P-229: Chromosomal Analysis of Parthenogenetic Mouse Embryos Generated from In Vitro Activated Oocytes by Hydrostatic Pressure and Ethanol and Cytochalasin B

Background: Studies of preimplantation stage embryos by classic cytogenetic techniques have limitations, starting with the need for good metaphase stage when only one third of all analyzed embryos may show good quality metaphases. The incidence of chromosome anomalies in embryos produced by in vitro procedures is generally higher than that of embryos formed in vivo. Pressure specifically affects centriole microtubules and that proximal microtubuletriplets are more resistant than distal microtubule doublets. The present study has been carried out to investigate the effects of hydrostatic pressure in the absence or presenc of ethanol on the improvement of chromosomal complements in 2-cell parthenogenetic embryos. Materials and Methods: 6 to 8-week-old female NMRI mouse were superovulated by an injection of 10 IU of PMSG, followed by 10 IU HCG 48 hours later. Metaphase П oocytes were collected from oviduct 14 hr after HCG injection. Oocytes transferred to T6 medium and randomly assigned to following groups: non- treatment (control), HP exposure (treatment I), ethanol exposure (treatment II) and ethanol 7% with HP exposure (treatment III). Groups of activated oocytes were further treated with 5 mg/ml cytochalasin B for 4 hours. About 24-hours post oocytes activation, Slides were prepared according to an ‘air drying’ technique and the chromosomal complement of 2-cell embryos was studied by Giemsa-staining. Results: Results indicated that in embryos treatmented by ethanol and HP observated the most incidence of normal chromosome (diploidy and tetraploidy) (64%) and the fewest incidence of anormal chromosome (haploidy, triploidy and mixopliody) (26%) compared with control group and other treatments (p<0/05). Conclusion: Induction of Ca fluxes promotes the resumption of meiosis and extrusion of the second polar body, but not pronuclear formation. Treatment with a diploidization agent is required to either suppress second polar body extrusion (CB). Hydrostatic pressure probably by affecting on centriole microtubules could improve chromosomal complements.