Interactions between oocytes and cumulus cells during in vitro maturation of porcine cumulus-oocyte complexes in a chemically defined medium: Effect of denuded oocytes on cumulus expansion and oocyte maturation
Interactions between oocytes and cumulus cells during in vitro maturation of porcine cumulus-oocyte complexes in a chemically defined medium: Effect of denuded oocytes on cumulus expansion and oocyte maturation
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DOI：
10.1016/j.theriogenology.2014.10.026
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发表时间：
2015-03-01
期刊：
影响因子：
2.8
通讯作者：
Van Soom, A.
中科院分区：
文献类型：
--
作者：
Appeltant, R.;Somfai, T.;Van Soom, A.
本研究的目的是利用一种允许单个卵母细胞跟踪的系统，在化学定义的培养基中，在卵丘-卵母细胞复合物（COCs）的IVM中，阐明卵母细胞和卵丘细胞（CCs）在卵丘扩张和卵母细胞成熟水平上的相互作用。特别是，通过添加去角质卵母细胞（DOs）来研究卵母细胞分泌因子的影响，作为改善IVM系统的可能途径。基本成熟培养基为猪卵母细胞培养基，仅在IVM的前20小时添加促性腺激素。在体外培养过程中，用Cell-Tak涂层将COCs固定在培养皿的底部，这样可以在体外培养过程中对COCs进行个体跟踪。通过数字图像分析测量IVM期间COCs的大小变化。在完整COCs的猪卵母细胞培养基中，积云膨胀在IVM的20小时内呈典型的增加趋势，随后在IVM的48小时内减小（P < 0.05）。通过卵母细胞切除从COCs中去除卵母细胞，可在一定程度上使cc扩增，但其扩增能力低于COCs （P < 0.05）。添加DOs （COCs与DOs之比为9:16）并没有改善完整COCs的积云扩张和卵母细胞成熟率（P < 0.05），但却增强了卵母细胞去中心化复合物的积云扩张（P < 0.05）。此外，与COCs相比，在IVM前去除CCs可提高卵母细胞成熟率（分别为523%和32.9%）（P < 0.05），在IVM培养基中加入间隙连接抑制剂卡贝诺洛酮对COCs也有类似的效果：在IVM 20小时时，卡贝诺洛酮抑制COCs的扩增。综上所述，猪卵母细胞通过间隙连接通讯和卵母细胞分泌因子的产生促进了积云的扩大。然而，卵母细胞的存在并不是这个过程的先决条件。作为回报，在最初的培养过程中，CCs通过间隙连接维持卵母细胞的减数分裂停滞。在此基础上，未来的研究将进一步探讨在体外受精过程中与DOs共培养是否有利于受精和胚胎发育。(C) 2015爱思唯尔公司版权所有。
The aim of the present study was to clarify interactions between oocytes and cumulus cells (CCs) on the level of cumulus expansion and oocyte maturation during IVM of cumulus-oocyte complexes (COCs) in a chemically defined medium using a system that allows individual tracking of oocytes. Especially, the influence of oocyte-secreted factors was investigated by the aid of addition of denuded oocytes (DOs) as a possible approach to improve the IVM system. The basic maturation medium was porcine oocyte medium with addition of gonadotropins only during the first 20 hours of IVM. During IVM, COCs were kept fixed to the bottom of culture dish by adhesive Cell-Tak coating, which enabled individual tracking of COCs during IVM. Size changes in COCs during IVM were measured by digital image analysis. Cumulus expansion in a porcine oocyte medium of intact COCs increased in a typical manner until 20 hours and decreased in size subsequently until 48 hours of IVM (P < 0.05). Removal of oocytes from COCs by oocytectomy allowed the expansion of CCs to some extent, although their expansion ability was lower than that of COCs (P < 0.05). Addition of DOs (COCs to DOs ratio of 9:16) did not improve cumulus expansion and oocyte maturation rates of intact COCs (P > 0.05) but did enhance cumulus expansion of oocytectomized complexes (P < 0.05). Furthermore, removal of CCs before IVM increased oocyte maturation rates compared with COCs (523% and 32.9%, respectively) (P < 0.05) and a similar effect was observed in COCs when the gap junction inhibitor carbenoxolone was added to the IVM medium: carbenoxolone repressed the expansion of COCs at 20 hours of IVM. In conclusion, the porcine oocyte enhances cumulus expansion both by gap junctional communications and presumably by oocyte-secreted factor production. Nevertheless, the presence of oocytes is not a prerequisite for this process. In return, CCs maintain meiotic arrest in cumulus-enclosed oocytes during the initial culture through gap junctions. On the basis of these findings, future research could investigate if coculture with DOs during IVM is beneficial for fertilization and embryo development. (C) 2015 Elsevier Inc. All rights reserved.