title: In vitro fertilization of ovine oocytes vitrified by solid surface vitrification at germinal vesicle stage.
abstract: Cryopreservation of immature oocytes at germinal vesicle (GV) stage would provide a readily available source of oocytes for use in research and allow experiments to be performed irrespective of seasonality or other constraints. This study was designed to evaluate the recovery, viability, maturation status, fertilization events and subsequent development of ovine oocytes vitrified at GV stage using solid surface vitrification (SSV). Cumulus oocyte complexes (COCs) obtained from mature ewes were randomly divided into three groups (1) SSV (oocytes were vitrified using SSV), (2) EXP (oocytes were exposed to vitrification and warming solutions without vitrification) or (3) Untreated (control). Following vitrification and warming, viable oocytes were matured in vitro for 24h. After that, nuclear maturation was evaluated using orcein staining. Matured oocytes were fertilized and cultured in vitro for 7days. Following SSV, 75.7% 143/189 oocytes were recovered. Of those oocytes recovered 74.8%, 107/143 were morphologically normal (viable). Frequencies of in vitro maturation were significantly (P<0.01) decreased in SSV and EXP groups as compared to control. In vitro fertilization rates were significantly (P<0.01) decreased in SSV (39.3%) group as compared to EXP (56.4%) and control (64.7%) groups. Cleavage at 48h post insemination (pi) and development to the blastocyst stage on day 7 pi were significantly (P<0.001) decreased in SSV oocytes as compared to EXP and control groups. In conclusion, immature ovine oocytes vitrified using SSV as a simple and rapid procedure can survive and subsequently be matured, fertilized and cultured in vitro up to the blastocyst stage, although the frequency of development is low.
authors.affiliation: Animal Development and Biotechnology Group, School of Biosciences, The University of Nottingham, Sutton Bonington Campus, Loughborough, Leics, LE12 5RD, UK.
authors.forename: Adel R
authors.initials: AR
authors.lastname: Moawad
authors.forename: Patricia
authors.initials: P
authors.lastname: Fisher
authors.forename: Jie
authors.initials: J
authors.lastname: Zhu
authors.forename: Inchul
authors.initials: I
authors.lastname: Choi
authors.forename: Zsuzsanna
authors.initials: Z
authors.lastname: Polgar
authors.forename: Andras
authors.initials: A
authors.lastname: Dinnyes
authors.forename: Keith H S
authors.initials: KH
authors.lastname: Campbell
mesh_terms: D000818:Animals; D002470:Cell Survival; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D005260:Female; D005307:Fertilization in Vitro / Q000379:methods / Q000662:veterinary*; D008297:Male; D008540:Meiosis; D009865:Oocytes / Q000166:cytology / Q000254:growth & development*; D009866:Oogenesis; D011897:Random Allocation; D012756:Sheep / Q000196:embryology*; D058989:Vitrification