Department of Obstetrics, Gynecology, and Reproductive Biology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Cryobiology. 2012 Oct;65(2):79-87. doi: 10.1016/j.cryobiol.2012.06.001. Epub 2012 Jun 9.
Freezing unfertilized oocytes is an option for females without a partner, either to preserve their fertility prior to sterilizing cancer treatment or for social reasons. Our study considered whether it is best to freeze immature human oocytes at the germinal vesicle (GV) stage, prior to in vitro maturation (IVM) or at metaphase-II (M-II), after IVM. Sibling GV-stage oocytes from stimulated ICSI cycles were allocated to freezing either prior to (n=109) or after (n=107) IVM. Cumulus-free oocytes were cryopreserved using a choline-substituted slow-freezing protocol and matured in a defined medium, with analysis of chromatin, microtubules, and microfilaments by three-dimensional imaging. Cryopreserved oocytes were compared with oocytes matured in vitro but never frozen (n=114). Survival was similar between oocytes frozen before or after IVM (69.7% vs. 70.5%). Polar body extrusion after IVM was lower in oocytes frozen at the GV stage versus those matured and then frozen (51.3% vs. 75.7%) or not frozen (75.4%). Stratification by patient age (<36 and ⩾36year) showed no difference in oocyte survival or maturation. Oocytes frozen as GVs showed elevated proportions of spontaneous activation (with or without polar body), an effect augmented by patient age. Spindle and chromosome configurations were disrupted to similar extents in both groups of frozen oocytes, with no further detrimental effect of patient age. The length, width, and volume of bipolar M-II spindles were comparable in all three groups. When frozen as GVs, oocytes exhibited decreased maturation and increased spontaneous activation, suggesting that it is best to freeze oocytes at M-II.
冷冻未受精的卵母细胞是没有伴侣的女性的一种选择，无论是在进行绝育癌症治疗前保存生育能力，还是出于社会原因。我们的研究考虑了在体外成熟（IVM）之前将不成熟的人类卵母细胞在生发泡（GV）阶段冷冻，还是在 IVM 之后在中期 II（M-II）阶段冷冻。从刺激的 ICSI 周期中获得的姐妹 GV 期卵母细胞被分配在 IVM 之前（n=109）或之后（n=107）冷冻。使用胆碱替代的缓慢冷冻方案对无卵丘卵母细胞进行冷冻保存，并在定义的培养基中成熟，通过三维成像分析染色质、微管和微丝。将冷冻保存的卵母细胞与从未冷冻过的体外成熟（n=114）的卵母细胞进行比较。在 IVM 之前或之后冷冻的卵母细胞的存活率相似（69.7%对 70.5%）。与成熟后冷冻（51.3%对 75.7%）或未冷冻（75.4%）的卵母细胞相比，在 GV 期冷冻的卵母细胞在 IVM 后排出极体的比例较低（51.3%对 75.7%）。按患者年龄（<36 岁和 ⩾36 岁）分层，卵母细胞的存活率或成熟率无差异。GV 期冷冻的卵母细胞自发激活（有或无极体）的比例升高，这种效应随着患者年龄的增加而增强。两组冷冻卵母细胞的纺锤体和染色体构型均受到类似程度的破坏，且患者年龄无进一步的不良影响。所有三组的双极 M-II 纺锤体的长度、宽度和体积均相似。当作为 GV 冷冻时，卵母细胞表现出成熟度降低和自发激活增加，这表明最好在 M-II 阶段冷冻卵母细胞。
