Effect of chilling on porcine germinal vesicle stage oocytes at the subcellular level
Effect of chilling on porcine germinal vesicle stage oocytes at the subcellular level
复制标题
冷冻对猪生发囊期卵母细胞亚细胞水平的影响
DOI：
10.1016/j.cryobiol.2009.04.005
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发表时间：
2009-08-01
期刊：
影响因子：
2.7
通讯作者：
Liu Zhong-Hua
中科院分区：
文献类型：
--
作者：
Gerelchimeg, Bou;Liu Li-Qing;Liu Zhong-Hua
研究了冷冻对猪生发泡(GV)期卵母细胞的亚细胞影响。体外成熟前，取直径3~6 mm的有腔卵泡卵丘-卵母细胞复合体(COCs)，随机分为4组，分别在39℃(对照组)、23℃(室温)、15℃(15℃)和10℃(10℃)的PVA-TL-HEPES培养液中孵育10min。在39℃下体外培养42h后，检测存活率。23℃冷冻组和对照组的存活率(77.92%和91.89%)无显著差异，但15℃和10℃冷冻组的存活率(46.34%和4.81%)显著低于对照组(P<；0.01)。15℃组的进一步实验表明，大多数卵母细胞在体外成熟后2~4h死亡。为了在亚细胞水平上研究冷冻对卵母细胞的影响，制备了对照组和15℃冷冻组的卵母细胞，分别固定在体外培养的不同时间点(体外培养2、2.5、3、3.5和4h)，进行透射电子显微镜观察。结果表明，与对照组相比，15℃处理组超微结构形态发生了两个显著变化：(1)异质脂显著减少，(2)线粒体-内质网-脂泡(M-E-L)结合紊乱。这些结果表明，15℃是猪GV期卵母细胞的临界冷藏温度，细胞化学成分的改变和M-E-L组合的破坏可能是该阶段猪卵母细胞冷害的原因。(C)2009 Elsevier Inc.保留所有权利。
The potential subcellular consequence of chilling on porcine germinal vesicle (GV) stage oocytes was examined. Prior to in vitro maturation (IVM), Cumulus-oocyte complexes (COCs) freshly collected from antral follicles (3-6 mm in diameter) were evenly divided into four groups and immediately incubated in PVA-TL-HEPES medium at the temperature of 39 degrees C (control group), 23 degrees C (room temperature), 15 degrees C and 10 degrees C for 10 min, respectively. Following 42 h of IVM at 39 degrees C, the survival rates were examined. There was no significant difference between the survival rate of 23 degrees C chilled group and control group (77.92 and 91.89%), but the survival rate of 15 and 10 degrees C chilled group were significantly decreased (46.34 and 4.81%, P < 0.01). A further experiment on15 degrees C group showed that most oocytes died from 2 to 4 h of IVM. In order to investigate the effects of chilling on oocytes at the subcellular level, the control and 15 degrees C chilled group COCs fixed at different time points of the IVM cultures (2, 2.5, 3, 3.5 and 4 h of IVM) were prepared for transmission electron microscope (TEM) observation. As the result, compared with the control group, there were two significant changes in the ultrastructural morphology of 15 degrees C treatment group: (1) dramatic reduction of heterogeneous lipid, (2) disorganized mitochondria-endoplasmic reticulum-lipid vesicles (M-E-L) combination. These results indicate that 15 degrees C is a critical chilling temperature for porcine GV stage oocyte and the alteration of cellular chemical composition and the destruction of M-E-L combination maybe responsible for chilling injury of porcine oocyte at this stage. (C) 2009 Elsevier Inc. All rights reserved.