Dietary exposure to 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) induces oxidative damage promoting cell apoptosis primarily via mitochondrial pathway in the hepatopancreas of carp, Cyprinus carpio.

To investigate the mechanisms of BDE-47 on hepatotoxicity in fish, this study examined the effects of dietary exposure to BDE-47 (40 and 4000 ng/g) on carp for 42 days. The results showed that BDE-47 significantly increased carp's condition factor and hepatosomatic index. Pathological results revealed unclear hepatic cord structure, hepatocytes swelling, cellular vacuolization, and inflammatory cell infiltration in the hepatopancreas of carp. Further investigation showed that ROS levels significantly increased on days 7, 14, and 42. Moreover, the activities of antioxidant enzymes SOD, GSH, CAT, and GST increased significantly from 1 to 7 days, and the transcription levels of antioxidant enzymes CAT, Cu-Zn SOD, Mn-SOD, GST, and GPX, and antioxidant pathway genes Keap1, Nrf2, and HO-1 changed significantly at multiple time-points during the 42 days. The results of apoptosis pathway genes showed that the mitochondrial pathway genes Bax, Casp3, and Casp9 were significantly upregulated and Bcl2 was significantly downregulated, while the transcription levels of FADD and PERK were significantly enhanced. These results indicate that BDE-47 induced oxidative damage in hepatopancreas, then it promoted cell apoptosis mainly through the mitochondrial pathway. This study provides a foundation for analyzing the mechanism of hepatotoxicity induced by BDE-47 on fish.

2,2',4,4'-Tetrabromodiphenyl ether and cadmium co-exposure activates aryl hydrocarbon receptor pathway to induce ROS and GSDME-dependent pyroptosis in renal tubular epithelial cells.

We have previously found that a mixture exposure of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) and cadmium (Cd) causes kidney damage; however, the mechanism was not fully understood. The aryl hydrocarbon receptor (AhR) is a ligand-receptor transcription factor that plays an important role in the adaptive response or metabolic detoxification of environmental toxins. Thus, this study aimed to examine the role of AhR in kidney toxicity. BDE-47 (50 µM) or Cd (5 µM) exposure reduced cell viability in renal tubular epithelial cells (HKC), with a larger effect observed in co-treatment. The cell morphology presented pyroptotic changes, including swollen cells, large bubbles, and plasma membrane pore formation. The gene expressions of AhR, heat shock protein 90 (Hsp90), AhR nuclear translocator (ARNT), and cytochrome P450 1B1 (CYP1B1) were increased, while CYP1A1 was decreased. Reactive oxygen species (ROS) were generated, which was reduced by the AhR antagonist CH223191. The apoptosis, necrosis, and intracellular lactated hydrogenase (LDH) release was elevated, and this was attenuated by N-acetylcysteine (NAC). Furthermore, the pyroptosis pathway was activated with increased protein levels of cleaved-caspase-3 and gasdermin E N-terminal (GSDME-NT), while caspase-8, caspase-3, and GSDME were decreased. These effects were alleviated by NAC and CH223191. Our data demonstrate a combined effect of BDE-47 and Cd on nephrotoxicity by activating AhR to induce ROS contributing to GSDME-dependent pyroptosis, and retardation of the AhR pathway could reduce this toxicity.

Maternal Exposure to Low-Dose BDE-47 Induced Weight Gain and Impaired Insulin Sensitivity in the Offspring.

Polybrominated diphenyl ethers (PBDEs), commonly used as synthetic flame retardants, are present in a variety of consumer products, including electronics, polyurethane foams, textiles, and building materials. Initial evidence from epidemiological and experimental studies suggests that maternal PBDE exposure may be associated with a higher BMI in children, with disturbance of energy metabolism and an increased risk of Type 2 diabetes. However, the causality between early exposure to real-life PBDE concentrations and increased weight as well as mechanisms underlying impaired metabolic pathways in the offspring remain elusive. Here, using a mouse model we examined the effect of maternal exposure to 2,2',4,4'-tetrabrominated diphenyl ether (BDE-47), the most abundant congener in human samples, on offspring weight gain and energy homeostasis using a mouse model. Maternal exposure to BDE-47 at low dose resulted in weight gain in female offspring together with an impaired glucose and insulin tolerance in both female and male mice. In vitro and in vivo data suggest increased adipogenesis induced by BDE-47, possibly mediated by DNA hypermethylation. Furthermore, mRNA data suggest that neuronal dysregulation of energy homeostasis, driven via a disturbed leptin signaling may contribute to the observed weight gain as well as impaired insulin and glucose tolerance.

