In comparison with analytical tools, bioassays provide higher sensitivity and more complex evaluation of environmental samples and are indispensable tools for monitoring increasing in anthropogenic ...pollution. Nevertheless, the disadvantage in cellular assays stems from the material variability used within the assays, and an interlaboratory adaptation does not usually lead to satisfactory test sensitivities. The aim of this study was to evaluate the influence of material variability on CXCL12 secretion by T47D cells, the outcome of the CXCL‐test (estrogenic activity assay). For this purpose, the cell line sources, sera suppliers, experimental and seeding media, and the amount of cell/well were tested. The multivariable linear model (MLM), employed as an innovative approach in this field for parameter evaluation, identified that all the tested parameters had significant effects. Knowledge of the contributions of each parameter has permitted step‐by‐step optimization. The most beneficial approach was seeding 20,000 cells/well directly in treatment medium and using DMEM for the treatment. Great differences in both basal and maximal cytokine secretions among the three tested cell lines and different impacts of each serum were also observed. Altogether, both these biologically based and highly variable inputs were additionally assessed by MLM and a subsequent two‐step evaluation, which revealed a lower variability and satisfactory reproducibility of the test. This analysis showed that not only parameter and procedure optimization but also the evaluation methodology must be considered from the perspective of interlaboratory method adaptation. This overall methodology could be applied to all bioanalytical methods for fast multiparameter and accurate analysis.
The CXCL‐test is a sensitive in vitro assay developed in T47D cell line to identify estrogenic properties of anthropogenic chemicals and to monitor pollution caused by endocrine disruptors. As the test sensitivity can be modified by several factors, multivariable linear model was employed as an innovative statistical approach for fast evaluation of contributing factors. It revealed that cell lines, media, sera, cell number, and cell density significantly affected CXCL12 secretion. This knowledge enables new interpretation of the test results.
Endocrine-disrupting compounds (EDCs) are chemical compounds with the ability to alter the hormonal systems of organisms. Such compounds are used in several industrial and domestic activities and ...reach the aquatic environment via wastewater discharge. The aim of this study is to assess the occurrence of 30 EDCs and related compounds in the surface waters of central Spain and to determine the overall estrogenic activity of environmental samples. This study analyzed a large number of EDCs and other emergent or suspected compounds with endocrine-disrupting activity. The results have shown the presence of 19 EDCs at concentrations ranging from 2 to 5928ngL−1. Organophosphorus-based flame retardants, alkylphenolic compounds and anticorrosives were found at the highest concentrations. Furthermore, although insufficient data are available to calculate an average over time, these preliminary results show the need to monitor the waters in both rivers studied. Alkylphenolic compounds, particularly nonylphenol, were the main contributors to overall estrogenicity. A higher concentration of the compounds studied was detected in the river Jarama, although the estrogenicity expressed as estradiol equivalents (EEQs) was higher in the river Manzanares due to a higher concentration of nonylphenol. However, the total estrogenicity did not exceed 1ngL−1 (EEQ), which is the level that may cause estrogenic effects in aquatic organisms, in any of the samples. In conclusion, the potential estrogenic risk in both rivers is low, although organophosphorus-based flame retardants may increase this risk as they were found at high levels in all samples. Unfortunately, these compounds could not be taken into account when calculating the estrogenic activity due to the lack of activity data for them. For future investigations, it will be important to assess the estrogenicity provided by these flame retardants. Due to the significant concentrations of EDCs detected in both rivers, further studies in this region are required.
•Our work analyzes a total of 30 EDCs and others emergent or suspected EDCs.•Flame retardants, alkylphenolics and anticorrosives are the most abundant compounds.•NP and OP were detected at higher concentrations than the annual average EQS.•Alkylphenolics are the major estrogenicity contributors in all samples.•None of the samples exceeded the EEQ 1ng/L with estrogenic effects on organisms.
Danshen Si Wu is a Traditional Chinese Medicine used for menopausal complains. Beside tanshinone IIA (Tan IIA), Danshen also contains tanshinone I (Tan I), cryptotanshinone (CT) and dihydrotanshinone ...(DT). The aim of this study was to compare the biological activity of these tanshinones and to determine their cytotoxicity and genotoxicity. Purities and stabilities of the substances were analyzed by LC-DAD and LC-MS analyses. DT and CT concentrations decreased rapidly in dimethylsulfoxide and were converted to Tan I and Tan IIA, respectively. In aqueous solution concentration of all tanshinones decreased after 24 h. Tan I and Tan IIA showed dose-dependent bioactivity mediated by ERα and ERβ. No cytotoxic and genotoxic effects for Tan I and Tan IIA were detected. In a yeast transactivation assay Tan I and Tan IIA showed antiandrogenic activity. A significant anabolic activity in C2C12 cells could be detected for Tan I and Tan IIA. In conclusion our data provide evidence that Tan I and Tan IIA are the most relevant bioactive tanshinones in Danshen. Our finding that all tanshinones display a certain instability in aqueous solutions is relevant when discussing their potential therapeutic benefits in humans.
