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Ü卵子体外成熟对胞质线粒体DNA拷贝和功能的影响
葛红山* 李晓和 张帆 陈华 习海涛 黄检英 朱春芳 吕杰强
解剖学报 ›› 2013, Vol. 44 ›› Issue (5) : 680-684.
Ü卵子体外成熟对胞质线粒体DNA拷贝和功能的影响
Effects of maturation in vitro on mitochondrial DNA copy and functions in oocytes
目的 研究卵子体外成熟（IVM）过程对胞质内线粒体的数目、功能及卵子质量的影响，进而探讨IVM卵子低发育潜能的可能机制。方法 实验小鼠随机分为IVM组和体内成熟（IVO）组；应用Real-time-PCR、免疫荧光和荧光-荧光素酶测定法，分别检测IVM和IVO来源卵子的线粒体DNA（mtDNA）拷贝数、线粒体膜电位强度、ATP含量、线粒体分布、胞质ROS水平、卵子骨架和染色体结构。 结果 相对于IVO卵子，IVM卵子的mtDNA拷贝数显著降低；而胞质ROS生成和纺锤体及染色体结构异常率则显著增加。胞质线粒体分布模型和氧化磷酸化活性在IVM和IVO卵子间差异无显著意义。结论 非生理性IVM过程显著抑制了胞质mtDNA的复制，并增加了ROS生成和纺锤体和染色体结构异常，继而可能影响卵子细胞质的成熟进程，这可部分解释IVM卵子低发育潜能的机制。
Objectiv To determine whether in vitromaturation (IVM) has an impact on oocyte’s mitochondrial number, and functions, and on oocyte’s quality.
Methods The maturation mice oocytes were collected following either in vitro or in vivo (IVO) maturation process. The copies of mtDNA, the magnitude of mitochondrial membrane potential (ΔΨm) and oocyte adenosine triphosphate (ATP) content, pattern of mitochondrial distribution, reactive oxygen species（ROS）level, and the integrity of spindle and chromosome alignment were evaluated by RT-PCR, immunofluorescence and fluorescence-luciferase mensuration. Results Significant differences were detected in the copies of mtDNA，ROS level and integrity of cytoskeleton in oocytes between IVM and IVO.There were no significant differences in mitochondrial distribution, oxidative phosphorylation function in oocyte between IVM and IVO. Conclusion The in vitro non-physiological maturation may inhibit mtDNA replication, and increase ROS production and rates of abnormal spindle and chromosome alignment. This may partly explain the low development potential of IVM oocyte.
卵子 / 线粒体 / 活性氧 / 细胞骨架 / 体外成熟 / 实时荧光定量PCR / 免疫荧光 / 小鼠
Oocyte /
Mitochondria /
Rective oxygen species /
Cytoskeleton /
In vitromaturation /
Real-time PCR /
Immunofluorescence /
Mouse
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国家自然科学基金（30900516）;浙江省自然科学基金（Y2081011）;国家自然科学基金资助项目;自然科学基金资助项目
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