Temporal sex specific brain gene expression pattern during early rat embryonic development.

Background: The classical concept of brain sex differentiation suggests that steroid hormones released from the gonads program male and female brains differently. However, several studies indicate that steroid hormones are not the only determinant of brain sex differentiation and that genetic differences could also be involved. Methods: In this study, we have performed RNA sequencing of rat brains at embryonic days 12 (E12), E13, and E14. The aim was to identify differentially expressed genes between male and female rat brains during early development. Results: Analysis of genes expressed with the highest sex differences showed that Xist was highly expressed in females having XX genotype with an increasing expression over time. Analysis of genes expressed with the highest male expression identified three early genes, Sry2, Eif2s3y, and Ddx3y. Discussion: The observed sex-specific expression of genes at early development confirms that the rat brain is sexually dimorphic prior to gonadal action on the brain and identifies Sry2 and Eif2s3y as early genes contributing to male brain development.

Microglia Signaling in Health and Disease - Implications in Sex-Specific Brain Development and Plasticity.

Microglia, the intrinsic neuroimmune cells residing in the central nervous system (CNS), exert a pivotal influence on brain development, homeostasis, and functionality, encompassing critical roles during both aging and pathological states. Recent advancements in comprehending brain plasticity and functions have spotlighted conspicuous variances between male and female brains, notably in neurogenesis, neuronal myelination, axon fasciculation, and synaptogenesis. Nevertheless, the precise impact of microglia on sex-specific brain cell plasticity, sculpting diverse neural network architectures and circuits, remains largely unexplored. This article seeks to unravel the present understanding of microglial involvement in brain development, plasticity, and function, with a specific emphasis on microglial signaling in brain sex polymorphism. Commencing with an overview of microglia in the CNS and their associated signaling cascades, we subsequently probe recent revelations regarding molecular signaling by microglia in sex-dependent brain developmental plasticity, functions, and diseases. Notably, C-X3-C motif chemokine receptor 1 (CX3CR1), triggering receptors expressed on myeloid cells 2 (TREM2), calcium (Ca2+), and apolipoprotein E (APOE) emerge as molecular candidates significantly contributing to sex-dependent brain development and plasticity. In conclusion, we address burgeoning inquiries surrounding microglia's pivotal role in the functional diversity of developing and aging brains, contemplating their potential implications for gender-tailored therapeutic strategies in neurodegenerative diseases.

Assessing organism differences in mixed metal sensitivity.

Metal contamination of aquatic environments remains a major concern and has received significant attention in recent years. The present study aimed to evaluate the effects of metal mixtures of varying concentrations over time in a lake receiving runoff water from a decommissioned mine. By subjecting several organisms to this water, we aimed to identify the most susceptible species, thus enabling a comprehensive evaluation of the risk posed by different toxins to the biotic environment. We have evaluated the effects of mixed metal exposure on survival and stress gene expression in selected invertebrate and vertebrate model species. Our observations revealed differences in sensitivity among the invertebrate models Caenorhabditis elegans, Daphnia magna, Ceriodaphnia dubia, and Heterocypris incongruens, as well as in the vertebrate model Zebrafish (Danio rerio) and two cell lines; a zebrafish liver cell line (ZFL) and a human hepatocellular carcinoma cell line (HepG2). While the sensitivity shows great variation among the tested species, the expression of metallothionein was consistent with the levels of metals found in the mixed exposure media. Despite differences in acute toxicity, the universal induction of mt1/A and mt2/B genes make them important biomarkers for assessing the environmental risk of metals.

Utilization of Computer Classification Methods for Exposure Prediction and Gene Selection in Daphnia magna Toxicogenomics.

