College of Resources and Environment, Hunan Agricultural University, Changsha, 410128, PR China.
Department of Obstetrics and Gynecology, Xiangya Third Hospital, Central South University, Changsha City, Hunan Province, 410013, PR China.
Toxicology. 2021 Jun 15;457:152805. doi: 10.1016/j.tox.2021.152805. Epub 2021 May 4.
Niclosamide (NIC), a helminthic drug used widely for controlling schistosomiasis, can reportedly disrupt the endocrine system. However, its underlying mechanisms are still unclear. In this study, we revealed the potential endocrine disruption mechanism of NIC by activating estrogen receptors (ERs) and estrogen-related receptors (ERRs). The binding potency of NIC with ERα, ERβ and ERRγ were determined by fluorescence competitive binding assays, which shows an IC (the concentration of NIC needed to displace 50 % of the probe from the receptor) of 90 ± 4.1, 10 ± 1.7 nM and 0.59 ± 0.07 nM respectively. The IC for ERRγ is the lowest one among the three detected receptors, which is three orders of magnitude lower than the known agonist GSK4716.The transcriptional activities of NIC on ERs and ERRs were detected by MVLN cells (stably transfected with ERs reporter gene) and HeLa cells (transiently transfected with ERRs reporter gene)-based luciferase reporter gene assay. The lowest observable effective concentration (LOEC) ranked as follows: ERRγ (0.5 nM) < ERRα (10 nM) < ERs (100 nM). The maximum observed induction rate for ERRγ (294 %) was higher than that for ERRα (191 %). The maximum observed induction rate of NIC for ERs was 30 % relative to 17β-estradiol. In addition, we simulated the interactions of NIC with ERs and ERRs by molecular docking. NIC could dock into the ligand binding pockets of ERs and ERRs and form hydrogen bonds with different amino acids. The binding energy ranked as follows: ERRγ (-8.90 kcal/mol) < ERβ (-7.57 kcal/mol) < ERRα (-7.15 kcal/mol) < ERα (-6.53 kcal/mol), which implied that NIC bound to ERRγ with higher binding affinity than the other receptors. Overall, we clarify that ERRγ might be the dominant target for NIC in cells rather than ERRα and ERs. We reveal potential novel mechanisms for the endocrine disruption effects of NIC by activating both ERRs and ERs at environmentally-related nanomolar levels.
硝氯酚（NIC）是一种广泛用于控制血吸虫病的驱虫药物，据报道它可以破坏内分泌系统。然而，其潜在的作用机制尚不清楚。在这项研究中，我们通过激活雌激素受体（ERs）和雌激素相关受体（ERRs），揭示了 NIC 的潜在内分泌干扰机制。通过荧光竞争结合测定法测定了 NIC 与 ERα、ERβ 和 ERRγ 的结合效力，结果显示其对 ERα、ERβ 和 ERRγ 的半数抑制浓度（IC）分别为 90±4.1、10±1.7 和 0.59±0.07 nM。在检测到的三种受体中，NIC 与 ERRγ 的 IC 最低，比已知激动剂 GSK4716 低三个数量级。通过基于 MVLN 细胞（稳定转染 ER 报告基因）和 HeLa 细胞（瞬时转染 ERR 报告基因）的荧光素酶报告基因检测法，检测了 NIC 对 ERs 和 ERRs 的转录活性。最低可观察有效浓度（LOEC）的排序如下：ERRγ（0.5 nM）<ERRα（10 nM）<ERs（100 nM）。ERRγ 的最大观察诱导率（294%）高于 ERRα（191%）。与 17β-雌二醇相比，NIC 对 ERs 的最大观察诱导率为 30%。此外，我们通过分子对接模拟了 NIC 与 ERs 和 ERRs 的相互作用。NIC 可以与 ERs 和 ERRs 的配体结合口袋结合，并与不同的氨基酸形成氢键。结合能的排序如下：ERRγ（-8.90 kcal/mol）<ERβ（-7.57 kcal/mol）<ERRα（-7.15 kcal/mol）<ERα（-6.53 kcal/mol），这表明 NIC 与 ERRγ 的结合亲和力高于其他受体。总的来说，我们阐明了在细胞中，ERRγ 可能是 NIC 的主要靶标，而不是 ERRα 和 ERs。我们揭示了 NIC 在环境相关的纳摩尔水平上通过激活 ERRs 和 ERs 发挥内分泌干扰作用的潜在新机制。
