Yao Mengmeng, Ji Xiaoli, Zhang Yuqing, Mao Zhilei, Chi Xia
Institute of Toxicology, School of Public Health, Nanjing Medical University, Nanjing, China.
Department of Child Health Care, Women's Hospital of Nanjing Medical University, Nanjing Maternity and Child Health Care Hospital, Nanjing, China.
Ann Transl Med. 2022 Apr;10(7):402. doi: 10.21037/atm-22-508.
Cerium dioxide nanoparticles (CeO NPs) are increasingly used as diesel additive, causing a lot of concern about their toxic effects when released into the atmosphere. To date, there is little knowledge about the toxic effects of CeO NPs on the female reproductive system.
The morphology and size distribution of CeO NPs was observed by transmission electronic microscope (TEM) and Zetasizer Nano, respectively. The uptake of CeO NPs by cells was also observed by TEM after treatment. The cytotoxicity of CeO NPs was studied by Cell Counting Kit-8 (CCK-8), the cellular invasive and migratory ability was examined by transwell assay, the cell apoptosis and reactive oxygen species (ROS) were studied by flow cytometry (FCM), and the mRNAs and proteins expressions were revealed by quantitative real-time PCR (qRT-PCR) and western blotting. The cytoskeletons and autophagy levels were revealed by immunofluorescence. The target regulation of miR-99 to mammalian target of rapamycin (mTOR) was proved by dual luciferase reporter assay after transfection.
We studied the cytotoxic effects of CeO NPs on human trophoblastic cells (HTR-8/Svneo) and found that the invasive and migratory abilities of HTR-8/SVneo cells were decreased after CeO NPs exposure. Immunofluorescence assays showed that the cellular microtubule networks and microfilament arrangement were obviously altered, and although the expression of cytoskeleton proteins (α-tubulin, β-tubulin, actin) did not change, the protein levels of invasion- and migration-related factors [matrix metalloproteinase 2 (MMP2), protein kinases B (AKT), mTOR] were decreased in exposed cells. Accordingly, the expression level of miR-99 family members (miR-99a, miR-99b, miR-100), which can regulate mTOR, was significantly increased after CeO NPs exposure. Dual luciferase reporter assay indicated that the miR-99 family members directly targeted mTOR.
CeO NPs impaired the invasive and migratory abilities, which play an important role in embryo implantation, as well as determining placental function and embryonic development.
二氧化铈纳米颗粒（CeO NPs）越来越多地用作柴油添加剂，当其释放到大气中时，引发了人们对其毒性作用的诸多担忧。迄今为止，关于CeO NPs对女性生殖系统的毒性作用知之甚少。
分别通过透射电子显微镜（TEM）和纳米粒度分析仪观察CeO NPs的形态和大小分布。处理后，还通过TEM观察细胞对CeO NPs的摄取情况。采用细胞计数试剂盒-8（CCK-8）研究CeO NPs的细胞毒性，通过Transwell实验检测细胞侵袭和迁移能力，采用流式细胞术（FCM）研究细胞凋亡和活性氧（ROS），通过定量实时聚合酶链反应（qRT-PCR）和蛋白质印迹法揭示mRNA和蛋白质表达。通过免疫荧光揭示细胞骨架和自噬水平。转染后通过双荧光素酶报告基因检测证实miR-99对雷帕霉素哺乳动物靶蛋白（mTOR）的靶向调控。
我们研究了CeO NPs对人滋养层细胞（HTR-8/Svneo）的细胞毒性作用，发现暴露于CeO NPs后，HTR-8/SVneo细胞的侵袭和迁移能力降低。免疫荧光分析表明，细胞微管网络和微丝排列明显改变，尽管细胞骨架蛋白（α-微管蛋白、β-微管蛋白、肌动蛋白）的表达没有变化，但暴露细胞中侵袭和迁移相关因子[基质金属蛋白酶2（MMP2）、蛋白激酶B（AKT）、mTOR]的蛋白水平降低。因此，暴露于CeO NPs后，可调节mTOR的miR-99家族成员（miR-99a、miR-99b、miR-100）的表达水平显著增加。双荧光素酶报告基因检测表明，miR-99家族成员直接靶向mTOR。
CeO NPs损害了在胚胎着床、决定胎盘功能和胚胎发育中起重要作用的侵袭和迁移能力。
