RESUMEN
This study exposed adult Sydney rock oysters, of either sex or both, to the synthetic estrogen 17α-ethinylestradiol (EE2) at 50 ng/L for 21 days, followed by an examination of developmental endpoints and transcriptomic responses in unexposed larvae. Reduced survival was observed at 1 day post-fertilisation (dpf) in larvae from bi-parental exposure (FTMT). Motile larvae at 2 dpf were fewer from maternal (FTMC), paternal (FCMT), and FTMT exposures. Additionally, shell length at 7 dpf decreased in larvae from FTMC and FTMT parents. RNA sequencing (RNA-seq) revealed 1064 differentially expressed genes (DEGs) in 1-dpf larvae from FTMT parents, while fewer DEGs were detected in larvae from FTMC and FCMT parents, with 258 and 7, respectively. GO and KEGG analyses showed significant enrichment of DEGs in diverse terms and pathways, with limited overlap among treatment groups. IPA results indicated potential inhibition of pathways regulating energy production, larval development, transcription, and detoxification of reactive oxygen species in FTMT larvae. qRT-PCR validation confirmed significant downregulation of selected DEGs involved in these pathways and relevant biological processes, as identified in the RNA-seq dataset. Overall, our results suggest that the intergenerational toxicity of EE2 is primarily maternally transmitted, with bi-parental exposure amplifying these effects.
Asunto(s)
Etinilestradiol , Larva , Ostreidae , Transcriptoma , Contaminantes Químicos del Agua , Animales , Etinilestradiol/toxicidad , Larva/efectos de los fármacos , Larva/crecimiento & desarrollo , Transcriptoma/efectos de los fármacos , Ostreidae/efectos de los fármacos , Ostreidae/crecimiento & desarrollo , Ostreidae/genética , Femenino , Contaminantes Químicos del Agua/toxicidad , Masculino , Exposición Materna , Exposición Paterna/efectos adversosRESUMEN
OBJECTIVE: To characterise and compare the toxicity of estetrol (E4) and 17α-ethinylestradiol (EE2), and their respective mixture with the progestin drospirenone (DRSP) in zebrafish (Danio rerio) embryos. METHODS: Zebrafish embryos were exposed to E4, EE2, DRSP, E4+DRSP, and EE2+DRSP in a fish embryo acute toxicity (FET) test. A second test examined behavioural responses and, using label-free proteomics, identified changes in protein expression in response to hormonal treatments, across a range of concentrations, including those that are considered to be environmentally relevant. RESULTS: In the FET test, no effects were found from E4 at concentrations ≤100 mg/L, while EE2 induced mortality and morphological abnormalities at concentrations of 1-2 mg/L. In the behavioural test, exposure to 30 ng/L EE2 (â¼200 × predicted environmental concentration - PEC) resulted in hypoactivity in fish larvae and exposure to 0.3 ng/L EE2 (â¼2 × PEC) led to quantitative changes in protein abundance, revealing potential impacts on RNA processing and protein synthesis machinery. Exposure to E4 did not alter behaviour, but several groups of proteins were modulated, mainly at 710 ng/L (â¼200 × PEC), including proteins involved in oxidative phosphorylation. When combined with DRSP, EE2 induced reduced effects on behaviour and proteomic responses, suggesting an antagonistic effect of DRSP. E4+DRSP induced no significant effects on behaviour or proteomic profiles at tested concentrations. CONCLUSIONS: These findings suggest that E4-based combined oral contraceptives present a more favourable environmental profile than EE2-based contraceptives, particularly during the early developmental stages of fish.
Asunto(s)
Androstenos , Conducta Animal , Etinilestradiol , Larva , Proteómica , Contaminantes Químicos del Agua , Pez Cebra , Animales , Etinilestradiol/toxicidad , Contaminantes Químicos del Agua/toxicidad , Androstenos/toxicidad , Conducta Animal/efectos de los fármacos , Larva/efectos de los fármacos , Embrión no Mamífero/efectos de los fármacosRESUMEN
Numerous pharmaceutical and industrial chemicals are classified as endocrine-disrupting chemicals (EDCs) that interfere with hormonal homeostasis, leading to developmental disorders and other pathologies. The synthetic estrogen 17α-ethynylestradiol (EE2) is used in oral contraceptives and other hormone therapies. EE2 and other estrogens are inadvertently introduced into aquatic environments through municipal wastewater and agricultural effluents. Exposure of male fish to estrogens increases expression of the egg yolk precursor protein vitellogenin (Vtg), which is used as a molecular marker of exposure to estrogenic EDCs. The mechanisms behind Vtg induction are not fully known, and we hypothesized that it is regulated via DNA methylation. Adult zebrafish were exposed to either dimethyl sulfoxide or 20 ng/L EE2 for 14 days. Messenger RNA (mRNA) expression and DNA methylation were assessed in male zebrafish livers at 0, 0.25, 0.5, 1, 4, 7, and 14 days of exposure; and those of females were assessed at 13 days (n ≥ 4/group/time point). To test the persistence of any changes, we included a recovery group that received EE2 for 7 days and did not receive any for the following 7 days, in the total 14-day study. Methylation of DNA at the vtg1 promoter was assessed with targeted gene bisulfite sequencing in livers of adult male and female zebrafish. A significant increase in vtg1 mRNA was observed in the EE2-exposed male fish as early as 6 h. Interestingly, DNA methylation changes were observed at 4 days. Decreases in the overall methylation of the vtg1 promoter in exposed males resulted in levels comparable to those in female controls, suggesting feminization. Importantly, DNA methylation levels in males remained significantly impacted after 7 days post-EE2 removal, unlike mRNA levels. These data identify an epigenetic mark of feminization that may serve as an indicator of not only estrogenic exposure but also previous exposure to EE2. Environ Toxicol Chem 2024;43:1547-1556. © 2024 SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.
