Organophosphate flame retardants (OPFRs), as alternatives of polybrominated diphenyl ethers (PBDEs), have been frequently detected in the environment and biota, and could pose adverse effects on organisms. However, information on the potential endocrine disruption of OPFRs, especially their effects on steroid hormone receptors, such as glucocorticoid and mineralocorticoid receptors (GR/MR), is limited. In this study, the dual-luciferase reporter gene assay via GR/MR and a H295R steroidogenesis assay were employed to evaluate the endocrine disruption of nine OPFRs. We found TMPP, TPHP, and TDBPP exhibited both GR and MR antagonistic activities, while TNBP and TDCIPP only showed MR antagonistic property within a concentration range of 10–8 to 10–5 mol/L(M). In the H295R steroidogenesis assay, the fold changes of eight steroidogenic genes in response to OPFRs were further studied. We found CYP17,CYP21, and CYP11B1 expression were significantly down-regulated following TMPP, TPHP, or TDBPP exposure at a concentration of 2 × 10–6 M. Meanwhile TMPP decreased the production of cortisol and TDBPP down-regulated the secretion of aldosterone. Our results indicate that some OPFRs can interact with GR and MR, and have the potential to disturb steroidogenesis. Data provided here will be helpful to comprehensively understand the potential endocrine disruption of OPFRs.

Troposphere ozone, which is from secondary formation processes, has been increasing dramatically during the last decades in China, inducing high health risks. In this study, temporal and spatial distribution of O3 was studied among 13 sites of three cities during 2014–2016. The objectives were to clarify the characteristics of the ambient pollution of O3 under the influence from other pollutants and meteorological parameters and the health outcomes from exposure to O3. The concentrations of O3 during summer were much higher than those during winter, and the concentrations in downtown areas were higher than in rural or mountain areas. PM2.5, NO2, SO2, and wind speed (WS) were negatively correlated with O3, and CO, temperature (T), and relative humidity (RH) were positively correlated with O3. In multivariable analysis, two separate factors—solar radiation and atmospheric diffusion status, affected the O3 levels. The concentrations of O3 reached the highest level at 15:00 and the lowest value at about 6:00–8:00, with the similar trend to T and WS, and opposite to RH. According to the dose-response model, relative risks (RRs) and population attributable fractions (PAFs) with confidence intervals (CIs) for chronic obstructive pulmonary disease (COPD) from exposure to O3 were 1.0612 (CI 1.0607–1.0616) and 5.32% (CI 5.29–5.36%), respectively, attributable to 2000 deaths in Zhejiang Province in 2014.

•The toxicity of these SPs to zebrafish embryos can be ranked as LCT > FEN > PM.•The T3 content in larvae were both decreased after exposed to 500 nM LCT and FEN.•The expressions of NRs genes caused by LCT, FEN and PM were disrupted.Synthetic pyrethroids (SPs) are one of the most widely used pesticides and frequently detected in the aquatic environment. Previous studies have shown that SPs posed high aquatic toxicity, but information on the developmental toxicity and endocrine disruption on zebrafish (Danio rerio) at environmentally relevant concentrations is limited. In this study, zebrafish embryos were employed to examine the adverse effects of λ-cyhalothrin (LCT), fenvalerate (FEN), and permethrin (PM) at 2.5, 10, 25, 125, 500 nM for 96 h. The results showed these 3 SPs caused dose-dependent mortality, malformation rate, and hatching rate. Thyroid hormone triiodothyronine (T3) levels were significantly decreased after exposure to LCT and FEN. Quantitative real-time PCR analysis was then performed on a series of nuclear receptors (NRs) genes involved in the hypothalamic–pituitary–gonadal (HPG), hypothalamic–pituitary–thyroid (HPT), hypothalamic–pituitary–adrenocortical (HPA) axes, and oxidative-stress-related system. Our results showed that LCT, FEN, and PM downregulated AR expression while upregulated ER1 expression, and caused alteration to ER2a and ER2b expression. As for the expression of TRα and TRβ, they were both decreased following exposure to the 3 SPs. LCT and PM downregulated the MR expression and FEN induced MR expression. In addition, the expression of GR was increased after treating with LCT, while it was suppressed after exposure to FEN and PM. The 3 SPs also caused various alterations to the expression of genes including AhRs, PPARα, and PXR. These findings suggest that these 3 SPs may cause developmental toxicity to zebrafish larvae by disrupting endocrine signaling at environmentally relevant concentrations.

