Science.gov

Sample records for ah receptor ahr

  1. TCDD and omeprazole prime platelets through the aryl hydrocarbon receptor (AhR) non-genomic pathway.

    PubMed

    Pombo, Mónica; Lamé, Michael W; Walker, Naomi J; Huynh, Danh H; Tablin, Fern

    2015-05-19

    The role of the aryl hydrocarbon receptor (AhR) in hemostasis has recently gained increased attention. Here, we demonstrate, by qRT-PCR and western blot, that human platelets express both AhR mRNA and AhR protein. AhR protein levels increase in a dose dependent manner when incubated with either 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or omeprazole. Treatment of platelets with puromycin blocks increased AhR protein synthesis in the presence of AhR activators. Additionally, treatment of platelets with either activator results in phosphorylation of p38MAPK and cPLA2, two key signaling molecules in platelet activation pathways. Using the AhR competitive inhibitors alpha naphthoflavone and CH-223191, we show that phosphorylation of p38MAPK is AhR dependent. Further, inhibition of p38MAPK blocks downstream cPLA2 phosphorylation induced by TCDD or omeprazole. Treatment with AhR activators results in platelet priming, as demonstrated by increased platelet aggregation, which is inhibited by AhR antagonists. Our data support a model of the platelet AhR non-genomic pathway in which treatment with AhR activators results in increased expression of the AhR, phosphorylation of p38MAPK and cPLA2, leading to platelet priming in response to agonist. PMID:25797602

  2. Identification and expression of aryl hydrocarbon receptors (AhR1 and AhR2) provide insight in an evolutionary context regarding sensitivity of white sturgeon (Acipenser transmontanus) to dioxin-like compounds.

    PubMed

    Doering, Jon A; Wiseman, Steve; Beitel, Shawn C; Giesy, John P; Hecker, Markus

    2014-05-01

    Sturgeons are ancient fishes, which are endangered in many parts of the world. Due to their benthic nature and longevity, sturgeon are at great risk of exposure to bioaccumulative contaminants such as dioxin-like compounds (DLCs). Despite their endangered status, little research has been conducted to characterize the relative sensitivity of sturgeons to DLCs. Proper assessment of risk of DLCs posed to these fishes therefore, requires a better understanding of this sensitivity and the factors that are driving it. Adverse effects associated with exposure to DLCs are mediated by the aryl hydrocarbon receptor (AhR). This study identified and characterized two distinct AhRs, AhR1 and AhR2, in white sturgeon (Acipenser transmontanus) for the first time as a first step in studying the relative sensitivities of sturgeons to DLCs. Furthermore, tissue-specific expression of both AhRs under basal conditions and in response to exposure to the model DLC, β-naphthoflavone (βNF), was determined. The sequence of amino acids of AhR1 of white sturgeon had greater similarity to AhRs of tetrapods, including amphibians, birds, and mammals, than to AhR1s of other fishes. The sequence of amino acids in the ligand binding domain of the AhR1 had greater than 80% similarity to AhRs known to bind DLCs and was less similar to AhRs not known to bind DLCs. AhR2 of white sturgeon had greatest similarity to AhR2 of other fishes. Profiles of expression of AhR1 and AhR2 in white sturgeon were distinct from those known in other fishes and appear more similar to profiles observed in birds. Expressions of both AhR1 and AhR2 of white sturgeon were greatest in liver and heart, which are target organs for DLCs. Furthermore, abundances of transcripts of AhR1 and AhR2 in all tissues from white sturgeon were greater than controls (up to 35-fold) following exposure to βNF. Based upon both AhRs having similar abundances of transcript in target organs of DLC toxicity, both AhRs being up-regulated following

  3. Identification and expression of aryl hydrocarbon receptors (AhR1 and AhR2) provide insight in an evolutionary context regarding sensitivity of white sturgeon (Acipenser transmontanus) to dioxin-like compounds.

    PubMed

    Doering, Jon A; Wiseman, Steve; Beitel, Shawn C; Giesy, John P; Hecker, Markus

    2014-05-01

    Sturgeons are ancient fishes, which are endangered in many parts of the world. Due to their benthic nature and longevity, sturgeon are at great risk of exposure to bioaccumulative contaminants such as dioxin-like compounds (DLCs). Despite their endangered status, little research has been conducted to characterize the relative sensitivity of sturgeons to DLCs. Proper assessment of risk of DLCs posed to these fishes therefore, requires a better understanding of this sensitivity and the factors that are driving it. Adverse effects associated with exposure to DLCs are mediated by the aryl hydrocarbon receptor (AhR). This study identified and characterized two distinct AhRs, AhR1 and AhR2, in white sturgeon (Acipenser transmontanus) for the first time as a first step in studying the relative sensitivities of sturgeons to DLCs. Furthermore, tissue-specific expression of both AhRs under basal conditions and in response to exposure to the model DLC, β-naphthoflavone (βNF), was determined. The sequence of amino acids of AhR1 of white sturgeon had greater similarity to AhRs of tetrapods, including amphibians, birds, and mammals, than to AhR1s of other fishes. The sequence of amino acids in the ligand binding domain of the AhR1 had greater than 80% similarity to AhRs known to bind DLCs and was less similar to AhRs not known to bind DLCs. AhR2 of white sturgeon had greatest similarity to AhR2 of other fishes. Profiles of expression of AhR1 and AhR2 in white sturgeon were distinct from those known in other fishes and appear more similar to profiles observed in birds. Expressions of both AhR1 and AhR2 of white sturgeon were greatest in liver and heart, which are target organs for DLCs. Furthermore, abundances of transcripts of AhR1 and AhR2 in all tissues from white sturgeon were greater than controls (up to 35-fold) following exposure to βNF. Based upon both AhRs having similar abundances of transcript in target organs of DLC toxicity, both AhRs being up-regulated following