Asunto(s)
Islas de CpG , Metilación de ADN , Etinilestradiol , Regiones Promotoras Genéticas , Vitelogeninas , Contaminantes Químicos del Agua , Pez Cebra , Animales , Pez Cebra/genética , Masculino , Etinilestradiol/toxicidad , Metilación de ADN/efectos de los fármacos , Vitelogeninas/genética , Vitelogeninas/metabolismo , Femenino , Contaminantes Químicos del Agua/toxicidad , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismo , Disruptores Endocrinos/toxicidadRESUMEN
Combined oral contraceptives, comprising of both an oestrogen and a progestin component, are released in aquatic environments and potentially pose a risk to aquatic wildlife by their capacity to disrupt physiological mechanisms. In this study, the endocrine disruptive potential of two mixtures, 17α-ethinylestradiol (EE2), a synthetic oestrogen, or estetrol (E4), a natural oestrogen, with the progestin drospirenone (DRSP) have been characterised in three generations of zebrafish, according to an adapted Medaka Extended One Generation Reproduction Test. Zebrafish (Danio rerio) were exposed to a range of concentrations of EE2/DRSP and E4/DRSP (â¼1×, â¼3×, â¼10× and â¼30× predicted environmental concentration, PEC). Survival, growth, hatching success, fecundity, fertilisation success, vitellogenin (VTG), gonad histopathology, sex differentiation, and transcriptional analysis of genes related to gonadal sex steroid hormones synthesis were assessed. In the F0 generation, exposure to EE2/DRSP at â¼10 and â¼30× PEC decreased fecundity and increased male VTG concentrations. The highest concentration of EE2/DRSP also affected VTG concentrations in female zebrafish and the expression of genes implicated in steroid hormones synthesis. In the F1 generation, sex determination was impaired in fish exposed to EE2/DRSP at concentrations as low as â¼3× PEC. Decreased fecundity and fertility, and abnormal gonadal histopathology were also observed. No effects were observed in the F2 generation. In contrast, E4/DRSP induced only minor histopathological changes and an increase in the proportion of males, at the highest concentration tested (â¼30× PEC) in the F1 generation and had no effect on hatching success of F2 generation. Overall, this study suggests that the combination E4/DRSP has a more favourable environmental profile than EE2/DRSP.
Asunto(s)
Androstenos , Disruptores Endocrinos , Etinilestradiol , Pez Cebra , Animales , Pez Cebra/fisiología , Etinilestradiol/toxicidad , Androstenos/toxicidad , Disruptores Endocrinos/toxicidad , Femenino , Masculino , Contaminantes Químicos del Agua/toxicidad , Vitelogeninas/metabolismo , Reproducción/efectos de los fármacosRESUMEN
In avian embryos, xenoestrogens induce abnormalities in reproductive organs, particularly the testes and Müllerian ducts (MDs). However, the molecular mechanisms remain poorly understood. We investigated the effects of ethynylestradiol (EE2) exposure on gene expression associated with reproductive organ development in Japanese quail embryos. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis revealed that the left testis containing ovary-like tissues following EE2 exposure highly expressed the genes for steroidogenic enzymes (P450scc, P45017α, lyase, and 3ß-HSD) and estrogen receptor-ß, compared to the right testis. No asymmetry was found in these gene expression without EE2. EE2 induced hypertrophy in female MDs and suppressed atrophy in male MDs on both sides. RNA sequencing analysis of female MDs showed 1,366 differentially expressed genes between developing left MD and atrophied right MD in the absence of EE2, and these genes were enriched in Gene Ontology terms related to organogenesis, including cell proliferation, migration and differentiation, and angiogenesis. However, EE2 reduced asymmetrically expressed genes to 21. RT-qPCR analysis indicated that genes promoting cell cycle progression and oncogenesis were more highly expressed in the left MD than in the right MD, but EE2 eliminated such asymmetric gene expression by increasing levels on the right side. EE2-exposed males showed overexpression of these genes in both MDs. This study reveals part of the molecular basis of xenoestrogen-induced abnormalities in avian reproductive organs, where EE2 may partly feminize gene expression in the left testis, developing as the ovotestis, and induce bilateral MD malformation by canceling asymmetric gene expression underlying MD development.
