Generation of rats from vitrified oocytes with surrounding cumulus cells via in vitro fertilization with cryopreserved sperm
Generation of rats from vitrified oocytes with surrounding cumulus cells via in vitro fertilization with cryopreserved sperm
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DOI：
10.1111/asj.12666
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发表时间：
2017-01-01
影响因子：
2
通讯作者：
Kashiwazaki, Naomi
中科院分区：
文献类型：
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作者：
Fujiwara, Katsuyoshi;Kamoshita, Maki;Kashiwazaki, Naomi
本研究的目的是评估大鼠冷冻精子体外受精（IVF）后玻璃化冷冻卵母细胞的生育力和足月发育。用30%乙二醇+0.5mol/L蔗糖+20%胎牛血清玻璃化冷冻卵母细胞。将温热的卵母细胞与精子共培养。尽管裸露/玻璃化冷冻的卵母细胞未受精，但部分卵丘细胞玻璃化冷冻的卵母细胞在IVF后与新鲜精子受精（32.7%）。当用冷冻保存的精子进行IVF时，玻璃化冷冻或新鲜卵母细胞与卵丘细胞受精（分别为62.9%或41.1%）。此外，为了证实冷冻精子IVF后玻璃化冷冻卵母细胞与周围卵丘细胞的足月发育，将108个具有两个原核（2 PN）的玻璃化冷冻卵母细胞转移到8只假孕雌性中，并从3只受体中获得8只幼崽。目前的工作表明，卵丘细胞包围的玻璃化冷冻大鼠卵母细胞可以在体外受精与冷冻保存的精子，并从冷冻保存的配子衍生的2 PN胚胎可以发育到足月。据我们所知，这是第一次报告的成功产生的大鼠后代来自玻璃化冷冻卵母细胞与冷冻精子体外受精。
The objective of this study was to evaluate fertility and full-term development of rat vitrified oocytes after in vitro fertilization (IVF) with cryopreserved sperm. Oocytes with or without surrounding cumulus cells were vitrified with 30% ethylene glycol+0.5mol/L sucrose+20% fetal calf serum by using the Cryotop method. The warmed oocytes were co-cultured with sperm. Although the denuded/vitrified oocytes were not fertilized, some of the oocytes vitrified with cumulus cells were fertilized (32.7%) after IVF with fresh sperm. When IVF was performed with cryopreserved sperm, vitrified or fresh oocytes with cumulus cells were fertilized (62.9% or 41.1%, respectively). In addition, to confirm the full-term development of the vitrified oocytes with surrounding cumulus cells after IVF with cryopreserved sperm, 108 vitrified oocytes with two pronuclei (2PN) were transferred into eight pseudopregnant females, and eight pups were obtained from three recipients. The present work demonstrates that vitrified rat oocytes surrounded by cumulus cells can be fertilized in vitro with cryopreserved sperm, and that 2PN embryos derived from cryopreserved gametes can develop to term. To our knowledge, this is the first report of successful generation of rat offspring derived from vitrified oocytes that were fertilized in vitro with cryopreserved sperm.