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In this study, we present a method for ion-associated liquid phase (IALP) separation and concentration of analytes from an aqueous matrix into an IALP formed in situ by the charge neutralization reaction of organic cations and anions, without centrifugation. The effects of various factors on the extraction efficiency and other parameters are investigated, whereas no instrumental stirring, such as vortexing or ultrasonics, is required because the solvent (IALP) is formed in situ. The organic cation and anion used are ethylhexyloxypropylammonium and dodecyl sulfate, respectively. The developed in situ IALP microextraction method for phase separation without centrifugation is tested using the thymol blue dye and several endocrine disruptors. The tested endocrine disruptors (bisphenol A, 17β-estradiol, 17α-ethinylestradiol, and estrone) are analyzed via high-performance liquid chromatography/fluorescence detection, with respective detection limits of 0.02, 0.02, 0.02, and 0.4 μg L−1, and the corresponding enrichment factor ranging from 47 to 71. This IALP microextraction method can be used to separate and concentrate environmental water samples of different matrices. The employed IALP is fast and easy to use, enables an approximately 100-fold analyte concentration, and has a high affinity for estrogens, thus holding promise for the separation, concentration, and quantitation of diverse trace analytes. [ABSTRACT FROM AUTHOR]

Objective: Bisphenol A (BPA) is an endocrine disruptor that interferes with estrogen (E2) levels, while Foeniculum vulgare L. (FV), a plant with phytoestrogen compounds and medicinal properties, has been suggested as a potential remedy against endocrine disruptors. This study aimed to evaluate the effect of FV extract on changes in estrogen levels, total antioxidant capacity (TAC), nitric oxide (NO), and ovarian structure in rats induced by BPA. Methods: The study used 30 female rats that were divided into different treatment groups, including a control group, a BPA group, and 3 BPA-FV-treated groups with varying doses of FV (250, 500, and 1000 mg/kg). Blood samples were taken from the rats to measure estrogen, TAC, and NO levels, while their ovaries were analyzed. Results: The results showed that BPA exposure caused a reduction in E2, TAC, and the number of ovarian follicles while increasing NO levels. However, treatment with FV extract resulted in increased E2 and TAC levels and the number of ovarian follicles while reducing NO levels. Conclusion: FV extract could be used as a natural supplement to mitigate the harmful effects of BPA. [ABSTRACT FROM AUTHOR]

Endocrine disruptors (EDs) are ubiquitous substances both in the environment and everyday products that interfere with the hormonal system. Growing evidence demonstrates their adverse effects on the organism, including the reproductive system and the prostate, owing to their (anti)estrogenic or antiandrogenic effects. Since EDs can interact with steroid hormone actions on-site, understanding the levels of intraprostatic EDs in conjunction with steroids may hold particular significance. The aim of this study was to develop and validate a method for determining estrogens, various groups of EDs (bisphenols, parabens, oxybenzone and nonylphenol) and phytoestrogens in their unconjugated and conjugated forms in prostate tissue by liquid chromatography-tandem mass spectrometry, and subsequently analyze 20 human prostate tissue samples. The method enabled 20 compounds to be analyzed: estrogens (estrone, estradiol, estriol), bisphenols (bisphenol A-BPA, BPS, BPF, BPAF, BPAP, BPZ, BPP), parabens (methyl-, ethyl-, propyl-, butyl-, benzyl-paraben), oxybenzone, nonylphenol and phytoestrogens (daidzein, genistein, equol) with LLOQs between 0.017-2.86 pg/mg of tissue. The most frequently detected EDs in prostate tissues were propylparaben (conjugated and unconjugated forms in 100 % of tissues), methylparaben (unconjugated in 45 % and conjugated in 100 %), ethylparaben (unconjugated in 25 % and conjugated in 100 %), BPA (unconjugated in 35 % and conjugated in 60 %) and oxybenzone (both forms in 45 %). To the best of our knowledge, this is the first study detecting EDs, phytoestrogens and estriol conjugate (E3C) in the prostate. E3C was the most abundant estrogen in prostatic tissue. This highlights the need for further explorations into estrogen metabolism within the prostate. [ABSTRACT FROM AUTHOR]

2,2‐Bis(4‐hydroxyphenyl)propane (bisphenol A; BPA) is an environmental endocrine‐disrupting chemical. It mimics the effects of estrogen at multiple levels by activating estrogen receptors (ERs); however, BPA also affects the proliferation of human breast cancer cells independent of ERs. Although BPA inhibits progesterone (P4) signaling, the toxicological significance of its effects remain unknown. Tripartite motif‐containing 22 (TRIM22) has been identified as a P4‐responsive and apoptosis‐related gene. Nevertheless, it has not yet been established whether exogenous chemicals change TRIM22 gene levels. Therefore, the present study investigated the effects of BPA on P4 signaling and TRIM22 and TP53 expression in human breast carcinoma MCF‐7 cells. In MCF‐7 cells incubated with various concentrations of P4, TRIM22 messenger RNA (mRNA) levels increased in a dose‐dependent manner. P4 induced apoptosis and decreased viability in MCF‐7 cells. The knockdown of TRIM22 abolished P4‐induced decreases in cell viability and P4‐induced apoptosis. P4 increased TP53 mRNA expression and p53 knockdown decrease the basal level of TRIM22 and P4 increased TRIM22 mRNA expression independent of p53 expression. BPA attenuated P4‐induced increases in the ratio of cell apoptosis in a concentration‐dependent manner, and the P4‐induced decreases in cell viability was abolished in the presence of 100 nM and higher BPA concentrations. Furthermore, BPA inhibited P4‐induced TRIM22 and TP53 expression. In conclusion, BPA inhibited P4‐induced apoptosis in MCF‐7 cells via the inhibition of P4 receptor transactivation. TRIM22 gene has potential as a biomarker for investigating the disruption of P4 signaling by chemicals. [ABSTRACT FROM AUTHOR]

Developmental exposure to endocrine disrupting chemicals (EDCs), e.g., bisphenol A (BPA) or genistein (GEN), causes longstanding epigenome effects. MicroRNAs (miRs) regulate which mRNAs will be translated to proteins and thereby serve as the final checkpoint in epigenetic control. Scant amount is known, however, whether EDCs affect neural miRNA (miR) patterns. We aimed to test the hypothesis that developmental exposure of California mice (Peromyscus californicus) to GEN, BPA, or both chemicals influences hypothalamic miR/small RNA profiles and ascertain the extent such biomolecular alterations correlate with behavioral and metabolic changes. California mice were developmentally exposed to GEN (250 mg/kg feed weight, FW), GEN (250 mg/kg FW)+BPA (5 mg/kg FW), low dose (LD) BPA (5 mg/kg FW), or upper dose (UD) BPA (50 mg/kg FW). Adult offspring were tested in a battery of behavioral and metabolic tests; whereupon, mice were euthanized, brains were collected and frozen, small RNAs were isolated from hypothalamic punches, and subsequently sequenced. California mice exposed to one or both EDCs engaged in one or more repetitive behaviors. GEN, LD BPA, and UD BPA altered aspects of ultrasonic and audible vocalizations. Each EDC exposure led to sex-dependent differences in differentially expressed miR/small RNAs with miR7-2, miR146, and miR148a being increased in all female and male EDC exposed groups. Current findings reveal that developmental exposure to GEN and/or BPA affects hypothalamic miR/small RNA expression patterns, and such changes correlate with EDC-induced behavioral and metabolic alterations. miR146 is likely an important mediator and biomarker of EDC exposure in mammals, including humans.

Trace of 17β‐estradiol (E2) contamination in food has been a concern for its negative impacts on human health, leading to the need for an E2‐monitoring system. This work reported a new simple, sensitive, and colorimetric E2 detection based on the designed repetitive‐loop aptamer and gold nanoparticles (AuNPs). The designed aptamers (L2–L5) exhibited a higher binding capability to E2 than the original truncated aptamer (L1). Although L3–L5 aptamers exhibited the highest binding capability, only L3‐aptasensor demonstrated the sensitive detection of E2 in a range of 0.05−0.8 nM, with the limit of detection at 13.1 pM. The developed L3‐aptasensor was 7.7‐folds more sensitive for E2 detection than the L1‐aptasensor. It selectively detected E2, but not the other tested chemicals with similar structures: progesterone, genistein, diethylstilbestrol, bisphenol A, and chloramphenicol. The L3‐aptasensor efficiently detected E2 spiked in milk samples within the precision acceptance criterion of recovery rates (100.1%−113.0%) and the relative standard deviations (5.24%−11.06%). These results demonstrated the development of a new aptasensor based on the designed repetitive‐loop aptamer that could enhance E2‐detection sensitivity and be potentially used for detecting E2 in milk samples with high accuracy and reliability. [ABSTRACT FROM AUTHOR]

