To 2,3,7,8-tetrachlorodibenzo-p-dioxin Research Articles

Effect of exposure to 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) and polychlorinated biphenyls (PCBs) on mitochondrial DNA (mtDNA) copy number in rats

Mitochondria are intracellular organelles responsible for biological oxidation and energy production. These organelles are susceptible to damage from oxidative stress and compensate for damage by increasing the number of copies of their own genome, mitochondrial DNA (mtDNA). Cancer and environmental exposure to some pollutants have also been associated with altered mtDNA copy number. Since exposures to polychlorinated biphenyls (PCBs) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) have been shown to increase oxidative stress, we hypothesize that mtDNA copy number will be altered with exposure to these compounds. mtDNA copy number was measured in DNA from archived frozen liver and lung specimens from the National Toxicology Program (NTP) study of female Harlan Sprague Dawley rats exposed to TCDD (3, 10, or 100 ng/kg/day), dioxin-like (DL) PCB 126 (10, 100, or 1000 ng/kg/day), non-DL PCB 153 (10, 100, or 1000 μg/kg/day), and PCB 126 + PCB 153 (10 ng/kg/day + 10 μg/kg/day, 100 ng/kg/day + 100 μg/kg/day, or 1000 ng/kg/day + 1000 μg/kg/day, respectively) for 13 and 52 weeks. An increase in mtDNA copy number was observed in the liver and lung of rats exposed to TCDD and the lung of rats exposed to the mixture of PCB 126 and PCB 153. A statistically significant positive dose-dependent trend was also observed in the lung of rats exposed to PCB 126 and a mixture of PCB 153 and PCB 126, although in neither case was the control copy number significantly exceeded at any dose level. These exposures produced a range of pathological responses in these organs in the two-year NTP studies. Conversely, there was a significant decrease or no change in mtDNA copy number in the liver and lung of rats exposed to non-DL PCB 153. This is consistent with a general lack of PCB 153 mediated liver or lung injury in the NTP study, with the exception of liver hypertrophy. Together, the results suggest that an increase in mtDNA copy number may serve as a sensitive, early biomarker of mitochondrial injury and oxidative stress that contributes to the development of the toxicity of dioxin-like compounds.

Toxicity studies of hexachlorobenzene administered by gavage to female Sprague Dawley (Hsd:Sprague Dawley SD) rats.

Despite the cessation of its production and use in many parts of the world, hexachlorobenzene (HCB) remains highly persistent in the environment, and chronic, low-dose exposure to HCB in humans continues. Its structural resemblance to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), ability to activate the aryl hydrocarbon (Ah) receptor, TCDD-like toxicities, and bioaccumulative nature suggest HCB be included in the toxic equivalency factor (TEF) methodology. Consequently, the National Toxicology Program conducted this subchronic study of HCB, including measurement of a variety of toxicological and biochemical endpoints, to allow comparison to TCDD data obtained in a previous 2-year bioassay. (Abstract Abridged).

Role of aryl hydrocarbon receptor (AHR) in overall retinoid metabolism: Response comparisons to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure between wild-type and AHR knockout mice

Young adult wild-type and aryl hydrocarbon receptor knockout (AHRKO) mice of both sexes and the C57BL/6J background were exposed to 10 weekly oral doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; total dose of 200 μg/kg bw) to further characterize the observed impacts of AHR as well as TCDD on the retinoid system. Unexposed AHRKO mice harboured heavier kidneys, lighter livers and lower serum all-trans retinoic acid (ATRA) and retinol (REOH) concentrations than wild-type mice. Results from the present study also point to a role for the murine AHR in the control of circulating REOH and ATRA concentrations. In wild-type mice, TCDD elevated liver weight and reduced thymus weight, and drastically reduced the hepatic concentrations of 9-cis-4-oxo-13,14-dihydro-retinoic acid (CORA) and retinyl palmitate (REPA). In female wild-type mice, TCDD increased the hepatic concentration of ATRA as well as the renal and circulating REOH concentrations. Renal CORA concentrations were substantially diminished in wild-type male mice exclusively following TCDD-exposure, with a similar tendency in serum. In contrast, TCDD did not affect any of these toxicity or retinoid system parameters in AHRKO mice. Finally, a distinct sex difference occurred in kidney concentrations of all the analysed retinoid forms. Together, these results strengthen the evidence of a mandatory role of AHR in TCDD-induced retinoid disruption, and suggest that the previously reported accumulation of several retinoid forms in the liver of AHRKO mice is a line-specific phenomenon. Our data further support participation of AHR in the control of liver and kidney development in mice.

