Supplementation with spermine during in vitro maturation of porcine oocytes improves early embryonic development after parthenogenetic activation and somatic cell nuclear transfer
Supplementation with spermine during in vitro maturation of porcine oocytes improves early embryonic development after parthenogenetic activation and somatic cell nuclear transfer
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DOI：
10.2527/jas.2015-9761
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发表时间：
2016-03-01
影响因子：
3.3
通讯作者：
Lee, B. C.
中科院分区：
文献类型：
--
作者：
Jin, J. X.;Lee, S.;Lee, B. C.
精胺在保护细菌、酵母和哺乳动物细胞免受活性氧自由基（ROS）侵害方面起着重要作用，但关于精胺对猪卵母细胞成熟和胚胎发育的影响的研究较少。研究精胺对猪卵母细胞体外成熟（IVM）及其孤雌激活（PA）和体细胞核移植（SCNT）后发育能力的影响。我们评估了卵母细胞的核成熟、细胞内谷胱甘肽（GSH）和ROS水平，以及它们随后的胚胎发育，以及成熟卵母细胞、卵丘细胞和PA囊胚中的基因表达。IVM培养液中不同浓度精胺处理后，核成熟率无显著差异。然而，精胺处理组（10-500 μ M）与对照组相比，细胞内GSH水平显著升高，ROS水平显著降低（P < 0.05）。10 μ M精胺组的囊胚形成率显著高于对照组（P < 0.05）。根据PA结果的最佳条件，我们研究了10 μ M精胺对SCNT的影响，与对照组相比，它也显著提高了囊胚形成率（P < 0.05）。在评价10 μ M精胺对卵丘细胞基因表达的影响时，促凋亡基因（Bax）的表达显著降低，而抗凋亡基因（Bcl 2）的表达显著升高（P < 0.05）。精胺处理的卵母细胞中FGFR 2增加。POU 5 F1和Bcl 2的转录水平显着增加PA囊胚。总之，10 μ M精胺补充IVM期间提高了猪PA和SCNT胚胎的发展，通过增加细胞内GSH，清除ROS水平，并调节基因表达。
Spermine plays an important role in protection from reactive oxygen species (ROS) in bacteria, yeast, and mammalian cells, but there are few studies on the effects of spermine on porcine oocyte maturation and subsequent embryo development. The aim of this study was to determine the effects of spermine on in vitro maturation (IVM) of porcine oocytes and their developmental competence after parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT). We evaluated nuclear maturation, intracellular glutathione (GSH), and ROS levels in oocytes, and their subsequent embryonic development, as well as gene expression in mature oocytes, cumulus cells, and PA blastocysts. After treatment with various concentrations of spermine in IVM culture medium, there was no significant difference in nuclear maturation rate. However, spermine treatment groups (10-500 mu M) showed significantly increased intracellular GSH levels and decreased ROS levels compared to the control (P < 0.05). Furthermore, 10 mu M spermine supported significantly higher blastocyst formation rates after PA than the control group (P < 0.05). According to the optimal condition from the PA results, we investigated the effects of 10 mu M spermine on SCNT, and it also significantly improved blastocyst formation rates compared with the control group (P < 0.05). In evaluating the effects of 10 mu M spermine on gene expression, there was significantly lower expression of a proapoptotic gene (Bax) and higher expression of an antiapoptotic gene (Bcl2) in cumulus cells (P < 0.05). FGFR2 was increased in spermine-treated oocytes. Levels of transcription for POU5F1 and Bcl2 were significantly increased in PA blastocysts. In conclusion, 10 mu M spermine supplementation during IVM improved the development of porcine PA and SCNT embryos by increasing intracellular GSH, scavenging ROS levels, and regulating gene expression.