Bisphenol A (BPA) is a monomer commonly used in the production of epoxy resins, plastic bottles and dental filling materials. Due to its chemical structure, BPA and its derivates show activity similar to the endocrine hormones. It can bind to estrogen receptors and cause neurological disturbances, even at low doses. Therefore, it is important to determine BPA and its derivatives quickly and sensitively at low concentrations. In this study, a single amperometric tyrosinase enzyme biosensor was designed for the determination of the amount of BPA, bisphenol F (BPF) and bisphenol S (BPS) monomers. Tyrosinase was immobilized onto a modified carbon paste electrode by cross-linking with glutaraldehyde. The amount of BPA (BPS and BPF) was determined directly on the reduction of quinone compound released as a result of the enzymatic reaction at -0.15V. Km(app) value of the designed biosensor for BPA was found 0.00067 μM, the linear operating range was 0.001-0.005 μM (a) and 0.03-0.1 μM (b) and the lower detection limit was found 1 nM for each monomer. It is clear that designed biosensor enable the fast, efficient and precise determination of BPA and its derivatives released from materials used in dental materials. [ABSTRACT FROM AUTHOR]

Endogenous and exogenous estrogens are widely present in food and food packaging, and high levels of natural estrogens and the misuse or illegal use of synthetic estrogens can lead to endocrine disorders and even cancer in humans. Therefore, it is consequently important to accurately evaluate the presence of food-functional ingredients or toxins with estrogen-like effects. In this study, an electrochemical sensor based on G protein-coupled estrogen receptors (GPERs) was fabricated by self-assembly, modified by double-layered gold nanoparticles, and used to measure the sensing kinetics for five GPER ligands. The interconnected allosteric constants (Ka) of the sensor for 17β-estradiol, resveratrol, G-1, G-15, and bisphenol A were 8.90 × 10−17, 8.35 × 10−16, 8.00 × 10−15, 5.01 × 10−15, and 6.65 × 10−16 mol/L, respectively. The sensitivity of the sensor for the five ligands followed the order of 17β-estradiol > bisphenol A > resveratrol > G-15 > G-1. The receptor sensor also demonstrated higher sensor sensitivity for natural estrogens than exogenous estrogens. The results of molecular simulation docking showed that the residues Arg, Glu, His, and Asn of GPER mainly formed hydrogen bonds with -OH, C-O-C, or -NH-. In this study, simulating the intracellular receptor signaling cascade with an electrochemical signal amplification system enabled us to directly measure GPER–ligand interactions and explore the kinetics after the self-assembly of GPERs on a biosensor. This study also provides a novel platform for the accurate functional evaluation of food-functional components and toxins. [ABSTRACT FROM AUTHOR]

Bisphenol A (BPA) as an endocrine‐disrupting chemical (EDC) with low estrogenic activity increases oxidative stress and testicular damage. Bromelain is a mixture of different thiol endopeptidases and other components with many uses as a natural anti‐inflammatory enzyme. The present study aimed to evaluate the effect of bromelain on male reproductive failure induced by BPA. A total of 60 healthy adult male mice were randomly divided into six groups (n = 6), including control, bromelain (70 mg/kg), BPA (5 and 600 mg/kg), and BPA (5 and 600 mg/kg) + bromelain. BPA and bromelain were administrated orally for 35 days. Then, the epididymis and testes were removed to evaluate sperm parameters, oxidative stress markers, serum levels of testosterone concentrations, and oestrogen receptors expression. The BPA significantly (P < 0.05) decreased sperm count, motility, viability, and normal sperm morphology, as well as testosterone levels, oestrogen receptors alpha (ERα) and beta (ERβ), GPx, CAT, and SOD activity than control. Also, BPA significantly (P < 0.05) increased the sperm anomalies, and MDA concentration. Co‐administration of bromelain + BPA caused a significantly (P < 0.05) increase sperm count, normal sperm morphology, testosterone levels, expression of ERα and ERβ, and GPx, CAT, and SOD activity than the BPA group (P < 0.05). Also, Bromelain significantly (P < 0.05) decreased sperm anomalies and MDA concentration than control. Based on the results of this study, it appears that BPA causes side effects on male reproduction. While, bromelain has the potential to reduce the side effects of BPA on the male reproductive system. [ABSTRACT FROM AUTHOR]

G protein-coupled estrogen receptor (GPER) plays a prominent role in facilitating the rapid, non-genomic signaling of estrogens in breast cancer cells. Herein, a comprehensive overview of the role of GPER in ER-ɑ-negative breast cancer is provided. Activation of GPER affected proliferation, metastasis and epithelial mesenchymal transition in ER-ɑ negative breast cancer cells. Clinical studies have demonstrated that GPER positivity was strongly correlated with larger tumor size and advanced clinical stage, suggesting that GPER/ERK signaling may play a role in promoting tumor progression. Strong evidence existed that environmental contaminants like bisphenol A have a carcinogenic potential mediated by GPER activation. The complexity of the cross talk between GPER and other receptors including ER-β, ER-α36, Estrogen-related receptor α (ERRα) and androgen receptor has been discussed. The potential utility of small molecules and phytoestrogens targeting GPER, adds valuable insights into its therapeutic potential. This review holds promises in advancing our understanding of GPER role in ER-ɑ-negative breast cancer. Overall, the consequences of GPER activation are still an area of active research and the implication are not entirely clear. • Activation of GPER affected proliferation, metastasis and stemness. • Contaminants like bisphenol A have a carcinogenic potential mediated by GPER activation. • GPER positivity was strongly correlated with larger tumor size and advanced clinical stage. • There is a cross talk between GPER and ER-β, ER-α36, ERR α and androgen receptors. • Small molecules and phytoestrogens targeting GPER may have therapeutic potential. [ABSTRACT FROM AUTHOR]

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Recent increase in incidence of meningiomas suggests the need to search for new risk factors. Leptin, a potentially pro-angiogenic and proliferative agent, could be a candidate for this role, as its expression correlates with body mass index (BMI). Because development of meningioma has also been linked to sex hormones, bisphenol A (BPA), a known xenoestrogen, can also be taken into consideration as a potential risk factor. The aim of this study was to determine plasma concentrations of both substances in patients with meningiomas and to match it to patients with gliomas – a group of brain tumors less hormone- and BMI-dependent. Concentrations of BPA and leptin were measured in plasma of 24 patients with low grade meningioma and in 29 patients with glioma, using gas chromatography-mass spectrometry (GC-MS) and ELISA kits, respectively. The concentrations of both substances in patients with neoplasms were interpreted in relation to their concentration in healthy population, published in recent reports. Free and conjugated BPA were present in both meningioma and glioma patients. Moreover, their concentrations far exceeded those reported in the healthy population. Nevertheless, the level of leptin revealed to be significantly higher in meningioma patients than in glioma patients. Occurrence of both meningioma and glioma may be accompanied by increased concentrations of leptin and BPA. Further large-scale studies are needed to clarify whether the presence of both substances may play a role in pathogenesis or influence clinical course in patients with brain neoplasms. [ABSTRACT FROM AUTHOR]

Chemicals with androgenic or estrogenic activity induce the sex reversal and/or intersex condition in various teleost fish species. Previously, we reported that exposure to 17α‐methyltestosterone, bisphenol A, or 4‐nonylphenol induces changes in expression of the gonadal soma‐derived factor (gsdf) gene accompanied by disruption of gonadal differentiation in Japanese medaka (Oryzias latipes). These findings suggest that gsdf expression might be a useful biomarker for predicting the potential effect of chemicals on gonadal differentiation. We examined the gsdf expression in Japanese medaka exposed to chemicals with estrogenic or androgenic activity. Exposure to the androgenic steroid 17β‐trenbolone at 0.5–22.1 μg/L induced the development of ovotestis (presence of ovarian tissue with testicular tissue) and female‐to‐male sex reversal in XX embryos, and exposure at 6.32 and 22.1 μg/L significantly increased gsdf expression in XX embryos compared with controls at developmental stage 38 (1 day before hatching). In the present study, no statistically significant difference in gsdf mRNA expression was observed after exposure to 17β‐estradiol, 17α‐ethinylestradiol, and 4‐t‐octylphenol, which have estrogenic activity. In addition, antiandrogenic chemicals or chemicals without endocrine‐disrupting activity did not induce changes in gsdf expression in XX or XY embryos. Thus, an increase in gsdf expression after androgen exposure was observed in XX embryos. Together, these findings indicate that gsdf expression might be useful for predicting the adverse effect of chemicals on gonadal differentiation. Environ Toxicol Chem 2022;41:1875–1884. © 2022 SETAC [ABSTRACT FROM AUTHOR]

Endocrine disrupting chemicals (EDCs) exert significant effects on health and physiology, many traceable to effects on stem cell programming underlying development. Understanding risk of low-level, chronic EDC exposure will be enhanced by knowledge of effects on stem cells. We exposed rhesus monkey embryonic stem cells to low levels of five EDCs [bisphenol A (BPA), atrazine (ATR), tributyltin (TBT), perfluorooctanoic acid (PFOA), and di-(2-ethylhexyl) phthalate (DEHP)] for 28days, and evaluated effects on gene expression by RNAseq transcriptome profiling. We observed little effect of BPA, and small numbers of affected genes (≤119) with other EDCs. There was substantial overlap in effects across two, three, or four treatments. Ingenuity Pathway analysis indicated suppression of cell survival genes and genes downstream of several stress response mediators, activation of cell death genes, and modulations in several genes regulating pluripotency, differentiation, and germ layer development. Potential adverse effects of these changes on development are discussed.