2,3,7,8-Tetrachlorodibenzo-p-dioxin prompted differentiation to CD4+CD8−CD25+ and CD4+CD8+CD25+ Tregs and altered expression of immune-related genes in the thymus of chicken embryos

The chicken (Gallus gallus), which has three aryl hydrocarbon receptor (AHR) isoforms (ckAHR1, ckAHR2, and ckAHR1β) and two AHR nuclear translocator (ARNT) isoforms (ckARNT1 and ckARNT2), is highly sensitive to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and can serve as an avian model to gain an understanding of the mechanism underlying dioxin toxicity. To elucidate the mechanism of TCDD-induced immunotoxicity in avian species, we treated chicken embryos in ovo with graded concentrations of TCDD (1.5, 2.5, 3.0, 3.3, 3.5, and 4.0 μM). Initially, we measured mRNA expression levels of ckAHR and ckARNT isoforms and analyzed the T cell populations and transcriptome in the thymuses of TCDD-treated chicken embryos. Quantitative polymerase chain reaction analysis revealed that mRNA expressions of ckAHR1 and ckARNT2 were dominant in the thymus. Severe weight loss and thymus atrophy were observed in the TCDD-treated embryos. Immunophenotyping analyses demonstrated significant increases in CD4+CD8-CD25+ and CD4+CD8+CD25+ regulatory T cells (Tregs) populations following TCDD exposure, suggesting that TCDD suppresses T cell-mediated immune responses in chicken embryos. In addition, thymic transcriptome analyses intimated that alteration of the signaling pathways related to erb-b2 receptor tyrosine kinase 4 (ERBB4) and wnt family member 5A (WNT5A), and bone morphogenetic protein (BMP) may be associated with the TCDD-induced thymus atrophy. We also observed significantly altered expression levels of genes including interleukine 13 receptor subunit alpha 2 (IL13RA2), transforming growth factor beta 1 (TGFβ1), collagen type III alpha 1 chain (COL3A1), and collagen type IX alpha 3 chain (COL9A3), implying immunosuppression, fibrosis development, and collagen deposition. Collectively, these findings suggest that TCDD exposure activates the ckAHR1-ckARNT2 signaling pathway and suppresses immune responses through the prompted differentiation to CD4+CD8-CD25+ and CD4+CD8+CD25+ Tregs and altered expressions of immune-related genes in the thymus of chicken embryos.

Meta-Analysis of Transcriptome Data Detected New Potential Players in Response to Dioxin Exposure in Humans.

Dioxins are one of the most potent anthropogenic poisons, causing systemic disorders in embryonic development and pathologies in adults. The mechanism of dioxin action requires an aryl hydrocarbon receptor (AhR), but the downstream mechanisms are not yet precisely clear. Here, we performed a meta-analysis of all available transcriptome datasets taken from human cell cultures exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Differentially expressed genes from different experiments overlapped partially, but there were a number of those genes that were systematically affected by TCDD. Some of them have been linked to toxic dioxin effects, but we also identified other attractive targets. Among the genes that were affected by TCDD, there are functionally related gene groups that suggest an interplay between retinoic acid, AhR, and Wnt signaling pathways. Next, we analyzed the upstream regions of differentially expressed genes and identified potential transcription factor (TF) binding sites overrepresented in the genes responding to TCDD. Intriguingly, the dioxin-responsive element (DRE), the binding site of AhR, was not overrepresented as much as other cis-elements were. Bioinformatics analysis of the AhR binding profile unveils potential cooperation of AhR with E2F2, CTCFL, and ZBT14 TFs in the dioxin response. We discuss the potential implication of these predictions for further dioxin studies.