BDE-47 Causes Depression-like Effects in Zebrafish Larvae via a Non-Image-Forming Visual Mechanism.

Depression is a high-incidence mood disorder that is frequently accompanied by sleep disturbances, which can be triggered by the non-image-forming (NIF) visual system. Therefore, we hypothesize that polybrominated diphenyl ethers are known to induce visual impairment that could promote depression by disrupting the NIF visual pathway. In this study, zebrafish larvae were exposed to BDE-47 at environmentally relevant concentrations (2.5 and 25 µg/L). BDE-47 caused melanopsin genes that dominate the NIF visual system that fell at night (p < 0.05) but rose in the morning (p < 0.05). Such bidirectional difference transmitted to clock genes and neuropeptides in the suprachiasmatic nucleus and impacted the adjacent serotonin system. However, indicative factors of depression, including serta, htr1aa, and aanat2, were unidirectionally increased 1.3- to 1.6-fold (p < 0.05). They were consistent with the increase in nighttime thigmotaxis (p < 0.05) and circadian hypoactivity (p < 0.05). The results of melanopsin antagonism also indicated that these consequences were possibly due to the combination of direct photoentrainment by melanopsin and circadian disruption originating from melanopsin. Collectively, our findings revealed that BDE-47 exposure disrupted the NIF visual pathway and resulted in depression-like effects, which may further exert profound health effects like mood disorders.

Metabolomic insights into neurological effects of BDE-47 exposure in the sea cucumber Apostichopus japonicus.

The persistent organic pollutant 2,2',4,4'-Tetrabromodiphenyl ether (BDE-47), a prevalent congener among polybrominated diphenyl ethers (PBDEs), exhibits potent bioaccumulation and toxicity. Despite extensive research into the adverse effects of BDE-47, its neurotoxicity in sea cucumbers remains unexplored. Given the crucial role of the sea cucumber's nervous system in survival and adaptation, evaluating the impacts of BDE-47 is vital for sustainable aquaculture and consumption. In this study, we employed ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Triple-TOF-MS) to analyze metabolomic changes in neuro-related tissues of Apostichopus japonicus exposed to low (0.1 µg/L), medium (1.0 µg/L), and high (10.0 µg/L) BDE-47 concentrations. We identified significantly changed metabolites in each exposure group (87 in low, 79 in medium, and 102 in high), affecting a variety of physiological processes such as steroid hormone balance, nucleotide metabolism, energy metabolism, neurotransmitter levels, and neuroprotection. In addition, we identified concentration-dependent, common, and some other metabolic responses in the neuro-related tissues. Our findings reveal critical insights into the neurotoxic effects of BDE-47 in sea cucumbers and contribute to risk assessment related to BDE-47 exposure in the sea cucumber industry, paving the way for future neurotoxicological research in invertebrates.

Multi-omics analysis reveals BDE47 induces depression-like behaviors in mice by interfering with the 2-arachidonoyl glycerol-associated microbiota-gut-brain axis.

2,2',4,4'-tetrabromodiphenyl ether (BDE47) is a foodborne environmental risk factor for depression, but the pathogenic mechanism has yet to be fully characterized. In this study, we clarified the effect of BDE47 on depression in mice. The abnormal regulation of the microbiome-gut-brain axis is evidenced closely associated with the development of depression. Using RNA sequencing, metabolomics, and 16s rDNA amplicon sequencing, the role of the microbiome-gut-brain axis in depression was also explored. The results showed that BDE47 exposure increased depression-like behaviors in mice but inhibited the learning memory ability of mice. The RNA sequencing analysis showed that BDE47 exposure disrupted dopamine transmission in the brain of mice. Meanwhile, BDE47 exposure reduced protein levels of tyrosine hydroxylase (TH) and dopamine transporter (DAT), activated astrocytes and microglia cells, and increased protein levels of NLRP3, IL-6, IL-1ß, and TNF-α in the brain of mice. The 16 s rDNA sequencing analysis showed that BDE47 exposure disrupted microbiota communities in the intestinal contents of mice, and faecalibaculum was the most increased genus. Moreover, BDE47 exposure increased the levels of IL-6, IL-1ß, and TNF-α in the colon and serum of mice but decreased the levels of tight junction protein ZO-1 and Occludin in the colon and brain of mice. In addition, the metabolomic analysis revealed that BDE47 exposure induced metabolic disorders of arachidonic acid and neurotransmitter 2-Arachidonoyl glycerol (2-AG) was one of the most decreased metabolites. Correlation analysis further revealed gut microbial dysbiosis, particularly faecalibaculum, is associated with altered gut metabolites and serum cytokines in response to BDE47 exposure. Our results suggest that BDE47 might induce depression-like behavior in mice through gut microbial dysbiosis. The mechanism might be associated with the inhibited 2-AG signaling and increased inflammatory signaling in the gut-brain axis.

