The combination treatment of cholesterol-loaded methyl-β-cyclodextrin and methyl-β-cyclodextrin significantly improves the fertilization capacity of vitrified bovine oocytes by protecting fertilization protein JUNO
The combination treatment of cholesterol-loaded methyl-β-cyclodextrin and methyl-β-cyclodextrin significantly improves the fertilization capacity of vitrified bovine oocytes by protecting fertilization protein JUNO
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DOI：
10.1111/rda.13890
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发表时间：
2021-01-18
影响因子：
1.7
通讯作者：
Zhao, Xueming
中科院分区：
文献类型：
--
作者：
Hao, Tong;Zhang, Peipei;Zhao, Xueming
许多实验表明，玻璃化冷冻会显着降低哺乳动物卵母细胞的受精能力，限制了玻璃化冷冻卵母细胞的应用。已证明JUNO蛋白在哺乳动物卵母细胞受精过程中发挥着至关重要的作用。然而，关于玻璃化冷冻对牛卵母细胞中 JUNO 蛋白的影响以及保护该蛋白的程序的信息很少。本研究旨在研究玻璃化冷冻对牛卵母细胞中 JUNO 蛋白水平的影响。在这项研究中，MII 卵母细胞在玻璃化前用负载胆固醇的甲基-β-环糊精（CLC；0、10、15、20 mM）处理 45 分钟，并在解冻（38-39 摄氏度）后用甲基-β-环糊精（M beta CD；0、2.25、4.25、6.25 mM）处理 45 分钟。然后检测JUNO蛋白的表达水平和功能、膜内胆固醇水平、磷脂酰丝氨酸的外化、精子结合能力和玻璃化牛卵母细胞的发育能力。我们的结果表明，玻璃化冷冻显着降低了JUNO蛋白水平、胆固醇水平、精子结合能力、发育能力，并增加了JUNO基因启动子甲基化水平和牛卵母细胞凋亡水平。此外，15 mM CLC + 4.25 mM M beta CD 处理显着改善了胆固醇水平，并增加了玻璃化牛卵母细胞的精子结合和发育能力。总之，负载胆固醇的甲基-β-环糊精和甲基-β-环糊精联合处理通过保护受精蛋白JUNO，显着提高玻璃化牛卵母细胞的受精能力。
Many experiments show that vitrification significantly reduces the fertilization capacity of mammalian oocytes, restricting the application of vitrified oocytes. It has been proven that the JUNO protein plays a vital role in mammalian oocytes fertilization. However, little information is available about the effects of vitrification on the JUNO protein and the procedure to protect it in bovine oocytes. Here, the present study was designed to investigate the effect of vitrification on the JUNO protein level in bovine oocytes. In this study, MII oocytes were treated with cholesterol-loaded methyl-beta-cyclodextrin (CLC; 0, 10, 15, 20 mM) for 45 min before vitrification and methyl-beta-cyclodextrin (M beta CD; 0, 2.25, 4.25, 6.25 mM) for 45 min after thawing (38-39 degrees C). Then, the expression level and function of JUNO protein, cholesterol level in the membrane, the externalization of phosphatidylserine, sperm binding capacity and the developmental ability of vitrified bovine oocytes were examined. Our results showed that vitrification significantly decreased the JUNO protein level, cholesterol level, sperm binding capacity, development ability, and increased the promoter methylation level of the JUNO gene and apoptosis level of bovine oocytes. Furthermore, 15 mM CLC + 4.25 mM M beta CD treatment significantly improved the cholesterol level and increased sperm binding and development ability of vitrified bovine oocytes. In conclusion, the combination treatment of cholesterol-loaded methyl-beta-cyclodextrin and methyl-beta-cyclodextrin significantly improves the fertilization capacity of vitrified bovine oocytes by protecting fertilization protein JUNO.