266 THE SHORT TIME TREATMENT WITH SODIUM BUTYRATE ON GERMINAL VESICLE STAGE OOCYTES IMPROVES OOCYTE QUALITY AND DEVELOPMENTAL COMPETENCE IN PIGS

Abstract

Oocyte maturation is a complex process during which the epigenetic modifications are dramatically changed, especially the histone acetylation and phosphorylation. Sodium butyrate (NaBu) is a histone deacetylase inhibitor that results in a more open structure of DNA. The aim of the present study was to analyse the role of NaBu in the meiosis of porcine oocytes and the subsequent embryonic developmental competence. Cumulus–oocyte complexes (COC) were collected from ovaries obtained at a local slaughterhouse. The COC were randomly divided into 3 groups and matured in vitro in medium (Hao et al. 2006) supplemented with 1 μM NaBu for 2 h [germinal vesicle (GV) stage, group 1] or for 22 h [GV to GV breakdown (GVBD) stage, group 2] or without treatment (control, group 3). After 44 h of in vitro maturation, the oocytes were denuded by 0.2% hyaluronidase, and oocytes with evenly dark ooplasm and visible first polar bodies were considered matured. The cortical granule distribution of matured oocytes was examined with immunostaining. The relative expression of CyclinB and Cdc2 of 3 group oocytes was determined with real-time PCR. Some matured oocytes from each group were collected and stimulated with electric pulse (2 direct current pulses of 1.2 kV cm^–1 for 30 μs). The rate of parthenogenetic blastocyst was recorded, and cell number of each blastocyst was determined under an inverted fluorescence microscope after staining with 10 μg mL^–1 of Hoechst 33342. The following results were found. 1) Compared with the control group (n = 70, 67.74 ± 1.64), oocyte maturation rates of group 1 and group 2 decreased significantly along with the extended treatments (n = 70, 59.57 ± 5.29 and 46.99 ± 1.22, respectively; P < 0.05). 2) The long time (22 h) treatment with NaBu inhibited the developmental competence (blastocyst rate) of oocytes (n = 30, 15.33 ± 3.47 v. 27.16 ± 2.10 P < 0.05), and the short time (2 h) treatment with NaBu on GV-stage oocytes inhibited the meiotic process slightly but improved the blastocyst rate (n = 30, 33.93 ± 2.51 v. 27.16 ± 2.10; P < 0.05). 3) The short time (2 h) treatment resulted in the migration of more cortical granules into the plasmasmic membrane and formed a monolayer with the membrane (compared with the control). 4) The exposure to NaBu from GV to GVBD stage induced the expression of the CyclinB and Cdc2 in the matured oocytes (4.68 ± 0.45 and 5.80 ± 0.58, respectively; P < 0.05) compared with the control. 5) Short time (2 h) exposure to NaBu on GV-stage oocytes inhibited the expression of the Cdc2 but increased the expression of the CylinB in the matured oocytes (0.43 ± 0.06 and 1.65 ± 0.26, respectively; P < 0.05) compared with the control. In conclusion, results of this study demonstrate that exposure to NaBu inhibits porcine oocyte meiosis in proportion to treatment length. However, a 2-h treatment with 1 μM NaBu improves oocyte developmental competence to the blastocyst stage. These results are useful for improving the developmental competent of oocytes for IVF and in vitro embryo production.