•Glufosinate promoted the growth of M. aeruginosa at 0.5 and 1 mg/L.•Total-MC-LR levels were significantly increased at 1 mg/L of glufosinate.•1 mg/L of glufosinate may induce the potential risk by elevating the MC-LR levels.Harmful algal blooms (HABs) have been emerging as a worldwide issue due to their detrimental effect to water quality and the threat to ecological health. However, there were scarcely reports on whether existing agrochemicals such as herbicides have potential effects on Microcystis aeruginosa (M. aeruginosa) which may form large surface blooms. In this study, we aimed to demonstrate the effects of glufosinate, a widely use herbicide in the world, on the growth of M. aeruginosa. The change of the content of intracellular (intra-) and extracellular (extra-) levels of microcystin-LR (MC-LR) were also determined by HPLC. The mRNA level of mcyA, mcyD, mcyH related to the production of MC-LR was detected after 48 h exposure to glufosinate. Our results revealed that low concentrations of glufosinate exposure (0.5 mg/L, 1 mg/L) would promote the growth of M. aeruginosa. It was worth noting intra-MC-LR contents of algal cells were also significantly increased at low concentrations of glufosinate exposure (1 mg/L). While at high concentrations (5 mg/L, 10 mg/L), glufosinate could induce the MDA level and inhibit the growth of M. aeruginosa that would limit the production of MC-LR to the water body. At last, the transcription levels of algae toxin synthesis related genes mcyA and mcyD were increased significantly at environmentally relevant concentrations of glufosinate, which may affect the MC production. This study not only illuminated effects of glufosinate on growth of and MC-LR synthesis of M. aeruginosa at environmentally relevant concentrations, but also reminding us the potential risk caused by herbicide use should arouse concerns.Download high-res image (209KB)Download full-size image

The use of glyphosate, which is a well-known sterilant herbicide, has been growing rapidly because the area under the cultivation of genetically modified crops that are tolerant to this herbicide has increased. Glyphosate can enter into aquatic systems through many different ways. However, information on the potential risks of glyphosate at environmentally relevant levels to aquatic systems is still limited. In this study, we selected the cyanobacterium Microcystis aeruginosa FACHB-905 (M. aeruginosa) as a model organism to evaluate the effects of glyphosate at environmentally relevant concentrations on the former’s growth and microcystin (MC) production. Our results show that low levels of glyphosate stimulate the growth of M. aeruginosa. Subsequently, there was significant increase in the total MC-LR and intracellular MC-LR, but not in extracellular MC-LR, after exposure to 0.1–2 mg/L of glyphosate. The increase in total MC-LR is mainly due to the effects of glyphosate on the cell density of M. aeruginosa. The data provided here show that low level of glyphosate in a water body is a potential environmental risk factor that stimulates the growth and enhances MC production in M. aeruginosa, which should arouse great concern.