Asunto(s)
Coturnix , Etinilestradiol , Regulación del Desarrollo de la Expresión Génica , Conductos Paramesonéfricos , Testículo , Animales , Masculino , Testículo/efectos de los fármacos , Testículo/metabolismo , Testículo/embriología , Testículo/patología , Coturnix/embriología , Coturnix/genética , Etinilestradiol/toxicidad , Conductos Paramesonéfricos/efectos de los fármacos , Conductos Paramesonéfricos/embriología , Conductos Paramesonéfricos/anomalías , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Embrión no Mamífero/efectos de los fármacos , Feminización/inducido químicamente , Feminización/genéticaRESUMEN
Tetra-amido macrocyclic ligands (TAMLs) are catalysts designed to mimic endogenous peroxidases that can degrade pollutants. Before TAMLs gain widespread use, it is first important to determine if they have endocrine disrupting properties. In this study, we evaluated the effects of the iron TAML, NT7, on hormone-sensitive outcomes in mice exposed during pregnancy and lactation, and on their litters prior to weaning. We administered NT7 at one of three doses to mice via drinking water prior to and then throughout pregnancy and lactation. Two hormonally active pharmaceuticals, ethinyl estradiol (EE2) and flutamide (FLUT), a known estrogen receptor agonist and androgen receptor antagonist, respectively, were also included. In the females, we measured pre- and post-parturition weight, length of pregnancy, organ weights at necropsy, and morphology of the mammary gland at the end of the lactational period. We also quantified maternal behaviors at three stages of lactation. For the offspring, we measured litter size, litter weights, and the achievement of other developmental milestones. We observed only one statistically significant effect of NT7, a decrease in the percentage of pups with ear opening at postnatal day 5. This contrasts with the numerous effects of EE2 on both the mother and the litter, as well as several modest effects of FLUT. The approach taken in this study could provide guidance for future studies that aim to evaluate novel compounds for endocrine disrupting properties.
Asunto(s)
Estrógenos , Lactancia , Embarazo , Femenino , Animales , Ratones , Estrógenos/farmacología , Flutamida , Tamaño de la Camada , Etinilestradiol/toxicidadRESUMEN
The interactions between estrogen and androgen in aquatic animals remain largely unknown. In this study, two generations (F0 and F1) of western mosquitofish (Gambusia affinis) were continuously exposed to 17α-ethinylestradiol (EE2, 10 ng/L), methyltestosterone (MT, 10 ng/L (MTL); 50 ng/L (MTH)), and mixtures (EE2+MTL and EE2+MTH). Various endpoints, including sex ratio (phenotypic and genetic), secondary sex characteristics, gonadal histology, and transcriptional profile of genes, were examined. The results showed that G. affinis exposed to MTH and EE2+MTH had a > 89.7 % of phenotypic males in F1 generation, with 34.5 and 50.0 % of these males originated from genetic females, respectively. Moreover, females from F0 and F1 generations exposed to MTH and EE2+MTH exhibited masculinized anal fins and skeletons. The combined effect of MT and EE2 on most endpoints was dependent on MT. Furthermore, significant transcriptional alterations in certain target genes were observed in both the F0 and F1 generations by EE2 and MT alone and by mixtures, showing some degree of interactions. These findings that the effects of EE2+MTH were primarily on the phenotypic sex of G. affinis in offspring generation suggest that G. affinis under chronic exposure to the binary mixture contaminated water could have sex-biased populations.
Asunto(s)
Ciprinodontiformes , Contaminantes Químicos del Agua , Masculino , Femenino , Animales , Etinilestradiol/toxicidad , Metiltestosterona/toxicidad , Contaminantes Químicos del Agua/toxicidad , Estrógenos , Ciprinodontiformes/genéticaRESUMEN
Estrogen-induced intrahepatic cholestasis (IHC) is a mild but potentially serious risk and urges for new therapeutic targets and effective treatment. Our previous study demonstrated that RORγt and CXCR3 signaling pathway of invariant natural killer T (iNKT) 17 cells play pathogenic roles in 17α-ethinylestradiol (EE)-induced IHC. Ursodeoxycholic acid (UDCA) and 18ß-glycyrrhetinic acid (GA) present a protective effect on IHC partially due to their immunomodulatory properties. Hence in present study, we aim to investigate the effectiveness of UDCA and 18ß-GA in vitro and verify the accessibility of the above targets. Biochemical index measurement indicated that UDCA and 18ß-GA presented efficacy to alleviate EE-induced cholestatic cytotoxicity. Both UDCA and 18ß-GA exhibited suppression on the CXCL9/10-CXCR3 axis, and significantly restrained the expression of RORγt in vitro. In conclusion, our observations provide new therapeutic targets of UDCA and 18ß-GA, and 18ß-GA as an alternative treatment for EE-induced cholestasis.
