title: Vitrification of porcine immature oocytes: Association of equilibration manners with warming procedures, and permeating cryoprotectants effects under two temperatures.
abstract: The aim of this study was to evaluate the association of equilibration manners with warming procedures, and the different permeating cryoprotectants (pCPAs) effects under two temperatures, in terms of survival, maturation and subsequent parthenogenetic development of porcine immature oocytes after Cryotop vitrification. In Experiment 1, oocytes were equilibrated by exposure to 5% (v/v) ethylene glycol (EG) for 10 min (EM1) or stepwise to 7.5% (v/v) and 15% (v/v) EG for 2.5 min respectively (EM2). Warming procedures were performed in 1.0 M sucrose for 1 min, then in 0.5 and 0.25 M sucrose for 2.5 min respectively (WP1), or in 0.5, 0.25 and 0.125 M sucrose each step for 2 min (WP2), or in 0.25, 0.125 and 0.063 M sucrose each step for 2 min (WP3). After 2 h of warming, the survival rate of oocytes treated by EM1 and WP1 was significantly higher (P < 0.05) than that of the other groups. Moreover, a similar proportion of survival and nuclear maturation in all vitrified groups was obtained after completion of the IVM. No significant difference in blastocyst development was observed among vitrified groups except the group treated by EM2 and WP3. In Experiment 2, oocytes were vitrified by using EG alone, EG combined with dimethyl sulphoxide (EG + DMSO) or propylene glycol (EG + PROH) as pCPAs under 25 °C and 39 °C. The percentages of cryosurvival and nuclear maturation were similar in all vitrified groups. Under 25 °C, the embryo development and total cell numbers of blastocysts were not significantly different among EG, EG + DMSO and EG + PROH groups. However, the application of EG + PROH at 39 °C resulted in significantly decreased both cleavage and blastocyst formation rates. In conclusion, our data showed that equilibration manner and warming procedure affect the cryosurvival of porcine immature oocytes, and the combination of pCPAs cannot give a better cryopreservation outcome whether 25 °C or 39 °C. Notably, the Cryotop vitrification accompanied by our modified strategy for porcine immature oocytes could achieve high survival and respectable blastocyst production.
authors.affiliation: Yunnan Provincial Engineering Laboratory of Animal Genetic Resource Conservation and Germplasm Enhancement, Yunnan Animal Science and Veterinary Institute, Kunming, Yunnan 650224, People's Republic of China.
authors.forename: Guoquan
authors.initials: G
authors.lastname: Wu
authors.affiliation: College of Animal Science and Technology, Yunnan Agricultural University, Kunming, Yunnan 650201, People's Republic of China.
authors.forename: Baoyu
authors.initials: B
authors.lastname: Jia
authors.affiliation: Yunnan Provincial Engineering Laboratory of Animal Genetic Resource Conservation and Germplasm Enhancement, Yunnan Animal Science and Veterinary Institute, Kunming, Yunnan 650224, People's Republic of China.
authors.forename: Guobo
authors.initials: G
authors.lastname: Quan
authors.affiliation: Yunnan Provincial Engineering Laboratory of Animal Genetic Resource Conservation and Germplasm Enhancement, Yunnan Animal Science and Veterinary Institute, Kunming, Yunnan 650224, People's Republic of China.
authors.forename: Decai
authors.initials: D
authors.lastname: Xiang
authors.affiliation: Yunnan Provincial Engineering Laboratory of Animal Genetic Resource Conservation and Germplasm Enhancement, Yunnan Animal Science and Veterinary Institute, Kunming, Yunnan 650224, People's Republic of China.
authors.forename: Bin
authors.initials: B
authors.lastname: Zhang
authors.affiliation: Yunnan Provincial Engineering Laboratory of Animal Genetic Resource Conservation and Germplasm Enhancement, Yunnan Animal Science and Veterinary Institute, Kunming, Yunnan 650224, People's Republic of China.
authors.forename: Qingyong
authors.initials: Q
authors.lastname: Shao
authors.affiliation: Yunnan Provincial Engineering Laboratory of Animal Genetic Resource Conservation and Germplasm Enhancement, Yunnan Animal Science and Veterinary Institute, Kunming, Yunnan 650224, People's Republic of China. Electronic address: qionghuahong@163.com.
authors.forename: Qionghua
authors.initials: Q
authors.lastname: Hong
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000187:drug effects; D015925:Cryopreservation / Q000379:methods* / Q000662:veterinary; D003451:Cryoprotective Agents / Q000494:pharmacology*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D047108:Embryonic Development / Q000187:drug effects; D019855:Ethylene Glycol / Q000494:pharmacology; D005260:Female; D009865:Oocytes; D010312:Parthenogenesis; D019946:Propylene Glycol / Q000494:pharmacology; D013395:Sucrose / Q000494:pharmacology; D013552:Swine; D013696:Temperature; D058989:Vitrification
chemical_list: D003451:Cryoprotective Agents; D013395:Sucrose; D019946:Propylene Glycol; D019855:Ethylene Glycol; D004121:Dimethyl Sulfoxide
keywords: Cryoprotectants; Equilibration; Porcine oocyte; Temperature; Vitrification; Warming