Bioanalytical and instrumental analysis of thyroid hormone disrupting compounds in water sources along the Yangtze River

Thyroid hormone (TH) agonist and antagonist activities of water sources along the Yangtze River in China were surveyed by a green monkey kidney fibroblast (CV-1) cell-based TH reporter gene assay. Instrumental analysis was conducted to identify the responsible thyroid-active compounds. Instrumentally derived l-3,5,3′-triiodothyronine (T₃) equivalents (T₃-EQs) and thyroid receptor (TR) antagonist activity equivalents referring to dibutyl phthalate (DBP-EQs) were calculated from the concentrations of individual congeners. The reporter gene assay demonstrated that three out of eleven water sources contained TR agonist activity equivalents (TR-EQs), ranging from 286 to 293 ng T₃/L. Anti-thyroid hormone activities were found in all water sources with the TR antagonist activity equivalents referring to DBP (Ant-TR-EQs), ranging from 51.5 to 555.3 μg/L. Comparisons of the equivalents from instrumental and biological assays suggested that high concentrations of DBP and di-2-ethylhexyl phthalate (DEHP) were responsible for the observed TR antagonist activities at some locations along the Yangtze River.     

Flow‐Biology Relationships Based on Fish Habitat Guilds in North Carolina

The health of freshwater biota is dependent on streamflow, yet identification of the flow regimes required to maintain ecological integrity remains challenging to states in the United States seeking to establish ecological flows. We tested the relationship between decreases in streamflow and Shannon‐Weaver diversity index of fish species for four flow‐based habitat guilds: riffle, riffle‐run, pool‐run, and pool in North Carolina. We found species that prefer shallow habitats, such as riffles and riffle‐runs were the most sensitive to decreases in streamflow; whereas no significant relationships were found for pool or pool‐run species. The sensitivity to decreases in streamflow was greatest during summer and fall, when streams are naturally lower. When all fish habitat guilds were included in the assessment of flow‐biology relationships, there were no significant relationships to decreases in streamflow. As the sensitivity of fish to reductions in streamflow is not constant across habitat guilds, combining all fish species together for flow‐biology analyses may greatly underestimate the response of fish species to decreases in flow and should be acknowledged when establishing ecological flows.     

Integral assessment of estrogenic potentials in sediment-associated samples

Background, aim, and scope  The enzyme-linked receptor assay (ELRA) detects estrogenic and anti-estrogenic effects at the molecular level of receptor binding and is a useful tool for the integrative assessment of ecotoxicological potentials caused by hormonally active agents (HAA) and endocrine disrupting compounds (EDC). The main advantage of the ELRA is its high sample throughput and its robustness against cytotoxicity and microbial contamination. After a methodological adaptation to salinity of the ELRA, according to the first part of this study, which increased its salinity tolerance and sensitivity for 17-β-estradiol, the optimised ELRA was used to investigate 13 native sediments characterised by different levels of salinity and chemical contamination. The applicability of the ELRA for routine analysis in environmental assessment was evaluated. Salinity is often a critical factor for bioassays in ecotoxicological sediment assessment. Therefore, salinity of the samples was additionally adjusted to different levels to characterise its influence on elution and binding processes of receptor-binding substances. Materials and methods  The ELRA was carried out with the human estrogen receptor α (ER) in a 96-well microplate format using the experimental setup known from the competitive immunoassay based on ligand–protein interaction. It is an important improvement that a physiologically relevant receptor was used as a linking protein instead of an antibody. The microplates were coated with a 17-β-estradiol-BSA conjugate, and dilution series of estradiol and of native sediment samples were added and incubated with the ER. After a washing step, a biotinylated mouse anti-ER antibody was added to each well. Receptor binding to estradiol, agonistic and antagonistic receptor binding, were determined by a streptavidin-POD-biotin complex with subsequent measurement of the peroxidase activity at the wavelength of 450 nm using a commercial ELISA multiplate reader. The sediment elutriates and pore water samples of sediments were tested in a dilution series to evaluate at which dilution step the receptor-binding potential ends. In the elution process (see Section 2.1 to 2.2), a method was developed to adjust the salinity to the levels of the reference testings, which offers an appropriate option to adjust the salinity in both directions. Statistical evaluation was made with a combination of the Mann–Whitney U test and the pT-method. Results  This part of the study characterised the environmental factor ‘salinity’ for prospective applications of the ELRA. Using reference substances such as 17-β-estradiol, the ELRA showed sigmoid concentration-effect relations over a broad range from 0.05 μg/l to 100 μg/l under physiological conditions. After methodological optimisation, both sensitivity and tolerance of the assay against salinity could be significantly raised, and the ELRA became applicable under salinity conditions up to concentrations of 20.5‰. The mean relative inter-test error (n = 3) was around 11% with reference substances and below 5% for single sediments elutriates in three replicates each. For sediment testings, the pore water and different salinity-adjusted elutriates of 13 sediments were used. A clear differentiation of the receptor-binding potential could be reached by application of the pT-method. Thereby, pT-values from one to six could be assigned to the sediments, and the deviation caused by the different salinity conditions was one pT-value. The mean standard deviation in the salinity adaptation procedure of the elutriates was below 5%. Discussion  Although the ELRA has already been used for assessments of wastewater, sludge and soil, its applicability for samples to different salinity levels has not been investigated so far. Even if the ELRA is not as sensitive as the E-screen or the YES-assay, with regard to reference substances like 17-β-estradiol, it is a very useful tool for pre-screening, because it is able to integrate both estrogenic as well as anti-estrogenic receptor-binding effects. According to the results of sediment testing, and given the integrative power to detect different directions of effects, the ELRA shows sufficient sensitivity and salinity tolerance to discriminate receptor-binding potentials in environmental samples. Conclusions  The optimised ELRA assay is a fast, cost-effective, reliable and highly reproducible tool that can be used for high-throughput screening in a microplate format in detecting both estrogenic and anti-estrogenic effects. Additionally, the ELRA is robust against microbial contaminations, and is not susceptible towards cytotoxic interferences like the common cell-culture methods. The general applicability and sufficient sensitivity of the ELRA was shown in freshwater environments. Marine and brackish samples can be measured up to salinity levels of 20.5‰. Recommendations and perspectives  In view of the proven sensitivity, functionality and the fastness of the ELRA, it is recommendable to standardise the test method. At the moment, no adequate in vitro test procedure exists which is standardised to DIN or ISO levels. The E-screen and the yeast estrogen/androgen screens (YES/YAS) sometimes underlie strong cytotoxic effects, as reported in the first part of this study. Further development of an ELRA assay using human androgen receptors appears to be very promising to gain information about androgenic and anti-androgenic effects, too. This would offer a possibility to use the ELRA as a fast and reliable pre-screening tool for the detection of endocrine potentials, thus minimising time and cost-expensive animal experiments.     

