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通过玻璃化冻融后延长培养恢复牛卵母细胞的纺锤体形态和线粒体功能。

Recovery of spindle morphology and mitochondrial function through extended culture after vitrification-warming of bovine oocytes.

机构信息

School of Animal Sciences, Louisiana State University Agricultural Center, Baton Rouge, LA, USA.

出版信息

Theriogenology. 2022 Sep 1;189:192-198. doi: 10.1016/j.theriogenology.2022.06.021. Epub 2022 Jun 24.

Abstract

The present study aimed to determine the effects of vitrification on the meiotic spindle and mitochondrial function of bovine oocytes submitted to different times of post-warming culture. Partially denuded cumulus-oocyte complexes were vitrified at different maturation times (18-, 20-, and 24-h) using a two-step cryoprotectant addition protocol and submitted to 6-, 4-, or 0-h of post-warming extended culture in maturation medium. Microtubule configuration and chromosomal arrangement were analyzed after 0- and 6-h of extended culture, whereas mitochondrial membrane potential and ATP content were measured at 0-, 4-, and 6-h of post-warming recovery. Results of meiotic spindle integrity revealed that vitrified-warmed oocytes that underwent 6-h of culture had similar incidence of normal microtubule configuration and chromosomal arrangement as compared to fresh oocytes, but higher than oocytes in the vitrification control group (no culture). Mitochondrial membrane potential was not different in all the vitrification groups, but the oocytes that were cultured for 4-h after warming had similar levels compared to fresh oocytes. ATP concentration in all vitrification groups was lower than the control group. However, oocytes cultured for 6-h had the lowest rate of ATP depleted oocytes among the vitrification groups. The results of this study indicate that extended culture after warming promotes the recovery of the meiotic spindle and, to some extent, mitochondrial function of vitrified-warmed metaphase II bovine oocytes.

摘要

本研究旨在确定玻璃化对不同解冻后培养时间的牛卵母细胞的减数分裂纺锤体和线粒体功能的影响。部分去卵丘的卵母细胞复合物在不同成熟时间(18、20 和 24 小时)使用两步法添加冷冻保护剂进行玻璃化,并在成熟培养基中进行 6、4 或 0 小时的解冻后延长培养。在延长培养的 0 和 6 小时后分析微管构型和染色体排列,而在解冻后恢复的 0、4 和 6 小时测量线粒体膜电位和 ATP 含量。减数分裂纺锤体完整性的结果表明,与新鲜卵母细胞相比,经过 6 小时培养的玻璃化解冻卵母细胞具有相似的正常微管构型和染色体排列的发生率,但高于玻璃化对照组(无培养)。所有玻璃化组的线粒体膜电位没有差异,但在解冻后培养 4 小时的卵母细胞与新鲜卵母细胞具有相似的水平。所有玻璃化组的 ATP 浓度均低于对照组。然而,培养 6 小时的卵母细胞在玻璃化组中具有最低的 ATP 耗竭卵母细胞率。本研究的结果表明,解冻后延长培养促进了玻璃化解冻中期牛卵母细胞的减数分裂纺锤体和线粒体功能的恢复。

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