturation when COC were cultured in the absence of GLUC or with excessive GLUC. Leptin (10 ng mL^–1) tended (P = 0.1) to promote nuclear maturation (46.2%) in the absence of GLUC (0 mm), compared with 0 ng mL^–1 LEP (32.3%). In high-GLUC (50 mm) medium, 1 ng mL^–1 LEP had a positive effect (P < 0.05) on nuclear maturation (87.8%) compared with 0 ng mL^–1 (66.7%) and 10 ng mL^–1 LEP (72.3%). LEP (0 or 10 ng mL^–1) inhibited (P < 0.05) nuclear maturation in the presence of high (50 mm) GLUC compared with that in 5 mm GLUC. These data demonstrate that addition of 10 ng mL^–1 LEP in maturation media may mitigate the negative effect of maturation in the absence of GLUC. Moreover, in the presence of excessive GLUC, 1 ng mL^–1 LEP stimulates nuclear maturation. These results suggest that LEP and GLUC may interact to regulate oocyte nuclear maturation in obese or diabetic individuals, or both.