Preincubation with glutathione ethyl ester improves the developmental competence of vitrified mouse oocytes
Preincubation with glutathione ethyl ester improves the developmental competence of vitrified mouse oocytes
复制标题
谷胱甘肽乙酯预孵育可提高玻璃化小鼠卵母细胞的发育能力
DOI：
10.1007/s10815-018-1215-4
复制
发表时间：
2018-07-01
影响因子：
3.1
通讯作者：
Sun, Yijuan
中科院分区：
文献类型：
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作者：
Li, Zhichao;Gu, Ruihuan;Sun, Yijuan
目的卵母细胞玻璃化冷冻是目前用于人类生育力保存的方法。然而，玻璃化损伤是一个问题所造成的卵浆谷胱甘肽（GSH）水平下降。GSH供体谷胱甘肽乙酯（GSH-OEt）能显著提高卵母细胞中GSH的含量。然而，从女性体内获得卵母细胞是困难的。为了克服这一点，我们使用小鼠卵母细胞，以取代人类卵母细胞作为study.MethodsB6D2F1小鼠卵母细胞预孵育30分钟与2.5 mmol/L的GSH-OEt（GSH-OEt组），没有GSH-OEt预孵育前玻璃化冷冻（对照玻璃化冷冻组）或在非玻璃化冷冻卵母细胞（新鲜组）。解冻后，卵母细胞进行受精，以评估胚胎在体外和体内的发育能力。采用免疫荧光、Polscope设备和定量逆转录聚合酶链反应（RT-qPCR）分析损伤，包括线粒体分布、活性氧（ROS）水平、纺锤体形态和基因表达水平结果受精率、3-4细胞率、囊胚形成和扩张囊胚明显较高（p< 0.05）在GSH-OEt组中（90.4%; 91.1%; 88.9%和63.0%）比在对照组中（80.0%; 81.4%; 77.7%和50.5%）。如果胚胎克服了2细胞阻滞并发育到囊胚期，所有组的出生率相似。玻璃化改变线粒体分布，增加ROS水平，并造成异常的纺锤体形态，GSH-OEt预孵育可以改善这种损害。RT-qPCR结果显示，对照组Bcl-2表达低于GSH-OEt组，BAX和MnSoD表达高于GSH-OEt组（p<0. 05）。
PurposeOocyte vitrification is currently used for human fertility preservation. However, vitrification damage is a problem caused by decreasing ooplasmic levels of glutathione (GSH). The GSH donor glutathione ethyl ester (GSH-OEt) can significantly increase the GSH content in oocytes. However, it is difficult to obtain oocyte from woman. To overcome this, we used mouse oocytes to replace human oocytes as a model of study.MethodsOocytes from B6D2F1 mice were preincubated for 30 min with 2.5 mmol/L GSH-OEt (GSH-OEt group), without GSH-OEt preincubation before vitrification (control vitrification group) or in nonvitrified oocytes (fresh group). After thawing, oocytes were fertilized for evaluating the developmental competence of embryos in vitro and in vivo. Immunofluorescence, Polscope equipment and quantitative reverse transcription polymerase chain reaction (RT–qPCR) were used to analyze damage, including mitochondrial distribution, reactive oxygen species (ROS) levels, spindle morphology, and gene expression levels (Bcl-2, BAX, and MnSOD).ResultsThe rates of fertilization, 3–4 cell, blastocyst formation and expanded blastocysts were significantly higher (p< 0.05) in the GSH-OEt group (90.4%; 91.1%; 88.9% and 63.0%) than in the control (80.0%; 81.4%; 77.7% and 50.5%). Provided embryos overcame the 2-cell block and developed to the blastocyst stage, birth rates of all groups were similar. Vitrification altered mitochondrial distribution, increased ROS levels, and caused abnormal spindle morphology; GSH-OEt preincubation could improve such damage. RT–qPCR showed that the expression of Bcl-2 was lower in the control group compared with the GSH-OEt group; BAX and MnSoD expression levels were higher in the control group than in the GSH-OEt group (p< 0.05).ConclusionsThe beneficial effect of GSH-OEt preincubation occurred before the 2-cell stage.