Age at menarche in Seveso daughters exposed in utero to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

In 2014, nearly 40 years after the explosion, we enrolled postexplosion offspring, 2 to 39 years of age, in the Seveso Second Generation Study. Age at onset of menarche (years) was collected for 316 daughters by maternal report or self-report at interview. In utero TCDD exposure was defined by maternal TCDD serum concentrations extrapolated to the pregnancy. At interview, 287 daughters were postmenarche and reported age at menarche averaged 12.1 years (±1.3 years). Overall, we found no change in risk of menarche onset with a 10-fold increase in in utero TCDD exposure (hazard ratio [HR] = 0.86; 95% confidence interval [CI] = 0.71, 1.04). When we considered maternal menarche status in 1976 as a potentially sensitive developmental exposure window, in utero TCDD (log10) was associated with later age at menarche among daughters whose mothers were premenarche (HR = 0.71; 95% CI = 0.52, 0.97) but not postmenarche (HR = 0.89; 95% CI = 0.71, 1.12) at the time of the explosion (P int = 0.24). These results suggest that in utero TCDD exposure may alter pubertal timing among daughters of women who were prepubescent at the time of the Seveso accident.

Gestational dioxin exposure suppresses prolactin-stimulated nursing in lactating dam rats to impair development of postnatal offspring

A number of epidemiological studies have implicated environmental chemicals including dioxins in the induction of negative effects on child development. To clarify the underlying mechanisms, almost all toxicologists have concentrated on effects on the offspring themselves. We examined an alternative hypothesis that gestational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a highly-toxic dioxin, targets factors related to maternal childcare to disturb offspring development. Oral administration of TCDD (1 µg/kg) to pregnant rats on gestational day 15 suppressed maternal licking behavior, a nursing behavior, and mammary gland maturation during the lactational stage, as well as the body weight and short-term memory of postnatal offspring. In support of these findings, maternal production of prolactin, a pituitary hormone essential for nursing including milk production, was decreased during the same period. Intracerebroventricular infusion of prolactin to dioxin-exposed dams restored or tended to restore many of the above defects observed both in mothers and offspring. The TCDD-dependent defects in maternal nursing behaviors can be due to a direct action on aryl hydrocarbon receptor (AHR) of lactating dams, because they did not emerge in AHR-knockout dams or control dams with TCDD-exposed offspring. Further examinations revealed that TCDD induces transforming growth factor β1 expression, which suppresses prolactin-producing cell proliferation, in a nursing period-specific manner. In agreement with this, the number of prolactin-positive cells in nursing dams was decreased by TCDD. These results provide novel evidence that gestational dioxin exposure attenuates prolactin-stimulated nursing in lactating dams to impair offspring development, and that immaturity of prolactin-producing cells can contribute to them.

Aryl hydrocarbon receptor-induced activation of the human IGH hs1.2 enhancer: Mutational analysis of putative regulatory binding motifs

