Rossier Nadine Madeleine, Chew Geraldine, Zhang Kun, Riva Francesco, Fent Karl
University of Applied Sciences and Arts Northwestern Switzerland, School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz, Switzerland; University of Basel, Division of Molecular and Systems Toxicology, Department of Pharmaceutical Sciences, Klingelbergstrasse 50, 4056 Basel, Switzerland.
University of Applied Sciences and Arts Northwestern Switzerland, School of Life Sciences, Gründenstrasse 40, CH-4132 Muttenz, Switzerland.
Aquat Toxicol. 2016 May;174:109-22. doi: 10.1016/j.aquatox.2016.02.005. Epub 2016 Feb 22.
Despite potential exposure of aquatic organisms to mixtures of steroid hormones, very little is known on their joint activity in fish. Drospirenone (DRS) is a new synthetic progestin used in contraceptive pills in combination with 17α-ethinylestradiol (EE2). Here we systematically analyzed effects of DRS in binary mixtures with progesterone (P4) and EE2. First, we determined the in vitro activity of single compounds in recombinant yeast assays that express the human progesterone, androgen, or estrogen receptor, followed by determination of mixture activities of DRS and P4, DRS and EE2, as well as medroxyprogesterone acetate (MPA) and dydrogesterone (DDG). Mixtures of DRS and P4, as well as of DRS and EE2 showed additive progestogenic and androgenic activities. However, DDG and MPA showed non-additive progestogenic and androgenic activities. We then analyzed the in vivo activity of single compounds and mixtures of DRS and P4, as well as DRS and EE2, by assessing transcriptional changes of up to 14 selected target genes in zebrafish embryos at 48h post fertilization (hpf), and in eleuthero-embryos at 96hpf and 144hpf. DRS, P4, and EE2 led to significant transcriptional alteration of genes, including those encoding hormone receptors (pgr, esr1), a steroidogenic enzyme (hsd17b3), and estrogenic markers (vtg1, cyp19b), in particular at 144 hpf. In general, DRS showed stronger transcriptional changes than P4. In mixtures of DRS and P4, they were mainly non-additive (antagonistic interaction). In mixtures of DRS and EE2, transcriptional responses of esr1, vtg1 and cyp19b were dominated by EE2, suggesting an antagonistic interaction or independent action. Equi-effective mixtures of DRS and EE2, based on progesterone receptor transcripts, showed antagonistic interactions. Our data suggest that interactions in mixtures assessed in vitro in recombinant yeast cannot be translated to the in vivo situation. The receptor-based responses did not correspond well to the transcriptional responses in embryos which are much more complex due to the interplay between hormonal pathways, receptor crosstalk, and hormonal feedback loops.
尽管水生生物可能会接触到类固醇激素混合物，但我们对它们在鱼类中的联合活性却知之甚少。屈螺酮（DRS）是一种新型合成孕激素，与17α-乙炔雌二醇（EE2）联合用于避孕药中。在此，我们系统地分析了DRS与孕酮（P4）和EE2的二元混合物的作用。首先，我们在表达人孕酮、雄激素或雌激素受体的重组酵母试验中测定了单一化合物的体外活性，随后测定了DRS与P4、DRS与EE2以及醋酸甲羟孕酮（MPA）和地屈孕酮（DDG）的混合物活性。DRS与P4以及DRS与EE2的混合物表现出相加的孕激素和雄激素活性。然而，DDG和MPA表现出非相加的孕激素和雄激素活性。然后，我们通过评估受精后48小时（hpf）斑马鱼胚胎以及96hpf和144hpf的孵化后胚胎中多达14个选定靶基因的转录变化，分析了单一化合物以及DRS与P4、DRS与EE2混合物的体内活性。DRS、P4和EE2导致基因发生显著的转录改变，包括那些编码激素受体（pgr、esr1）、一种类固醇生成酶（hsd17b3）和雌激素标志物（vtg1、cyp19b）的基因，特别是在144 hpf时。总体而言，DRS表现出比P4更强的转录变化。在DRS与P4的混合物中，它们主要是非相加的（拮抗相互作用）。在DRS与EE2的混合物中，esr1、vtg1和cyp19b的转录反应主要由EE2主导，表明存在拮抗相互作用或独立作用。基于孕酮受体转录本的DRS与EE2等效混合物表现出拮抗相互作用。我们的数据表明，在重组酵母中体外评估的混合物中的相互作用不能直接应用于体内情况。基于受体的反应与胚胎中的转录反应不太相符，由于激素途径、受体串扰和激素反馈回路之间的相互作用，胚胎中的转录反应要复杂得多。
