Aged mouse oocytes fail to readjust intracellular adenosine triphosphates at fertilization
Aged mouse oocytes fail to readjust intracellular adenosine triphosphates at fertilization
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DOI：
10.1095/biolreprod.104.034926
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发表时间：
2005-05-01
影响因子：
3.6
通讯作者：
Kurachi, H
中科院分区：
文献类型：
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作者：
Igarashi, H;Takahashi, T;Kurachi, H
排卵后卵母细胞衰老对胚胎发育有显著影响。此外，Ca2+振荡模式的改变可以观察到在受精，衰老的小鼠卵母细胞。由于Ca2+振荡依赖于内质网Ca2+的释放和再摄取，而后者依赖于ATP的可用性，我们同时测量了新鲜和衰老小鼠卵母细胞中细胞内ATP浓度（[ATP](i)）和Ca2+振荡的变化。我们通过荧光染料镁绿（MgG）测量细胞内游离Mg2+浓度来持续评估[ATP]i的变化，同时通过Fura-PE3监测细胞内Ca2+浓度（[Ca2+](i)）。在受精时，MgG荧光随着[Ca2+](i)的第一次短暂升高而短暂增加，表明[ATP](i)相对减少。在新鲜卵母细胞中，它很快就会显著下降到基线以下，表明[ATP]i相对增加。相反，在衰老的卵母细胞中，没有观察到MgG荧光的这种下降。在单独的实验中，用荧光素-荧光素酶法测定新鲜和老化卵母细胞中ATP的含量。体外测定的细胞内ATP含量在未受精的新鲜卵母细胞和衰老卵母细胞中是相当的。受精后5 h，两种卵母细胞的细胞内ATP含量均有所增加，其中新鲜卵母细胞的细胞内ATP含量明显高于衰老卵母细胞。这些发现表明，老年小鼠卵母细胞在受精时不能重新调节细胞内ATP的水平。受精时ATP的相对缺乏可能导致Ca2+振荡模式的改变和发育效力差，这在衰老的卵母细胞中很常见。
Postovulatory aging of oocytes significantly affects embryonic development. Also, altered Ca2+ oscillation patterns can be observed in fertilized, aged mouse oocytes. Because Ca2+ oscillations depend on Ca2+ release and reuptake in the endoplasmic reticulum, and the latter relies on ATP availability, we simultaneously measured changes in intracellular ATP concentration ([ATP](i)) and Ca2+ oscillations in fresh and aged mouse oocytes. We continuously assessed changes in [ATP]i from intracellular free Mg2+ concentration measured by fluorescent dye Magnesium Green (MgG) while intracellular Ca2+ concentration ([Ca2+](i)) was monitored by Fura-PE3. At fertilization, MgG fluorescence was transiently increased concomitant with the first transient elevation in [Ca2+](i), indicating a relative decrease in [ATP](i). In fresh oocytes, it was quickly followed by a significant decrease below baseline, indicating a relative increase in [ATP]i. In contrast, in aged oocytes, such a decrease in MgG fluorescence was not observed. In a separate experiment, ATP content in fresh and aged oocytes was determined in vitro by the luciferin-luciferase assay. Intracellular ATP contents measured in vitro were comparable in unfertilized fresh and aged oocytes. Intracellular ATP content at 5 h after fertilization was increased in both oocytes, where fresh oocytes showed a significantly higher intracellular value than aged oocytes. These findings suggest that aged mouse oocytes fail to readjust the level of intracellular ATP at fertilization. Relative deficiencies of ATP at fertilization might lead to an altered Ca2+ oscillation pattern and poor developmental potency, which is commonly noted in aged oocytes.