Cadmium (Cd) and 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) co-exposure induces acute kidney injury through oxidative stress and RIPK3-dependent necroptosis.

Environmental pollution is complex, and co-exposure can accurately reflect the true environmental conditions that are important for assessment of human health. Cadmium (Cd) is a widespread toxicant that can cause acute kidney injury (AKI), while its combined effect with 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) is not fully understood. Thus, we used an in vivo model where C57BL/6J mice were treated with low dietary intake of Cd (5 mg/kg/day) and/or BDE-47 (1 mg/kg/day) for 28 days to examine AKI, and in vitro experiments to investigate the possible mechanism. Results showed that Cd or BDE-47 caused pathological kidney damage, accompanied by elevated urea nitrogen (BUN) and urinary creatinine, as well as increased interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α), and reduced IL-10 in kidney tissues. In vitro Cd or BDE-47 exposure decreased cell viability and induced cell swelling and blebbing of human embryonic kidney 293 (HEK-293) and renal tubular epithelial cell lines (HKCs), and changes in co-exposure was larger than that in Cd and BDE-47 treatment. Oxidative stress indicators of the reactive oxygen species (ROS) and malondialdehyde (MDA) were elevated, while the antioxidant superoxide dismutase (SOD) was decreased. Necrosis occurred with increased lactate dehydrogenase (LDH) release and propidium iodide (PI) staining, which was attenuated by the ROS scavenger N-acetyl-L-cysteine (NAC). Furthermore, necroptotic genes of receptor-interacting protein kinase-3 (RIPK3), classical mixed lineage kinase domain-like protein-dependent (MLKL), IL-1ß and TNF-α were up-regulated, whereas RIPK1 was down-regulated, which was attenuated by the RIPK3 inhibitor GSK872. These findings demonstrate that Cd or BDE-47 alone produces kidney toxicities, and co-exposure poses an additive effect, resulting in AKI via inducing oxidative stress and regulating RIPK3-dependent necroptosis, which offers a further mechanistic understanding for kidney damage, and the combined effect of environmental pollutants should be noticed.

Purine Metabolism and Pyrimidine Metabolism Alteration Is a Potential Mechanism of BDE-47-Induced Apoptosis in Marine Rotifer Brachionus plicatilis.

This present study was conducted to provide evidence and an explanation for the apoptosis that occurs in the marine rotifer Brachionus plicatilis when facing 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) stress. Metabolomics analysis showed that aminoacyl-tRNA biosynthesis, valine, leucine and isoleucine biosynthesis, and arginine biosynthesis were the top three sensitive pathways to BDE-47 exposure, which resulted in the reduction in the amino acid pool level. Pyrimidine metabolism and purine metabolism pathways were also significantly influenced, and the purine and pyrimidine content were obviously reduced in the low (0.02 mg/L) and middle (0.1 mg/L) concentration groups while increased in the high (0.5 mg/L) concentration group, evidencing the disorder of nucleotide synthesis and decomposition in B. plicatilis. The biochemical detection of the key enzymes in purine metabolism and pyrimidine metabolism showed the downregulation of Glutamine Synthetase (GS) protein expression and the elevation of Xanthine Oxidase (XOD) activity, which suggested the impaired DNA repair and ROS overproduction. The content of DNA damage biomarker (8-OHdG) increased in treatment groups, and the p53 signaling pathway was found to be activated, as indicated by the elevation of the p53 protein expression and Bax/Bcl-2 ratio. The ROS scavenger (N-acetyl-L-cysteine, NAC) addition effectively alleviated not only ROS overproduction but also DNA damage as well as the activation of apoptosis. The combined results backed up the speculation that purine metabolism and pyrimidine metabolism alteration play a pivotal role in BDE-47-induced ROS overproduction and DNA damage, and the consequent activation of the p53 signaling pathway led to the observed apoptosis in B. plicatilis.

In vitro immunotoxicity and possible mechanisms of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) on Ruditapes philippinarum hemocytes.

