Grupen Christopher G, Armstrong David T
Faculty of Veterinary Science, The University of Sydney, Camden, NSW 2570, Australia.
Reprod Fertil Dev. 2010;22(7):1100-9. doi: 10.1071/RD09307.
The objective of the present study was to determine the temporal effects of sow follicular fluid (FF) in vitro on cumulus cell viability and function, as well as oocyte nuclear and cytoplasmic maturation. Cumulus-oocyte complexes (COCs) recovered from the ovaries of prepubertal pigs were matured in medium with (+FF) or without (-FF) follicular fluid for the first 22 h of IVM. At 22 h of IVM, each group of COCs was then transferred to medium with or without FF and matured for another 22 h, forming four treatment groups (-FF/-FF, -FF/+FF, +FF/-FF and +FF/+FF). The concentration of progesterone in spent IVM medium and the incidence of cumulus cell apoptosis in individual COCs were determined at 22 and 44 h of IVM. Cumulus expansion was also recorded at 44 h of IVM. Finally, the ability of oocytes to complete meiosis to the MII stage and form blastocysts after IVF and embryo culture was assessed. Maturation with FF for part or the whole of IVM increased cumulus expansion and progesterone production and decreased the incidence of cumulus cell apoptosis compared with the -FF/-FF group (P < 0.05). The changes were greatest for the +FF/+FF group and intermediate for the -FF/+FF and +FF/-FF groups. Regression analysis revealed a negative association between cumulus cell progesterone production and the incidence of cumulus cell apoptosis (P < 0.001). Meiotic maturation was enhanced when FF was present during the first half of IVM. Oocytes matured in the presence of FF during the first and/or second half of IVM displayed an increased ability to form blastocysts compared with the -FF/-FF group (P < 0.05). The extent of the increase was similar for all FF-supplemented groups. The results show that FF exerts several beneficial effects at different times during IVM and suggest that a major role of FF is to provide protection from oxidative stress. We propose that the incidence of cumulus cell apoptosis in COCs must be kept below a certain threshold to ensure adequate functionality, including steroidogenic activity, is maintained for the acquisition of oocyte developmental competence.
本研究的目的是确定体外培养的母猪卵泡液（FF）对卵丘细胞活力和功能以及卵母细胞核与细胞质成熟的时间效应。从青春期前猪的卵巢中回收的卵丘-卵母细胞复合体（COCs）在体外成熟培养（IVM）的前22小时培养于添加（+FF）或不添加（-FF）卵泡液的培养基中。在IVM 22小时时，将每组COCs转移至添加或不添加FF的培养基中再培养22小时，形成四个处理组（-FF/-FF、-FF/+FF、+FF/-FF和+FF/+FF）。在IVM 22小时和44小时时测定用过的IVM培养基中孕酮的浓度以及单个COCs中卵丘细胞凋亡的发生率。在IVM 44小时时也记录卵丘扩展情况。最后，评估卵母细胞在体外受精和胚胎培养后完成减数分裂至MII期并形成囊胚的能力。与-FF/-FF组相比，在IVM的部分或全部时间用FF培养可增加卵丘扩展和孕酮生成，并降低卵丘细胞凋亡的发生率（P<0.05）。+FF/+FF组的变化最大，-FF/+FF和+FF/-FF组变化居中。回归分析显示卵丘细胞孕酮生成与卵丘细胞凋亡发生率之间呈负相关（P<0.001）。当IVM上半程存在FF时，减数分裂成熟得到增强。与-FF/-FF组相比，在IVM上半程和/或下半程存在FF时成熟的卵母细胞形成囊胚的能力增强（P<0.05）。所有添加FF的组增加的程度相似。结果表明，FF在IVM的不同时间发挥多种有益作用，并提示FF的主要作用是提供抗氧化应激的保护。我们提出，COCs中卵丘细胞凋亡的发生率必须保持在一定阈值以下，以确保维持包括类固醇生成活性在内的足够功能，从而获得卵母细胞发育能力。
