Somfai Tamás, Nakai Michiko, Tanihara Fuminori, Noguchi Junko, Kaneko Hiroyuki, Kashiwazaki Naomi, Egerszegi István, Nagai Takashi, Kikuchi Kazuhiro
NARO Institute of Livestock and Grassland Science, Ibaraki 305-0901, Japan.
J Reprod Dev. 2013;59(4):378-84. doi: 10.1262/jrd.2013-015. Epub 2013 May 10.
Our aim was to optimize a cryoprotectant treatment for vitrification of immature porcine cumulus-oocyte complexes (COCs). Immature COCs were vitrified either in 35% ethylene glycol (EG), 35% propylene glycol (PG) or a combination of 17.5% EG and 17.5% PG. After warming, the COCs were in vitro matured (IVM), and surviving oocytes were in vitro fertilized (IVF) and cultured. The mean survival rate of vitrified oocytes in 35% PG (73.9%) was higher (P<0.05) than that in 35% EG (27.8%). Oocyte maturation rates did not differ among vitrified and non-vitrified control groups. Blastocyst formation in the vitrified EG group (10.8%) was higher (P<0.05) than that in the vitrified PG group (2.0%) but was lower than that in the control group (25.0%). Treatment of oocytes with 35% of each cryoprotectant without vitrification revealed a higher toxicity of PG on subsequent blastocyst development compared with EG. The combination of EG and PG resulted in 42.6% survival after vitrification. The maturation and fertilization rates of the surviving oocytes were similar in the vitrified, control and toxicity control (TC; treated with EG+PG combination without cooling) groups. Blastocyst development in the vitrified group was lower (P<0.05) than that in the control and TC groups, which in turn had similar development rates (10.7%, 18.1% and 23.3%, respectively). In conclusion, 35% PG enabled a higher oocyte survival rate after vitrification compared with 35% EG. However, PG was greatly toxic to oocytes. The combination of 17.5% EG and 17.5% PG yielded reasonable survival rates without toxic effects on embryo development.
我们的目标是优化一种用于未成熟猪卵丘-卵母细胞复合体（COCs）玻璃化冷冻的冷冻保护剂处理方法。未成熟COCs分别在35%乙二醇（EG）、35%丙二醇（PG）或17.5% EG与17.5% PG的组合中进行玻璃化冷冻。解冻后，COCs进行体外成熟（IVM），存活的卵母细胞进行体外受精（IVF）并培养。35% PG组玻璃化冷冻卵母细胞的平均存活率（73.9%）高于35% EG组（27.8%）（P<0.05）。玻璃化冷冻组和未玻璃化冷冻的对照组之间的卵母细胞成熟率没有差异。玻璃化冷冻EG组的囊胚形成率（10.8%）高于玻璃化冷冻PG组（2.0%）（P<0.05），但低于对照组（25.0%）。用35%的每种冷冻保护剂处理卵母细胞而不进行玻璃化冷冻显示，与EG相比，PG对后续囊胚发育的毒性更高。EG和PG组合在玻璃化冷冻后存活率为42.6%。玻璃化冷冻组、对照组和毒性对照组（TC；用EG+PG组合处理但未冷却）中存活卵母细胞的成熟率和受精率相似。玻璃化冷冻组的囊胚发育率低于对照组和TC组（P<0.05），而对照组和TC组的发育率相似（分别为10.7%、18.1%和23.3%）。总之，与35% EG相比，35% PG在玻璃化冷冻后能使卵母细胞存活率更高。然而，PG对卵母细胞毒性很大。17.5% EG和17.5% PG的组合产生了合理的存活率，且对胚胎发育没有毒性作用。