Effects of biosurfactants on assays of PCB congeners in transgenic arabidopsis plants carrying a recombinant guinea pig AhR-mediated GUS reporter gene expression system

The transgenic Arabidopsis plants carrying a recombinant guinea pig (g) aryl hydrocarbon receptor (AhR)-mediated β-glucuronidase (GUS) reporter gene expression system were generated for assays of polychlorinated biphenyl (PCB) congeners. The selected transgenic Arabidopsis plant XgD2V11-6 exhibited a correlation between uptake of PCB126 and PCB126-induced GUS activity. Also, the plants showed induced GUS activity towards the supplemental indole 3-acetic acid (IAA). Thus, the GUS assay may reflect induction by both endogenous and exogenous AhR ligands. When biosurfactants, MEL-B, produced in the culture of yeast isolated from plants were used for assays of PCB congeners in the transgenic Arabidopsis plants, they showed marked PCB126 dose-dependent and toxic equivalency factor (TEF) dependent GUS activities. The effects of biosurfactants were clearer when the plants were cultivated on soils containing PCB congeners for 7 days as compared with on soils for 3 days as well as in the medium for 3 days. Threfore, it was estimated that biosurfactants form micellae with PCB congeners, which are easily uptaken by the plants in a mode of passive diffusion, transport into the aerial parts and then induce GUS activity.     

Dioxin-like activity in environmental and human samples from Greenland and Denmark

Dioxins and dioxin-like (DL) compounds are some of the most toxic chemicals being highly persistent in the environment. The toxicological effects of dioxins are mediated via the aryl hydrocarbon receptor (AhR). Compounds of diverse structure and lipophility can bind and activate AhR. The AhR transactivation bioassay is utilized in an array of projects to study the AhR-mediated activities of individual chemicals and mixtures and for epidemiological purposes.This review summarizes a series of studies regarding the DL-activity of single compounds and complex compound mixtures in the environment and humans.We found that some pesticides, plasticizers and phytoestrogens can activate the AhR, and the combined effect of compounds with no or weak AhR potency cannot be ignored.The significant DL-activity in the wastewater effluent indicates the treatment is not sufficient to prevent contamination of surface waters with dioxins. Our results from human studies suggest that the serum DL-activity reflect the complex mixture of persistent organic pollutants (POPs). Greenlandic Inuit had lower serum DL-activity level compared to Europeans, probably due to long distance from the dioxin sources and UV degradation of the high potent dioxin and/or the inhibitory effect of the high level of non-DL POPs. Selective bioaccumulation of PCBs in the food chain may contribute to the negative correlation between serum POPs and DL-activity observed in Greenlandic Inuit.Hence the AhR transactivation bioassay provides a cost-effective and integrated screening tool for measurement of the DL-activity in human, environmental and commercial samples.     