Asunto(s)
Colestasis , Ácido Glicirretínico , Células T Asesinas Naturales , Receptores CXCR3 , Ácido Ursodesoxicólico , Colestasis/inducido químicamente , Colestasis/tratamiento farmacológico , Etinilestradiol/toxicidad , Ácido Glicirretínico/análogos & derivados , Ácido Glicirretínico/farmacología , Ácido Glicirretínico/uso terapéutico , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Receptores CXCR3/genética , Receptores CXCR3/metabolismo , Transducción de Señal , Ácido Ursodesoxicólico/farmacología , Ácido Ursodesoxicólico/uso terapéutico , Animales , RatonesRESUMEN
Endocrine disruptors, such as estrogen, are chemical substances with the potential to alter the hormonal balance of organisms. Their origin can be natural or artificial, and they can act at very low doses. The estrogen 17α-ethinylestradiol (EE2) is used worldwide as an oral contraceptive and is a potential contaminant in aquatic ecosystems. It is well documented that these environmental pollutants can act directly or indirectly on the reproductive system, impairing development and fertility. However, little is known about the alteration of the cell oxidative status induced by EE2. The main objective of this study was to evaluate the effect on the gill cells of adult zebrafish exposed in vivo to EE2, analyzing cell histology, DNA damage and the expression levels of genes encoding the main enzymes involved in oxidative stress pathways. The histological study showed that EE2 produces moderate to high damage to the gill tissue, an increase in gill cell DNA damage and the mRNA levels of the genes corresponding to the manganese superoxide dismutase (Mn-sod) and catalase (cat) after exposure to 5 ng/L EE2. The results indicate that EE2 causes tissue alterations, DNA damage and oxidative stress. EE2 produced important alterations in the gills, a fundamental organ for the survival of fish. There is a clear need for further research on the ecological consequences of EDCs on non-target organisms.
Asunto(s)
Contaminantes Químicos del Agua , Pez Cebra , Animales , Pez Cebra/genética , Antioxidantes/farmacología , Branquias , Ecosistema , Etinilestradiol/toxicidad , Estrógenos/farmacología , Daño del ADN , Contaminantes Químicos del Agua/toxicidadRESUMEN
Estrogenic chemicals are common pollutants in wastewater and current effluent treatment processes are not typically effective in removing these compounds. Tetra-amido macrocyclic ligands (TAMLs) are catalysts that mimic endogenous peroxidases that may provide a solution to remove environmental pollutants including low concentrations of estrogenic compounds. Yet relatively little is known about the toxicity of TAMLs, and few studies have evaluated whether they may have endocrine disrupting properties. We administered one of three doses of a TAML, NT7, to mice via drinking water throughout pregnancy and lactation. Two pharmacologically active compounds, ethinyl estradiol (EE2) and flutamide were also included to give comparator data for estrogen receptor agonist and androgen receptor antagonist activities. Male pups were evaluated for several outcomes at weaning, puberty, and early adulthood. We found that EE2 exposures during gestation and the perinatal period induced numerous effects that were observed across the three ages including changes to spleen and testis weight and drastic effects on the morphology of the mammary gland. Flutamide had fewer effects but altered anogenital distance at weaning as well as spleen, liver, and kidney weight. In contrast, relatively few effects of NT7 were observed, but included alterations to spleen weight and modest changes to adult testis weight and morphology of the mammary gland at weaning. Collectively, these results provide some of the first evidence suggesting that NT7 may alter some hormone-sensitive outcomes, but that the effects were distinct from either EE2 or flutamide. Additional studies are needed to characterize the biological activity of this and other TAML catalysts.