Zinc (Zn) is an essential element that influences many cellular functions. Depending on bioavailability, Zn can cause both deficiency and toxicity. Zn bioavailability is influenced by water hardness. Therefore, water quality analysis for health-risk assessment should consider both Zn concentration and water hardness. However, exposure media selection for traditional toxicology tests are set to defined hardness levels and do not represent the diverse water chemistry compositions observed in nature. Moreover, these tests commonly use whole organism endpoints, such as survival and reproduction, which require high numbers of test animals and are labor intensive. Gene expression stands out as a promising alternative to provide insight into molecular events that can be used for risk assessment. In this work, we apply machine learning techniques to classify the Zn concentrations and water hardness from Daphnia magna gene expression by using quantitative PCR. A method for gene ranking was explored using techniques from game theory, namely, Shapley values. The results show that standard machine learning classifiers can classify both Zn concentration and water hardness simultaneously, and that Shapley values are a versatile and useful alternative for gene ranking that can provide insight about the importance of individual genes.

Per- and polyfluoroalkyl substances enhance Staphylococcus aureus pathogenicity and impair host immune response.

Per- and Poly-fluoroalkyl substances (PFAS) are major persistent environmental contaminants. Epidemiological studies have linked PFAS exposures to altered immunity and increased occurrence of infections in children. However, the mechanisms leading to immune susceptibility to bacterial infections remains unclear. To elucidate the mechanism, transcriptional alteration in the Caenorhabditis elegans model caused by a PFAS contaminated environmental water and two reconstituted PFAS solutions were evaluated using RNA-sequencing. PFAS affected the expression of several genes involved in C. elegans immune surveillance to Gram-positive bacteria (cpr-2, tag-38, spp-1, spp-5, clec-7, clec-172). The combined exposure to PFAS and Staphylococcus aureus significantly reduced C. elegans survival and increased intestinal membrane permeability. Furthermore, the growth of S. aureus in the presence of PFAS increased the expression of virulence genes, specifically, the virulence gene regulator saeR and α-hemolysin, hla, which resulted in increased hemolytic activity. The present study demonstrated that PFAS exposure not only increased C. elegans susceptibility to pathogens by reducing host immunity and increasing intestinal membrane permeability, but also increased bacteria virulence. This presents a broader implication for humans and other animals, where environmental contaminants simultaneously reduce host resilience, while, increasing microbial pathogenicity.

Carbapenemase-producing Aeromonas species isolated from the urban-impacted Akaki river in Ethiopia.

Carbapenemase-producing Aeromonas species are an emerging health threat. This study aimed to determine carbapenemase-mediated resistance among Aeromonas isolates from the Akaki river, Ethiopia during the dry and wet seasons in 2019-2020. Antimicrobial susceptibility to carbapenems and cephalosporins was determined and carbapenemase production was confirmed. Of 163 isolates, the majority were human pathogens Aeromonas caviae (62), Aeromonas hydrophila (33) and Aeromonas veronii (49). These isolates were resistant to carbapenem and cephalosporin antibiotics, with the highest resistance to cefotaxime 86 (59.7%), ertapenem 71 (49.3%) and imipenem 65 (45.1%). Resistance to carbapenem antibiotics varied between species, where most A. veronii 37 (75.5%) and A. hydrophila 28 (84.8%) were resistant to imipenem and all A. caviae were sensitive. A. veronii, A. caviae and A. hydrophila resistance to meropenem was 31 (63.3%), 3 (4.8%) and 19 (57.6%), respectively. Of isolates resistant to carbapenem, 82.1% A. hydrophila and 94.4% A. veronii were carbapenemase producers. Cephalosporin resistance also varied among the different species. The highest resistance to carbapenem antibiotics was in isolates collected during the wet season (p<0.05); however, it was not consistent across all classes of antibiotics tested. The rivers in megacities could be reservoirs of carbapenemase-producing Aeromonas spp.

Antimicrobial resistance genes in microbiota associated with sediments and water from the Akaki river in Ethiopia.