Coffee brewed from capsule machines may contain estrogenic chemicals migrated from plastic, but the estrogenic activity of capsule coffee has not been evaluated. This study evaluated the estrogenic activity of capsule coffee using the VM7Luc4E2 estrogen receptor transcriptional activation assay. Estrogenic potentials of six capsule coffee samples were calculated using relative maximum amplitude response of E2 (>15%RME2 indicative of estrogenic activity) and estradiol equivalent factor (EEF). Estrogenic chemical content was determined using ultra-performance liquid chromatography with tandem mass spectrometry. All capsule coffee samples possessed estrogenic activity (48–56%RME2). EEFs were 6–7 orders of magnitude lower than that of E2, (1.2 × 10−7−1.7 × 10−6), indicating substantially weaker estrogenic potencies. Bisphenol A, bisphenol F, benzophenone, 4-nonylphenol, dibutyl phthalate, and dimethyl terephthalate were detected in capsule coffee. Capsule coffee exhibited estrogenic activity in vitro, and its estrogenic chemical content is likely driving its estrogenicity, warranting further investigations to fully understand the degree to which they are related and to predict the estrogenic potential based on the concentration of estrogenic chemicals.

Bisphenol A (BPA) is an endocrine-disrupting chemical (EDC), associated with obesity and insulin resistance. The FDA prohibited the use of BPA-based polycarbonate resins in infant formula packaging; thus, its analogs, viz. Bisphenol S (BPS) and Bisphenol F (BPF) were considered alternatives in epoxy resins, plastics, and food cans. As these analogs might evoke a similar response, we investigated the role of Bisphenols (BPA, BPF, and BPS), on insulin signaling in CHO-HIRc-myc-GLUT4eGFP cells at environmentally relevant concentrations of 2 nM and 200 nM. Insulin signaling demonstrated that Bisphenols reduced phosphorylation of IR and AKT2, GLUT4 translocation, and glucose uptake. This was accompanied by increased oxidative stress. Furthermore, SWATH-MS-based proteomics of 3T3-L1 cells demonstrated that Bisphenol-treated cells regulate proteins in insulin resistance, adipogenesis, and fatty acid metabolism pathways differently. All three Bisphenols induced differentially expressed proteins enriched similar pathways, although their abundance differed for each Bisphenol. This might be due to their varying toxicity level, structural differences, and estrogen-mimetic activity. This study has important implications in addressing health concerns related to EDCs. Given that the analogs of BPA are considered alternatives to BPA, the findings of this study suggest they are equally potent in altering fatty acid metabolism and inducing insulin resistance. The probable molecular mechanism involved in Bisphenols-induced disruption in insulin signaling and fatty acid metabolism. [Display omitted] • BPA, BPF, BPS induced insulin resistance as reflected by reduced p -IR and p -AKT2, glucose uptake, and GLUT4 translocation. • BPA, BPF, BPS increased NADPH oxidase expression and ROS. • Proteomic analysis suggested BPA, BPF, and BPS disrupt insulin signaling and fatty acid metabolism. • BPA, BPF, and BPS increased lipid accumulation, thus inducing lipotoxicity. [ABSTRACT FROM AUTHOR]

The effects of environmental chemicals on health outcomes may be underestimated due to deficiency of knowledge regarding the actions of compounds on toxico-pathogenic mechanisms underlying biological systems outcomes. In this regard, the current study aimed to explore the potential target-pathway-disease relationship attributed to bisphenol A (BPA) responses in target tissues. Computational methods including reverse pharmacophore mapping approach, structural similarity based search and kinome wide interaction profiling were employed with molecular docking validation. Gene ontology (GO) enrichment analysis and protein-protein interaction (PPI) network based illustrations were utilized to prioritize target-pathway and disease relationships. Data illustrated that BPA possessed multi-target nature since this chemical potentially interacted with various protein targets where many of these were validated through docking. Potential BPA targets were significantly enriched to various cellular signaling pathways including steroid biosynthesis, peroxisome proliferator-activated receptor gamma (PPARℽ) and cancer. Further, hypertension was prioritized as disease target. In addition, BPA targeted 17 cell signaling kinases encompassed in the human kinome. In addition, inflammatory (5-LO) and apoptosis regulators (Bcl-X and Bcl-2) were also explored as novel targets. Evidence indicates that the multi-target nature and plausible mechanisms underlying BPA actions in a system wide manner aids toward understanding of adverse effects. This observation may lead us to more precise method to elucidate the toxico-pathogenic mechanisms of BPA with an environmental health perspective. [ABSTRACT FROM AUTHOR]

Endocrine disrupting chemicals (EDC) are those compounds that can interrupt the biosynthesis, metabolism, and functioning of endogenous hormones. Bisphenol A and S are well-known EDCs that can mimic an endogenous hormone estrogen. Both the chemicals have been detected in commercially available food products as used in polycarbonate plastic bags or within canned linings. BPS is more likely to accumulate in the aquatic environment or in the food chain; also detected in human samples. Due to its estrogen-like activity, it is able to interfere with the processes of growth and development such as reproductive development, bone development and soon. One such process that can be altered by BPA is neurogenesis. Neurogenesis continues to occur in adults in a limited fashion which is enabled by the action of genes involved ingrowth and development such as, Wnt3a, BMP2 and Sox2. Wnt3a promotes neurogenesis while BMP2 represses the neurogenesis but also promotes the fate specification and survival of neuroblasts, andSox2 is responsible for themaintenance of quiescent neural stem cells (NSCs), these genes together act to enable neurogenesis in a controlled manner.BPA and BPS have the potential to disturb neuron expressions as well as their fate, as it is directly linked with the nervous system. This review dealt the effect of EDCs, BPA and BPS in neurogenesis, via exogenous and endogenous factors thereby, aid in understanding the basic cross-talk among these pathways during neuronal development and dysregulation. [ABSTRACT FROM AUTHOR]

Compounds that trigger breast cancer onset and establishment are of great interest in biological research. Endocrine disruptors are relevant because they initiate carcinogenesis by changing endocrine pathways. Bisphenol A (BPA), as a ubiquitous xenoestrogen, is largely associated with dysfunctions in the female reproductive system and associated organs. This study proposes an investigation of the mammary gland (MG) in aged Mongolian gerbil (Meriones unguiculatus) mothers after their exposure to BPA in two windows of morphophysiological plasticity: pregnancy and lactation. A low dose (50 µg/kg) and a high dose (5000 µg/kg) of BPA were considered, and results showed few differences between them. As expected, we observed contrasts among control and BPAexposed MG. The control groups presented a regressive phase with high apoptotic activity and elastic stroma. However, BPA damaged mammary tissue and provoked multifocal carcinoma development supported by an apparent epithelial-mesenchymal transition (EMT) and reactive stroma establishment. BPA remodeled stromal fibers deposition and enhanced the recruitment of tumor-associated cells, contributing to a tumoral microenvironment. Overexpression of TGF-ß1 was induced by BPA in the epithelial compartment of exposed MG, and increased expression of metalloproteinases (MMP-2, MMP-3, MMP-9) was present in carcinoma cells. In conclusion, exposure of mothers to BPA during the gestational/lactational window of susceptibility leads to carcinogenic impacts with aging. [ABSTRACT FROM AUTHOR]

The G protein‐coupled estrogen receptor 1 (Gper1) is a membrane‐bound estrogen receptor that mediates non‐genomic action of estrogens. A Gper1‐mediating pathway has been implicated in reproductive activities in fish, including oocyte growth, but Gper1 has been characterized in only a very limited number of fish species. In this study, we cloned and characterized two genes encoding medaka (Oryzias latipes) Gper1s, namely, Gper1a and Gper1b, and phylogenic and synteny analyses suggest that these genes originate through a teleost‐specific whole genome duplication event. We found that Gper1a induced phosphorylation of mitogen‐activated protein kinase (MAPK) in 293T cells transfected with medaka Gper1s on exposure to the natural estrogen, 17β‐estradiol (E2) and a synthetic Gper1 agonist (G‐1), and treatment with both E2 and G‐1 also decreased the rate of spontaneous maturation in medaka oocytes. These findings show that the processes for oocyte growth and maturation are sensitive to estrogens and are possibly mediated through Gper1a in medaka. We also show that 17α‐ethinylestradiol (EE2), one of the most potent estrogenic endocrine‐disrupting chemicals, and bisphenol A (BPA, a weak environmental estrogen) augmented phosphorylation of MAPK through medaka Gper1s in 293T cells. Interestingly, however, treatment with EE2 or BPA did not attenuate maturation of medaka oocytes. Our findings support that Gper1‐mediated effects on oocytes are conserved among fish species, but effects of estrogenic endocrine‐disrupting chemicals on oocytes acting through Gper1 may be divergent among fish species. We cloned and characterized medaka Gper1a and Gper1b, which encode membrane‐bound estrogen receptors. Gper1a induced phosphorylation of MAPK in 293T cells transfected with medaka Gper1s on exposure to 17β‐estradiol (E2) and G‐1, and treatment with both E2 and G‐1 also decreased the rate of spontaneous maturation in medaka oocytes. Estrogenic endocrine‐disrupting chemicals, such as 17α‐ethinylestradiol and bisphenol A, also, at least in part, activated MAPK. The current results provide advance understanding on the membrane‐bound estrogen receptor in fish. [ABSTRACT FROM AUTHOR]