Asunto(s)
Flutamida , Maduración Sexual , Embarazo , Femenino , Ratones , Animales , Masculino , Flutamida/toxicidad , Estrógenos/toxicidad , Etinilestradiol/toxicidad , LactanciaRESUMEN
Understanding species differences in sensitivity to toxicants is a critical issue in ecotoxicology. We recently established that double-crested cormorant (DCCO) embryos are more sensitive than Japanese quail (JQ) to the developmental effects of ethinylestradiol (EE2). We explored how this difference in sensitivity between species is reflected at a transcriptomic level. The EE2 was dissolved in dimethyl sulfoxide and injected into the air cell of eggs prior to incubation at nominal concentrations of 0, 3.33, and 33.3 µg/g egg weight. At midincubation (JQ 9 days; DCCO 16 days), livers were collected from five embryos/treatment group for RNA sequencing. Data were processed and analyzed using EcoOmicsAnalyst and ExpressAnalyst. The EE2 exposure dysregulated 238 and 1,987 genes in JQ and DCCO, respectively, with 78 genes in common between the two species. These included classic biomarkers of estrogen exposure such as vitellogenin and apovitellenin. We also report DCCO-specific dysregulation of Phase I/II enzyme-coding genes and species-specific transcriptional ontogeny of vitellogenin-2. Twelve Kyoto Encyclopedia of Genes and Genomes pathways and two EcoToxModules were dysregulated in common in both species including the peroxisome proliferator-activated receptor (PPAR) signaling pathway and fatty acid metabolism. Similar to previously reported differences at the organismal level, DCCO were more responsive to EE2 exposure than JQ at the gene expression level. Our description of differences in transcriptional responses to EE2 in early life stage birds may contribute to a better understanding of the molecular basis for species differences. Environ Toxicol Chem 2024;43:772-783. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.
Asunto(s)
Coturnix , Etinilestradiol , Animales , Etinilestradiol/toxicidad , Coturnix/genética , Vitelogeninas , Perfilación de la Expresión Génica , HígadoRESUMEN
Non-steroidal anti-inflammatory drugs (NSAIDs) and 17α-ethinylestradiol (EE2) are extensively used in human and veterinary medicine. Due to their partial removal by wastewater treatment plants, they are frequent environmental contaminants, particularly in drinking water. Here, we investigated the adverse outcomes of chronic exposure to mixtures of NSAIDs (ibuprofen, 2hydroxy-ibuprofen, diclofenac) and EE2 at two environmentally relevant doses in drinking water, on the reproductive organ development and fertility in F1-exposed male and female mice and in their F2 offspring. In male and female F1 mice, which were exposed to these mixtures, reproductive organ maturation, estrous cyclicity, and spermiogenesis were altered. These defects were observed also in F2 animals, in addition to some specific sperm parameter alterations in F2 males. Transcriptomic analysis revealed significant changes in gene expression patterns and associated pathways implicated in testis and ovarian physiology. Chronic exposure of mice to NSAID and EE2 mixtures at environmental doses intergenerationally affected male and female fertility (i.e. total number of pups and time between litters). Our study provides new insights into the adverse effects of these pharmaceuticals on the reproductive health and will facilitate the implementation of a future regulatory environmental risk assessment of NSAIDs and EE2 for human health.
Asunto(s)
Agua Potable , Contaminantes Químicos del Agua , Humanos , Masculino , Animales , Ratones , Etinilestradiol/toxicidad , Reproducción , Ibuprofeno/farmacología , Semen , Fertilidad , Antiinflamatorios no Esteroideos/toxicidad , Contaminantes Químicos del Agua/toxicidadRESUMEN
Several adverse outcome pathways (AOPs) have linked molecular initiating events like aromatase inhibition, androgen receptor (AR) agonism, and estrogen receptor (ER) antagonism to reproductive impairment in adult fish. Estrogen receptor agonists can also cause adverse reproductive effects, however, the early key events (KEs) in an AOP leading to this are mostly unknown. The primary aim of this study was to develop hypotheses regarding the potential mechanisms through which exposure to ER agonists might lead to reproductive impairment in female fish. Mature fathead minnows were exposed to 1 or 10 ng 17α-ethynylestradiol (EE2)/L or 10 or 100 µg bisphenol A (BPA)/L for 14 d. The response to EE2 and BPA was contrasted with the effects of 500 ng/L of 17ß-trenbolone (TRB), an AR agonist, as well as TRB combined with the low and high concentrations of EE2 or BPA tested individually. Exposure to 10 ng EE2/L, 100 µg BPA/L, TRB, or the various mixtures with TRB caused significant decreases in plasma concentrations of 17ß-estradiol. Exposure to TRB alone caused a significant reduction in plasma vitellogenin (VTG), but VTG was unaffected or even increased in females exposed to EE2 or BPA alone or, in most cases, in mixtures with TRB. Over the course of the 14-d exposure, the only treatments that clearly did not affect egg production were 1 ng EE2/L and 10 µg BPA/L. Based on these results and knowledge of hypothalamic-pituitary-gonadal axis function, we hypothesize an AOP whereby decreased production of maturation-inducing steroid leading to impaired oocyte maturation and ovulation, possibly due to negative feedback or direct inhibitory effects of membrane ER activation, could be responsible for causing adverse reproductive impacts in female fish exposed to ER agonists.