Marine bivalves can accumulate large amounts of pollutants from sea water, sediments and microalgae due to their filter-feeding habits. BDE-47 is often the most highly concentrated congener in bivalves. BDE-47 has been found to have toxic effects on bivalves, however, the immunotoxicity and the underlying mechanisms of BDE-47 on bivalves are not well understood yet. In this study, isolated hemocytes of Manila clam Ruditapes philippinarum were exposed to five concentrations of BDE-47 (6.25 µM, 12.5 µM, 25 µM, 50 µM, 100 µM), the effects of BDE-47 on hemocyte survival rate, cell viability, granulocyte ratio, phagocytosis, bacteriolytic activity, reactive oxygen species (ROS), lysosomal membrane permeability (LMP), superoxide dismutase (SOD), and phosphorylation state of extracellular regulated protein kinase (ERK) and p38 at 2 h, 6 h and 12 h were studied. The results indicated that BDE-47 exposure declined the hemocyte cell viability, reduced the granulocyte ratio, hampered the hemocyte phagocytosis and bacteriolytic activity, elevated the ROS levels, increased the LMP, significantly changed SOD expression and depressed the phosphorylation levels of ERK and p38. Taken together, the results demonstrated that BDE-47 had significant toxic effects on the immune function, and the immunotoxicity may partly via the overproduction of ROS and the alteration of MAPK signaling pathways.

Toxic effects of polystyrene nanoplastics and polybrominated diphenyl ethers to zebrafish (Danio rerio).

Nanoplastics (NPs) are good carriers of persistent organic pollutants (POPs) such as polybrominated diphenyl ethers (PBDEs), and can alter their bioavailability and toxic impacts to aquatic organisms. This study highlights the single and combined toxic effects of polystyrene nanoplastics (PS-NPs) and 2,2',4,4'-tetrabromodiphenyl ether (BDE-47, one of the dominant congeners of PBDEs) on zebrafish embryos after an exposure duration of up to 120 hpf. Results showed that PS-NPs and BDE-47 co-exposure exacerbated the morphological deformities in terms of pericardial edema, yolk sac edema and curved tail in zebrafish larvae. Compared to BDE-47 single exposure, the combined exposure caused lower survival rates, shorter body lengths, and accelerated spontaneous movements. Further, PS-NPs were quickly aggregated on the surface of the embryonic chorions covered almost the entire membrane at 12 and 48 hpf, and concentration dependent accumulation was also found in the brain, mouth, trunk, gills, heart, liver and gastrointestinal tract at the larval stages. During the recovery period (7 days), PS-NPs were released from all the organs, with the highest elimination from the gastrointestinal tract. Histopathological examination revealed that co-exposure caused greater damage to retinal structures, muscle fibers and cartilage tissues. Responses of hypothalamic-pituitary-thyroid axis (CRH, TSHß, NIS, TTR, Dio2, TG, TRα and TRß) and reproduction (Esr2 and Vtg1) related genes were also investigated, and results showed that the co-exposure induced more significant upregulated expressions of TSHß, TG, Doi 2, and TRß, compared to BDE-47 single exposure. In conclusion, co-exposure to NPs and BDE-47 exacerbated developmental and thyroid toxicity in zebrafish, generally elucidating the toxicological effects mediated by complex chemical interactions between NPs with POPs in the freshwater environment.

Behavioral changes and transcriptomic effects at embryonic and post-embryonic stages reveal the toxic effects of 2,2',4,4'-tetrabromodiphenyl ether on neurodevelopment in zebrafish (Danio rerio).

Polybrominated biphenyl ethers (PBDEs) are new persistent pollutants that are widely exist in the environment and have many toxic effects. However, their toxicity mechanisms on neurodevelopment are still unclear. In this study, zebrafish embryos were exposed to 2, 2', 4, 4'-tetrabromodiphenyl ether (BDE-47) (control, 10, 50 and 100 µg/L) at 2 h postfertilization (hpf) - 7 dpf. Locomotion analysis indicated that BDE-47 increased spontaneous coiling activity in zebrafish embryos under high-intensity light stimuli and decreased locomotor in zebrafish larvae. RNA-Seq analysis revealed that most of the up-regulated pathways were related to the metabolism of cells and tissues, while the down-regulated pathways were related to neurodevelopment. Consistent with the locomotion and KEGG results, BDE-47 affected the expression of genes for central nervous system (gfap, mbpa, bdnf & pomcb), early neurogenesis (neurog1 & elavl3), and axonal development (tuba1a, tuba1b, tuba1c, syn2a, gap43 & shha). Furthermore, BDE-47 interfered with gene expression of the Wnt signaling pathway, especially during embryonic stages, suggesting that the mechanisms of BDE-47 toxicity to zebrafish at various stages of neurodevelopment may be different. In summary, early neurodevelopment effects and metabolic disturbances may have contributed to the abnormal neurobehavioral changes induced by BDE-47 in zebrafish embryos/larvae, suggesting the neurodevelopmental toxicity of BDE-47.