The human hs1.2 enhancer within the Ig heavy chain gene (IGH) is polymorphic and associated with a number of autoimmune diseases. The polymorphic region is characterized by tandem repeats of an ∼53-bp invariant sequence containing possible binding sites for several transcription factors. Our previous studies suggest the human hs1.2 enhancer is sensitive to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), an environmental toxicant and high affinity ligand of the aryl hydrocarbon receptor (AhR). TCDD induced hs1.2 enhancer activity in an AhR-dependent manner and the number of invariant sequences influenced the magnitude of activity. To better understand the regulation of human hs1.2 enhancer activity, the objective of the current study was to utilize mutational analysis and luciferase reporter constructs to evaluate the contribution of putative transcription factor binding sites to overall hs1.2 enhancer activity and modulation by TCDD. Basal and LPS-induced activity of the hs1.2 enhancer appeared to be most affected by mutation of sites outside of the invariant sequence or deletion of the entire invariant sequence; whereas sites influencing the effect of TCDD were dependent on the cellular activation state (i.e. unstimulated vs. LPS stimulation) and relatively independent of the putative AhR binding site within the invariant sequence. These results suggest that AhR activation affects human hs1.2 activity through an as yet undetermined non-canonical pathway. A better understanding regarding the role of the hs1.2 enhancer in human Ig expression and how AhR ligands modulate its activity may lead to insights into overall Ig regulation and mechanisms of dysfunction.

Dioxin male rat reproductive toxicity mode of action and relative potency of 2,3,7,8-tetrachlorodibenzo-p-dioxin and 2,3,7,8-tetrachlorodibenzofuran characterized by fetal pituitary and testis transcriptome profiling

Fetal rat exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) reduces epididymal sperm number involving altered pituitary-testicular hormonal signaling as the proposed mode-of-action (MOA). To evaluate this MOA and compare TCDD to 2,3,7,8-tetrachlorodibenzofuran (TCDF), an in utero rat exposure and study was conducted. Endpoints included congener tissue levels and transcriptomes of maternal liver and fetal liver, testis, and pituitary. Decreased gonadotropin subunit mRNAs levels (Lhb and Fshb) and enriched signaling pathways including GNRH Signaling and Calcium Signaling were observed in fetal pituitary after TCDD (but not TCDF) exposure. TCDD (but not TCDF) decreased fetal testis cholesterologenic and steroidogenic pathway genes. TCDD tissue concentrations in dam liver, dam adipose, and whole fetus were approximately 3- to 6-fold higher than TCDF. These results support a MOA for dioxin-induced rat male reproductive toxicity involving key events in both the fetal pituitary (e.g., reduced gonadotropin production) and fetal testis (e.g., reduced Leydig cell cholesterologenesis and steroidogenesis).

Prenatal dioxin exposure and thyroid hormone levels in the Seveso second generation study

BackgroundIn animal studies, perinatal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) alters thyroid homoeostasis and thyroid hormone concentrations; epidemiologic evidence is limited. ObjectivesWe aimed to determine the association of prenatal exposure to TCDD with thyroid hormone concentrations in the Seveso Second Generation Study, a unique cohort of children born to TCDD-exposed women resulting from a 1976 chemical factory explosion in Seveso, Italy. MethodsWe included 570 children (288 female, 282 male) with complete follow-up data, including a fasting blood draw. Serum levels of total and free thyroxine (T4), free triiodothyronine (T3), and thyroid stimulating hormone (TSH) were measured using immunoassays. We defined prenatal TCDD exposure as: 1) maternal initial TCDD concentration measured in serum collected soon after the explosion and 2) maternal TCDD estimated at pregnancy. ResultsCompared to the lowest quartile (Q1), maternal initial serum TCDD was associated with lower free T3 (Q2: adj-β = −0.13, 95%CI -0.26, 0.00; Q3: adj-β = −0.22, 95%CI -0.35, −0.09; Q4: adj-β = −0.14, 95%CI -0.28, 0.00; p-trend = 0.02). In participants with high thyroid antibody status, inverse associations between maternal initial serum TCDD and free T3 were significantly stronger than in participants with normal antibody status (p-interaction = 0.02). We also observed a positive association between maternal initial serum TCDD and TSH concentrations in participants with high thyroid antibody status (Q2: adj-β = 11.4%, 95%CI -25.2, 66.1; Q3: adj-β = 49.0%, 95%CI 3.0, 115.5; Q4: adj-β = 105.5, 95%CI 36.6, 209.2; p-trend < 0.01) but not in those participants with normal antibody status (p-interaction < 0.01). Similar results were found for TCDD estimated at pregnancy. DiscussionOur results suggest prenatal exposure to TCDD, a potent endocrine-disrupting compound, may alter thyroid function later in life. Populations with additional thyroid stress may be particularly susceptible to in utero exposure of thyroid disrupting chemicals.