  4. The immune phenotype of AhR null mouse mutants: not a simple mirror of xenobiotic receptor over-activation.

    PubMed

    Esser, Charlotte

    2009-02-15

    Intrinsic and induced cell differentiation and the cellular response to endogenous and exogenous signals are hallmarks of the immune system. Specific and common signalling cascades ensure a highly flexible and adapted response. Increasing evidence suggests that gene modulation by the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor, is an important part of these processes. For decades the AhR has been studied mainly for its toxic effects after artificial activation by man-made chemical pollutants such as dioxins. These studies gave important, albeit to some extent skewed, evidence for a mechanistic link between the AhR and the immune system. AhR null mutants and other mutants of the AhR signalling pathway have been generated and used to analyse the physiological function of the AhR, including for the developing and antigen-responding immune system. In this review I look at the natural immunological function(s) of the AhR.

  5. Arylhydrocarbon receptor (AhR) activation in airway epithelial cells induces MUC5AC via reactive oxygen species (ROS) production.

    PubMed

    Chiba, Takahito; Uchi, Hiroshi; Tsuji, Gaku; Gondo, Hisaki; Moroi, Yoichi; Furue, Masutaka

    2011-02-01

    The dioxins and dioxin-like compounds in cigarette smoke regulate various immunological responses via the arylhydrocarbon receptor (AhR). These environmental toxicants are known to cause bronchitis, asthma, chronic obstructive pulmonary disease (COPD), and lung cancer. Recent studies have demonstrated that AhR activation upregulates the expression of mucin 5AC, oligomeric mucus/gel-forming (MUC5AC) in the airway epithelial cell line. However, the mechanism for the production of mucin has not been clarified. In this study, we investigated the role and pathway of AhR in airway epithelial cells by using selective agonists and antagonists. After stimulation with or without benzopyrene (B[a]P), an AhR agonist, MUC5AC expression was measured by real-time RT-PCR. The mechanism of AhR-induced MUC5AC expression in airway epithelial cells was studied in terms of the production of cytokine and reactive oxygen species (ROS). Treatment with B[a]P increased ROS generation in NCI-H₂₉₂ cells. Furthermore, B[a]P-induced MUC5AC upregulation and mucin production were inhibited by AhR siRNA or the use of an antioxidative agent. These results suggest that the AhR-induced increase of mucin production is partially mediated by ROS generation. An antioxidant therapy approach may help to cure AhR-induced mucus hypersecretory diseases. PMID:20709182

  6. AhR signalling and dioxin toxicity.

    PubMed

    Sorg, Olivier

    2014-10-15

    Dioxins are a family of molecules associated to several industrial accidents such as Ludwigshafen in 1953 or Seveso in 1976, to the Agent Orange used during the war of Vietnam, and more recently to the poisoning of the former president of Ukraine, Victor Yushchenko. These persistent organic pollutants are by-products of industrial activity and bind to an intracellular receptor, AhR, with a high potency. In humans, exposure to dioxins, in particular 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces a cutaneous syndrome known as chloracne, consisting in the development of many small skin lesions (hamartoma), lasting for 2-5 years. Although TCDD has been classified by the WHO as a human carcinogen, its carcinogenic potential to humans is not clearly demonstrated. It was first believed that AhR activation accounted for most, if not all, biological properties of dioxins. However, certain AhR agonists found in vegetables do not induce chloracne, and other chemicals, in particular certain therapeutic agents, may induce a chloracne-like syndrome without activating AhR. It is time to rethink the mechanism of dioxin toxicity and analyse in more details the biological events following exposure to these compounds and other AhR agonists, some of which have a very different chemical structure than TCDD. In particular various food-containing AhR agonists are non-toxic and may on the contrary have beneficial properties to human health. PMID:24239782

  7. Inhibition of constitutive aryl hydrocarbon receptor (AhR) signaling attenuates androgen independent signaling and growth in (C4-2) prostate cancer cells

    PubMed Central

    Tran, Cindy; Richmond, Oliver; Aaron, LaTayia; Powell, Joann B.