Gestational bisphenol A (BPA) exposure induced multiple programmed diseases in the adult offsprings. Thus, this study targeted exploring the physiological impacts of melatonin (MEL) as a reprogramming strategy against in utero BPA exposure on reproductive capacity of adult F1 female rat offspring. Forty adult pregnant albino female rats were divided equally into 5 groups (n = 8): group I (control), group II (low-dose BPA; 25 μg BPA/kg B.w.t.), group III (low-dose BPA + 10 mg MEL/kg B.w.t.), group IV (high-dose BPA; 250 μg/kg B.w.t.), and group V (high-dose BPA + MEL). Treatments were given daily by subcutaneous (s/c) injection from the fourth day of pregnancy until full term. After delivery, female offspring were selected, and on postnatal day 60, adult offspring were examined for estrus regularity and then were sacrificed at estrus to collect blood and tissue samples. Findings clarified that in utero BPA exposure (both doses) increased significantly (P < 0.05) the ovarian weights and the serum levels of estrogen but decreased that of triiodothyronine (T3) compared to control groups. Significant increasing of serum malondialdehyde (MDA) and decreasing of total antioxidant capacity (TAC) were also detected. Both doses of BPA disturbed remarkably the estrus cycles and caused marked aberrations in ovarian and uterine tissues. Interestingly, prenatal MEL co-treatment with BPA mitigated significantly all of these degenerative changes. Thus, this study first demonstrated that prenatal MEL therapy could be used as a potent reprogramming intervention against BPA-induced reproductive disorders in the adult F1 female rat offspring. [ABSTRACT FROM AUTHOR]

BackgroundChemicals that have estrogenic activity (EA) can potentially cause adverse health effects in mammals including humans, sometimes at low doses in fetal through juvenile stages with effects detected in adults. Polycarbonate (PC) thermoplastic resins made from bisphenol A (BPA), a chemical that has EA, are now often avoided in products used by babies. Other BPA-free thermoplastic resins, some hypothesized or advertised to be EA-free, are replacing PC resins used to make reusable hard and clear thermoplastic products such as baby bottles.MethodsWe used two very sensitive and accurate in vitro assays (MCF-7 and BG1Luc human cell lines) to quantify the EA of chemicals leached into ethanol or water/saline extracts of fourteen unstressed or stressed (autoclaving, microwaving, UV radiation) thermoplastic resins. Estrogen receptor (ER)-dependent agonist responses were confirmed by their inhibition with the ER antagonist ICI 182,780.ResultsOur data showed that some (4/14) unstressed and stressed BPA-free thermoplastic resins leached chemicals having significant levels of EA, including one polystyrene (PS), and three Tritan™ resins, the latter reportedly EA-free. Exposure to UV radiation in natural sunlight resulted in an increased release of EA from Tritan™ resins. Triphenyl-phosphate (TPP), an additive used to manufacture some thermoplastic resins such as Tritan™, exhibited EA in both MCF-7 and BG1Luc assays. Ten unstressed or stressed glycol-modified polyethylene terephthalate (PETG), cyclic olefin polymer (COP) or copolymer (COC) thermoplastic resins did not release chemicals with detectable EA under any test condition.ConclusionsThis hazard survey study assessed the release of chemicals exhibiting EA as detected by two sensitive, widely used and accepted, human cell line in vitro assays. Four PC replacement resins (Tritan™ and PS) released chemicals having EA. However, ten other PC-replacement resins did not leach chemicals having EA (EA-free-resins). These results indicate that PC-replacement plastic products could be made from EA-free resins (if appropriate EA-free additives are chosen) that maintain advantages of re-usable plastic items (price, weight, shatter resistance) without releasing chemicals having EA that potentially produce adverse health effects on current or future generations.

Mounting evidence suggests that environmental factors-in particular, those that we are exposed to during perinatal life-can dramatically shape the organism's risk for later diseases, including neurobehavioral disorders. However, depending on the environmental insult, one sex may demonstrate greater vulnerability than the other sex. Herein, we focus on two well-defined extrinsic environmental factors that lead to sexually dimorphic behavioral differences in animal models and linkage in human epidemiological studies. These include maternal or psychosocial stress (such as social stress) and exposure to endocrine-disrupting compounds (such as one of the most prevalent, bisphenol A [BPA]). In general, the evidence suggests that early environmental exposures, such as BPA and stress, lead to more pronounced behavioral deficits in males than in females, whereas female neurobehavioral patterns are more vulnerable to later in life stress. These findings highlight the importance of considering sex differences and developmental timing when examining the effects of environmental factors on later neurobehavioral outcomes.

Endocrine disrupting chemicals (EDCs) pose a significant threat to human health, society, and the environment. Many EDCs elicit their toxic effects through nuclear hormone receptors, like the estrogen receptor α (ERα). In silico models can be used to prioritize chemicals for toxicological evaluation to reduce the amount of costly pharmacological testing and enable early alerts for newly designed compounds. However, many of the current computational models are overly dependent on the chemistry of known modulators and perform poorly for novel chemical scaffolds. Herein we describe the development of computational, three-dimensional multi-conformational pocket-field docking, and chemical-field docking models for the identification of novel EDCs that act via the ligand-binding domain of ERα. These models were highly accurate in the retrospective task of distinguishing known high-affinity ERα modulators from inactive or decoy molecules, with minimal training. To illustrate the utility of the models in prospective in silico compound screening, we screened a database of over 6000 environmental chemicals and evaluated the 24 top-ranked hits in an ERα transcriptional activation assay and a differential scanning fluorimetry-based ERα binding assay. Promisingly, six chemicals displayed ERα agonist activity (32nM-3.98μM) and two chemicals had moderately stabilizing effects on ERα. Two newly identified active compounds were chemically related β-adrenergic receptor (βAR) agonists, dobutamine, and ractopamine (a feed additive that promotes leanness in cattle and poultry), which are the first βAR agonists identified as activators of ERα-mediated gene transcription. This approach can be applied to other receptors implicated in endocrine disruption.

This study evaluated the effects of bisphenol A (BPA) on human endometrial stromal fibroblast (ESF) differentiation and expression of genes involved in oestrogen metabolism. Human ESF from eight hysterectomy specimens were cultured and treated with 5-100 μmol/l of BPA ± oestradiol or 8-br-cAMP for 48 h. mRNA expression was analysed by real-time reverse-transcription PCR. 8-br-cAMP-induced human ESF decidualization was confirmed by expression of insulin-like growth factor binding protein-1 (IGFBP1) and prolactin secretion. Short-term exposure (48 h) decreased human ESF proliferation (P

BackgroundBisphenol A (BPA), a chemical used as a plasticizer, is a potent endocrine disruptor that, even in low concentrations, disturbs normal development and functions of reproductive organs in different species.ObjectivesWe investigated whether BPA affects human ovarian granulosa cell function.MethodsWe treated KGN granulosa cells and granulosa cells from subjects undergoing in vitro fertilization (IVF) with follicle-stimulating hormone (FSH), BPA, or BPA plus FSH in a dose- and time-dependent manner. We then evaluated expression of insulin-like growth factor 1 (IGF-1), aromatase, and transcription factors known to mediate aromatase induction by FSH [including steroidogenic factor-1 (SF-1), GATA4, cAMP response element binding protein-1 (CREB-1), and peroxisome proliferator-activated receptor-gamma (PPARgamma)], as well as 17beta-estradiol (E2) secretion. KGN cells were transfected with a PPARgamma-containing vector, followed by assessment of aromatase and IGF-I expression.ResultsBPA reduced FSH-induced IGF-1 and aromatase expression and E2 secretion in a dose-dependent fashion. Similar effects on aromatase were observed in IVF granulosa cells. SF-1 and GATA4, but not CREB-1, were reduced after BPA treatment, although PPARgamma, an inhibitor of aromatase, was significantly up-regulated by BPA in a dose-dependent manner, with simultaneous decrease of aromatase. Overexpression of PPARgamma in KGN cells reduced FSH-stimulated aromatase and IGF-1 mRNAs, with increasing concentrations of the transfected expression vector, mimicking BPA action. Also, BPA reduced granulosa cell DNA synthesis without changing DNA fragmentation, suggesting that BPA does not induce apoptosis.ConclusionsOverall, the data demonstrate that BPA induces PPARgamma, which mediates down-regulation of FSH-stimulated IGF-1, SF-1, GATA4, aromatase, and E2 in human granulosa cells. These observations support a potential role of altered steroidogenesis and proliferation within the ovarian follicular compartment due to this endocrine disruptor.