Asunto(s)
Rutas de Resultados Adversos , Cyprinidae , Contaminantes Químicos del Agua , Animales , Femenino , Andrógenos/metabolismo , Contaminantes Químicos del Agua/toxicidad , Estrógenos/toxicidad , Estrógenos/metabolismo , Etinilestradiol/toxicidad , Etinilestradiol/metabolismo , Cyprinidae/metabolismo , Vitelogeninas/metabolismoRESUMEN
Estrogenic endocrine disrupting chemicals (EEDC) have been suspected to impact offspring in a transgenerational manner via modifications of the germline epigenome in the directly exposed generations. A holistic assessment of the concentration/ exposure duration-response, threshold level, and critical exposure windows (parental gametogenesis and embryogenesis) for the transgenerational evaluation of reproduction and immune compromise concomitantly will inform the overall EEDC exposure risk. We conducted a multigenerational study using the environmental estrogen, 17α-ethinylestradiol (EE2), and the marine laboratory model fish Oryzias melastigma (adult, F0) and their offspring (F1-F4) to identify transgenerationally altered offspring generations and phenotype persistence. Three exposure scenarios were used: short parental exposure, long parental exposure, and a combined parental and embryonic exposure using two concentrations of EE2 (33ng/L, 113ng/L). The reproductive fitness of fish was evaluated by assessing fecundity, fertilization rate, hatching success, and sex ratio. Immune competence was assessed in adults via a host-resistance assay. EE2 exposure during both parental gametogenesis and embryogenesis was found to induce concentration/ exposure duration-dependent transgenerational reproductive effects in the unexposed F4 offspring. Furthermore, embryonic exposure to 113 ng/L EE2 induced feminization of the directly exposed F1 generation, followed by subsequent masculinization of the F2 and F3 generations. A sex difference was found in the transgenerationally impaired reproductive output with F4 females being sensitive to the lowest concentration of EE2 (33 ng/L) upon long-term ancestral parent exposure (21 days). Conversely, F4 males were affected by ancestral embryonic EE2 exposure. No definitive transgenerational impacts on immune competence were identified in male or female offspring. In combination, these results indicate that EEDCs can be transgenerational toxicants that may negatively impact the reproductive success and population sustainability of fish populations.
Asunto(s)
Oryzias , Contaminantes Químicos del Agua , Animales , Femenino , Masculino , Oryzias/fisiología , Aptitud Genética , Contaminantes Químicos del Agua/toxicidad , Reproducción , Fertilidad , Etinilestradiol/toxicidadRESUMEN
Tributyltin (TBT)-binding protein type 1 in Japanese medaka (Oryzias latipes) (O.latTBT-bp1) is a fish lipocalin implicated in TBT binding and detoxification. We purified recombinant O.latTBT-bp1 (rO.latTBT-bp1; ca. 30 kDa) by using a baculovirus expression system and His- and Strep-tag chromatography process. Then, we examined O.latTBT-bp1 binding to several endo/exogenous steroid hormones by means of competitive binding assay. The dissociation constants for the binding of rO.latTBT-bp1 to DAUDA and ANS, two fluorescent ligands of lipocalin, were 7.06 and 13.6 µM, respectively. Multiple model validations indicated that a single-binding-site model was the most appropriate for evaluating rO.latTBT-bp1 binding. In the competitive binding assay, testosterone, 11-ketotestosterone, and 17ß-estradiol were each bound by rO.latTBT-bp1; rO.latTBT-bp1 showed the strongest affinity for testosterone (inhibition constant, Ki = 3.47 µM). Endocrine-disrupting chemical (synthetic steroid) also bound to rO.latTBT-bp1; the affinity for ethinylestradiol (Ki = 9.29 µM) was stronger than that for 17ß-estradiol (Ki = 30.0 µM). To determine the function of O.latTBT-bp1, we produced TBT-bp1 knockout medaka (TBT-bp1 KO), which we exposed to ethinylestradiol for 28 days. After exposure, the number of papillary processes in TBT-bp1 KO genotypic male medaka was significantly fewer (3.5), compared to that in wild-type male medaka (22). Thus, TBT-bp1 KO medaka were more sensitive to the anti-androgenic effects of ethinylestradiol than wild-type medaka. These results indicate that O.latTBT-bp1 may bind to steroids and act as a gatekeeper of ethinylestradiol action by regulating the androgen-estrogen balance.