The spread of antimicrobial-resistant pathogens is a global health concern. Most studies report high levels of antimicrobial resistance genes (ARGs) in the aquatic environment; however, levels associated with sediments are limited. This study aimed to investigate the distribution of ARGs in the sediments and water of the Akaki river in Addis Ababa, Ethiopia. The diversity and abundance of 84 ARGs and 116 clinically important bacteria were evaluated from the sediments and water collected from five sites in the Akaki river. Most of the ARGs were found in the city close to anthropogenic activities. Water samples collected in the middle catchment of the river contained 71-75% of targeted ARGs, with genes encoding aminoglycoside acetyltransferase (aac(6)-Ib-cr), aminoglycoside adenylyl transferase (aadA1), ß-lactamase (blaOXA-10), quinolone resistance S (qnrS), macrolide efflux protein A (mefA), and tetracycline resistance (tetA), were detected at all sampling sites. Much fewer ARGs were detected in all sediments, and those near the hospitals had the highest diversity and level. Despite the lower levels and diversity, there were no unique ARGs detected in the sediments that were also not detected in the waters. A wide range of clinically relevant pathogens were also detected in the Akaki river. The findings suggest that the water phase, rather than the sediments in the Akaki river, is a potential conduit for the spread of ARGs and antibiotic-resistant bacteria.

Transcriptional responses of Daphnia magna exposed to Akaki river water.

Pollution of the aquatic environment is a global problem, with industrial waste, farming effluents, sewage, and wastewater as the main contributors. Many pollutants are biologically active at low concentrations, resulting in sublethal effects, which makes it a highly complex situation and difficult to assess. In many places, such as the Akaki river in Ethiopia, the pollution situation has resulted in streams with minimal presence of invertebrates or vertebrates. As it is difficult to perform a complete chemical analysis of the waters, the present study focused on using gene expression analysis as a biological end point to determine the effects of Akaki river contaminants. The present study was conducted using the small planktonic crustacean Daphnia magna with toxicogenomic molecular markers. Daphnia magna neonates were exposed to Akaki water samples collected from two different sites on the river and analyzed for mortality and expression of genes involved in different biological pathways. Despite the poor quality of Akaki river water, 48 h acute toxicity tests showed no mortality. Interestingly, analysis of sublethal toxicogenomic responses showed that exposure to Akaki water altered the expression of 25 out of 37 genes involved in metal regulation, immune response, oxidative stress, respiration, reproduction, and development. The toxicogenomic data gives insight into the mechanisms involved in causing potential adverse effects to aquatic biota harboring the Akaki river system.

Lysinibacillus sphaericus mediates stress responses and attenuates arsenic toxicity in Caenorhabditis elegans.

Exposure to toxic metals alters host response and that leads to disease development. Studies have revealed the effects of metals on microbial physiology, however, the role of metal resistant bacteria on host response to metals is unclear. The hypothesis that xenobiotic interactions between gut microbes and arsenic influence the host physiology and toxicity was assessed in a Caenorhabditis elegans model. The arsenic-resistant Lysinibacillus sphaericus B1CDA was fed to C. elegans to determine the host responses to arsenic in comparison to Escherichia coli OP50 food. L. sphaericus diet extended C. elegans lifespan compared to E. coli diet, with an increased expression of genes involved in lifespan, stress response and immunity (hif-1, hsp-16.2, mtl-2, abf-2, clec-60), as well as reduced fat accumulation. Arsenic-exposed worms fed L. sphaericus also had a longer lifespan than those fed E. coli and had an increased expression of genes involved in cytoprotection, stress resistance (mtl-1, mtl-2) and oxidative stress response (cyp-35A2, isp-1, ctl-2, sod-1), together with a decreased accumulation of reactive oxygen species (ROS). In comparison with E. coli, L. sphaericus B1CDA diet increased C. elegans fitness while detoxifying arsenic induced ROS and extending lifespan.

Influence of water hardness on zinc toxicity in Daphnia magna.