Objective(s): Since bisphenol A (BPA) induces bone loss and phytoestrogens enhance the osteoblastogenesis by binding to the non-classical and classical oestrogen receptors, respectively, the present study was aimed to observe the osteoprotective effect of phytoestrogens on BPA-induced osteoblasts in hFOB 1.19 cells. Materials and Methods: All groups of hFOB 1.19 cells were induced with 12.5 μg/ml of BPA except the control (Ctrl) group. Meanwhile, treated groups received phytoestrogens; Daidzein (Dz), Genistein (Gt), Equol (Eq) and 17β-oestradiol (Est) in different concentrations for 24 hr duration. Results: We found that the protein expression of non-classical oestrogen-related receptor (ERRG) was highly expressed in BPA group, whereas classical oestrogen receptor alpha (ERα) and oestrogen receptor beta (ERβ) were relatively increased with phytoestrogens treatment under BPA exposure. The dense actin cytoskeletal filaments were also observed. qRT-PCR showed up-regulation of mitogen-activated protein kinase 3 (MAPK3) and G protein-coupled receptor 30 (GPR30) expressions; significant downregulation of ERRG and up-regulation of ERα and ERβ were observed in phytoestrogens-treated cells, which was supported by the increased expressions of oestrogen receptor 1 (ESR1) and oestrogen receptor 2 (ESR2). Conclusion: Phytoestrogens improved the deteriorative effect of BPA via down-regulation of ERRG in hFOB 1.19 cells. This study showed that the efficacy of consumption of phytoestrogens in rendering them as potential therapeutic strategy in combating the adverse bone effects of BPA. [ABSTRACT FROM AUTHOR]

Food contact materials (FCM) may contain complex mixtures of estrogenic chemicals. A yeast estrogen screen performed on high performance thin-layer chromatography plates (planar-YES, P-YES) is promising for analysis of such mixtures, as it could allow for better elucidation of effects compared with established methods in microtiter plates. However, the P-YES has not been directly compared with established methods. We compared the performance of a microtiter plate YES (lyticase-YES, L-YES) to P-YES on silica gel HPTLC plates using 17β-estradiol (E2), 20 chemicals representative of migrants from plastic FCM, and three migrates of coated metal food cans. Effective doses (ED10, ED50) and estradiol equivalencies were calculated for each chemical. Thirteen chemicals had calculable EDs in the L-YES or P-YES, with average EDs 13-fold (range 0.63–36) more potent in P-YES than in the L-YES. Normalized to E2, the median estrogenicity was within 1.5-fold (0.43–8.8) between the assays. Therefore, P-YES was as or more sensitive than L-YES but potencies relative to E2 were comparable between assays. With chromatography, the P-YES detected estrogenicity in coated metal cans, effects that were unmeasurable in L-YES. With the sample preparation methods used in this study, both YES assays are sufficiently sensitive to detect bisphenol A below the specific migration limit for plastic packaging (0.05 mg/kg food). This study demonstrates that P-YES outperforms L-YES because it is more sensitive, provides comparable estradiol equivalents, and circumvents confounding mixture effects. The P-YES will be useful for routine monitoring of FCM and toxicant identification in problematic materials. [ABSTRACT FROM AUTHOR]

Bisphenol A (BPA), a xenoestrogen commonly used in plastics, may act as an endocrine disruptor, which indicates that BPA might be a public health risk. The present study aimed to investigate the effect of BPA on 17β-estradiol (E2)-mediated protection against liver ischemia/reperfusion (I/R) injury, and to identify the underlying mechanisms using a rat model. A total of 56 male Sprague Dawley rats were randomly divided into the following seven groups: i) Sham; ii) I/R; iii) Sham + BPA; iv) I/R + BPA; v) I/R + E2; vi) I/R + E2 + BPA; and vii) I/R + E2 + BPA + losartan [LOS; an angiotensin II (Ang II) type I receptor (ATIR) antagonist]. A rat model of hepatic I/R injury was established by inducing hepatic ischemia for 60 min followed by reperfusion for 24 h. When ischemia was induced, rats were treated with vehicle, E2, BPA or LOS. After 24 h of reperfusion, blood samples and hepatic tissues were collected for histopathological and biochemical examinations. The results suggested that 4 mg/kg BPA did not significantly alter the liver function, or Ang II and AT1R expression levels in the Sham and I/R groups. However, 4 mg/kg BPA inhibited E2-mediated hepatic protection by enhancing hepatic necrosis, and increasing the release of alanine transaminase, alkaline phosphatase and total bilirubin (P<0.05). Moreover, BPA increased serum and hepatic Ang II levels, as well as AT1R protein expression levels in the E2-treated rat model of liver I/R injury (P<0.05). LOS treatment reversed the negative effects of BPA on hepatic necrosis and liver serum marker levels, although it did not reverse BPA-mediated upregulation of serum and hepatic Ang II levels, or hepatic AT1R expression. Therefore, the present study suggested that BPA disrupted E2-mediated hepatic protection following I/R injury, but did not significantly affect healthy or I/R-injured livers; therefore, the mechanism underlying the effects of BPA may be associated with upregulation of the Ang II/AT1R signaling pathway. [ABSTRACT FROM AUTHOR]

Thermochromic inks, materials that change color at a certain temperature, are increasingly used on papers and other materials in the areas of design, commercials, and security printing. Pseudo-estrogen bisphenol A (BPA) may be one of their main compounds present in mass fractions of up to several percent. In this work, the mass fractions of BPA in thermochromic prints on seven types of paper were determined. Migration of BPA from the surface of the thermochromic print to artificial sweat solutions was investigated as well. Total amount of BPA in papers with thermochromic prints was determined by an HPLC-UV method with ultrasonic-assisted extraction in methanol developed and validated in this work. Total amount of BPA, which ranged from 0.126 to 0.778 mg/g, was compared with the amounts extracted under the same conditions in two artificial human sweat solutions, which differed in chemical composition, ionic strength, and pH-value. Mass fractions of BPA extracted with artificial sweat solutions were from 0.047 to 0.175 mg/g with respect to the mass of the paper. On average, the mass fraction of BPA was four times less when extracted with artificial sweat solutions than the maximal amount extracted with methanol. The amounts of extracted BPA raise a concern of health risk through dermal exposure to BPA from thermochromic prints on paper. [ABSTRACT FROM AUTHOR]

Purpose of Review: Cardiovascular disease (CVD) is a non-subsiding disease that remains a leading cause of morbidity and mortality. CVD has been associated with endocrine disruptors, such as bisphenol A (BPA). This review critically summarizes existing findings on BPA and hypertension, with particular attention to genomic, non-genomic, molecular, and cellular mechanisms of action that render BPA as a cardiovascular estrogenic disruptor. Recent Findings: Owing to its similar estrogenic structure, BPA has been shown to affect various phenotypes that are regulated by the natural hormone, estrogen. Indeed, BPA has been shown to interact with estrogen receptors, located both in the cell membrane and in the cytoplasm/nucleus. Given that estrogen plays an important role in cardiovascular physiology, a contributing role for BPA in CVD would not be unexpected. Existing literature, though limited, established BPA as a source of disruption in cardiovascular health, particularly hypertension. However, effects of BPA are largely dependent on the dose, patient gender, tissue, and developmental stage of the exposed tissue/organ. Summary: Accumulating evidence argues for an adverse effect of BPA on blood pressure, with this effect being gender, dose, and time specific. Thus, comprehensive studies which take these factors and other parameters, like epigenetic factors, into account are warranted before a thorough understanding is at hand. [ABSTRACT FROM AUTHOR]