The o,p′-dichlorodiphenyltrichloroethane (DDT) with a chiral center possesses enantioselective estrogenic activity, in which R-(−)-o,p′-DDT exerts a more potent estrogenic effect than S-(+)-o,p′-DDT. Although concern regarding DDT exposure and breast cancer has increased in recent decades, the mode of enantioselective action of o,p′-DDT in breast cancer development is still unknown. Herein, we conducted a systematic study of the effect of o,p′-DDT on stereoselective breast tumor cell progression in a widely used in vitro breast tumor cell model, MCF-7 cells. We demonstrated that R-(−)-o,p′-DDT promoted more cancer cell invasion mediated by the human estrogen receptor (ER) by inducing invasion-promoted genes (matrix metalloproteinase-2 and -9 and human telomerase reverse transcriptase) and inhibiting invasion-inhibited genes (tissue inhibitor of metalloproteinase-1 and -4). Molecular docking verified that the binding affinity between R-(−)-o,p′-DDT and human ER was stronger than that of S-(+)-o,p′-DDT. The enantioselective-induced decrease in cell-to-cell adhesion may involve the downregulation of adhesion-promoted genes (E-cadherin and β-catenin). For the first time, these results reveal that estrogenic-like chiral compounds are of significant concern in the progression of human cancers and that human health risk assessment of chiral chemicals should consider enantioselectivity.

Polychlorinated biphenyls (PCBs), with 209 congeners, are a large family of persistent organic pollutants. PCBs elicit a wide range of toxicities, such as neurotoxicity, hepatoxicity, oncogenicity, and endocrine-disrupting effects. However, an understanding of the potential disruption of systematic iron metabolism by PCBs is still limited. To maintain iron homeostasis, the hepcidin–ferroportin (FPN) axis is critically important, and hepcidin is the central governor in guiding dietary iron absorption and iron egress from macrophages. Hepcidin is secreted by hepatocytes and binds to FPN to promote its degradation. Dysregulation of hepcidin gives rise to disordered iron homeostasis, associated with diverse diseases including anemia and β-thalassemia. Our previous study demonstrated that there is an estrogen response element (ERE) within the promoter of hepcidin gene and that its expression is regulated by estrogen. In the current study, we demonstrated that both PCB153 and PCB126 greatly suppress hepcidin expression in HepG2 cells, with a greater repression occurring in cells upon PCB126 treatment. Further studies uncovered that both PCB153 and PCB126 harbor estrogenic activity and that the estrogenic activity of PCB126 was stronger than that of PCB153 in HepG2 cells. Mechanistic investigation revealed that PCBs suppress hepcidin transcription through a functional ERE within the hepcidin promoter, analogous to the action of 17β-estradiol. Moreover, hepatic hepcidin was downregulated in wild-type mice upon PCB126 administration, coupled with elevated serum iron content as well as reduced hepatic and splenic iron mass. These changes were not replicated in hepcidin-deficient mice upon PCB administration. Additionally, hepatocytes were observed with severe accumulation of lipid droplets in the livers of mice challenged with PCB126, irrespective of the presence of hepcidin. To summarize, our results have deciphered a suppressive role of PCBs in restraining the expression of hepcidin through mimicking estrogenic activity and revealed a novel property of PCBs in disrupting systemic iron metabolism. This study also unearthed a PCB-mediated connection linking estrogen-like activity, iron effects, and lipid homeostasis.

As the substitute of polybrominated diphenyl ethers (PBDEs), further assessments about the potential ecological safety and health risks of phosphorus-containing flame retardants (PFRs) are required because the worldwide demand for PFRs has been increasing every year. In this study, we examined the agonistic/antagonistic activity of a group of PFRs by three in vitro models (luciferase reporter gene assay, yeast two-hybrid assay, and E-screen assay). Molecule docking was used to further explain the interactions between ERα and PFRs. Data from luciferase reporter gene analysis showed three members of the nine tested PFRs significantly induced estrogenic effects, with the order of TPP > TCP > TDCPP, while TCEP and TEHP have remarkable antiestrogenic properties with calculated REC20 and RIC20 values of 10–6 M or lower. Results from the luciferase reporter gene method are generally consistent with results obtained from the yeast two-hybrid assay and E-screen, except for the positive estrogenic activity of TBP in E-screen testing. Docking results showed that binding between ligands and ERα was stabilized by hydrophobic interactions. As a proposed alternative for brominated flame retardant, PFRs may have anti/estrogenic activity via ERα at the low dose typical of residue in environmental matrix or animals. PFRs with a short chain, halogen, and benzene ring in the substituent group tend to be estrogenic. Our research suggests that comprehensive evaluations, including health and ecological assessments, are required in determining whether PFRs are preferable as an emerging industrial substitute.