Asunto(s)
Etinilestradiol , Oryzias , Animales , Masculino , Etinilestradiol/toxicidad , Etinilestradiol/metabolismo , Peces/metabolismo , Lipocalinas/química , Lipocalinas/metabolismo , Estradiol/metabolismo , Testosterona/metabolismo , Oryzias/metabolismoRESUMEN
Natural and synthetic oestrogens are commonly found in aquatic ecosystems. The synthetic oestrogen 17α-ethinylestradiol (EE2) is widely used in oral contraceptives and its ecotoxicological effects on aquatic organisms have been widely reported. The natural oestrogen estetrol (E4) was recently approved for use in a new combined oral contraceptive and, after therapeutic use, is likely to be found in the aquatic environment. However, its potential effects on non-target species such as fish is unknown. In order to characterize and compare the endocrine disruptive potential of E4 with EE2, zebrafish (Danio rerio) were exposed to E4 or EE2 in a fish short-term reproduction assay conducted according to OECD Test Guideline 229. Sexually mature male and female fish were exposed to a range of concentrations, including environmentally relevant concentrations of E4 and EE2, for 21 days. Endpoints included fecundity, fertilization success, gonad histopathology, head/tail vitellogenin concentrations, as well as transcriptional analysis of genes related to ovarian sex steroid hormones synthesis. Our data confirmed the strong impact of EE2 on several parameters including an inhibition of fecundity, an induction of vitellogenin both in male and female fish, an alteration of gonadal structures and the modulation of genes involved in sex steroid hormone synthesis in female fish. In contrast, only few significant effects were observed with E4 with no impact on fecundity. The results suggest that the natural oestrogen, E4, presents a more favorable environmental profile than EE2 and is less likely to affect fish reproductive capacity.
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Estetrol , Contaminantes Químicos del Agua , Animales , Masculino , Femenino , Pez Cebra/fisiología , Etinilestradiol/toxicidad , Estetrol/farmacología , Vitelogeninas , Ecosistema , Contaminantes Químicos del Agua/toxicidad , Reproducción , Estrógenos/toxicidadRESUMEN
This study was conducted for the first time to investigate the long-term impacts of sublethal concentrations of 17 α-ethinylestradiol (EE2) on growth, survival, and reproductive performances in a model shrimp, the red cherry (Neocaridina davidi), females during five successive spawning steps for 7.5 months. Females were distributed in eighteen aquariums and continuously exposed to EE2 at six nominal concentrations of 0 (control), 0.02, 0.2, 2, 20, and 200 µg/L. Growth indices increased up to 0.2 µg/L and then sharply declined up to 200 µg/L. Most reproductive indices significantly decreased at levels > 0.02-0.2 µg/L with increasing EE2 levels. The highest absolute, relative, and actual fecundity values were recorded in the control, with the lowest value at 200 µg/L. With increasing EE2 levels, mean egg volume showed an increasing trend from the third spawning event onwards. Except for the time required to reach the first spawning, inter-spawning intervals considerably decreased with increasing EE2 levels at > 0.2 µg/L, especially from the third spawning stage onwards. Survival of exposed females significantly decreased with increasing EE2 levels. Unlike the body size, the juvenile's survival rates in all exposed treatments were considerably lower than the control. Females at concentrations 0.02-0.2 µg/L gained more body weight and length but produced fewer eggs with lower hatching percentages during five consecutive spawns. The results suggest that EE2 depending on the concentrations can cause unbalanced growth, reduce reproductive performances, especially from the third stage of spawning onwards, and reduce survival rates in brooders and subsequent offspring. In terms of growth, survival, and reproductive indices over successive spawns in ecotoxicology studies, the concentrations of 0.02-0.2 µg/L can be considered as chronic levels, but higher levels may have detrimental effects.
Asunto(s)
Decápodos , Penaeidae , Contaminantes Químicos del Agua , Animales , Femenino , Contaminantes Químicos del Agua/toxicidad , Reproducción , Etinilestradiol/toxicidad , FertilidadRESUMEN
Untargeted Nuclear Magnetic Resonance metabolomics was employed to study the effects of warming conditions (17-21 °C) and exposure to 17-α-ethinylestradiol (EE2) on the polar metabolome of Ruditapes philippinarum clams, to identify metabolic markers for monitoring/prediction of deviant environmental conditions. Warming alone triggered changes in alanine/aspartate/glutamate, aromatic amino acids, taurine/hypotaurine and homarine/trigonelline pathways, as well as in energy metabolism, suggesting osmoregulatory adaptations and glycolytic/tricarboxylic acid (TCA) cycle activation, possibly accompanied to some extent by gluconeogenesis to preserve glycogen reserves. At 17 °C, the lowest EE2 concentration (5 ng/L) specifically engaged branched-chain and aromatic amino acids to activate the glycolysis/TCA cycle. Notably, a partial metabolic recovery was observed at 25 ng/L, whereas higher EE2 concentrations (125 and 625 ng/L) again induced significant metabolic disturbances. These included enhanced glycogen biosynthesis and increased lipid reserves, sustained by low-level glutathione-based antioxidative mechanisms that seemed active. At 21 °C, response to EE2 was notably weak at low/intermediate concentrations, becoming particularly significant at the highest EE2 concentration (625 ng/L), suggesting higher protection capacity of Ruditapes philippinarum clams under warming conditions. At 625 ng/L, disturbances in alanine/aspartate/glutamate and taurine/hypotaurine metabolisms were observed, with no evidence of enhanced carbohydrate/protein catabolism. This low energy function profile was accompanied by marked antioxidative mechanisms and choline compounds modulation for cell membrane protection/repair. These results help monitor clams´ response to temperature rise and EE2 exposure, paving the way for future effective guidance and prediction of environmental damaging effects.