4-Hexylresorcinol (4HR) has been used as a food additive and antiseptic. The aim of this study was to evaluate whether the application of 4HR in breast cancer cells and ovariectomized rats showed estrogen-like effects. MCF-7 and SK-BR-3 cells were treated by solvent, 1–100 µM bisphenol-A (BPA), or 1–100 µM 4HR, respectively. 3-(4, 5-Dimethylthiazole-2-yl)- 2,5-diphenyltetrazolium bromide (MTT) assay and Western blot for extracellular signal-regulated kinase-1/-2 (ERK1/2), phosphorylated ERK1/2 (p-ERK1/2), estrogen receptor-α (ERα), and ERβ were done. As an in vivo study, ovariectomized rats (n = 15) received solvent, 125 mg/kg of 4HR, or 10 µg of 17-β estradiol via daily subcutaneous injection for 7 days. Blood samples were obtained for evaluation of prolactin levels. Pituitary glands and uteruses were biopsied for histological evaluation and Western blot analysis. Compared with the control group, the application of 4HR decreased the proliferation of MCF-7 and SK-BR-3 cells, while the application of BPA increased (p < 0.05). The application of BPA increased the expression of ERα, ERβ, and p-ERK1/2, but 4HR did not change the expression of ERα, ERβ, or p-ERK1/2 in MCF-7 cells. In an animal model, the 4HR group showed similar levels of ERα, ERβ, and prolactin expression in the pituitary gland compared to the solvent only group, while the estradiol group showed higher levels. Serum prolactin levels were similar between the 4HR and solvent only groups. Taken together, 1–100 µM 4HR did not show BPA-like behavior in MCF-7 cells, and 125 mg/kg of 4HR daily subcutaneous injection for 7 days did not demonstrate estradiol-like effects in ovariectomized rats. Collectively, 4HR has no estrogen-like effects on both ERα-positive cells and estrogen-deficient rat models. [ABSTRACT FROM AUTHOR]

• In mouse testes with Leydig cell tumor expression of GPER and PPAR is altered. • Treatment of MA-10 cells with BPA and its derivatives effects on GPER and PPAR expression. • There is a relation between expression of INSL3 and RLN and estrogen level in Leydig cell tumor. • PI3K-Akt-mTOR pathway and/or only mTOR is involved in GPER and PPAR interaction. The etiology and molecular characteristics of Leydig cell tumor (LCT) are scarcely known. From the research data stems that estrogen can be implicated in LCT induction and development, however it is not investigated in detail. Considering the above, herein we analyzed the relation between G-protein coupled membrane estrogen receptor, peroxisome proliferator-activated receptor and insulin-like family peptides (insulin-like 3 peptide; INSL3 and relaxin; RLN) expressions as well as estrogen level with impact of xenoestrogen (bisphenol A; BPA, tetrabromobisphenol A; TBBPA, and tetrachlorobisphenol A; TCBPA). While in our previous studies altered GPER-PPAR partnership was found in human LCT being a possible cause and/or additionally effecting on LCT development, here mouse testes with experimentally induced LCT and mouse tumor Leydig cell (MA-10) treated with BPA chemicals were examined. We revealed either diverse changes in expression or co-expression of GPER and PPAR in mouse LCT as well as in MA-10 cells after BPA analogues when compared to human LCT. Relationships between expression of INSL3, RLN, including co-expression, and estrogen level in human LCT, mouse LCT and MA-10 cells xenoestrogen-treated were found. Moreover, involvement of PI3K-Akt-mTOR pathway or only mTOR in the interactions of examined receptors and hormones was showed. Taken together, species, cell of origin, experimental system used and type of used chemical differences may result in diverse molecular characteristics of LCT. Estrogen/xenoestrogen may play a role in tumor Leydig cell proliferation and biochemical nature but this issue requires further studies. Experimentally-induced LCT in mouse testis and MA-10 cells after BPA exposure seem to be additional models for understanding some aspects of human LCT biology. [ABSTRACT FROM AUTHOR]

Resin-based dental composites have been developed to restore decayed teeth or modify tooth color due to their excellent physical and chemical properties. Such composites may have intrinsic toxicity due to components released into the mouth during the early stage of polymerization, and afterward as a result of erosion or material decomposition. In addition, resin-based dental composites have potential environmental pollutant by elution of monomers and degradation. Since certain monomers of resin matrices are synthesized from bisphenol A (BPA), which acts as an estrogenic endocrine disruptor, these resin matrices may have estrogenic activity. Therefore, the estrogenic endocrine-disrupting activity of various dental composites should be evaluated. In this study, we evaluated the estrogenic endocrine-disrupting activity of 10 resin composites by using a BRET-based estrogen receptor (ER)α and ERβ dimerization assays and ER transactivation assay. BPA, BisDMA, BisGMA, BisEMA, TEGDMA, HMBP, and DMPA mediated ERα dimerization, and BPA, BisDMA, and DMPA also mediated ERβ dimerization. Except for UDMA and CQ, all the compounds were identified as estrogen agonists or antagonists. In-depth information for the safe use of dental composites was acquired, and it was confirmed how the component of dental composites acts in the ER signaling pathway. Further studies on the low-dose and long-term release of these compounds are needed to ensure the safe use of these resin-based dental composites. [Display omitted] • A comprehensive study into the estrogen signaling pathway of resin composites. • Resin composite components have the potential to disrupt the estrogenic system. • Most of the resin composite components induced estrogen receptor dimerization. • Most of the resin composite components affected estrogen receptor transactivation. [ABSTRACT FROM AUTHOR]

Despite Bisphenol-A (BPA) being subject to extensive study, a thorough understanding of molecular mechanism remains elusive. Here we show that using weighted gene correlation network analysis (WGCNA), which takes advantage of a graph theoretical approach to understanding correlations amongst genes and grouping genes into modules that typically have co-ordinated biological functions and regulatory mechanisms, that despite some commonality in altered genes, there is minimal overlap between BPA and estrogen in terms of network topology. We confirmed previous findings that ZNF217 and TFAP2C are involved in the estrogen pathway, and are implicated in BPA as well, although for BPA they appear to be active in the absence of canonical estrogen-receptor driven gene expression. Furthermore, our study suggested that PADI4 and RACK7/ZMYNDB8 may be involved in the overlap in gene expression between estradiol and BPA. Lastly, we demonstrated that even at low doses there are unique transcription factors that appear to be driving the biology of BPA, such as SREBF1. Overall, our data is consistent with other reports that BPA leads to subtle gene changes rather than profound aberrations of a conserved estrogen signaling (or other) pathways.

Understanding estrogen contamination characteristics of organic fertilizers derived from livestock manures is important for organic fertilizer production and contamination control. In the present study, 17 organic fertilizers derived from chicken, swine, and cow manures were collected from a market. Organic matter, nitrogen, phosphorus, and potassium contents of organic fertilizers were determined. The concentration of estriol（E3）, 17β-estradiol（17β-E2）, bisphenol A（BPA）, and ethinyl estradiol（EE2）were detected by SPE-HPLC/FLD. At the same time, we compared the estrogen concentrations in organic fertilizers with fresh livestock manures. Results showed that organic fertilizers with low organic matter were detected in the organic fertilizer samples derived from chicken, swine, and cow manures. The average value（dry weight）of E3, 17β-E2, and BPA in organic fertilizers derived from chicken manures were 270.22, 2.71 μg·kg-1, and 87.93 μg· kg-1, respectively. The average values of E3, 17β-E2, BPA, and EE2 in organic fertilizers derived from swine manures were 609.71, 41.84, 32.69 μg·kg-1, and 46.74 μg·kg-1, respectively. The average value of E3 in organic fertilizers derived from cow manures was 50.96 μg· kg-1. Organic fertilizers had a large risk of estrogen contamination, and the risk of E3 contamination was higher than that of the other estro⁃ gens. Additionally, the concentration and detection rates of estrogen in cow organic fertilizers were lower and safer. The detection rates of E3, 17β-E2, BPA, and EE2 in organic fertilizers were 47.06%, 11.76%, 17.65%, and 5.88%, respectively, while detection rates were 67.74%, 70.97%, 70.97%, and 58.06% in fresh livestock manures, respectively. The detection rates of estrogen in organic fertilizers were lower than that of fresh livestock manures. The conclusion is that there are estrogen residues in organic fertilizers derived from livestock manures, which needs further attention. [ABSTRACT FROM AUTHOR]

Traditional approaches for chemical risk assessment cannot keep pace with the number of substances requiring assessment. Thus, in a global effort to expedite and modernize chemical risk assessment, New Approach Methodologies (NAMs) are being explored and developed. Included in this effort is the OECD Integrated Approaches for Testing and Assessment (IATA) program, which provides a forum for OECD member countries to develop and present case studies illustrating the application of NAM in various risk assessment contexts. Here, we present an IATA case study for the prediction of estrogenic potential of three target phenols: 4-tert-butylphenol, 2,4-di-tert-butylphenol and octabenzone. Key features of this IATA include the use of two computational approaches for analogue selection for read-across, data collected from traditional and NAM sources, and a workflow to generate predictions regarding the targets' ability to bind the estrogen receptor (ER). Endocrine disruption can occur when a chemical substance mimics the activity of natural estrogen by binding to the ER and, if potency and exposure are sufficient, alters the function of the endocrine system to cause adverse effects. The data indicated that of the three target substances that were considered herein, 4-tert-butylphenol is a potential endocrine disruptor. Further, this IATA illustrates that the NAM approach explored is health protective when compared to in vivo endpoints traditionally used for human health risk assessment. • The use of New Approach Methodologies (NAMs) in chemical risk assessment is explored. • A workflow for the prediction of estrogen receptor activation by substituted phenols. • The workflow is applied to: 4-tert-butylphenol, 2,4-di-tert-butylphenol, octabenzone. • Two computational approaches for analogue selection were used and evaluated. • When compared to in vivo endpoints, the NAM approach was health protective. [ABSTRACT FROM AUTHOR]

