Department of Biotechnology, College of Engineering, Daegu University, 201 Daegudae-ro, Jillyang, Gyeongsan, Gyeongbuk 38453, Korea.
Institute of Infertility, Daegu University, 201 Daegudae-ro, Jillyang, Gyeongsan, Gyeongbuk 38453, Korea.
Int J Mol Sci. 2020 Apr 26;21(9):3050. doi: 10.3390/ijms21093050.
While triclosan (TCS) exerts detrimental effects on female reproduction, the effect of TCS-derived toxins on porcine oocytes during in vitro maturation (IVM) is unclear. This study investigated the effects of TCS on mitochondrion-derived reactive oxygen species (ROS) production and apoptosis pathways during porcine oocyte maturation. Porcine oocytes were treated with TCS (1, 10, and 100 μM) and triphenylphosphonium chloride (Mito-TEMPO; 0.1 μM), and matured cumulus oocyte complexes (COCs) were stained with orcein, dichlorofluorescein diacetate (DCF-DA), and Mito-SOX. Proteins and mRNA levels of factors related to cumulus expansion and mitochondrion-mediated apoptosis and antioxidant enzymes were analyzed by western blotting and reverse-transcription polymerase chain reaction (RT-PCR), respectively. Meiotic maturation and cumulus cell expansion significantly decreased for COCs after TCS treatment along with an increase in mitochondrial superoxide levels at 44 h of IVM. Further, mitochondrion-related antioxidant enzymes and apoptosis markers were significantly elevated in porcine COCs following TCS-mediated oxidative damage. The protective effect of Mito-TEMPO as a specific superoxide scavenger from TCS toxin improved the maturation capacity of porcine COCs. Mito-TEMPO downregulated the mitochondrial apoptosis of TCS-exposed porcine COCs by reducing superoxide level. In conclusion, our data demonstrate that TCS mediates toxicity during porcine oocyte maturation through superoxide production and mitochondrion-mediated apoptosis.
虽然三氯生（TCS）对女性生殖系统有不良影响，但 TCS 衍生毒素对猪卵母细胞体外成熟（IVM）的影响尚不清楚。本研究探讨了 TCS 对猪卵母细胞成熟过程中线粒体衍生活性氧（ROS）产生和凋亡途径的影响。用 TCS（1、10 和 100 μM）和三苯基膦氯化物（Mito-TEMPO；0.1 μM）处理猪卵母细胞，并对成熟的卵丘-卵母细胞复合物（COC）进行染色，用曙红、二氯荧光素二乙酸酯（DCF-DA）和 Mito-SOX。通过 Western blot 和逆转录聚合酶链反应（RT-PCR）分别分析与卵丘扩张和线粒体介导的凋亡以及抗氧化酶相关的因子的蛋白质和 mRNA 水平。TCS 处理后，COC 的减数分裂成熟和卵丘细胞扩张明显减少，IVM44 小时时线粒体中超氧化物水平增加。此外，TCS 介导的氧化损伤后，猪 COC 中的线粒体相关抗氧化酶和凋亡标志物显著升高。Mito-TEMPO 作为 TCS 毒素的特异性超氧化物清除剂，可改善猪 COC 的成熟能力。Mito-TEMPO 通过降低超氧化物水平来下调 TCS 暴露的猪 COC 的线粒体凋亡。总之，我们的数据表明，TCS 通过产生超氧阴离子和线粒体介导的细胞凋亡来介导猪卵母细胞成熟过程中的毒性。
