Anguita B, Vandaele L, Mateusen B, Maes D, Van Soom A
Departament de Ciència Animal i dels Aliments, Facultat de Veterinària, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain.
Theriogenology. 2007 Feb;67(3):537-49. doi: 10.1016/j.theriogenology.2006.09.004. Epub 2006 Sep 27.
The number of follicles undergoing atresia in an ovary is very high, and isolation of cumulus-oocyte complexes (COCs) from such atretic follicles may impair subsequent embryo development in vitro. Our aim was to study if stringent selection by morphological assessment of COCs can improve embryo development, and to evaluate whether oocyte diameter is related with apoptotic ratio in oocytes and blastocysts. COCs from slaughtered cattle were recovered by follicle aspiration and classified depending on oocyte diameter: (A) <110 microm; (B) 110-120 microm; (C) >120 microm. COCs were matured, fertilized and cultured in vitro. Early and late stages of apoptosis were detected by Annexin-V and TUNEL staining, respectively, in denuded oocytes, COCs and blastocysts. Immature oocytes from Group A showed higher apoptotic ratio assessed by TUNEL assay, and the COCs corresponding to this group also showed a higher proportion of apoptotic cumulus cells. After maturation, no differences were present in the incidence of apoptosis among oocytes from different groups, but COCs corresponding to the largest diameter showed less apoptotic cumulus cells. In addition, the percentage of apoptotic oocytes decreased during in vitro maturation in all groups. Apoptotic cell ratio (ACR) in blastocysts was not related to oocyte diameter. In conclusion, oocyte selection and oocyte morphological evaluation prior to maturation was not sufficient to select non-atretic oocytes. When oocyte diameter was used as an additional selection the embryonic developmental potential increased together with oocyte diameter, but this improvement was not related to a lower incidence of apoptosis in the largest oocytes.
卵巢中发生闭锁的卵泡数量非常多,从这些闭锁卵泡中分离卵丘 - 卵母细胞复合体(COCs)可能会损害随后的体外胚胎发育。我们的目的是研究通过对COCs进行形态学评估进行严格筛选是否能改善胚胎发育,并评估卵母细胞直径是否与卵母细胞和囊胚中的凋亡率相关。通过卵泡抽吸从屠宰牛中回收COCs,并根据卵母细胞直径进行分类:(A)<110微米;(B)110 - 120微米;(C)>120微米。将COCs进行体外成熟、受精和培养。分别通过Annexin - V和TUNEL染色检测裸卵、COCs和囊胚中的早期和晚期凋亡阶段。通过TUNEL检测,A组未成熟卵母细胞显示出更高的凋亡率,并且该组对应的COCs也显示出更高比例的凋亡卵丘细胞。成熟后,不同组卵母细胞的凋亡发生率没有差异,但直径最大的COCs对应的卵丘细胞凋亡较少。此外,所有组在体外成熟过程中凋亡卵母细胞的百分比均下降。囊胚中的凋亡细胞比率(ACR)与卵母细胞直径无关。总之,在成熟前进行卵母细胞选择和卵母细胞形态学评估不足以选择非闭锁卵母细胞。当将卵母细胞直径用作额外的选择标准时,胚胎发育潜力随卵母细胞直径增加而增加,但这种改善与最大卵母细胞中较低的凋亡发生率无关。