p,p′-DDE, the main metabolite of DDT, is notorious for its persistent and bioaccumulation. It has detrimental effects on the nervous system, while the mechanism is unclear. We sought to investigate the mechanism of p,p′-DDE-induced neurocytic apoptosis in PC12 cells by cytoflow and screen the potential target gene by microarray and ELISA. Co-incubation with antagonist and SiRNA were applied to confirm the effect of the selected molecular. Results were also confirmed in zebrafish embryo. Results showed that p,p′-DDE induced apoptosis in PC12 cells at a concentration of ≥2 × 10–5 mol/L. Microarray results indicate that the TNF family plays a key role in p,p′-DDE-induced apoptosis among 84-apoptotic genes. In particular, the protein level of TNFα increased 4-fold. When incubated with TNFα antibody (infliximab), the number of apoptotic cells attenuated by 50%, and both activities of caspases 8 and 9 decreased. SiRNA silencing of TNFα showed similar trends. Furthermore, p,p′-DDE induced neuronal apoptosis in zebrafish embryos in a dose-dependent manner.This effect was partially reversed by infliximab, too. Overall, the present study herein indicated that the TNFα signaling pathway is involved in p,p′-DDE-induced neurocyte apoptosis. These data could be expanded to other cases of OCP-induced apoptosis and would support the need for scientific intervention to address the neurotoxicity of these chemicals.

The increased release of chiral persistent organic pollutants (POPs) into the environment has resulted in more attention to the role of enantioselectivity in the fate and ecotoxicological effects of these compounds. Although the enantioselectivity of chiral POPs has been considered in previous studies, little effort has been expended to discern the enantiospecific effects of chiral POPs metabolites, which may impede comprehensive risk assessments of these chemicals. In the present study, o,p′-DDD, the chiral metabolite of o,p′-DDT, was used as a model chiral metabolite. First, a preferential chiral separation at 100% ethanol was employed to obtain a pure enantiomer. The enantioselective cytotoxicity of o,p′-DDD in rat cells (PC12) was evaluated by detecting activation of the cellular apoptosis and oxidative stress systems and microarray analysis. We have documented for the first time that R-(+)-o,p′-DDD increases apoptosis by selectively disturbing the oxidative system (enzymes and molecules) and regulating the transcription of Aven, Bid, Cideb and Tp53. By comparing the data from the present study to data derived from the parent compound, we concluded that the R-enantiomer is the more detrimental stereostructure for both o,p′-DDT and o,p′-DDD. This observed stereostructural effect is in line with the structure–activity relationship formulated at other structural levels. Biological activities of the chiral metabolites are likely to occur in the same absolute configuration between chiral POPs and their metabolites provided that they have the similar stereostructures.

Diclofop-methyl (DM) is a widely used chiral herbicide, which rapidly hydrolyzes to its major metabolite diclofop acid (DC) after application. With a carbon chiral center, DC not only is an important ingredient of herbicidal activity, but also has a long half-life in soil. Studies so far have only considered the activity of racemic DM in target organisms, and the enantioselective toxicity in nontarget plants of DM and DC has yet to be explored. In this study, the enantioselective phytotoxicity of DC mediated by oxidative stress and the key enzyme ACCase in the fatty acid synthesis system on the model plant Arabidopsis thaliana was investigated. Significant differences between the two enantiomers were observed in phytotoxicity including growth inhibition, oxidative damage and alteration of key genes expression of ACCase, with R-DC showing greater toxicity to Arabidopsis thaliana than S-DC. The results of molecular docking showed that there was a stronger affinity between R-DC and the target enzyme carboxyltransferase domain of ACCase, likely leading to the enantioselective phytotoxicity of DC. This study suggested that chirality of both parent compounds and metabolites should be considered to improve our understanding of the environmental fate and risks of chiral pesticides.