In this study mouse Leydig cell (MA-10) were treated with G-protein coupled membrane estrogen receptor antagonist (G-15; 10 nM). Cells were analyzed by Western blotting for expression of estrogen-related receptors (ERRα, β and γ), steroidogenic markers (lutropin receptor; LHR and 3β-hydroxysteroid dehydrogenase; 3β-HSD) and lipid droplet markers (perilipin; PLIN and microtubule-associated protein 1 A/1B-light chain 3; LC3). Concomitantly, microscopic analyses by light microscope (immunofluorescent staining for lipid droplets, PLIN and LC3) as well as by electron microscope (for lipid droplet ultrastructure) were utilized. For analysis of cholesterol content, cAMP level and progesterone secretion, G-15, estrogen receptor (ER) antagonist (ICI 182,780; 10 μM), 17β-estradiol (10 mM) and, bisphenol A (BPA; 10 nM) were used alone or in combinations. We revealed no changes in ERRs expression but alterations in ERRβ and γ localization in G-15-treated cells when compared to control. Partial translocation of ERRβ and γ from the cell nucleus to cytoplasm was observed. Decreased expression of LHR, 3β-HSD, PLIN and LC3 was detected. Moreover, in treated cells large lipid droplets and differences in their distribution were found. Very strong signal of co-localization for PLIN and LC3 was found in treated cells when compared to control. In ultrastructure of treated cells, degenerating lipid droplets and double membrane indicating on presence of lipophagosome were observed. We found, that only (i) BPA and G-15 did not effect on cholesterol content, (ii) BPA, G-15 and ICI did not effect on cAMP level and (iii) BPA, ICI alone and in combination, and BPA with G-15 did not modulate progesterone secretion. These findings showed complex and diverse estrogen effects on mouse Leydig cells at various steps of steroid hormone production (cholesterol storage, release and processing). Lipid homeostasis and metabolism in these cells were affected by endogenous and exogenous estrogen, interactions of receptors (GPER, ER and ERR) and GPER and ER antagonists. [ABSTRACT FROM AUTHOR]

• Bisphenol A and 17-alpha ethinylestradiol induce trascriptional alterations in the medaka gill. • Early life exposure led to sexually dimorphic alterations in expression patterns of genes involved in osmoregulation. • Embryonic exposure led to transcriptional alterations in the gills four generations later. Embryonic bisphenol A (BPA) and 17α-ethinylestradiol (EE2) exposure can have far reaching health effects in fish, including adult onset transgenerational reproductive abnormalities, anxiety, and cardiac disorders. It is unknown whether these two environmental estrogens can induce transgenerational abnormalities in the gill. The present study examined transgenerational effects of BPA or EE2 exposure on genes that are critical for osmoregulation in fish. Medaka (Oryzias latipes) embryos were exposed to either BPA (100 μg/L) or EE2 (0.05 μg/L) for the first 7 days of embryonic development and never thereafter for the remainder of that generation (F0) and in subsequent generations of this study (F1, F2, and F3). Expression of osmoregulatory genes (NKAα1a , NKAα1b , NKAα1c , NKAα3a , NKAα3b , NKCC1a, and CFTR) were examined in gills of the first-generation (F0) adults which were directly exposed as embryo and in the fourth-generation adults (F3), which were never exposed to either of these environmental estrogens. Significant alterations in expression of osmoregulatory genes were observed in both F0 and F3 generations. Within the F0 generation, a sex-specific expression pattern was observed with a downregulation of osmoregulatory genes in males and an upregulation of osmoregulatory genes in females. At the F3 generation, this pattern reversed with the majority of the osmoregulatory genes upregulated in males and downregulated in females, suggesting that exposure to BPA and EE2 during embryonic development induced transgenerational impairment in molecular events associated with osmoregulatory functions in subsequent generations. These adverse outcomes may have impacts on physiological functions related to osmoregulation of fish inhabiting contaminated aquatic environments. [ABSTRACT FROM AUTHOR]

Abstract: This study determined and evaluated the estrogenic potentials on hepatic vitellogenin gene (vtg1) of adult male zebrafish which were exposed to low level concentrations (ng/L-μg/L levels) of individual and binary mixtures of 17β-estradiol (E 2), bisphenol A (BPA) and nonylphenol (NP) through the use of reduced life expectancy (RLE) model and molecular docking analysis. The order of in vivo estrogenic potentials of individual and binary exposure of E 2 , BPA and NP was as follows: E 2 +BPA＞E 2 ＞E 2 +NP＞BPA＞BPA + NP >>>NP. Binary mixtures of E 2 , BPA and NP had weak synergistic effects under the exposure concentration ranges. With the expression of hepatic vtg1 gene, the hepatic toxicity was analyzed by using the RLE model. All plots of the linear RLE model had negative slopes indicating that EC 50 was negatively correlated with the natural logarithm of exposure time (ln ET 50). The RLE model analyses can be useful to evaluate the exposure time effects of zebrafish by using EC 50 as toxicity endpoint. Molecular docking analysis revealed that the interaction potential of E 2 (Binding energy: −10.1 kcal/mol) for the zebrafish ERα-LBD was the most potent (stable), followed by BPA (−8.0 kcal/mol) and NP (−6.8 kcal/mol). Molecular docking analysis can be useful to understand interactive effects of E 2 , BPA and NP. Graphical abstract Image 10 Highlights • Binary mixtures of E 2 , BPA and NP have weak synergistic effects on hepatic vtg1 gene of zebrafish. • The RLE model for lethal toxicity was extended to study the effect of exposure time on EC 50 of vtg1 levels. • A new interpretation for interaction potential of binary mixtures of E 2 , BPA and NP was given through molecular docking. [ABSTRACT FROM AUTHOR]

Abstract Information on the occurrence and endocrine potencies of analogues of bisphenol A (BPA) and diglycidyl ester derivatives (BDGEs) of BPA and BPF is limited. Such information is, however, important as the current debate on BPA and the lowered BPA migration limit in Europe may provide an incentive for application of structural analogues. A new sensitive multi-analyte LC-ESI-MS/MS method was developed to measure 17 bisphenols (BPs) and 6 BDGEs in food, beverages and drinkware. Yeast based bioassays were used to determine the in vitro (anti)estrogenic and (anti)androgenic properties of these and 7 additional BPs and BDGEs. Drinkware of polycarbonate and other materials were analysed for BPs and BDGEs. Only BPA and BPS and both at trace levels were found in a few containers. A limited number of (canned) foods and beverages were also analysed. BPA was the most frequently detected BP (ranged from 0.03 ng mL−1 in a beverage sample to 68 ng g−1 in food). Other BPs detected were BPS, 2,2-BPF and 4,4-BPF. In addition BADGE, BADGE.HCl, BADGE.H 2 O and BADGE.2H 2 O were detected from 0.08 ng mL−1 in a beverage sample to 3.3 ng g−1 in food. In vitro testing showed that most BPs exhibited an equal or higher estrogenic potency than BPA and most of them also showed a higher anti-androgenic potency, i.e. BPB, BPCl, BPC, BPE, 4,4-BPF, BPP, BPAF, and BPTMC. Some BPs and BDGEs were not estrogenic, but showed an anti-estrogenic effect and were anti-androgenic too. BPS was only weakly estrogenic and BADGE.2H 2 O and BFDGE.2H 2 O showed no in vitro activity. The present data show that in addition to BPA, other BPs and BDGEs can be present in food and drinks, some displaying in vitro endocrine activities. Graphical abstract Image 1 Highlights • A multi-analyte LC-MS/MS method including 23 BPA and BADGE analogues. • A high method sensitivity by electrospray ionisation under alkaline conditions. • In-vitro (anti)estrogenic and (anti)androgenic potencies for 30 BPA/BADGE analogues. [ABSTRACT FROM AUTHOR]

Abstract Paperboard used as packaging, a non-inert material, can transfer chemicals into food. Over the years, endocrine disrupting compounds (EDCs), such as NonylPhenols (NPs), BisPhenol A (BPA) and phthalates have been shown to migrate from packaging materials into food. Due to chronic exposure and mixture effects of these EDCs, they could cause health effects even at very low doses. Many EDCs are still unknown and many more are still unregulated. The ERE-CALUX bioassay was used as a bioanalytical tool to investigate estrogenic activities of paperboard food packaging and its characteristics, including recycling rate and printing ink. A "worst case" scenario with full extraction is compared to a dry food migration experiment. By measuring an overall estrogenic activity, known and unknown estrogenic chemicals and mixture effects are taken into account and the data are compared to molecule specific analysis. Estrogenic activities ranged from 682 ± 66 pg E2 eq./dm2 to 3250 ± 400 pg E2 eq./dm2 for "worst case" extraction and from 347 ± 30 pg E2 eq./dm2 to 1350 ± 70 pg E2 eq./dm2 for migration experiments. A two-factor ANOVA revealed a relationship between estrogenic activity and the recycling rate of the paperboard, but no significant difference with printing ink was observed for these paperboard samples. Bis(2-ethylhexyl)phthalate (DEHP), dibutyl phthalate (DBP), butyl benzyl phthalate (BBP) and 1,2-cyclohexane dicarboxylic acid diisononyl ester (DINCH) were determined in all extraction and migrations experiment samples. A Spearman rank correlation analysis showed a relationship between the estrogenic activity and the total phthalates as well as with each compound individually. Highlights • Estrogenic activities of paperboard in "worst case" extractions and in migrations. • Activities have relative standard deviations below 18% with a median value of 9.7%. • Recycling rate of paperboard food packaging highly related to estrogenic activity. • Estrogenic activities are compared to molecule specific analysis of plasticizers. • A relationship was shown between the estrogenic activity and the phthalates. [ABSTRACT FROM AUTHOR]

