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Microcystin-LR exposure results in aberrant spindles and induces apoptosis in porcine oocytes
Theriogenology ( IF 2.4 ) Pub Date : 2020-12-01 , DOI: 10.1016/j.theriogenology.2020.09.031
Fengyao Shi 1 , Wenhui Li 1 , Hongyu Zhao 1 , Yijing He 1 , Yao Jiang 1 , Jun Ni 1 , Benazir Abbasi 1 , Rong Rui 1 , Shiqiang Ju 1
Affiliation  

Microcystin-LR (MC-LR), as a well-known hepatotoxin, was recently found to accumulate in gonads and induce a variety of reproductive damages in zebrafish, mice and other model organisms, however, little information is available on whether MC-LR has toxic effects on the mammalian oocytes, especially in livestock species. In this study, the effects of MC-LR on meiotic maturation of porcine oocytes were investigated, and the potential mechanism of MC-LR toxicity was explored. Germinal vesicle (GV)-stage oocytes were exposed to 0, 20, 40 and 60 μM MC-LR, respectively, during the in vitro maturation for 44 h, and the results showed that the first polar body (PbI) extrusion rate of the oocytes decreased significantly when the MC-LR concentration reached 40 (P < 0.01) or 60 μM (P < 0.001). After treated with 60 μM MC-LR for 44 h, a significant higher percentage of the oocytes arrested at anaphase-telophase I (ATI) stage (P < 0.01). Laser scanning confocal results further confirmed that a significantly larger proportion of the 60 μM MC-LR-treated oocytes exhibited aberrant spindles and misaligned chromosomes, suggesting a failure of spindle assembly and homologous chromosome segregation during the ATI stage. Furthermore, the ROS levels in the 60 μM MC-LR-exposed oocytes were significantly higher than the control group (P < 0.01), while the expression of antioxidant related genes (SOD1, CAT and GPX) were much lower compared with control group, indicating that oxidative stress was induced and the antioxidant capacity of oocytes was depleted by 60 μM MC-LR treatment. Additionally, markedly decreased mitochondrial membrane potential (MMP) (P < 0.01) and significantly higher incidence of early apoptosis (P < 0.01) were observed in the 60 μM MC-LR-treated oocytes, suggesting that MC-LR exposure induced apoptosis in porcine oocytes. Moreover, the protein expression of PP2A was remarkably inhibited, whereas the expression of p53, BAX, Caspase3 and Cleaved-caspase3 were prominently increased in the 60 μM MC-LR-exposed oocytes. Together, these results suggested that 60 μM of MC-LR exposure can induce oxidative stress, and lead to aberrant spindles, impaired MMP, and trigger apoptosis, which eventually result in failure of porcine oocyte maturation.

中文翻译:

微囊藻毒素-LR 暴露导致异常纺锤体并诱导猪卵母细胞凋亡

微囊藻毒素-LR (MC-LR) 作为一种众所周知的肝毒素,最近被发现在性腺中积累并在斑马鱼、小鼠和其他模式生物中引起多种生殖损伤,但是关于 MC-LR 是否存在的信息很少对哺乳动物卵母细胞有毒性作用,尤其是对家畜物种。本研究研究了MC-LR对猪卵母细胞减数分裂成熟的影响,并探讨了MC-LR毒性的潜在机制。生殖囊泡 (GV) 期卵母细胞分别暴露于 0、20、40 和 60 μM MC-LR,在体外成熟 44 小时,结果表明,第一极体 (PbI) 挤出率当 MC-LR 浓度达到 40 (P < 0.01) 或 60 μM (P < 0.001) 时,卵母细胞显着减少。用 60 μM MC-LR 处理 44 h 后,在后期-末期 I (ATI) 阶段停滞的卵母细胞百分比显着更高 (P < 0.01)。激光扫描共聚焦结果进一步证实,60 μM MC-LR 处理的卵母细胞中显着更大比例的纺锤体和染色体错位,表明在 ATI 阶段纺锤体组装和同源染色体分离失败。此外,60 μM MC-LR 暴露的卵母细胞中 ROS 水平显着高于对照组(P < 0.01),而抗氧化相关基因(SOD1、CAT 和 GPX)的表达量远低于对照组,表明氧化应激被诱导,卵母细胞的抗氧化能力被 60 μM MC-LR 处理耗尽。此外,线粒体膜电位(MMP)显着降低(P < 0. 01) 和显着更高的早期凋亡发生率 (P < 0.01) 在 60 μM MC-LR 处理的卵母细胞中观察到,表明 MC-LR 暴露诱导猪卵母细胞凋亡。此外,PP2A 的蛋白质表达受到显着抑制,而 p53、BAX、Caspase3 和 Cleaved-caspase3 的表达在 60 μM MC-LR 暴露的卵母细胞中显着增加。总之,这些结果表明 60 μM 的 MC-LR 暴露可诱导氧化应激,并导致异常纺锤体、MMP 受损并引发细胞凋亡,最终导致猪卵母细胞成熟失败。而 p53、BAX、Caspase3 和 Cleaved-caspase3 的表达在 60 μM MC-LR 暴露的卵母细胞中显着增加。总之,这些结果表明 60 μM 的 MC-LR 暴露可诱导氧化应激,并导致异常纺锤体、MMP 受损并引发细胞凋亡,最终导致猪卵母细胞成熟失败。而 p53、BAX、Caspase3 和 Cleaved-caspase3 的表达在 60 μM MC-LR 暴露的卵母细胞中显着增加。总之,这些结果表明 60 μM 的 MC-LR 暴露可诱导氧化应激,并导致异常纺锤体、MMP 受损并引发细胞凋亡,最终导致猪卵母细胞成熟失败。
更新日期:2020-12-01
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