Mitochondrial dysfunction is considered a crucial factor aggravating oocyte viability after vitrification-warming. To clarify the role of mitophagy in mitochondrial extinction of vitrified porcine oocytes, mitochondrial function, ultrastructural characteristics, mitochondria-lysosomes colocalization, and mitophagic proteins were detected with or without chloroquine (CQ) treatment. The results showed that vitrification caused mitochondrial dysfunction, including increasing reactive oxygen species production, decreasing mitochondrial membrane potential, and mitochondrial DNA copy number. Damaged mitochondrial cristae and mitophagosomes were observed in vitrified oocytes. A highly fused fluorescence distribution of mitochondria and lysosomes was also observed. In the detection of mitophagic flux, mitophagy was demonstrated as increasing fluorescence aggregation of microtubule-associated protein light chain 3B (LC3B), enhanced colocalization between LC3B, and voltage-dependent anion channels 1 (VDAC1), and upregulated LC3B-II/I protein expression ratio. CQ inhibited the degradation of mitophagosomes in vitrified oocytes, manifested as decreased mitochondria-lysosomes colocalization, increased fluorescence fraction of VDAC1 overlapping LC3B, increased LC3B-II/I protein expression ratio, and p62 accumulation. The inhibition of mitophagosomes degradation by CQ aggravated mitochondrial dysfunction, including increased oxidative damage, reduced mitochondrial function, and further led to loss of oocyte viability and developmental potentiality. In conclusion, mitophagy is involved in the regulation of mitochondrial function during porcine oocyte vitrification.

中文翻译：

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线粒体自噬参与玻璃化猪卵母细胞的线粒体功能障碍

线粒体功能障碍被认为是玻璃化加热后加重卵母细胞活力的关键因素。为了阐明线粒体自噬在玻璃化猪卵母细胞线粒体消退中的作用，在有或没有氯喹 (CQ) 处理的情况下检测线粒体功能、超微结构特征、线粒体-溶酶体共定位和线粒体蛋白。结果表明，玻璃化导致线粒体功能障碍，包括增加活性氧的产生、降低线粒体膜电位和线粒体 DNA 拷贝数。在玻璃化卵母细胞中观察到受损的线粒体嵴和线粒体吞噬体。还观察到线粒体和溶酶体的高度融合的荧光分布。在检测线粒体通量时，线粒体自噬被证明为增加微管相关蛋白轻链 3B (LC3B) 的荧光聚集，增强 LC3B 和电压依赖性阴离子通道 1 (VDAC1) 之间的共定位，以及上调 LC3B-II/I 蛋白表达比。CQ 抑制玻璃化卵母细胞中线粒体吞噬体的降解，表现为线粒体-溶酶体共定位减少，VDAC1 与 LC3B 重叠的荧光分数增加，LC3B-II/I 蛋白表达比增加和 p62 积累。CQ 抑制线粒体降解会加重线粒体功能障碍，包括增加氧化损伤、降低线粒体功能，并进一步导致卵母细胞活力和发育潜力的丧失。综上所述，