Molecular and Functional Properties of the Atlantic Cod (Gadus morhua) Aryl Hydrocarbon Receptors Ahr1a and Ahr2a.

The aryl hydrocarbon receptor (Ahr) is a ligand-activated transcription factor that mediates the toxicity of halogenated and polycyclic aromatic hydrocarbons in vertebrates. Atlantic cod (Gadus morhua) has recently emerged as a model organism in environmental toxicology studies, and increased knowledge of Ahr-mediated responses to xenobiotics is imperative. Genome mining and phylogenetic analyses revealed two Ahr-encoding genes in the Atlantic cod genome, gmahr1a and gmahr2a. In vitro binding assays showed that both gmAhr proteins bind to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), but stronger binding to gmAhr1a was observed. Transactivation studies with a reporter gene assay revealed that gmAhr1a is one order of magnitude more sensitive to TCDD than gmAhr2a, but the maximal responses of the receptors were similar. Other well-known Ahr agonists, such as β-naphthoflavone (BNF), 3,3',4,4',5-pentachlorobiphenyl (PCB126), and 6-formylindolo[3,2-b]carbazole (FICZ), also activated the gmAhr proteins, but gmAhr1a was, in general, the more sensitive receptor and produced the highest efficacies. The induction of cyp1a in exposed precision-cut cod liver slices confirmed the activation of the Ahr signaling pathway ex vivo. In conclusion, the differences in transcriptional activation by gmAhr's with various agonists, the distinct binding properties with TCDD and BNF, and the distinct tissue-specific expression profiles indicate different functional specializations of the Atlantic cod Ahr's.

A Biomimetic, One-Step Transformation of Simple Indolic Compounds to Malassezia-Related Alkaloids with High AhR Potency and Efficacy.

Malassezia furfur isolates from diseased skin preferentially biosynthesize compounds which are among the most active known aryl-hydrocarbon receptor (AhR) inducers, such as indirubin, tryptanthrin, indolo[3,2-b]carbazole, and 6-formylindolo[3,2-b]carbazole. In our effort to study their production from Malassezia spp., we investigated the role of indole-3-carbaldehyde (I3A), the most abundant metabolite of Malassezia when grown on tryptophan agar, as a possible starting material for the biosynthesis of the alkaloids. Treatment of I3A with H2O2 and use of catalysts like diphenyldiselenide resulted in the simultaneous one-step transformation of I3A to indirubin and tryptanthrin in good yields. The same reaction was first applied on simple indole and then on substituted indoles and indole-3-carbaldehydes, leading to a series of mono- and bisubstituted indirubins and tryptanthrins bearing halogens, alkyl, or carbomethoxy groups. Afterward, they were evaluated for their AhR agonist activity in recombinant human and mouse hepatoma cell lines containing a stably transfected AhR-response luciferase reporter gene. Among them, 3,9-dibromotryptanthrin was found to be equipotent to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) as an AhR agonist, and 3-bromotryptanthrin was 10-times more potent than TCDD in the human HG2L7.5c1 cell line. In contrast, 3,9-dibromotryptanthrin and 3-bromotryptanthrin were ∼4000 and >10,000 times less potent than TCDD in the mouse H1L7.5c3 cell line, respectively, demonstrating that they are species-specific AhR agonists. Involvement of the AhR in the action of 3-bromotryptanthrin was confirmed by the ability of the AhR antagonists CH223191 and SR1 to inhibit 3-bromotryptanthrin-dependent reporter gene induction in human HG2L7.5c1 cells. In conclusion, I3A can be the starting material used by Malassezia for the production of both indirubin and tryptanthrin through an oxidation mechanism, and modification of these compounds can produce some highly potent, efficacious and species-selective AhR agonists.

