Vitrification of immature porcine oocytes: Effects of lipid droplets, temperature, cytoskeleton, and addition and removal of cryoprotectant
Vitrification of immature porcine oocytes: Effects of lipid droplets, temperature, cytoskeleton, and addition and removal of cryoprotectant
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DOI：
10.1006/cryo.1998.2079
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发表时间：
1998-05-01
期刊：
影响因子：
2.7
通讯作者：
Grishchenko, V
中科院分区：
文献类型：
--
作者：
Isachenko, V;Soler, C;Grishchenko, V
通过三个实验研究了冷冻保护剂、温度和细胞骨架松弛剂的单步暴露和逐步去除对生发泡猪卵母细胞发育到M-II期的影响。在实验一中，未冷却的卵丘-卵母细胞复合体(COCs)被一步/逐步暴露在乙二醇(EG)中并在23℃或42℃下去除，发现逐步暴露于EG中并在42℃下稀释对COC的发育速度有积极的影响。在实验II中，同样在没有冷却的情况下，在42摄氏度下使用细胞松弛素B处理COCs，采用单步和分步方案暴露和去除EG。细胞松弛素B对玻璃化冷冻但不冷却的COCs无影响。在实验三中，将COCs分为两个处理组和一个对照组。将第1组COCs在42℃逐步暴露于EG中，然后直接插入液氮中进行玻璃化冷冻。5-S浸泡在50℃的水浴中升温，冷冻保护剂为。在42℃下暴露于一系列分级的蔗糖溶液中取出，第2组COCs用同样的程序玻璃化冷冻，然后用细胞松弛素B预处理。第3组COCs为对照(未处理)卵母细胞。经过复温和蔗糖传代后，培养48h，培养结果表明，冷冻保护剂的逐渐饱和/去除、温度的升高和细胞骨架抑制剂细胞松弛素B的预处理对GV-猪卵母细胞的玻璃化冷冻有积极的影响。(C)1998年学术出版社。
Three experiments were conducted to investigate the effects of single-step and stepwise exposure to and 0 removal of cryoprotectant, of temperature, and of a cytoskeletal relaxant on the development of germinal vesicle porcine oocytes to the M-II stage. In experiment I, noncooled cumulus-oocyte complexes (COCs) were treated using single-step/stepwise exposure to ethylene glycol (EG) and removal at 23 or 42 degrees C. Stepwise exposure to EG and dilution at 42 degrees C were found to have a positive effect on the COC developmental rate. In experiment II, also without cooling, COCs were treated with Cytochalasin B at 42 degrees C using single-step and stepwise protocols of exposure to and removal of EG. No effects of Cytochalasin B were noticed on the COCs pretreated for vitrification but not cooled. In experiment III, COCs were divided into two treatment groups and one control group. Group 1 COCs were vitrified by stepwise exposure to EG at 42 degrees C and direct plunging into liquid nitrogen. Warming was carried out by 5-s immersion in a water bath at 50 degrees C, and the cryoprotectant was. removed by exposure to a graded series of sucrose solutions at 42 degrees C. Group 2 COCs were vitrified using the same procedure bur following pretreatment with Cytochalasin B. Group 3 COCs were control (untreated) oocytes. After rewarming and passage through the sucrose series, the experimental COCs were cultured for 48 h. The results of cultivation (progression to MII stage) suggest that the gradual saturation/removal of cryoprotectant, elevated temperature, and pretreatment with cytoskeletal inhibitor Cytochalasin B have a positive effect on vitrification of GV-porcine oocytes. (C) 1998 Academic Press.