Gu Ruihuan, Feng Yun, Guo Song, Zhao Shen, Lu Xiaowei, Fu Jing, Sun Xiaoxi, Sun Yijuan
Shanghai Ji Ai Genetics & IVF Institute, Obstetrics & Gynecology Hospital, Fudan University, Shanghai, 200011, China.
Reproductive Medical Center, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, 200025, China.
Cryobiology. 2017 Apr;75:144-150. doi: 10.1016/j.cryobiol.2016.12.009. Epub 2017 Jan 3.
At present, the metaphase II (MII) oocytes have a very special structure that leads to complex difficulties associated with its vitrification, and their efficacy still needs a large amount of study to observe. The present study was to investigate whether transient hydrostatic pressure (THP), which was utilized for oocytes before vitrification, had positive effect on the oocytes' developmental ability and reactive oxygen species, and had no damage on meiotic spindle, zona pellucida, and DNA copy number.
All the immature oocytes used in this research were collected between February 2015 and December 2015 in Shanghai Ji Ai Genetics & IVF Institute. The MII oocytes, which were originated from metaphase I (MI) oocytes, were randomly distributed into three groups: A) fresh oocytes; B) vitrification; and C) vitrification after THP treatment. The embryo developmental outcome was evaluated after intracytoplasmic sperm injection and embryo culture. Furthermore, the meiotic spindle behavior, reactive oxygen species (ROS), zona pellucida (ZP), and DNA copy number variation were evaluated and compared among the three groups.
A total of 568 MII oocytes were included in the study. Embryos from group B had fewer cells on day 3 compared with group A and C (5.01 ± 2.11 for group A, 3.89 ± 2.21 for group B, and 4.69 ± 2.05 for group C). The developmental feature of blastocyst in groups A and C were superior to those of group B. The MII oocytes were manipulated with THP before vitrification, and the equilibration time was significantly shorter in the vitrification procedure (244.9 ± 30.1 vs. 181.5 ± 10.1). The ROS, ZP of vitrified/warmed oocytes in group C were improved with THP before vitrification. THP had no influence on the meiotic spindle and DNA copy number variation of vitrified/warmed oocytes.
The results of the study indicated that sublethal THP treatment before vitrification increased the developmental competence of human in vitro matured oocytes, reduced vitrification-related changes in the ROS, which occurred during oocyte vitrification, and did not damage the meiotic spindle, ZP and DNA copy number variation.
目前，中期 II（MII）卵母细胞具有非常特殊的结构，这导致其玻璃化存在复杂的困难，其效果仍需要大量研究来观察。本研究旨在探讨玻璃化前用于卵母细胞的瞬时静水压力（THP）是否对卵母细胞的发育能力和活性氧有积极影响，并且对减数分裂纺锤体、透明带和 DNA 拷贝数无损害。
本研究中使用的所有未成熟卵母细胞于 2015 年 2 月至 2015 年 12 月在上海集爱遗传与不育诊疗中心收集。源自中期 I（MI）卵母细胞的 MII 卵母细胞被随机分为三组：A）新鲜卵母细胞；B）玻璃化组；C）THP 处理后玻璃化组。在卵胞浆内单精子注射和胚胎培养后评估胚胎发育结果。此外，对三组卵母细胞的减数分裂纺锤体行为、活性氧（ROS）、透明带（ZP）和 DNA 拷贝数变异进行评估和比较。
本研究共纳入 568 个 MII 卵母细胞。与 A 组和 C 组相比，B 组胚胎在第 3 天的细胞数量较少（A 组为 5.01±2.11，B 组为 3.89±2.21，C 组为 4.69±2.05）。A 组和 C 组囊胚的发育特征优于 B 组。MII 卵母细胞在玻璃化前用 THP 处理，玻璃化过程中的平衡时间明显缩短（244.9±30.1 对 181.5±10.1）。C 组玻璃化/解冻卵母细胞的 ROS、ZP 在玻璃化前用 THP 处理后得到改善。THP 对玻璃化/解冻卵母细胞的减数分裂纺锤体和 DNA 拷贝数变异没有影响。
研究结果表明，玻璃化前亚致死性 THP 处理可提高人类体外成熟卵母细胞的发育能力，减少卵母细胞玻璃化过程中发生的与玻璃化相关的 ROS 变化，并且不会损害减数分裂纺锤体、ZP 和 DNA 拷贝数变异。