Zinc is an essential trace metal required for the maintenance of multiple physiological functions. Due to this, organisms can experience both zinc deficiency and toxicity. Hardness is recognized as one of the main modifying physiochemical factors regulating zinc bioavailability. Therefore, the present study analyzed the effect of hardness on zinc toxicity using Daphnia magna. Endpoint parameters were acute-toxicity, development, reproduction, and expression data for genes involved in metal regulation and oxidative stress. In addition, the temporal expression profiles of genes during the initiation of reproduction and molting were investigated. Water hardness influenced the survival in response to exposures to zinc. A zinc concentration of 50 µg/l in soft water (50 mg CaCO3 /L) caused 73% mortality after 96 h exposure, whereas the same zinc concentration in the hardest water did not cause any significant mortality. Moreover, increasing water hardness from 100 to 200 mg CaCO3 /L resulted in a reduced number of offspring. Fecundity was higher at first brood for groups exposed to higher Zn concentrations. The survival data were used to assess the precision of the bioavailability models (Bio-met) and the geochemical model (Visual MINTEQ). As the Bio-met risk predictions overestimated the Zn toxicity, a competition-based model to describe the effects of hardness on zinc toxicity is proposed. This approach can be used to minimize differences in setting environmental quality standards. Moreover, gene expression data showed that using the toxicogenomic approach was more sensitive than the physiological endpoints. Therefore, data presented in the study can be used to improve risk assessment for zinc toxicity.

In silico and in vitro assessment of androgen receptor antagonists.

There is a growing concern for male reproductive health as studies suggest that there is a sharp increase in prostate cancer and other fertility related problems. Apart from lifestyle, pollutants are also known to negatively affect the reproductive system. In addition to many other compounds that have been shown to alter androgen signaling, several environmental pollutants are known to disrupt androgen signaling via binding to androgen receptor (AR) or indirectly affecting the androgen synthesis. We analyzed here the molecular mechanism of the interaction between the human AR Ligand Binding Domain (hAR-LBD) and two environmental pollutants, linuron (a herbicide) and procymidone (a pesticide), and compared with the steroid agonist dihydrotestosterone (DHT) and well-known hAR antagonists bicalutamide and enzalutamide. Using molecular docking and dynamics simulations, we showed that the co-activator interaction site of the hAR-LBD is disrupted in different ways by different ligands. Binding free energies of the ligands were also ordered in increasing order as follows: linuron, procymidone, DHT, bicalutamide, and enzalutamide. These data were confirmed by in vitro assays. Reporter assay with MDA-kb2 cells showed that linuron, procymidone, bicalutamide and enzalutamide can inhibit androgen mediated activation of luciferase activity. Gene expression analysis further showed that these compounds can inhibit the expression of prostate specific antigen (PSA) and microseminoprotein beta (MSMB) in prostate cell line LNCaP. Comparative analysis showed that procymidone is more potent than linuron in inhibiting AR activity. Furthermore, procymidone at 10 µM dose showed equivalent and higher activity to AR inhibitor enzalutamide and bicalutamide respectively.

Sublethal effects of DBE-DBCH diastereomers on physiology, behavior, and gene expression of Daphnia magna.

1,2-dibromo-4-(1,2-dibromoethyl)-cyclohexane (DBE-DBCH) is a brominated flame retardant used in commercial and industrial applications. The use of DBE-DBCH containing products has resulted in an increased release into the environment. However, limited information is available on the long-term effects of DBE-DBCH and its effects in aquatic invertebrates. Thus, the present study was aimed at determining how DBE-DBCH diastereomers (αß and γδ) affects aquatic invertebrates using Daphnia magna as a model organism. Survival, reproduction, feeding, swimming behavior and toxicogenomic responses to environmental relevant concentrations of DBE-DBCH were analyzed. Chronic exposure to DBE-DBCH resulted in decreased lifespan, and reduced fecundity. Expression of genes involved in reproductive processes, vtg1 and jhe, were also inhibited. DBE-DBCH also induced hypoxia by inhibiting the transcription of genes involved in heme biosynthesis and oxygen transport. Furthermore, DBE-DBCH also inhibited feeding resulting in emptiness of the alimentary canal. Increased expression of the stress response biomarkers was observed following DBE-DBCH exposure. In addition, DBE-DBCH diastereomers also altered the swimming behavior of Daphnia magna. The present study demonstrates that DBE-DBCH cause multiple deleterious effects on Daphnia magna, including effects on reproduction and hormonal systems. These endocrine disrupting effects are in agreement with effects observed on vertebrates. Furthermore, as is the case in vertebrates, DBE-DBCH γδ exerted stronger effects than DBE-DBCH αß on Daphnia magna. This indicate that DBE-DBCH γδ has properties making it more toxic to all so far studied animals than DBE-DBCH αß.