Transcriptomics-based analysis of co-exposure of cadmium (Cd) and 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) indicates mitochondrial dysfunction induces NLRP3 inflammasome and inflammatory cell death in renal tubular epithelial cells.

Environmental pollution often releases multiple contaminants resulting in as yet largely uncharacterized additive toxicities. Cadmium (Cd) is a widespread pollutant that induces nephrotoxicity in animal models and humans. However, the combined effect of Cd in causing nephrotoxicity with 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), a typical congener of polybrominated diphenyl ethers (PBDEs), has not been evaluated and mechanisms are not completely clear. Here, we applied transcriptome sequencing analysis to investigate the combined toxicity of Cd and BDE-47 in the renal tubular epithelial cell lines HKCs. Cd or BDE-47 exposure decreased cell viability in a dose-dependent manner, and exhibited cell swelling and rounding similar to necrosis, which was exacerbated by co-exposure. Transcriptomic analysis revealed 2191, 1331 and 3787 differentially-expressed genes following treatment with Cd, BDE-47 and co-exposure, respectively. Interestingly, functional annotation and enrichment analyses showed involvement of pathways for oxidative stress, NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome and inflammatory cell death for all three treatments. Examination of indices of mitochondrial function and oxidative stress in HKC cells showed that the levels of reactive oxygen species (ROS), malondialdehyde (MDA) and intracellular calcium ion concentration [Ca2+]i were elevated, while superoxide dismutase (SOD) and mitochondrial membrane potential (MMP) were decreased. The ratio of apoptotic and necrotic cells and intracellular lactate dehydrogenase (LDH) release were increased by Cd or BDE-47 exposure, and was aggravated by co-exposure, and was attenuated by ROS scavenger N-Acetyl-L-cysteine (NAC). NLRP3 inflammasome and pyroptosis pathway-related genes of NLRP3, adaptor molecule apoptosis-associated speck-like protein (ASC), caspase-1, interleukin-18 (IL-18) and IL-1ß were elevated, while gasdermin D (GSDMD) was down-regulated, and protein levels of NLRP3, cleaved caspase-1 and cleaved GSDMD were increased, most of which were relieved by NAC. Our data demonstrate that exposure to Cd and BDE-47 induces mitochondrial dysfunction and triggers NLRP3 inflammasome and GSDMD-dependent pyroptosis leading to nephrotoxicity, and co-exposure exacerbates this effect, which could be attenuated by inhibiting ROS. This study provides a further mechanistic understanding of kidney damage, and co-exposure impact is worthy of concern and should be considered to improve the accuracy of environmental health assessment.

Melatonin relieves 2,2,4,4-tetrabromodiphenyl ether (BDE-47)-induced apoptosis and mitochondrial dysfunction through the AMPK-Sirt1-PGC-1α axis in fish kidney cells (CIK).

Polybrominated diphenyl ethers (PBDEs) exist in aquatic environments with nephrotoxicity to non-target aquatic species. Melatonin (MT) exhibits an inhibitory effect of oxidative stress and apoptosis in various diseases. 2, 2', 4, 4'-tetrabromodiphenyl ether (BDE-47) is the main homolog of PBDE samples. Therefore, we investigated the toxic mechanism of BDE-47 and the alleviation effect of MT, the ctenopharyngodon idellus kidney (CIK) cells were treated with BDE-47 (100 µM) and/or MT (60 µM) for 24 h. Firstly, BDE-47 exposure could inhibit oxidative stress-related antioxidant enzymes (T-AOC, SOD, CAT and GPx) and increase the content of malondialdehyde (MDA) to cause oxidative stress. Secondly, BDE-47 enhanced mitochondrial division and inhibited fusion to induce mitochondrial membrane potential in CIK cells. BDE-47 enhanced the mRNA and protein levels of mitochondrial-pathway apoptosis related genes (Cas 3, Cyt-c, and BAX). Thirdly, BDE-47 treatment decreased the expression levels of mitochondrial-related regulatory factors AMPK-Sirt1-PGC-1α signal pathway. Intriguingly, BDE-47-induced oxidative stress, mitochondrial pathway apoptosis and mitochondrial dynamics disorder could be alleviated by MT treatment. Overall, we concluded that MT could relieve BDE-47-induced oxidative stress, mitochondrial dysfunction and apoptosis through the AMPK-Sirt1-PGC-1α axis. These results enrich the mechanisms of BDE-47 poisoning and reveal that MT treatment may be a potential strategy for solving BDE-47 poisoning.