Suppression of estrogenic activity of 17-beta-estradiol by beta-cyclodextrin

The suppressive effects of cyclodextrins (CDs) on the strong estrogenic activity of 17β-estradiol (E2) in water environments were investigated in this study. Cyclodextrins are doughnut-shaped molecules that possess a hydrophobic cavity and a hydrophilic exterior. The cavity can incorporate nonpolar molecules as guests to form inclusion complexes. β-CD and 2-hydroxypropyl-β-CD (HP-β-CD) were the most successful in forming a complex with E2 and improving its low aqueous solubility. The E2/CDs complexes bound to the estrogen receptor in a cell-free system as determined by ELISA and suppressed the hormone activities as measured by a yeast two-hybrid assay. These results indicate that hydrophobic E2 is easily transported through the lipid zone of the plasma membrane into the target cell and can bind to the nuclear receptor. However, the hydrophilic E2/β-CD and E2/HP-β-CD complexes do not penetrate the membrane. Therefore, these CDs are able to suppress the hormone activity of E2 through complex formation.     

Two-Compartment Sorption of Phenanthrene on Eight Soils with Various Organic Carbon Contents

Sorption characteristics of phenanthrene (PHE) were studied on eight soils with organic carbon contents spanning over an order of magnitude using phase distribution relationships (PDRs) at 1 h, 48 h, and 720 h contact times. A new algebraic method was employed to describe the sorption characteristics at different time intervals (between 1 h and 48 h, and 1 h and 720 h). It was found that nonlinearity increased with increasing contact time and sorption that occurred in the subsequent time interval following the initial 1 h exhibited stronger isotherm nonlinearity. Sorption coefficients were positively correlated with the organic carbon contents of the soils. Detailed sorption dynamics were also examined on these soils. A two-compartment, first-order model was used to describe the sorption dynamics. The rate constants of the two compartments differed 18–170 times, suggesting the dissimilar sorption behaviors of the mathematically separated compartments. These two compartments were labeled fast and slow sorption compartment according to the rate constants. Calculation showed that the fast compartment accounted for over 80% of the overall sorption at the initial 1 h, while the slow compartment predominated the total sorption in the following 47 h. By combining the discussion of PDRs and sorption dynamics, the contributions of the two compartments to linear and nonlinear sorption were differentiated. The slow sorption compartment made a major contribution to nonlinear sorption and possibly to sequestration of organic pollutants by these soils.     

2,3,7,8-Tetrachlorodibenzo-p-dioxin promotes migration ability of primary cultured rat astrocytes via aryl hydrocarbon receptor

Emerging evidence showed that 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) could induce expression of certain reactivation-associated genes in astrocytes, however, the consequent cellular effects and molecular mechanisms are still unclear. During the process of astrocyte reactivation, migration is a critical cellular event. In the present study, we employed wound-healing assay and Transwell® motility assay to explore the effects of TCDD on cell migration in primary cultured rat cortical astrocytes. We found that upon TCDD treatments at relative low concentrations (10^? 10 and/or 10^? 9?mol/L), the ability of primary astrocytes to migrate horizontally and vertically was promoted. In line with this cellular effect, the mRNA expression of two pro-migratory genes, including cell division cycle 42 (CDC42) and matrix metalloproteinase 2 (MMP2) was induced by TCDD treatment. Dioxin exerts its toxic effects mainly through aryl hydrocarbon receptor (AhR) pathway. So the role of AhR pathway in the pro-migratory effects of TCDD was examined using an AhR antagonist, CH223191. We found that application of CH223191 significantly reversed the pro-migratory effects of TCDD. Interestingly, the basal ability of horizontal migration as well as basal levels of CDC42 and MMP2 expression were dramatically reduced suggesting a possible physiological role of AhR in maintaining the endogenous migration ability of the primary astrocytes. These findings support the notion that dioxin promotes astrocyte reactivation at molecular and cellular levels.     

Three lessons for gender equity in biodiversity conservation

Amid a growing global agenda, biodiversity conservation has embraced gender equity as a pillar of equitable and effective practice. Gender equity has become enshrined in the global environment and development agenda through global commitments, policy and funding. However, for various reasons, conservation biodiversity often takes a simplistic view of gender as synonymous with women or as a dualism between women and men. This narrow view risks promoting inequitable processes and ineffective outcomes. Deeper engagement with feminist theory, and feminist political ecology in particular, could help advance biodiversity conservation's approach to how gender is understood, framed and integrated. Engaging with lessons from feminist political ecology can help advance gender equity in conservation through attention to power dynamics, intersectionality, and subjectivity.