The brominated flame retardants TBECH and DPTE alter prostate growth, histology and gene expression patterns in the mouse.

The brominated flame retardants (BFRs), 1,2-dibromo-4-(1,2 dibromoethyl)cyclohexane (TBECH) and 2,3-dibromopropyl-2,4,6-tribromophenyl ether (DPTE) bind to the androgen receptor (AR). in vitro bioassays have shown that TBECH is a potent androgen agonist while DPTE is a potent AR antagonist. Both TBECH and DPTE alter gene expression associated with AR regulation. However, it remains to be determined if TBECH and DPTE can affect the prostate. For this reason, we exposed CD1 mice to a 1:1 mixture of TBECH diastereomers α and ß, a 1:1 mixture of γ and δ, and to DPTE, and tested their effects on prostate growth, histology and gene expression profiles. Castrated mice were used to study the androgenic effects of TBECHαß and TBECHγδ while the antagonistic effects of DPTE were studied in non-castrated mice. We observed that testosterone and TBECHγδ increased body and prostate weights while TBECHαß affected neither of them; and that DPTE had no effect on body weight but reduced prostate weight drastically. Histomorphometric analysis of the prostate revealed epithelial and glandular alterations in the TBECHγδ group comparable to those in testosterone group while alterations in the TBECHαß group were less pronounced. DPTE displayed androgen antagonist activity reminiscent of castration. The transcription profile of the prostate was altered by castration and exposure to testosterone and to TBECHγδ reversed several of these changes. Testosterone and TBECHγδ also regulated the expression of several androgen responsive genes implicated in prostate growth and cancer. While DPTE resulted in a drastic reduction in prostate weight, it only affected a small number of genes. The results indicate that TBECHγδ and DPTE are of high human health concern as they may contribute to changes in prostate growth, histology and function.

Sex differences in severity and mortality from COVID-19: are males more vulnerable?

Coronavirus disease 2019 (COVID-19) has shown high infection and mortality rates all over the world, and despite the global efforts, there is so far no specific therapy available for COVID-19. Interestingly, while the severity and mortality of COVID-19 are higher in males than in females, the underlying molecular mechanisms are unclear. In this review, we explore sex-related differences that may be contributing factors to the observed male-biased mortality from COVID-19. Males are considered the weaker sex in aspects related to endurance and infection control. Studies show that viral RNA clearance is delayed in males with COVID-19. A recent study has indicated that the testis can harbor coronavirus, and consequently, males show delayed viral clearance. However, the role of testis involvement in COVID-19 severity and mortality needs further research. Males and females show a distinct difference in immune system responses with females eliciting stronger immune responses to pathogens. This difference in immune system responses may be a major contributing factor to viral load, disease severity, and mortality. In addition, differences in sex hormone milieus could also be a determinant of viral infections as estrogen has immunoenhancing effects while testosterone has immunosuppressive effects. The sex-specific severity of COVID-19 infections indicates that further research on understanding the sex differences is needed. Inclusion of both males and females in basic research and clinical trials is required to provide critical information on sex-related differences that may help to better understand disease outcome and therapy.

Perfluorinated alkyl substances impede growth, reproduction, lipid metabolism and lifespan in Daphnia magna.