Asunto(s)
Bivalvos , Contaminantes Químicos del Agua , Animales , Temperatura , Ácido Aspártico , Antioxidantes/metabolismo , Taurina/farmacología , Bivalvos/metabolismo , Etinilestradiol/toxicidad , Etinilestradiol/metabolismo , Contaminantes Químicos del Agua/metabolismoRESUMEN
Ethinylestradiol (EE2) and sulfamethoxazole (SMX) are among pharmaceuticals and personal care products (PPCPs) and regarded as emerging contaminants in groundwater worldwide. However, the ecotoxicity and potential risk of these co-contaminants remain unknown. We investigated the effects of early-life long-term co-exposure to EE2 and SMX in groundwater on life-history traits of Caenorhabditis elegans and determined potential ecological risks in groundwater. L1 larvae of wild-type N2 C. elegans were exposed to measured concentrations of EE2 (0.001, 0.75, 5.1, 11.8 mg/L) or SMX (0.001, 1, 10, 100 mg/L) or co-exposed to EE2 (0.75 mg/L, no observed adverse effect level derived from its reproductive toxicity) and SMX (0.001, 1, 10, 100 mg/L) in groundwater. Growth and reproduction were monitored on days 0 - 6 of the exposure period. Toxicological data were analyzed using DEBtox modeling to determine the physiological modes of action (pMoAs) and the predicted no-effect concentrations (PNECs) to estimate ecological risks posed by EE2 and SMX in global groundwater. Early-life EE2 exposure significantly inhibited the growth and reproduction of C. elegans, with lowest observed adverse effect levels (LOAELs) of 11.8 and 5.1 mg/L, respectively. SMX exposure impaired the reproductive capacity of C. elegans (LOAEL = 0.001 mg/L). Co-exposure to EE2 and SMX exacerbated ecotoxicity (LOAELs of 1 mg/L SMX for growth, and 0.001 mg/L SMX for reproduction). DEBtox modeling showed that the pMoAs were increased growth and reproduction costs for EE2 and increased reproduction costs for SMX. The derived PNEC falls within the range of detected environmental levels of EE2 and SMX in groundwater worldwide. The pMoAs for EE2 and SMX combined were increased growth and reproduction costs, resulting in lower energy threshold values than single exposure. Based on global groundwater contamination data and energy threshold values, we calculated risk quotients for EE2 (0.1 - 123.0), SMX (0.2 - 91.3), and combination of EE2 and SMX (0.4 - 341.1). Our findings found that co-contamination by EE2 and SMX exacerbates toxicity and ecological risk to non-target organisms, suggesting that the ecotoxicity and ecological risk of co-contaminants of pharmaceuticals should be considered to sustainably manage groundwater and aquatic ecosystems.
Asunto(s)
Agua Subterránea , Contaminantes Químicos del Agua , Animales , Sulfametoxazol/toxicidad , Caenorhabditis elegans , Etinilestradiol/toxicidad , Ecosistema , Contaminantes Químicos del Agua/toxicidad , Preparaciones FarmacéuticasRESUMEN
The widespread use of 17α-ethinylestradiol (EE2), and other estrogenic endocrine disruptors, results in a continuous release of estrogenic compounds into aquatic environments. Xenoestrogens may interfere with the neuroendocrine system of aquatic organisms and may produce various adverse effects. The aim of the present study was to expose European sea bass larvae (Dicentrarchus labrax) to EE2 (0.5 and 50 nM) for 8 d and determine the expression levels of brain aromatase (cyp19a1b), gonadotropin-releasing hormones (gnrh1, gnrh2, gnrh3), kisspeptins (kiss1, kiss2) and estrogen receptors (esr1, esr2a, esr2b, gpera, gperb). Growth and behavior of larvae as evidenced by locomotor activity and anxiety-like behaviors were measured 8 d after EE2 treatment and a depuration period of 20 d. Exposure to 0.5 nM EE2 induced a significant increase in cyp19a1b expression levels, while upregulation of gnrh2, kiss1, and cyp19a1b expression was noted after 8 d at 50 nM EE2. Standard length at the end of the exposure phase was significantly lower in larvae exposed to 50 nM EE2 than in control; however, this effect was no longer observed after the depuration phase. The upregulation of gnrh2, kiss1, and cyp19a1b expression levels was found in conjunction with elevation in locomotor activity and anxiety-like behaviors in larvae. Behavioral alterations were still detected at the end of the depuration phase. Evidence indicates that the long-lasting effects of EE2 on behavior might impact normal development and subsequent fitness of exposed fish.