P-102: Evaluation of G6PDH Enzyme Activity in GV Oocytes Cytoplasm to Recompense The Effects of Vitrification Process

Background: Process of oocyte freezing, seriously damages the molecular structure of the oocyte into the life and impair its ability to develop. It seems Oocytes can partially recompense those damages. The pentose phosphate pathway (PPP) constitutes the major source of NADPH required for the neutralization of reactive oxygen species (ROS). Glucose-6-phosphate dehydrogenase (G6PDH) is the first enzyme of the oxidative part of the PPP. This study was conducted to determine whether BCB staining performed on immature bovine oocytes to assess G6pdh level has an impact on quality bovine embryos product from GV vitrified oocyte. Materials and Methods: We selected growing and fully grown bovine oocytes based on G6PDH reaction with brilliant cresyl blue (BCB); Immature oocytes were exposed to 26 μM BCB concentration and classified according to their cytoplasm coloration as grown BCB+ (blue cytoplasm) and growing BCB- (colourless cytoplasm) After selection, oocytes were vitrified by using a crytop. GV oocytes (3-5) were suspended in equilibration solution (HM containing) ethylene glycol (EG) and dimethylsulfoxide (DMSO) for 10 minutes to allow initial shrinkage and recovery. Following equilibration, they were transferred to the vitrification solution (HM containing EG, DMSO and 0.5 M sucrose) for 45-60 seconds. After oocyte thawing the oocytes cultured in maturation media. For this study, biochemical and morphological events which were occurring during early embryonic development, parthenogenesis has been used. In order to evaluate the quality of blastocyst inner cell mass (ICM) and trophectoderm (TE), cell number between BCB+ and BCB- were determined by using a differential staining protocol.Results: Cleavage was significantly higher BCB+ vitrified oocyte (p<0.05) in in compare BCB- oocytes (53.6 ± 6% vs 40.7 ± 4.2% respectively). The percentage of Parthenogenesis blastocyst development was higher in BCB+ and was significantly different between BCB+ (15.01 ± 9.03) and BCB- (3.19 ± 7.48) oocytes. Total cell number, TE cell number, ICM cell number: TE ratio of blastocysts were significantly higher in the BCB+ group compared with the BCB- group. Conclusion: The cytoplasm of BCB+ vitrified oocyte show more competency to repair damage of verification process and it make improve efficiently of gamete cryopreservation.