Nottola S A, Coticchio G, De Santis L, Macchiarelli G, Maione M, Bianchi S, Iaccarino M, Flamigni C, Borini A
Department of Anatomy, Sapienza University, Rome 00161, Italy.
Reprod Biomed Online. 2008 Sep;17(3):368-77. doi: 10.1016/s1472-6483(10)60220-9.
The morphological characteristics of frozen-thawed human mature oocytes (n = 12) were studied by light and transmission electron microscopy following cryopreservation using a slow cooling protocol including increasing concentrations of ethylene glycol (0.5-1.5 mol/l) and sucrose 0.2 mol/l in the freezing solution. Fresh human mature oocytes (n = 12) were used as controls. Fresh and frozen-thawed oocytes appeared rounded in section, with a homogeneous cytoplasm, an intact oolemma and a continuous zona pellucida. Disorganization of mitochondria-smooth endoplasmic reticulum aggregates and a decreased complement of microvilli and cortical granules were frequently observable in frozen-thawed oocytes. Increased density of the inner zona pellucida, possibly related to the occurrence of zona 'hardening', was sometimes found associated with a reduced amount of cortical granules. In addition, delamination of the zona pellucida was evident in some frozen-thawed samples. Finally, numerous vacuoles and secondary lysosomes were detected in the ooplasm of most frozen-thawed oocytes. In conclusion, frozen-thawed oocytes treated with ethylene glycol may show a variety of ultrastructural alterations, possibly related, at least in part, to the use of this cryoprotectant. Thus, the ethylene glycol-based protocol of slow cooling herein described does not seem to offer significant advantages in terms of oocyte structural preservation.
采用缓慢降温方案进行冷冻保存，冷冻液中乙二醇浓度递增（0.5 - 1.5 mol/l）并添加0.2 mol/l蔗糖，之后运用光学显微镜和透射电子显微镜研究了冻融人成熟卵母细胞（n = 12）的形态学特征。新鲜人成熟卵母细胞（n = 12）用作对照。新鲜及冻融卵母细胞在切片中呈圆形，细胞质均匀，卵膜完整，透明带连续。在冻融卵母细胞中经常可观察到线粒体 - 滑面内质网聚集体紊乱、微绒毛和皮质颗粒数量减少。有时发现透明带内层密度增加，可能与透明带“硬化”的发生有关，同时皮质颗粒数量减少。此外，在一些冻融样本中透明带分层明显。最后，在大多数冻融卵母细胞的卵质中检测到大量空泡和次级溶酶体。总之，用乙二醇处理的冻融卵母细胞可能会出现多种超微结构改变，这可能至少部分与这种冷冻保护剂的使用有关。因此，本文所述的基于乙二醇的缓慢降温方案在卵母细胞结构保存方面似乎没有显著优势。