•Dihydrotanshinone and cryptotanshinone is converted to tanshinone I and tanshinone IIA, respectively, in DMSO solutions.•In aqueous solutions all four tanshinones seem to show a certain instability.•No cytotoxic and genotoxic effects for tanshinone I and tanshinone IIA were detected.•Tanshinone I and tanshinone IIA showed dose-dependent bioactivity mediated by Estrogen receptor α and Estrogen receptor β.•Tanshinone I and tanshinone IIA showed antiandrogenic and anabolic activity.
As a recognized endocrine disruptor in the environment targeting estrogen receptors (ERs), Bisphenol A (BPA) and its bisphenol S (BPS) analogs are involved in the development of breast cancer. ...Epigenetic modifications are crucial in many biological processes, and DNA hydroxymethylation (DNAhm) coupled with histone methylation is implicated in epigenetic machinery covering cancer occurrence. Our previous study indicated that BPA/BPS induces breast cancer cell (BCC) proliferation with enhanced estrogenic transcriptional activity and causes the change of DNAhm depending on ten-eleven translocation 2 (TET2) dioxygenase. Herein, we investigated the interplay of KDM2A-mediated histone demethylation with ER-dependent estrogenic activity (EA) and identified their function in DNAhm catalyzed by TET2 for ER-positive (ER+) BCC proliferation induced by BPA/BPS. We found that BPA/BPS-treated ER+ BCCs presented increased KDM2A mRNA and protein levels but reduced TET2 and genomic DNAhm. Furthermore, KDM2A promoted H3K36me2 loss and suppressed TET2-dependent DNAhm by reducing its chromatin binding during BPA/BPS-induced cell proliferation. Results of Co-IP & ChIP assays suggested the direct interplay of KDM2A with ERα in multiple manners. KDM2A reduced the lysine methylation of ERα protein to increase its phosphorylated activation. On the other hand, ERα did not affect KDM2A expression, while KDM2A protein levels decreased after ERα deletion, indicating that ERα binding might maintain KDM2A protein stability. In conclusion, a potential feedback circuit of KDM2A/ERα-TET2-DNAhm was identified among ER+ BCCs with significant effects on regulating BPA/BPS-induced cell proliferation. These insights advanced the understanding of the relationship between histone methylation, DNAhm, and cancer cell proliferation with EA attributed to BPA/BPS exposure in the environment.
•KDM2A interplays with ERα in proliferation of BPA/BPS-treated ER+ breast cancer cells.•KDM2A diminishes DNA hydroxymethylation in BPA/BPS-treated ER+ breast cancer cells.•KDM2A-mediated H3K36 demethylation represses TET2-catalyzed DNA hydroxymethylation.•KDM2A mediates demethylation of ERα to promote its activation upon BPA/BPS exposure.•ERα maintains KDM2A protein stability to increase its expression upon BPA/BPS exposure.
There are many reports on the detection and removal of emerging pollutants in the wastewater effluents, while the fate of their chlorinated derivatives generated during chlorination is not well ...understood. Here we investigated the photodegradation of chlorinated derivatives of bisphenol A (CDBPAs), mainly including 3-chlorobisphenol A, 3,3′-dichlorobisphenol A, 3,5-dichlorobisphenol A, 3,3′,5-trichlorobisphenol A, and 3,3′,5,5′-tetrachlorobisphenol A, under simulated sunlight. Distinct from BPA, CDBPAs underwent rapid direct photodegradation due to a pronounced bathochromic shift of UV absorption. The photodegradation of CDBPAs was significantly enhanced by effluent organic matter (EfOM) from the wastewater effluent. A series of quenching experiments and laser flash photolysis analysis verified the contribution of triplet states of EfOM (3EfOM∗) for the indirect photodegradation of CDBPAs with rate constant of ∼109 M−1 s−1. Both direct and EfOM-induced indirect photodegradation of CDBPAs increased with a higher degree of chlorination. Furthermore, high-resolution mass spectrometry showed similar photoproducts for direct and indirect photodegradation of CDBPAs, mainly ascribed to the cleavage of C–Cl bond and hydroxylation with further cleavage of the benzene ring. The estrogenic activity of the photoproducts was diminished. These findings suggest that photodegradation is an important pathway for the removal and detoxication of CDBPAs from effluents and receiving natural waters under sunlight.
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•CDBPAs underwent rapid direct photodegradation under sunlight.•3EfOM∗ accounted for the photosensitized degradation of CDBPAs.•Photolysis of CDBPAs increased with a higher degree of chlorination.•Cleavage of C–Cl bond and hydroxylation were major photolysis processes.•The estrogenic activity of the photoproducts was diminished.