Effects of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) on reproductive and endocrine function in female zebrafish (Danio rerio).

BACKGROUND: Polybrominated diphenyl ethers (PBDEs), a group of brominated flame retardants (BFRs), were reported exist extensively in various ecological environmental. Studies have indicated that PBDEs induce reproductive toxic effects on human health, but the mechanisms remain poorly understood. In this study, the adult female zebrafish were used to investigate the effects of 2, 2', 4, 4'-tetrabromodiphenyl ether (BDE-47) on the reproductive endocrine system and its mechanism. METHODS: Female zebrafish (AB strains) were continuously exposed to BDE-47 at the concentrations of 0, 10, 50, 100 and 500 µg/L till 21 days. The morphology of ovary were stained and evaluated with hematoxylin-eosin (H&E), and levels of sex hormones including follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone (T) and 17ß-estradiol (E2) and the biomarkers of oxidative stress such as superoxide dismutase (SOD) and malondialdehyde (MDA), were measured via ELISA. Subsequently, the expression of genes along the hypothalamic pituitary-gonad (HPG) and oxidative stress were determined using quantitative real-time PCR (qRT-PCR). RESULT: The results showed that exposure to high level of BDE-47 reduced the index of condition factor (CF) and gonadosomatic index (GSI). Treatment with BDE-47 impaired the normal development and structure of oocytes in zebrafish ovary. Moreover, the steroid hormone of FSH, LH, T and E2 were significantly decreased in BDE-47 exposure group. A dose-dependent elevation in SOD activity and MDA levels were recorded. Meanwhile, the transcription level of cyp19a, cyp19b, fshß, lhß were up-regulated while the transcription of fshr, lhr, cyp17a, 17ßhsd were down-regulated in the gonad of female adult zebrafish. CONCLUSION: Exposure to BDE-47 have detrimental impact on the development of ovary, decreasing sex hormone levels, inducing oxidative damage as well as altering HPG axis-related genes.

Establishment of a comprehensive method to derive seawater quality criteria of BDE-47 in China.

2,2',4,4'-tetrabromodiphenyl ether (BDE-47) is an emerging toxic organic pollutant widely detected in territorial waters. Accordingly, establishing the seawater quality criteria (SWQC) for BDE-47 is of significant importance to protect the marine ecosystems. In this study, published ecotoxicity data of BDE-47 to aquatic species in China were collected, and acute and chronic toxicity tests were carried out on saltwater aquatic organisms from 5 phyla and 8 families widely existed in the Chinese marine environment. Static acute toxicity tests and renewed chronic toxicity tests were adopted. The scientific theories and technical methods of the United States Environmental Protection Agency (US EPA), Canada, the European Union (EU), and the Netherlands on water quality criteria guidelines, as well as the Chinese freshwater quality criteria guidelines were compared. Then an integrated method of SWQC derivation was introduced through comprehensive consideration. Afterward, the SWQC of BDE-47 was derived based on the ecotoxicity data. The SWQC includes short-term seawater quality criteria (S-SWQC), long-term seawater quality criteria (L-SWQC), and serious risk concentration for the ecosystem (SRCeco). And they were derived and recommended as 7.90 µg/L, 0.217 µg/L, and 3.65 µg/L, respectively. This study served as a specific example to quantitatively studies the differences between different scientific theories and technique methods. The derivation process and improvement of SWQC for BDE-47 provide support for the future revision of water quality criteria in China.

2, 2', 4, 4'-tetrabromodiphenyl ether induces placental toxicity via activation of p38 MAPK signaling pathway in vivo and in vitro.

