Liu Huiyu, Gao Yan, Zhai Bo, Jiang Hao, Ding Yu, Zhang Lianjiang, Li Changhong, Deng Qiong, Yu Xianfeng, Zhang Jiabao
Laboratory Animal Center, College of Animal Sciences, Jilin University, Changchun, China.
Cell Physiol Biochem. 2016;39(5):1735-1745. doi: 10.1159/000447874. Epub 2016 Oct 17.
This study aims to clarify the effects of polyadenylation status on M-phase promoting factors (MPFs) during in vitro porcine oocyte maturation.
In this study, porcine follicular oocytes from large follicles (> 5 millimeter (mm)) and small follicles (< 3 mm) were examined at different follicular developmental stages. The polyadenylation of maternal mRNAs was inhibited by the addition of 3'-deoxyadenosine (3'-da) during the germinal vesicle (GV)(0 h), GV breakdown (GVBD)(18 h), metaphase I (MI)(28 h), and metaphase II (MII) (44 h) stages. In addition, the expression levels and poly-(A) tail lengths of the maternal mRNAs Cyclin B1 and cell division cycle 2 (Cdc2) were determined by real-time quantitative PCR. Immunofluorescence was used to assess spindle formation and chromosome alignment in the examined oocytes.
In large-follicle oocytes, the effects of inhibiting polyadenylation caused the percentage of mature to be significantly lower for the treated group than for the untreated group (p < 0.01). 3'-da can significantly improve the rate of small oocyte maturation in vitro and inhibits Cdc2 polyadenylation. Cyclin B1 plays a significant role in promoting the maturation of large-follicle oocytes. Polyadenylation contributes to the formation of dominant follicles and facilitates the selection of dominant follicles. However, the inhibition of adenylation affected spindle formation-related propulsion and chromosome alignment in both large- and small-follicle oocytes. The first polar body could not be extruded in certain large follicles.
3'-da can significantly improve the rate of small oocyte maturation in vitro, but it can also affect spindle formation-related propulsion and chromosome alignment.
本研究旨在阐明体外猪卵母细胞成熟过程中多聚腺苷酸化状态对M期促进因子（MPF）的影响。
在本研究中，对来自大卵泡（>5毫米（mm））和小卵泡（<3毫米）的猪卵泡卵母细胞在不同卵泡发育阶段进行了检测。在生发泡（GV）（0小时）、生发泡破裂（GVBD）（18小时）、中期I（MI）（28小时）和中期II（MII）（44小时）阶段，通过添加3'-脱氧腺苷（3'-da）抑制母源mRNA的多聚腺苷酸化。此外，通过实时定量PCR测定母源mRNA细胞周期蛋白B1和细胞分裂周期2（Cdc2）的表达水平和多聚（A）尾长度。采用免疫荧光法评估所检测卵母细胞中的纺锤体形成和染色体排列。
在大卵泡卵母细胞中，抑制多聚腺苷酸化的处理组成熟率显著低于未处理组（p<0.01）。3'-da可显著提高小卵母细胞的体外成熟率，并抑制Cdc2的多聚腺苷酸化。细胞周期蛋白B1在促进大卵泡卵母细胞成熟中起重要作用。多聚腺苷酸化有助于优势卵泡的形成并促进优势卵泡的选择。然而，腺苷酸化的抑制影响了大、小卵泡卵母细胞中纺锤体形成相关的推进和染色体排列。在某些大卵泡中无法排出第一极体。
3'-da可显著提高小卵母细胞的体外成熟率，但也会影响纺锤体形成相关的推进和染色体排列。