•All three SCCPs exert potential estrogenic activities via estrogen receptor α.•The C11-43.20% showed glucocorticoid receptor-mediated antagonistic activity.•The SCCPs disturbed the genes involved in synthesis of steroid hormones.Short-chain chlorinated paraffins (SCCPs), which are candidate persistent organic pollutants (POPs) according to the Stockholm Convention, are of great concern because of their persistent bioaccumulation, long-range transport and potential adverse health effects. However, data on the endocrine-disrupting effects of SCCPs remain scarce. In this study, we first adopted two in vitro models (reporter gene assays and H295R cell line) to investigate the endocrine-disrupting effects of three SCCPs (C10-40.40%, C10-66.10% and C11-43.20%) via receptor mediated and non-receptor mediated pathway. The dual-luciferase reporter gene assay revealed that all test chemicals significantly induced estrogenic effects, which were mediated by estrogen receptor α (ERα), in the following order: C11-43.20% > C10-66.10% > C10-40.40%. Notably, C10-40.40% and C10-66.10% also demonstrated remarkable anti-estrogenic activities. Only C11-43.20% showed glucocorticoid receptor-mediated (GR) antagonistic activity, with a RIC20 value of 2.6 × 10− 8 mol/L. None of the SCCPs showed any agonistic or antagonistic activities against thyroid receptor β (TRβ). Meanwhile, all test SCCPs stimulated the secretion of 17β-estradiol (E2). Both C10-66.10% and C11-43.20% increased the production of cortisol at a high level in H295R cell lines. In order to explore the possible mechanism underlying the endocrine-disrupting effects of SCCPs through the non-receptor pathway, the mRNA levels of 9 steroidogenic genes were measured by real-time polymerase chain reaction (RT-PCR). StAR, 17βHSD, CYP11A1, CYP11B1, CYP19 and CYP21 were upregulated in a concentration-dependent manner by all chemicals. The data provided here emphasized that comprehensive assessments of the health and ecological risks of emerging contaminants, such as SCCPs, are of great concern and should be investigated further.Download high-res image (247KB)Download full-size image

Silver nanoparticles (nanosilver, AgNPs) have been shown to induce toxicity in vitro and in vivo; however, the molecular bases underlying the detrimental effects have not been thoroughly understood. Although there are numerous studies on its genotoxicity, only a few studies have investigated the epigenetic changes, even less on the changes of histone modifications by AgNPs. In the current study, we probed the AgNP-induced alterations to histone methylation that could be responsible for globin reduction in erythroid cells. AgNP treatment caused a significant reduction of global methylation level for histone 3 (H3) in erythroid MEL cells at sublethal concentrations, devoid of oxidative stress. The ChIP-PCR analyses demonstrated that methylation of H3 at lysine (Lys) 4 (H3K4) and Lys 79 (H3K79) on the β-globin locus was greatly reduced. The reduction in methylation could be attributed to decreased histone methyltransferase DOT-1L and MLL levels as well as the direct binding between AgNPs to H3/H4 that provide steric hindrance to prevent methylation as predicted by the all-atom molecular dynamics simulations. This direct interaction was further proved by AgNP-mediated pull-down assay and immunoprecipitation assay. These changes, together with decreased RNA polymerase II activity and chromatin binding at this locus, resulted in decreased hemoglobin production. By contrast, Ag ion-treated cells showed no alterations in histone methylation level. Taken together, these results showed a novel finding in which AgNPs could alter the methylation status of histone. Our study therefore opens a new avenue to study the biological effects of AgNPs at sublethal concentrations from the perspective of epigenetic mechanisms.