Gene-Dioxin Interactions and Time to Pregnancy in the Seveso Women’s Health Study

TPS 634: Health effects of pops, voc and other chemicals, Johan Friso Foyer, Floor 1, August 27, 2019, 3:00 PM - 4:30 PM Background/Aim: Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a widespread environmental contaminant, has been associated with reduced fertility in multiple animal species. TCDD is proposed to impact fertility via altered activity of the aryl hydrocarbon receptor (AhR) pathway, which regulates diverse biological and developmental processes, including xenobiotic metabolism. The Seveso Women’s Health Study (SWHS) followed up 981 women exposed to TCDD in a chemical factory explosion in 1976. Twenty years later, we reported longer time to pregnancy (TTP) associated with individual 1976 serum TCDD levels, but the youngest women had not yet attempted pregnancy. Now, 40 years after the explosion, when most women will have completed their families, we aim to re-examine the relationship of TCDD exposure with TTP and assess whether variations in AhR pathway genes modify this relationship. Methods: Of 617 women who had attempted pregnancy after the explosion, 367 delivered a live birth not associated with contraceptive failure. We examined the relationship of 1976 serum TCDD levels to TTP (parameterized as the monthly probability of conception within the first 12 months of trying) using discrete-time Cox proportional hazards regression. We also utilized genotype data (87 SNPs across 7 AhR pathway genes) to examine gene-environment interactions between AhR pathway variants and TCDD levels on TTP. Results: Women averaged 26.7 (±4.3) years at the time of attempting pregnancy. Median TTP was 2 months, with 17.2% taking 12 or more months to conceive. Consistent with our previous findings, a 10-fold increase in serum TCDD was associated with longer TTP (adjusted fecundability odds ratio = 0.80; 95%CI: 0.66, 0.98). In preliminary analyses, we found one SNP in AHRR which adversely modified the association between TCDD levels and TTP, implicating gene-environment interaction. Conclusions: These results suggest that TCDD exposure is associated with reduced fertility in the Seveso cohort; gene-environment interaction may play a role.

Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure on acetylcholinesterase during myogenic differentiation of contractile rat primary skeletal muscle cells

Emerging data indicate that prenatal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) could interfere with myogenic differentiation in vivo. Acetylcholinesterase (EC3.1.1.7; AChE), an enzyme critical for cholinergic neurotransmission, is abundantly expressed in neurons and mature myotubes, and we recently found that muscle AChE expression was suppressed in parallel with the inhibition of myogenic differentiation upon TCDD treatment in mouse C2C12 cells. This TCDD-induced suppression of muscle AChE was proposed to involve an aryl hydrocarbon receptor (AhR)-independent mechanism, but the precise underlying mechanism remains unclear. Considering the widely recognized role of muscular activity in AChE expression and its potential crosstalk with the AhR signaling pathway, we sought to investigate the effect of TCDD on muscle AChE expression in the presence of muscular activity. Therefore, we employed a highly contractile rat primary skeletal muscle culture system in which AChE activity and the expression of genes related to it (AChE T subunit and collagen Q (ColQ)) were increased during the myogenic differentiation process. Although TCDD treatment successfully induced the expression of genes regulated by AhR activation, the treatment exerted no notable effects on myogenic differentiation. Moreover, muscle AChE enzymatic activity and mRNA level remained unchanged following TCDD treatment, and only ColQ mRNA expression was slightly increased after 4-day treatment with TCDD (10−10 M). The compensatory role of muscle-contraction-related signaling pathways in this newly identified unresponsiveness of muscle AChE to TCDD warrants further investigation.