Per- and polyfluorinated alkyl substances (PFASs) are synthetic organofluorine compounds with unique stability accompanied with hydrophobic and lipophobic properties. Perfluorooctane sulfonate (PFOS) and Perfluorooctanoic acid (PFOA) are of high concern due to their wide application in consumer and industrial products, extreme persistence, abundant occurrence in the environment and their toxic effect to humans and animals. However, knowledge on the molecular mechanisms of toxicity and the effects on reproduction output remain scarce. In this study, we analyzed the effects of PFOS and PFOA on Daphnia magna. Acute toxicity, development, reproduction, lipid metabolism (lipid-accumulation) and lifespan was investigated, as well as the expression of genes related to these endpoints. Exposure of PFOS and PFOA at 1, 10 and 25 µM did not cause acute lethality. Hatching was reduced following exposure to both compounds, and lifespan was decreased following exposure to 25 µM PFOS. Body length of Daphnia magna was reduced significantly by 25 µM PFOS following 7 days exposure. Lipid staining revealed that all PFAS exposures increased lipid accumulation. qRT-PCR analysis of genes involved in lipid metabolism suggests that the increase in lipid content could be due to inhibition of genes involved on absorption and catabolism of fatty acids. Exposure to both PFOA and PFOS reduced the fecundity significantly. Downregulation of genes involved in development and reproductive process, including vtg2, vasa, EcRA, EcRB, usp, jhe, HR3, ftz-F1, E74 and E75 were observed. The alterations in developmental and reproductive genes as well as the disturbed lipid metabolism provides mechanistic insight into the possible causes for decreased fecundity and lifespan observed following exposure to both PFOS and PFOA.

Nonsteroidal anti-inflammatory drugs (NSAIDs) cause male-biased sex differentiation in zebrafish.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used pharmaceuticals to treat pain, fever and inflammation. NSAIDs are also known to have many side effects including adverse effects on reproduction in both humans and animals. As NSAIDs usage is not regulated they are frequently detected at high concentrations in the environment. In order to understand the effect of NSAIDs on zebrafish sex differentiation, we used seven different NSAIDs which were either Cox-1 selective, Cox-1 biased, non-selective or COX-2 selective. We show that at higher concentration, NSAIDs are toxic to zebrafish embryo as they lead to mortality and hatching delay. Gene expression analysis following short term exposure of NSAIDs led to downregulation of female specific genes including zp2, vtg2 foxl2 and wnt4. Long term exposure of larvae to environmentally relevant concentrations of Cox-2 selective and non-selective NSAIDs resulted in male-biased sex ratio which confirmed the qRT-PCR analysis. However, the Cox-1 selective acetylsalicylic acid and the Cox-1 biased ketoprofen did not alter sex ratio. The observed male-biased sex ratio could also be due to induction of apoptosis process as the genes including p21 and casp8 were significantly upregulated following exposure to the Cox-2 selective and the non-selective NSAIDs. The present study indicates that NSAIDs alter sex differentiation in zebrafish, primarily through inhibition of Cox-2. This study clearly demonstrates that the use of NSAIDs and their release into the aquatic environment should be carefully monitored to avoid adverse effects to the aquatic organisms.

Zebrafish cyp17a1 knockout reveals that androgen-mediated signaling is important for male brain sex differentiation.

Brain sex differentiation is a complex process, wherein genes and steroid hormones act to induce specific gender brain differentiation. Testosterone (T) derived from the gonads has been linked to neural circuit modeling in a sex-specific manner. Previously, we have shown that cyp17a1 knockout (KO) zebrafish have low plasma androgen levels, and display compromised male-typical mating behaviors. In this study, we demonstrated that treatment of cyp17a1 KO males with T or 11-ketotestosterone (11-KT) is sufficient to rescue mating impairment by restoring the male-typical secondary sex characters (SSCs) and mating behaviors, confirming an essential role of androgen in maintaining SSCs and mating behaviors. Brain steroid hormone analysis revealed that cyp17a1 KO fish have reduced levels of T and 11-KT. We performed RNA sequencing on brain samples of control and cyp17a1 KO male zebrafish to get insights regarding the impact of cyp17a1 KO on gene expression pattern, and to correlate it with the observed disruption of male-typical mating behaviors. Transcriptome analysis of cyp17a1 KO males showed a differential gene expression when compared to control males. In total, 358 genes were differentially regulated between control males and KO males. Important genes including brain aromatase (cyp19a1b), progesterone receptor (pgr), deiodinase (dio2), and insulin-like growth factor 1 (igf1) that are involved in brain functions, as well as androgen response genes including igf1, frem1a, elovl1a, pax3a, mmp13b, hsc70, ogg1 were regulated. RT-qPCR analysis following rescue of cyp17a1 KO with T and 11-KT further suggested that androgen-mediated signaling is disrupted in the cyp17a1 KO fish. Our results indicated that cyp17a1 KO fish have an incomplete masculinization and altered brain gene expression, which could be due to decreased androgen levels.