•Potential liver injury of bifenthrin and its mechanism were investigated in vivo.•Classical clinical biomarkers of liver dysfunction showed no obvious change.•Oxidative stress-related genes were induced and the metabolic profile was disturbed.•Liver injury was induced via caspase-mediated mitochondrial-dependent cell death.•Multilevel evaluations of potential pyrethroid-induced liver toxicity are needed.The widespread use of pesticides, such as pyrethroids, increases health risks to non-target organisms. The potential toxicity of pyrethroids to the liver remains unclear and could be easily overlooked if only the common clinical indicators of liver disease are examined. In the present study, BALB/c mice were given intraperitoneal injections of 0, 2, 4, or 8 mg/kg bifenthrin (BF) for 7 days. The potential liver injury of BF and its underlying mechanism were then investigated through multilevel evaluations. Histological analyses and serum enzyme activities showed no obvious clinical evidence of liver damage. Oxidative stress was induced and caspases were activated in response to increased BF concentrations. Exposure to BF also significantly altered the expression levels of mitochondrial apoptosis-related genes in dose-dependent relationships. The microarray results showed that BF could disturb the metabolic profile and extensively induce genes related to oxidative stress, including the cytochrome P450 family, glutathione peroxidases, glutathione s-transferases and kinases. In the in vivo model, BF induced liver injury through caspase-mediated mitochondrial-dependent cell death, a process that is closely related to oxidative stress, even in the absence of classical clinical biomarkers of liver dysfunction. The results of this study suggest that classical evaluations are not adequate for liver toxicity of pyrethroids, and highlight the need for more comprehensive assessment of health risks of these widely used pesticides.Download full-size image

Previous studies have showed that some chiral pesticides with estrogenic activity possess enantioselectivity in endocrine disruption. Despite the assessment of enantioselectivity in the estrogenic potential of chiral pesticides, which deserve particular attention, there has been limited research into their molecular mechanisms of human health risk. In this study, the role of enantioselectivity in the endocrine disruption and potential human maternal–fetal health risk of acetofenate (AF), an organochlorine insecticide, were investigated in both MCF-7 and JEG-3 cells. The two in vitro assays showing a clear enantioselectivity in the estrogenic activity with S-(+)-AF showed stronger effects than R-(−)-AF and rac-AF. Moreover, the racemate's estrogenicity was in between that of enantiomers. Our results also demonstrated that S-(+)-AF possesses the strongest potential effects in disruption of hormone secretion, maternal immune tolerance, and steroidogenesis in the trophoblast. The results suggest that assessment of the health risk of chiral contaminants should consider the role of enantioselectivity.Highlights► We investigated the role of enantioselectivity of acetofenate in both MCF-7 and JEG-3 cells. ► The results showed a clear enantioselectivity in the estrogenic activity. S-(+)-AF exerted stronger effects than R-(−)-AF and rac-AF. ► These were expressed in disruption of hormone secretion and maternal immune tolerance.

According to some previous studies, titanium dioxide (TiO2) nanoparticles (NPs) would be of low to medium ecological toxicity in short-term tests. But the long-term risks of TiO2 NPs that is currently unknown. In this study, we used the zebrafish (Danio rerio) as in vivo model to assess the chronic toxicity of TiO2 NPs. Our results revealed that there are obvious adverse effect to zebrafish, including concentration-dependent and time-dependent inhibition of growth and decrease the liver weight ratio of zebrafish. Meanwhile, only the gills displayed histopathologic change including thickening of oedema and the gill lamellae. ICP-MS examination showed that TiO2 NPs were accumulated and distributed in gill, liver, heart as well as brain. Our present study suggested that the TiO2 NPs could translocate among organs and pass through the blood–brain and the blood–heart barrier after long-term exposure. Given the widespread use of TiO2 NPs, a systematic, coherent, and tested foundation for managing the uncertain health and environmental aspects of TiO2 NPs is necessary.