Transcriptomic analysis of Anabas testudineus and its defensive mechanisms in response to persistent organic pollutants exposure

The freshwater climbing perch (Anabas testudineus) can tolerate water environments contaminated with persistent organic pollutants (POPs). The mechanisms underlying this tolerance are unknown. We used de novo transcriptomic analysis to investigate the defensive mechanisms of A. testudineus against POPs based on its genetic features and biological responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure. Our results revealed a specific expansion of cytochrome P450 (CYP) 3A subfamily, which may be involved in the elimination of certain POPs. In xenobiotic responses, the aryl-hydrocarbon receptor (AhR) pathway represents a critical signaling mechanism, and we characterized four AhR and two AhR nuclear translocator homologs and one AhR repressor (AhRR) gene in A. testudineus. TCDD-induced AhRR and CYP1A mRNA upregulation suggests that negative-feedback regulation of AhR signaling through AhRR helps avoid excessive xenobiotic responses. Furthermore, liver and gill transcriptomic profiles were markedly altered after TCDD exposure, with some of the altered genes being related to common defensive responses reported in other species. Based on the newly identified TCDD-altered genes, several A. testudineus-specific responses are proposed, such as enhanced fatty acid β-oxidation. The genetic features of CYP3A subfamily and AhR pathway and the TCDD-induced defensive biological processes elucidated here enhance our understanding of A. testudineus defensive responses against POPs.

Interindividual variation in the cytochrome P4501A response to 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin in herring gull embryo hepatocytes

Exposure to dioxin-like compounds is consistently associated with concentration-dependent induction of cytochrome P4501A (CYP1A) enzymes in primary cultures of avian hepatocytes. We have previously demonstrated that the median effective concentration (EC50) for induction of this response is predictive of in vivo sensitivity to dioxin-like compounds in birds. We investigated sources of interindividual variation in the CYP1A response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in wild herring gulls and considered how this variation may complicate dioxin sensitivity estimates based on the CYP1A bioassay. Concentration-dependent effects of TCDD on CYP1A mRNA expression were characterized in 55 hepatocyte cultures prepared from individual herring gull embryos. A large degree of variability was observed among the hepatocyte culture preparations. For example, 1) basal CYP1A4 and CYP1A5 mRNA expression varied by 20- and 126-fold, respectively, among individuals, and 2) exposure to TCDD induced CYP1A4 mRNA expression by 57-fold in the most responsive sample but did not significantly induce CYP1A4 mRNA expression above baseline values in 42% of hepatocyte culture preparations. Environmental and genetic factors contributing to the observed variability are discussed. Despite the large amount of interindividual variation, we conclude that reproducible EC50-based estimates of species sensitivity can be obtained from the CYP1A cell culture bioassay when samples are collected from relatively uncontaminated colonies. Environ Toxicol Chem 2019;38:660-670. © 2019 SETAC.

The role of prostaglandin E2 receptor EP1 in 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced neonatal hydronephrosis in mice

Prostaglandin E2 (PGE2) is a critical factor in the pathogenesis of dioxin-induced neonatal hydronephrosis. Since the PGE2 receptor has four subtypes, EP1 − EP4, this study was aimed to challenge the hypothesis that at least one of the four subtypes is responsible for the pathogenesis of dioxin-induced hydronephrosis. To this end, we used mouse pups, with a C57BL/6 J background, genetically lacking EP1, EP2, or EP3, and wild-type pups in whom EP4 was suppressed by administering ONO-AE3-208 (ONO), an EP4 antagonist, from postnatal day 1 (PND 1) to PND 13. To expose the pups to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) via lactation, the dams were administered TCDD at an oral dose of 20 μg/kg on PND 1. The pups’ urine and kidneys were collected on PND 14 for urinalysis and histological examination, respectively. We found that the incidence of hydronephrosis was 80% in the EP1+/+ group, but was markedly reduced to 28.6% in the EP1–/– group despite the fact that PGE2 concentration in the urine was similarly increased in the both groups. In contrast, the incidence of hydronephrosis was 80% and 100% in the EP2+/+ and EP2–/–groups, respectively, and 88.9% and 100% in the EP3+/+ and EP3–/– groups, respectively. With regard to EP4, the incidence of hydronephrosis in vehicle (saline)-treated groups and ONO-treated was 88.9% and 100%, respectively. Therefore, we concluded that among PGE2 receptor subtypes, EP1 plays a predominant role in the onset of TCDD-induced neonatal hydronephrosis in mouse pups.