The food preservative ethoxyquin impairs zebrafish development, behavior and alters gene expression profile.

In the present study, we investigated the detrimental effects of ethoxyquin (EQ) on zebrafish embryonic development using different endpoints including lethality, malformations, locomotion and gene expression. EQ is primarily used as a preservative in animal feed and it has been shown to have negative impacts on different laboratory animals. However, studies on the adverse effects of EQ in aquatic animals are still limited. In this study, zebrafish eggs were exposed to different concentrations of EQ ranging from 1 to 100 µM for six days. In the 100 µM treated groups 95 and 100% mortality was observed at 24 and 48 h, respectively. Delayed development, decreased pigmentation and pericardial edema were observed in larvae. Behavioral analysis of larvae demonstrated a distinct locomotive pattern in response to EQ both in light and dark indicating a possible developmental neurotoxicity and deficits in locomotion. The expression levels of genes involved in several physiological pathways including stress response, cell cycle and DNA damage were altered by EQ. Our results demonstrate that EQ could cause developmental and physiological toxicity to aquatic organisms. Hence, its toxic effect should be further analyzed and its use and levels in the environment must be monitored carefully.

Species differences in ligand interaction and activation of estrogen receptors in fish and human.

Estrogen receptor (ER) sequences vary between species and this suggests that there are differences in the ligand-specificity, leading to species-specific effects. This would indicate that it is not possible to generalize effects across species. In this study, we investigated the differences in activation potencies and binding affinities of ER´s alpha (α) and beta (ß) in human, zebrafish and sea bream to elucidate species differences in response to estradiol, estrone, estriol and methyltestosterone. In vitro analysis showed that estradiol had the highest activity for all the ER´s except for human ERß and seabream ERß2. Alignment of the ligand binding domain and ligand binding pocket (LBP) residues of the three species showed that different residues were involved in the LBPs which led to differences in pocket volume, affected binding affinity and orientation of the ligands. By combining in silico and in vitro results, it was possible to identify the ligand specificities of ER´s. The results demonstrated that the human ER´s show lower resolution in ligand-dependent activation, suggesting higher promiscuity, than the zebrafish and seabream ER´s. These results show species-specificity of ER´s and suggest that species-specific differences must be taken into consideration when studying different exposure scenarios.

Distinct transcriptional response of Caenorhabditis elegans to different exposure routes of perfluorooctane sulfonic acid.

Although people are exposed daily to per- and polyfluorinated alkyl substances (PFASs), the biological consequences are poorly explored. The health risks associated with PFAS exposure are currently based on chemical analysis with a weak correlation to potential harmful effects in man and animals. In this study, we show that perfluorooctane sulfonic acid (PFOS), often the most enriched PFAS in the environment, can be transferred via bacteria to higher organisms such as Caenorhabditis elegans. C. elegans nematodes were exposed to PFOS directly in buffer or by feeding on bacteria pretreated with PFOS, and this led to distinct gene expression profiles. Specifically, heavy metal and heat shock associated genes were significantly, although inversely, expressed following the different PFOS exposures. The innate immunity receptor for microbial pathogens, clec-60, was shown for the first time to be down-regulated by PFOS. This is in line with a previous study indicating that PFOS is associated with children's susceptibility to certain infectious diseases. Furthermore, bar-1, a gene associated with various cancers was highly up-regulated only when C. elegans were exposed to PFOS pretreated live bacteria. Furthermore, dead bacterial biomass had higher binding capacity for linear and isomeric PFOS than live bacteria, which correlated to the higher levels of PFOS detected in C. elegans when fed the treated E. coli, respectively. These results reveal new aspects concerning trophic chain transport of PFOS.