Mycotoxins, including zearalenone (ZEA), can occur worldwide in cereals. They can enter the food chain and cause several health disorders. ZEA and its derivatives (α-zearalenol, α-ZOL and ...β-zearalenol, β-ZOL) have structural analogy to estrogen, thus they can bind to estrogen receptors (ERs). In order to characterize the estrogenic activity of ZEA, α-ZOL and β-ZOL, the proliferation of ER-positive human breast cancer cells (MCF-7) exposed to these mycotoxins was measured. After exposure at levels ranging from 6.25 to 25 µM, cell proliferation was evaluated by using the E-Screen bioassay. In accordance with previous studies, our results show the estrogenic activity of ZEA, α-ZOL and β-ZOL in MCF-7 cells. This effect is related to ZEA and its metabolites being flexible enough to bind to mammalian ERs. The relative proliferative effect (RPE) ranged from 10% to 91%. The α-ZOL induced the highest proliferative effect due to its higher affinity for the ERs compared to the other mycotoxins.
Humans are simultaneously exposed to complex chemical mixtures, and its combined effect can affect human health. As part of the HBM4EU project, the actual mixture of perfluoroalkylated substances ...(PFAS) in 25 human placenta samples was extracted by chromatographic methods and assessed for xeno-estrogenic activity using two in-vitro bioassays: the estrogen receptor transactivity and the E-Screen assay. Most of the PFAS extracts displayed xeno-estrogenic activity, in one or both assays. The xeno-estrogenic activities in the two bioassays were not correlated, but both assays showed an overall negative correlation with placenta concentrations of single PFAS. Xeno-estrogenic activities were significantly related to maternal characteristics; being higher in young, smokers and primiparous women, but not with fetal growth (birth weight, birth length, head circumference, gestational age, placenta weight). The presented extraction method can be used to study the combined effect of real-life mixtures of PFAS in relation to health outcomes in large-scale human biomonitoring studies.
•Mixture of perfluoroalkylated substances (PFAS) were extracted from human placentas.•Xeno-estrogenic activities of the extracts were assessed in two in-vitro bioassays.•Most of the PFAS extracts displayed xeno-estrogenic activity, in one or both assays.•Xeno-estrogenic activities of PFAS were related to maternal age, smoking and parity.•PFAS induced xeno-estrogenic activities were not associated with fetal growth.
Cancer arises from multiple genetic errors occurring in a single stem cell (clonality). Every time DNA replicates, mistakes occur. Thus, agents can increase the risk of cancer either by directly ...damaging DNA (DNA-reactive carcinogens) or increasing the number of DNA replications (increased cell proliferation). Increased cell proliferation can be achieved either by direct mitogenesis or cytotoxicity with regenerative proliferation. Human carcinogens have a mode of action of DNA reactivity, immunomodulation (mostly immunosuppression), increased estrogenic activity (mitogenesis), or cytotoxicity and regeneration. By focusing on screening for these four effects utilizing in silico , in vitro , and short-term in vivo assays, a biologically based screening for human chemical carcinogens can be accomplished with greater predictivity than the traditional 2-year bioassay with considerably less cost, less time, and the use of fewer animals.
This article reviews different photodegradation technologies used for the removal of four endocrine disrupting chemicals (EDCs): estrone (E1), 17β-estradiol (E2), estriol (E3) and ...17α-ethinylestradiol (EE2). The degradation efficiency is greater under UV than visible light; and increases with light intensity up to when mass transfer becomes the rate limiting step. Substantial rates are observed in the environmentally relevant range of pH7–8, though higher rates are obtained for pH above the pKa (~10.4) of the EDCs. The effects of dissolved organic matter (DOM) on EDC photodegradation are complex with both positive and negative impacts being reported. TiO2 remains the best catalyst due to its superior activity, chemical and photo stability, cheap commercial availability, capacity to function at ambient conditions and low toxicity. The optimum TiO2 loading is 0.05–1gl−1, while higher loadings have negative impact on EDC removal. The suspended catalysts prove to be more efficient in photocatalysis compared to the immobilised catalysts, while the latter are considered more suitable for commercial scale applications. Photodegradation mostly follows 1st or pseudo 1st order kinetics. Photodegradation typically eradicates or moderates estrogenic activity, though some intermediates are found to exhibit higher estrogenicity than the parent EDCs; the persistence of estrogenic activity is mainly attributed to the presence of the phenolic moiety in intermediates.
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•Photodegradation of E1, E2, E3 and EE2 follows 1st or pseudo 1st order kinetics.•Fastest removal rates occur at pH7–8, for a range relevant to natural environment.•The optimum TiO2 loadings are often between 0.05 and 1gl−1.•Estrogenicity persists if the intermediates possess the phenolic moiety.•Both positive and negative impacts from DOM on photodegradation are reported.
