Effects of vitrification of cumulus-enclosed porcine oocytes at the germinal vesicle stage on cumulus expansion, nuclear progression and cytoplasmic maturation
Effects of vitrification of cumulus-enclosed porcine oocytes at the germinal vesicle stage on cumulus expansion, nuclear progression and cytoplasmic maturation
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DOI：
10.1071/rd16386
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发表时间：
2017-10-01
影响因子：
1.9
通讯作者：
Kikuchi, Kazuhiro
中科院分区：
文献类型：
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作者：
Appeltant, Ruth;Somfai, Tamas;Kikuchi, Kazuhiro
虽然在生发泡(GV)阶段玻璃化冷冻的猪卵母细胞已经产生了后代，但胚胎发育率仍然很低。本研究比较了玻璃化冷冻卵丘-卵母细胞复合体(COCs)和对照COCs(未经冷冻保护剂处理和未冷却)以及毒性对照(未经冷冻)的核形态和进展、卵丘扩展、跨带投射(TZPs)、三磷酸腺苷(ATP)和谷胱甘肽(GSH)水平。玻璃化冷冻分别加入17.5%(v/v)乙二醇组和17.5%(v/v)丙二醇组。GV阶段的玻璃化冷冻导致减数分裂过早进展，表现为GV较早破裂和MII阶段过早达到。然而，细胞质成熟，通过测量ATP和GSH水平，以及卵丘扩张，尽管在玻璃化的COCs中可检测到TZPs的损伤，但仍正常进行。此外，冷冻保护剂的处理导致核仁前体断裂和F-肌动蛋白的形态变化，在随后的IVM培养中，卵母细胞能够从中恢复。卵母细胞发育能力下降的原因可能是核过早成熟导致卵母细胞老化，但也可以考虑其他机制，如启动细胞凋亡和细胞质mRNA的减少。还需要进一步的研究来阐明这些现象在未成熟的COCs玻璃化冷冻过程中的存在和影响。
Although offspring have been produced from porcine oocytes vitrified at the germinal vesicle (GV) stage, the rate of embryo development remains low. In the present study, nuclear morphology and progression, cumulus expansion, transzonal projections (TZPs), ATP and glutathione (GSH) levels were compared between vitrified cumulus-oocyte complexes (COCs) and control COCs (no cryoprotectant treatment and no cooling), as well as a toxicity control (no cooling). Vitrification was performed with 17.5% (v/v) ethylene glycol and 17.5% (v/v) propylene glycol. Vitrification at the GV stage caused premature meiotic progression, reflected by earlier GV breakdown and untimely attainment of the MII stage. However, cytoplasmic maturation, investigated by measurement of ATP and GSH levels, as well as cumulus expansion, proceeded normally despite detectable damage to TZPs in vitrified COCs. Moreover, treatment with cryoprotectants caused fragmentation of nucleolus precursor bodies and morphological changes in F-actin from which oocytes were able to recover during subsequent IVM culture. Reduced developmental competence may be explained by premature nuclear maturation leading to oocyte aging, although other mechanisms, such as initiation of apoptosis and reduction of cytoplasmic mRNA, can also be considered. Further research will be required to clarify the presence and effects of these phenomena during the vitrification of immature COCs.