Abstract Exposure to mixtures of endocrine disrupting compounds (EDCs) has been hypothesized to produce potential synergistic or antagonistic effects that can cause undesired effects that are not reflected by the individual compounds. In this study, the estrogenic activities of 11 EDCs of global environmental concern were systematically investigated using the yeast estrogen screen (YES). The contribution of the individual chemical to the total endocrine activity of environmentally relevant mixtures was evaluated. Compared to 17β-estradiol (E2) as a standard, estrone (E1), estriol (E3), ethinyl estradiol (EE2), bisphenol-A (BPA), and genistein (GEN) showed estrogenic effects, while dibutyl phthalate (DBP), n -butyl benzyl phthalate (BBP), Bis(2-ethylhexyl) phthalate (DEHP), nonyl phenol (NP) and 4- tert -octyl phenol (OP) showed anti-estrogenic effects. The 11 EDCs mixture at a constant environmentally relevant ratio also showed estrogenic activity. The mixtures data were fit to concentration addition (CA), response addition (RA) and interaction (IR) models, respectively. The IR model was not statistically different from the observed value and better predicted results than the CA model for mixtures of all 11 compounds. For the mixtures with the 6 estrogenic compounds only, additive effects were observed, and the data were well predicted by the CA and IR models. Further, in the 11 EDCs mixture the presence of EE2 at an environmentally relevant concentration did not increase the estrogenic activity as compared to a 10 EDCs mixture without EE2. Highlights • Additive, not synergistic, effects were observed for mixtures of estrogenic EDCs. • The Interaction Response (IR) model yielded a better prediction than the Concentration Addition (CA) model. • The IR model has the advantage in addressing interaction of mixtures of EDCs. • Contribution of EE2 to the mixture effects was clarified. [ABSTRACT FROM AUTHOR]

Abstract The study aimed to assess the occurrence and partitioning of estrogens (ß-estradiol, E2; estrone, E1; 17α-ethinylestradiol, EE2), phenolic compounds (nonylphenol, NP; octylphenol, OP; bisphenol A, BPA) and perfluorinated compounds (perfluorooctanoic acid, PFOA; perfluorooctane sulfonate, PFOS) in waters and sediments of the main rivers and the coastal lagoon of Pialassa Baiona, in the north-eastern part of Italy. Overall, water samples showed a wide distribution of the three classes of compounds in the study area, with the exception of estrogens which recorded only some local detections, at concentrations <10 ng/L. Sediments were found to be affected only by contamination of phenolic chemicals, namely BPA and NP, whereas perfluorinated compounds were almost undetected (

The occurrence of the endocrine disruptors bisphenol-A (BPA) and 17β-estradiol (E 2 ) in cultured populations of green mussel ( Perna viridis ) and water samples collected from selected fresh and marine coastal environments along the eastern coast of Thailand were investigated. Analysis found high levels of BPA in mussel tissues that correlated with levels found in coastal (maximum 37.13 ng/L) and freshwater (50.7 ng/L) sites situated near industrial and densely populated areas. By comparison, high levels of E 2 (62.99 ± 5.03 ng/L) were found in freshwater sites near to urban areas. Higher concentrations of BPA and E 2 were found in mature green mussels (>6 mo. old; 6.40 ± 0.52 cm shell length) than levels determined in juveniles (<2–3 mo. old; 2.29 ± 0.65 cm shell length). To evaluate the potential risks associated with the consumption of green mussels, the bioconcentration factor (BCF) for BPA was determined to be 1650 for adult bivalves and 283 for juveniles. As P. viridis can accumulate BPA from the environment, this raises concerns regarding the risks posed by consuming seafood sourced from zones near to major conurbations. Whether the high levels of E2 found in green mussels is due to accumulation or to de novo synthesis as seen in other molluscs, requires further investigation. While industrial and domestic wastewater may be important sources of BPA, E 2 contamination within the eastern part of the Gulf of Thailand has been linked to domestic waste. The study highlights the importance of the temporal and spatial monitoring of sentinel species, such green mussels, for environmental contaminants, the results of which can lead to the construction of regional risk maps helping to inform national strategies regarding aquaculture zoning and aquatic food safety. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRs) are small molecules important for regulation of transcription and translation. The objective was to identify hormonally regulated miRs in human endometrial stromal cells and to determine the impact of the endocrine disruptor, bisphenol A (BPA), on those miRs. miR microarray analysis and multiple confirmatory cell preparations treated with 17ß-estradiol (E2) and BPA altered miR-27b, let-7c, let-7e and miR-181b. Further, decidualization downregulated miR-27b. VEGFB and VEGFC were validated as targets of miR-27b. Identification of miR-27b target genes suggests that BPA and E2 downregulate miR-27b thereby leading to upregulation of genes important for vascularization and angiogenesis of the endometrium during the menstrual cycle and decidualization. [ABSTRACT FROM AUTHOR]

Despite Bisphenol-A (BPA) being subject to extensive study, a thorough understanding of molecular mechanism remains elusive. Here we show that using weighted gene correlation network analysis (WGCNA), which takes advantage of a graph theoretical approach to understanding correlations amongst genes and grouping genes into modules that typically have co-ordinated biological functions and regulatory mechanisms, that despite some commonality in altered genes, there is minimal overlap between BPA and estrogen in terms of network topology. We confirmed previous findings that ZNF217 and TFAP2C are involved in the estrogen pathway, and are implicated in BPA as well, although for BPA they appear to be active in the absence of canonical estrogen-receptor driven gene expression. Furthermore, our study suggested that PADI4 and RACK7/ZMYNDB8 may be involved in the overlap in gene expression between estradiol and BPA. Lastly, we demonstrated that even at low doses there are unique transcription factors that appear to be driving the biology of BPA, such as SREBF1. Overall, our data is consistent with other reports that BPA leads to subtle gene changes rather than profound aberrations of a conserved estrogen signaling (or other) pathways. [ABSTRACT FROM AUTHOR]

Abstract Proper cyclicity is essential to reach successful optimal fertility. In rats and mice, BPA exposure is repeatedly and reliably reported to show an adverse effect on the estrous cycle after exposures at different life stages. In humans, a possible association between modifications of menstrual cycle characteristics (e.g. length of the cycle, duration of menstrual bleeding) and sub-fecundity or spontaneous abortion has been observed. Alterations of ovarian cyclicity can therefore be definitely considered as an adverse health outcome. As a prerequisite for the EU REACH regulation to identify a substance as an endocrine disruptor and a SVHC, 1 the proof has to be established that the substance can have deleterious health effects resulting from an endocrine mode of action. This review provides an overview of the currently available data allowing to conclude that the adverse effects of BPA exposure on ovarian cyclicity is mediated by an endocrine mode of action. Highlights • BPA alters ovarian steroidogenesis in adult females via aromatase reduction. • BPA fetal exposure is associated to altered estrous cyclicity. • BPA fetal exposure alters neuroendocrine functions critical for female reproduction. [ABSTRACT FROM AUTHOR]

Abstract The extensive database on BPA provides strong evidence of its adverse effects on reproductive, neurobehavioural, metabolic functions and mammary gland. Disruption of estrogenic pathway is central in the mediation of these effects although other modes of action may be involved. BPA has a weak affinity for ERα/β but interaction with extranuclearly located pathways activated by estrogens such as ERRγ and GPER reveals how BPA can act at low doses. The effects are observed later in life after developmental exposure and are associated with pathologies of major societal concern in terms of severity, incidence, impact on quality of life, burden on public health system. The complexity of the dose response raise uncertainties on the possibility to establish safe levels and the scope of ED-mediated effects of BPA may be wider. These concerns fulfill the requirements for ED identification under REACH regulation. Highlights • BPA may alter reproductive, neurobehavioural, metabolic functions and mammary gland. • Disruption of estrogenic pathway is central in mediation of these effects. • Involved pathologies are of major societal and public health concern. • There are important uncertainties on dose-response and scope of BPA's ED effects. • These concerns justify application of a stringent regulation. [ABSTRACT FROM AUTHOR]