195 SODIUM SELENITE INCREASES DEVELOPMENTAL ABILITY AND DECREASES APOPTOSIS IN PORCINE PARTHENOTES

Abstract

Selenium is an essential component of serum required for cell proliferation and growth. It has also been shown to have a beneficial effect on in vitro maturation of oocytes. This study examined the development and apoptosis of porcine parthenotes treated with sodium selenite (SS). Immature cumulus–oocyte complexes were cultured in TCM-199 supplemented with 25 mM NaHCO₃, 10% (v/v) porcine follicular fluid, 0.57 mM cysteine, 0.22 µg mL⁻¹ sodium pyruvate, 25 µg mL⁻¹ gentamycin sulfate, 0.5 µg mL⁻¹ Follitropin V, 10 ng mL⁻¹ epidermal growth factor, and 1 µg mL⁻¹ estradiol-17β for 42–44 h. The matured oocytes were activated by electro-stimulation (single electrical pulse of 1.36 kV cm⁻¹ for 30 µs) and treated with 10 µg mL⁻¹ cytochalasin B for 4 h. Activated oocytes were cultured to the blastocyst stage in NCSU23 supplemented with 0.4% polyvinyl alcohol in the presence or absence of 25 ng mL⁻¹ SS for 7 days at 39°C in a humidified atmosphere of 5% CO₂ in air. Inner cell mass (ICM) and apoptosis rates in blastocysts were evaluated by differential double staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling method. In addition, relative abundance of Bax/Bcl_XL transcripts was quantified by real-time qRT-PCR. The activity of caspase-3 and MAP kinase in blastocysts was examined by western blotting. The cleavage rate, blastocyst rate, total cell number, apoptosis ratio, and ICM ratio of parthenotes in NCSU23 + PVA without/with SS is shown in Table 1. Expression level of Bax/Bcl_XL genes in blastocysts decreased 0.4 ± 0.1-fold in the presence of SS. Furthermore, activity of caspase-3 in blastocysts decreased and activity of MAP kinase in blastocysts increased with SS. Therefore, our data suggest that SS may play a critical role in increasing the development of porcine parthenotes and in reducing their apoptosis.

Table 1.  Developmental ability and apoptosis of porcine parthenotes cultured in the presence or absence of sodium selenite

This work was supported by the Research Project on the Production of Bio-organs (No. 200503030202), Ministry of Agriculture and Forestry, Republic of Korea.