Mitochondrial DNA quantity Counteracts ROS damage in oocytes during female reproductive aging

We investigated the contentious role of mitochondrial reactive oxygen species (ROS) on mitochondrial DNA (mtDNA) quality and quantity in female reproductive aging. By conditionally knocking out the Sod2 gene in female mouse germline, we observed increased mitochondrial ROS and decreased oocyte quality, primarily due to impacts on OXPHOS complex II and mtDNA encoded mRNA levels. Interestingly, we found no increased mtDNA mutations, but alterations in mtDNA quantity, indicating the susceptibility of mtDNA to the mitochondrial ROS during reproductive aging. Notably, when we further decreased the basal level of mtDNA quantity by deactivating the mtSSB protein in Sod2 conditional knockout females, we observed an exacerbation of reproductive aging effects. This highlights the crucial role of mtDNA quantity in mitigating the impact of oxidative stress on fertility. To examine the molecular alterations in oocytes caused by elevated levels of mitochondrial matrix ROS, we employed the Smart seq2 method to study the transcriptomic alterations in mature oocytes from approximately six-month-old females between L/L cre mice and L/L mice.本研究探究了线粒体活性氧（mitochondrial reactive oxygen species, ROS）在雌性生殖衰老过程中对线粒体DNA（mitochondrial DNA, mtDNA）质量与数量的争议性作用。我们通过在雌性小鼠生殖系中条件性敲除Sod2基因，观察到线粒体ROS水平升高且卵母细胞质量下降，该效应主要源于线粒体ROS对氧化磷酸化复合体II及线粒体DNA编码mRNA水平的影响。有趣的是，我们未检测到线粒体DNA突变水平升高，但观察到线粒体DNA数量发生改变，这表明在生殖衰老过程中，线粒体DNA对线粒体ROS具有易感性。值得注意的是，当我们在Sod2条件性敲除雌性小鼠中进一步通过失活线粒体单链DNA结合蛋白（mitochondrial single-stranded DNA-binding protein, mtSSB）以降低线粒体DNA的基础水平时，生殖衰老的表型出现了加重。这一结果凸显了线粒体DNA数量在缓解氧化应激对生育力的影响中所发挥的关键作用。为探究线粒体基质ROS水平升高所引发的卵母细胞分子改变，我们采用Smart-seq2方法，对约6月龄的L/L cre雌性小鼠与L/L雌性小鼠的成熟卵母细胞进行转录组学分析。