2, 2', 4, 4'-tetrabromodiphenyl ether (BDE-47) is one of the most important polybrominated diphenyl ethers (PBDEs) congeners, and epidemiological studies have shown that it can cause adverse pregnancy outcomes. The aim of our study was to investigate the role of placental injury in BDE-47-induced adverse pregnancy outcomes through in vivo and in vitro models. From day 0.5 to day 16.5 of pregnancy of ICR mice, BDE-47 oral doses of 0, 25, 50 and 100 mg/kg/day were administered. Immunohistochemical staining found that BDE-47 inhibited the expression of CD34 in mouse placenta, and ELISA results showed that BDE-47 reduced the levels of VEGF and PlGF in the serum of pregnant mice. Western blot assays found that the expression levels of VEGF-A and invasion-related factors were decreased in the placentas of BDE-47-treated group, which indicated that BDE-47 could impair placental angiogenesis. Furthermore, BDE-47 inhibited proliferation, increased apoptosis and autophagy, and activated p38 MAPK signaling pathway in mouse placental tissue. In vitro, HTR-8/SVneo cells were treated with 0, 5, 10, 20 µM BDE-47 for 24 h. Wound healing assays and Transwell assays showed that BDE-47 inhibited the migration and invasion ability of HTR-8/SVneo cells. We also found that BDE-47 inhibited the proliferation of HTR-8/SVneo cells and increased apoptosis and autophagy. BDE-47 activated p38 MAPK signaling pathway in HTR-8/SVneo cells, and inhibition of p38 MAPK signaling pathway in HTR-8/SVneo cells restored the effects caused by BDE-47. In conclusion, BDE-47 impairs placental angiogenesis by inhibiting cell migration and invasion, and induces placental toxicity by inhibiting proliferation, increasing apoptosis and autophagy. In vitro, activation of p38 MAPK signaling pathway is involved in the processes of placental injury by BDE-47.

2,2',4,4'-Tetrabromodiphenyl ether disrupts spermatogenesis in mice by interfering with the ER-Nrf1-Tfam-mitochondria pathway.

2,2',4,4' -tetrabromodiphenyl ether (BDE47), a well-known endocrine disruptor of the estrogen receptor (ER) is toxic to the mitochondria and spermatogenesis. This study aimed to explore the mechanism of BDE47 on spermatogenesis in mammals. Adult male Institute of Cancer Research (ICR) mice were gavaged daily with BDE47 (0, 1, or 10 mg/kg bw) for 8 weeks. Testicular weight, sperm production and motility, morphology of spermatogenic cells, nuclear respiratory factor 1 (Nrf1) level, and its expression in testes were determined. In vitro, cell viability, and key molecules in the ER-Nrf1-mitochondrial transcription factor A (Tfam)-mitochondria pathway in the immortalized mouse spermatogonia line (GC1) were determined at 48 h and 0-5 h after exposure; RNA interference (RNAi) was also performed to verify that the decreased Nrf1 was associated with mitochondrial dysfunction and the impaired viability of germ cells. The results indicated that BDE47 impaired testis weight and spermatogenesis, impaired mitochondria and germ cells, and decreased Nrf1 in the testes of mice. In vitro, after 48 h exposure, BDE47 reduced cell viability, Nrf1 protein, and mRNA of Nrf1, Tfam, ATP synthase subunit ß (Atp5b), and cytochrome c oxidase subunit I (mt-CO1) in GC1 while also reducing mRNA of Nrf1 and Tfam promptly (from 1 to 5 h) after exposure. Furthermore, Nrf1 RNA interference decreased viability and mitochondrial function in GC1. These results indicated that BDE47 disrupts spermatogenesis in mice, probably by interfering with the ER-Nrf1-Tfam-mitochondria pathway, and Nrf1 is a target molecule of BDE47 estrogen receptor.

Bidirectional regulation of BDE-47 on 3T3-L1 cell differentiation based on a restricted cubic spline model.

BDE-47 (2,2,4,4-tetrabromodiphenyl ether) is a polybrominated diphenyl ether (PBDE) congener, which has the characteristics of high biological detection rate, the highest content and strong biological toxicity, and is widely distributed in organisms. Many studies have found that BDE-47 may also be an environmental risk factor for metabolic diseases such as obesity, insulin resistance, type 2 diabetes, and hypertension. However, the way that PBDEs influence adipocyte differentiation remains unclear. The methylisobutylxanthine, dexamethasone, and insulin method was used to study the effect of BDE-47 on the differentiation of 3T3-L1 cells. The 3T3-L1 cells were exposed by different concentrations of BDE-47, and the effect of cell viability was detected at different stages. In addition, the lipid droplet aggregation of adipocytes was observed and the triglyceride (TG) levels in the cytoplasm were detected after differentiation. The relative mRNA expression levels of leptin, adiponectin, and PPARγ in cells were determined by RT-PCR, and differentially expressed genes were preliminarily screened by digital gene expression profile. Our study found that BDE-47 promoted the differentiation of 3T3-L1 cells. Restriction cubic spline analysis showed that BDE-47 bidirectionally. regulated the mRNA synthesis of TG, PPARγ, and leptin genes and the aggregation of lipid droplets. BDE-47 may induce adipocyte differentiation by activating PPARγ, resulting in the differential expression of genes related to the AMPK signaling pathway, insulin resistance, and other metabolic pathways. The highest and lowest-dose BDE-47 exposure groups had the greatest impact on adipocyte differentiation.

