Glutathione (GSH) Concentration and Developmental Competence of Korean Native Cow Oocytes Selected by Brilliant Cresyl Blue (BCB)

Abstract

This study was carried out to evaluate the nuclear, cytoplasmic maturation and developmental potential of bovine oocytes selected by brilliant cresyl blue (BCB) as indirect measurement of oocytes growth phase. Cumulus-oocyte complexes (COCs) were collected from 2 to 8 mm follicles from slaughterhouse Hanwoo ovaries. The COCs were divided into stained cytoplasm to blue (BCB+) and unstained (BCB-) according to their ooplasm BCB coloration stained by 26 μm of BCB after 90 min. Selected COCs were cultured in a TCM199 for 18 to 26 h. Nuclear maturation and total cell number was evaluated after in vitro maturation (IVM) or in vitro culture (IVC) using 10 μg/ml Hoechst 33342, and cytoplasmic maturation was evaluated by intracellular glutathione (GSH) assay before (0 h) and after (24 h) IVM. The oocyte diameters were not differed significantly between BCB+ (157.4±5.8 μm) and BCB- (149.0±31.0 μm) groups (p>0.05). However, the proportion of metaphase II oocytes in BCB+ group was significantly higher than BCB- group after IVM (p<0.05). GSH content of BCB+ group oocytes was significantly higher than that of BCB- group just after collection (7.3±0.6 vs. 4.8±0.6 pmol/oocyte, p<0.05), but not varied after IVM (13.1±0.9 and 12.6±2.5 pmol/oocytes for BCB+ and BCB- respectively; p>0.05). The proportion of blastocyst formation and total cell number in BCB+ group (23.5% and 105.5±28.6) was significantly higher than that in BCB- (9.8% and 72.4±26.1; p<0.05). The results indicate that BCB+ group oocytes may provide a cellular and functional basis for the greater developmental competence in Korean Native Cow (KNC) oocytes.