Follicular extracellular vesicles enhance meiotic resumption of domestic cat vitrified oocytes
Follicular extracellular vesicles enhance meiotic resumption of domestic cat vitrified oocytes
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DOI：
10.1038/s41598-020-65497-w
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发表时间：
2020-05-25
影响因子：
4.6
通讯作者：
Songsasen, Nucharin
中科院分区：
文献类型：
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作者：
Ferraz, Marcia de Almeida Monteiro Melo;Fujihara, Mayako;Songsasen, Nucharin
细胞外囊泡（EVs）包含多种调节细胞和组织功能的因子。然而，对它们对配子的影响，包括与卵母细胞的交流的了解仍然有限。在本研究中，我们对家猫（Felis catus）卵泡液EVs （ffEV）的蛋白质组进行了表征。为了确定卵泡液ev对配子低温存活和体外成熟能力的影响，在存在或不存在ffEV的情况下，使用Cryotop方法对猫卵母细胞进行玻璃化。将玻璃化的卵母细胞在有或没有ffev的情况下解冻，评估存活，体外培养26小时，然后评估活力和减数分裂状态。Cat ffev的平均尺寸为129.3 +/- 61.7 nm（平均+/- SD），透射电镜下具有典型的甜甜圈状圆形囊泡。ffEVs的蛋白质组学分析共鉴定出1974种蛋白质中的674种蛋白质组，这些蛋白质组被归类为参与氧化磷酸化、细胞外基质形成、卵母细胞减数分裂、胆固醇代谢、糖酵解/糖异生以及MAPK、PI3K-AKT、HIPPO和钙信号通路的调节。此外，还鉴定了几种与渗透和热胁迫反应相关的伴侣蛋白。新鲜卵母细胞与玻璃化卵母细胞存活率无显著差异；然而，在玻璃化和/或解冻培养基中添加ffev可增强冷冻解冻卵母细胞恢复减数分裂的能力。总之，本研究首次表征了猫ffev的蛋白质含量及其在维持冷冻保存的卵母细胞减数分裂能力中的潜在作用。
Extracellular vesicles (EVs) contain multiple factors that regulate cell and tissue function. However, understanding of their influence on gametes, including communication with the oocyte, remains limited. In the present study, we characterized the proteome of domestic cat (Felis catus) follicular fluid EVs (ffEV). To determine the influence of follicular fluid EVs on gamete cryosurvival and the ability to undergo in vitro maturation, cat oocytes were vitrified using the Cryotop method in the presence or absence of ffEV. Vitrified oocytes were thawed with or without ffEVs, assessed for survival, in vitro cultured for 26 hours and then evaluated for viability and meiotic status. Cat ffEVs had an average size of 129.3 +/- 61.7 nm (mean +/- SD) and characteristic doughnut shaped circular vesicles in transmission electron microscopy. Proteomic analyses of the ffEVs identified a total of 674 protein groups out of 1,974 proteins, which were classified as being involved in regulation of oxidative phosphorylation, extracellular matrix formation, oocyte meiosis, cholesterol metabolism, glycolysis/gluconeogenesis, and MAPK, PI3K-AKT, HIPPO and calcium signaling pathways. Furthermore, several chaperone proteins associated with the responses to osmotic and thermal stresses were also identified. There were no differences in the oocyte survival among fresh and vitrified oocyte; however, the addition of ffEVs to vitrification and/or thawing media enhanced the ability of frozen-thawed oocytes to resume meiosis. In summary, this study is the first to characterize protein content of cat ffEVs and their potential roles in sustaining meiotic competence of cryopreserved oocytes.