Certain ortho-hydroxylated brominated ethers are promiscuous kinase inhibitors that impair neuronal signaling and neurodevelopmental processes.

The developing nervous system is remarkably sensitive to environmental signals, including disruptive toxins, such as polybrominated diphenyl ethers (PBDEs). PBDEs are an environmentally pervasive class of brominated flame retardants whose neurodevelopmental toxicity mechanisms remain largely unclear. Using dissociated cortical neurons from embryonic Rattus norvegicus, we found here that chronic exposure to 6-OH-BDE-47, one of the most prevalent hydroxylated PBDE metabolites, suppresses both spontaneous and evoked neuronal electrical activity. On the basis of our previous work on mitogen-activated protein kinase (MAPK)/extracellular signal-related kinase (ERK) (MEK) biology and our observation that 6-OH-BDE-47 is structurally similar to kinase inhibitors, we hypothesized that certain hydroxylated PBDEs mediate neurotoxicity, at least in part, by impairing the MEK-ERK axis of MAPK signal transduction. We tested this hypothesis on three experimental platforms: 1) in silico, where modeling ligand-protein docking suggested that 6-OH-BDE-47 is a promiscuous ATP-competitive kinase inhibitor; 2) in vitro in dissociated neurons, where 6-OH-BDE-47 and another specific hydroxylated BDE metabolite similarly impaired phosphorylation of MEK/ERK1/2 and activity-induced transcription of a neuronal immediate early gene; and 3) in vivo in Drosophila melanogaster, where developmental exposures to 6-OH-BDE-47 and a MAPK inhibitor resulted in offspring displaying similarly increased frequency of mushroom-body ß-lobe midline crossing, a metric of axonal guidance. Taken together, our results support that certain ortho-hydroxylated PBDE metabolites are promiscuous kinase inhibitors and can cause disruptions of critical neurodevelopmental processes, including neuronal electrical activity, pre-synaptic functions, MEK-ERK signaling, and axonal guidance.

Interaction of BDE-47 with nuclear receptors (NRs) based on the cytotoxicity: In vitro investigation and molecular interaction.

Polybrominated diphenyl ethers (PBDEs) are endocrine-disrupting chemicals that possess neuroendocrine and reproductive toxicity to humans and disturb thyroid hormone homeostasis, neurobehavior, and development. The most predominant congener of PBDEs in humans and other organisms is 2,2',4,4'-tetrabromodiphenyl ether (BDE-47); however, the molecular mechanisms underlying its cytotoxicity remain largely unknown. Here, we evaluated the toxic effect and underlying mechanism of nuclear receptors (NRs) induced by BDE-47 in SK-N-SH human neuroblastoma cells. The CCK-8 cell viability assay showed that the proliferation of human SK-N-SH cells exposed to BDE-47 was significantly inhibited in time- and dose-dependent manners, and flow cytometry showed that cell cycle was arrested at the S phase after BDE-47 exposure. Moreover, compared with the control group, the expression of retinoic acid receptor alpha (RXRα), pregnane X receptor (PXR), thyroid hormone receptors (TRs), and peroxisome proliferator-activated receptors (PPARs) at the mRNA and protein levels was significantly increased, as determined by quantitative PCR and western blot analysis, demonstrating that BDE-47 activated the NRs in vitro. Moreover, BDE-47 could bind to all four NRs in the affinity order of PPARγ > PXR > TRß > RXRα under molecular dynamics. Because RXR is the promiscuous dimerization partner for a large number of NRs, ZDock was used to calculate its interaction with other three NRs. Taking the number of hydrogen bonds and ZDock scores into account, the rank of docking ability between RXRα and the NRs was PXR > TRß > PPARγ. Further analysis of the interaction between BDE-47 and dimerized-NRs, the affinity order was RXRα > TRß > PXR > PPARγ via Glide. The results of this study demonstrated that BDE-47 interfered the cross-talk among NRs, especially the promiscuous RXRα, which might be critical for the harmonized re-adjustment of cytotoxicity and biological regulation. Our findings provide a better understanding of the mechanisms underlying toxic effects and intermolecular interaction induced by BDE-47.