Induction of maxillary and mandibular squamous epithelial cell proliferation in mink (Neovison vison) by β-naphthoflavone.

A jaw lesion reported in mink exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and TCDD-like chemicals is considered a potential indicator of exposure to these chemicals. Many of the effects of TCDD-like chemicals are induced through interaction with the aryl hydrocarbon receptor. The present study indicates that mink dosed with β-naphthoflavone, which is an aryl hydrocarbon receptor ligand but not a TCDD-like chemical, also develop the lesion. Environ Toxicol Chem 2019;38:460-463. © 2018 SETAC.

Cardiotoxicity induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin exposure through lactation in mice.

Dioxins are a group of structurally related chemicals that persist in the environment. Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), the most toxic congener, is a suspected risk factor for cardiac diseases in humans. TCDD induces signs of cardiotoxicity in various animals. Mouse models of TCDD exposure suggest cardiotoxicity phenotypes develop differently depending on the timing and time-course of exposure. In order to clarify and characterize the TCDD-induced cardiotoxicity in the developing period, we utilized mouse pups exposed to TCDD. One day after delivery, groups of nursing C57BL/6J dams were orally administered TCDD at a dose of 0 (Control), 20 (TCDD-20), or 80 μg/kg (TCDD-80) body weight (BW). On postnatal days (PNDs) 7 and 21, pups' hearts were examined by histological and gene expression analyses. The TCDD-80 group was found to have a left ventricular remodeling on PND 7, and to develop heart hypertrophy on PND 21. It was accompanied by fibrosis and increased expression of associated genes, such as those for atrial natriuretic peptide (ANP), β-myosin heavy chain (β-MHC), and endothelin-1 (ET-1). These results revealed that TCDD directly induces cardiotoxicity in the postnatal period represented by progressive hypertrophy in which ANP, β-MHC, and ET-1 have potentials to mediate the cardiac hypertrophy and heart failure.

A Toxicogenomic Approach Reveals a Novel Gene Regulatory Network Active in In Vitro and In Vivo Models of Thyroid Carcinogenesis.

Epidemiological and experimental studies emphasize the link between environmental chemicals exposure and thyroid cancer. However, this association is strongly debated and the mechanisms of action of environmental thyroid carcinogens still need to be identified. The analysis of in vitro transcriptomic data developed to investigate the effects of chlorpyrifos on immortalized thyrocytes highlighted the impaired expression of genes involved in endodermal carcinogenesis. This endodermal carcinogenic gene-network (ECGN, including Zfp36l2, Dmbt1, Ddit4), was validated in cellular and mouse models of thyroid carcinogenesis, characterized by the constitutive activation of the mitogen-activated protein kinase (MAPK) pathway and in immortalized thyrocytes exposed to tetrachlorodibenzo-p-dioxin (TCDD) and chlorpyrifos (CPF). The mRNA levels of Zfp36l2, Dmbt1 and Ddit4 were increased in models characterized by MAPK activation or following TCDD exposure, whereas they were inhibited by CPF exposure. Overall, the ECGN transcripts identify a novel gene-regulatory network associated with thyroid carcinogenesis promoted by genetic mutation or by environmental carcinogens. The latter have opposite effects on the modulation of the ECGN transcripts according to their mechanisms of action in promoting carcinogenesis. Therefore, the analyses of ECGN might be helpful in discriminating compounds that promote cellular survival associated or not to proliferation of thyrocytes.