Abstract：
Exposure to persistent organic pollutants (POPs), such as dichlorodiphenyltrichloroethane (DDT) and polychlorinated biphenyls (PCBs), has historically been linked to population collapses in wildlife. Despite international regulations, these legacy chemicals are still currently detected in women of reproductive age, and their levels correlate with reduced ovarian reserve, longer time-to-pregnancy, and higher risk of infertility. However, the specific modes of action underlying these associations remain unclear. Here, we examined the effects of five commonly occurring POPs - hexachlorobenzene (HCB), p,p\'-dichlorodiphenyldichloroethylene (DDE), 2,3,3\',4,4\',5-hexachlorobiphenyl (PCB156), 2,2\',3,4,4\',5,5\'-heptachlorobiphenyl (PCB180), perfluorooctane sulfonate (PFOS) - and their mixture on human ovaries in vitro. We exposed human ovarian cancer cell lines COV434, KGN, and PA1 as well as primary ovarian cells for 24 h, and ovarian tissue containing unilaminar follicles for 6 days. RNA-sequencing of samples exposed to concentrations covering epidemiologically relevant levels revealed significant gene expression changes related to central energy metabolism in the exposed cells, indicating glycolysis, oxidative phosphorylation, fatty acid metabolism, and reactive oxygen species as potential shared targets of POP exposures in ovarian cells. Alpha-enolase (ENO1), lactate dehydrogenase A (LDHA), cytochrome C oxidase subunit 4I1 (COX4I1), ATP synthase F1 subunit alpha (ATP5A), and glutathione peroxidase 4 (GPX4) were validated as targets through qPCR in additional cell culture experiments in KGN. In ovarian tissue cultures, we observed significant effects of exposure on follicle growth and atresia as well as protein expression. All POP exposures, except PCB180, decreased unilaminar follicle proportion and increased follicle atresia. Immunostaining confirmed altered expression of LDHA, ATP5A, and GPX4 in the exposed tissues. Moreover, POP exposures modified ATP production in KGN and tissue culture. In conclusion, our results demonstrate the disruption of cellular energy metabolism as a novel mode of action underlying POP-mediated interference of follicle growth in human ovaries.
摘要：
接触持久性有机污染物(POPs)，如二氯二苯基三氯乙烷（DDT）和多氯联苯（PCB），历史上一直与野生动物种群崩溃有关。尽管有国际法规,这些遗留化学物质目前仍在育龄妇女中检测到，它们的水平与卵巢储备减少有关，怀孕时间更长，和更高的不孕风险。然而,这些关联的具体作用方式尚不清楚.这里,我们研究了五种常见的持久性有机污染物-六氯苯(HCB)的影响，p,对二氯二苯基二氯乙烯(DDE),2,3,3\'，4,4\'，5-六氯联苯(PCB156),2,2\'，3,4,4'，5,5'-七氯联苯(PCB180),全氟辛烷磺酸（PFOS）-及其混合物在体外人卵巢中的作用。我们暴露了人卵巢癌细胞系COV434，KGN，和PA1以及原代卵巢细胞24小时，和含有单层卵泡的卵巢组织持续6天。暴露于涵盖流行病学相关水平的浓度的样品的RNA测序显示，与暴露细胞中中枢能量代谢相关的显著基因表达变化。表明糖酵解，氧化磷酸化，脂肪酸代谢，和活性氧作为卵巢细胞中POP暴露的潜在共享目标。α-烯醇化酶(ENO1),乳酸脱氢酶A（LDHA），细胞色素C氧化酶亚基4I1（COX4I1），ATP合酶F1亚基α（ATP5A），在KGN的其他细胞培养实验中，通过qPCR验证了谷胱甘肽过氧化物酶4（GPX4）作为靶标。在卵巢组织培养中,我们观察到暴露对卵泡生长和闭锁以及蛋白质表达的显著影响。所有POP暴露,除PCB180外，单层卵泡比例降低，卵泡闭锁增加。免疫染色证实LDHA表达改变，ATP5A,和GPX4在暴露的组织中。此外，POP暴露改变了KGN和组织培养中的ATP产生。总之,我们的结果表明,细胞能量代谢的破坏是POP介导的人类卵巢卵泡生长干扰的一种新的作用方式.
