Page 7 - 南京医科大学自然版

第44卷第7期 南京医科大学学报（自然科学版）
2024年7月 Journal of Nanjing Medical University（Natural Sciences） ·891 ·

·基础研究·

腐胺通过线粒体相关内质网膜改善高龄卵母细胞质量

倪 曼，程 丽，郭 爽，腊博雅，严正杰，崔毓桂，覃莲菊 ，刘嘉茵 *
*
南京医科大学第一附属医院生殖医学中心，生殖医学与子代健康全国重点实验室，江苏 南京 210029

［摘 要］ 目的：评估体外成熟（in vitro maturation，IVM）培养液中添加腐胺是否可改善高龄小鼠卵母细胞质量，及腐胺对高龄
小鼠卵母细胞中线粒体相关内质网膜（mitochondria⁃associated endoplasmic reticulum membrane，MAM）的影响。方法：取生发泡
期（germinal vesicle，GV）小鼠卵母细胞进行体外成熟培养至减数分裂（meiosis，M）Ⅱ期，分别使用8周龄小鼠卵母细胞作为年
轻对照组，40周龄小鼠卵母细胞作为老龄对照组，IVM液中添加0.5 mmol/L腐胺的40周龄小鼠卵母细胞作为老龄实验组。通
过检测MⅡ期卵母细胞的第一极体排出率、二细胞率、囊胚率、皮质颗粒分布、纺锤体异常率和染色体异常率评估卵母细胞的
质量。使用透射电镜观察MⅡ期卵母细胞的MAM，qPCR检测MAM间Ca 传递的关键分子含量，Ca 探针测定线粒体、内质网和
2+
2+
胞质的Ca 水平，并通过检测线粒体膜电位、ATP含量和活性氧（reactive oxygen species，ROS）水平评估线粒体功能。结果：腐
2+
胺提高了高龄小鼠卵母细胞的第一极体排出率（P < 0.05）和囊胚率（P < 0.05），并显著降低皮质颗粒分布异常率（P < 0.01）、纺
锤体异常率（P < 0.01）和染色体异常率（P < 0.05）。腐胺可阻止高龄小鼠卵母细胞的MAM间距缩短变化（P < 0.001），降低其
间 Ca 传递的关键分子含量（P < 0.05），缓解由 MAM 间 Ca 快速传递引起的线粒体钙超载（P < 0.001），从而改善线粒体功能
2+
2+
（P < 0.05），降低细胞内ROS水平（P < 0.001）。结论：体外成熟培养液中添加腐胺可显著改善高龄小鼠卵母细胞质量，且腐胺
可通过影响MAM间Ca 传递缓解线粒体钙超载，改善线粒体功能。
2+
［关键词］ 腐胺；线粒体相关内质网膜；卵母细胞；高龄
［中图分类号］ R321.1 ［文献标志码］ A ［文章编号］ 1007⁃4368（2024）07⁃891⁃10
doi：10.7655/NYDXBNSN240332

Putrescine improves the quality of aged oocytes through mitochondria ⁃ associated
endoplasmic reticulum membrane
* *
NI Man，CHENG Li，GUO Shuang，LA Boya，YAN Zhengjie，CUI Yugui，QIN Lianju ，LIU Jiayin
State Key Laboratory of Reproductive Medicine and Offspring Health，Clinical Center of Reproductive Medicine，the
First Affiliated Hospital of Nanjing Medical University，Nanjing 210029，China

［Abstract］ Objective：The aim of this study is to evaluate the impact of putrescine on oocyte quality in aged mice during in vitro
maturation（IVM）. Additionally，the effect of putrescine on mitochondria⁃associated endoplasmic reticulum membrane（MAM）in aged
mouse oocytes will be investigated. Methods：Germinal vesical（GV）stage mouse oocytes were cultured in vitro until they reached the
Meiosis（M）Ⅱ stage. The young control group consisted of 8⁃week⁃old mice，while the old control group consisted of 40⁃week⁃old mice.
The old experimental group consisted of 40⁃week⁃old mouse oocytes supplemented with 0.5 mmol/L putrescine in the IVM solution. The
oocyte quality was assessed by detecting the first polar body（PB1）extrusion rate，2⁃cell rate，blastocyst formation rate，cortical granule
distribution，spindle abnormality rate，and chromosome abnormality rate of oocytes at stage M Ⅱ. Additionally，MAM in MⅡ⁃stage
2+
oocyte was observed using transmission electron microscopy，the content of key molecules for Ca transfer was detected by qPCR，and
2 +
2 +
the Ca levels in mitochondria，endoplasmic reticulum，and cytoplasm were determined using Ca probes. Mitochondrial function was
assessed by measuring mitochondrial membrane potential，ATP content，and reactive oxygen species（ROS）levels. Results：Putrescine
increased the rate of PB1 extrusion（P < 0.05）and the blastocyst rate（P < 0.05）in oocytes of aged mice and significantly reduced the
rate of cortical granule distribution abnormalities（P < 0.01），spindle abnormalities（P < 0.01），and chromosomal abnormalities（P <
0.05）. The oocytes contained MAM structures. Putrescine prevented the shortening of MAM spacing（P < 0.001）in oocytes of aged mice

［基金项目］ 国家重点研发项目（2021YFC2700404，2021YFC2700501）
∗
通信作者（Corresponding author），E⁃mail：jyliu_nj@126.com；ljqin@njmu.edu.cn