    2013-01-01

    The aryl hydrocarbon receptor is a member of the basic-helix-loop-helix family of transcription factors. AhR mediates the biochemical and toxic effects of a number of polyaromatic hydrocarbons such as 2,3,7,8,-tetrachloro-dibenzo-p-dioxin (TCDD). AhR is widely known for regulating the transcription of drug metabolizing enzymes involved in the xenobiotic metabolism of carcinogens and therapeutic agents, such as cytochrome P450-1B1 (CYP1B1). Additionally, AhR has also been reported to interact with multiple signaling pathways during prostate development. Here we investigate the effect of sustained AhR signaling on androgen receptor function in prostate cancer cells. Immunoblot analysis shows that AhR expression is increased in androgen independent (C4-2) prostate cancer cells when compared to androgen sensitive (LNCaP) cells. RT-PCR studies revealed constitutive AhR signaling in C4-2 cells without the ligand induced activation required in LNCaP cells. A reduction of AhR activity by short RNA mediated silencing in C4-2 cells reduced expression of both AhR and androgen responsive genes. The decrease in androgen responsive genes correlates to a decrease in phosphorylated androgen receptor and androgen receptor expression in the nucleus. Furthermore, the forced decrease in AhR expression resulted in a 50% decline in the growth rate of C4-2 cells. These data indicates that AhR is required to maintain hormone independent signaling and growth by the androgen receptor in C4-2 cells. Collectively, these data provide evidence of a direct role for AhR in androgen independent signaling and provides insight into the molecular mechanisms responsible for sustained androgen receptor signaling in hormone refractory prostate cancer. PMID:23266674

  8. Functionality of aryl hydrocarbon receptors (AhR1 and AhR2) of white sturgeon (Acipenser transmontanus) and implications for the risk assessment of dioxin-like compounds.

    PubMed

    Doering, Jon A; Farmahin, Reza; Wiseman, Steve; Kennedy, Sean W; Giesy, John P; Hecker, Markus

    2014-07-15

    Worldwide, populations of sturgeons are endangered, and it is hypothesized that anthropogenic chemicals, including dioxin-like compounds (DLCs), might be contributing to the observed declines in populations. DLCs elicit their toxic action through activation of the aryl hydrocarbon receptor (AhR), which is believed to regulate most, if not all, adverse effects associated with exposure to these chemicals. Currently, risk assessment of DLCs in fishes uses toxic equivalency factors (TEFs) developed for the World Health Organization (WHO) that are based on studies of embryo-lethality with salmonids. However, there is a lack of knowledge of the sensitivity of sturgeons to DLCs, and it is uncertain whether TEFs developed by the WHO are protective of these fishes. Sturgeons are evolutionarily distinct from salmonids, and the AhRs of sturgeons differ from those of salmonids. Therefore, this study investigated the sensitivity of white sturgeon (Acipenser transmontanus) to DLCs in vitro via the use of luciferase reporter gene assays using COS-7 cells transfected with AhR1 or AhR2 of white sturgeon. Specifically, activation and relative potencies (RePs) of 2,3,7,8-tetrachloro-dibenzo-p-dioxin (TCDD), 2,3,4,7,8-pentachloro-dibenzofuran, 2,3,7,8-tetrachloro-dibenzofuran, 3,3',4,4',5-pentachlorobiphenyl, 3,3',4,4'-tetrachlorobiphenyl, and 2,3,3',4,4'-pentachlorobiphenyl were determined for each AhR. It was demonstrated that white sturgeon expresses AhR1s and AhR2s that are both activated by DLCs with EC50 values for 2,3,7,8-TCDD that are lower than those of any other AhR of vertebrates tested to date. Both AhRs of white sturgeon had RePs for polychlorinated dibenzofurans more similar to TEFs for birds, while RePs for polychlorinated biphenyls were most similar to TEFs for fishes. Measured concentrations of select DLCs in tissues of white sturgeon from British Columbia, Canada, were used to calculate toxic equivalents (TEQs) by use of TEFs for fishes used by the WHO and TCDD

  9. Fibroblast growth factor (Fgf) 21 is a novel target gene of the aryl hydrocarbon receptor (AhR)

    SciTech Connect

    Cheng, Xingguo; Vispute, Saurabh G.; Liu, Jie; Cheng, Christine; Kharitonenkov, Alexei; Klaassen, Curtis D.

    2014-07-01

    The toxic effects of dioxins, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), mainly through activation of the aryl hydrocarbon receptor (AhR) are well documented. Fibroblast growth factor (Fgf) 21 plays critical roles in metabolic adaptation to fasting by increasing lipid oxidation and ketogenesis in the liver. The present study was performed to determine whether activation of the AhR induces Fgf21 expression. In mouse liver, TCDD increased Fgf21 mRNA in both dose- and time-dependent manners. In addition, TCDD markedly increased Fgf21 mRNA expression in cultured mouse and human hepatocytes. Moreover, TCDD increased mRNA (in liver) and protein levels (in both liver and serum) of Fgf21 in wild-type mice, but not in AhR-null mice. Chromatin immunoprecipitation assays showed that TCDD increased AhR protein binding to the Fgf21 promoter (− 105/+ 1 base pair). Fgf21-null mice administered 200 μg/kg of TCDD died within 20 days, whereas wild-type mice receiving the same treatment were still alive at one month after administration. This indicates that TCDD-induced Fgf21 expression protects against TCDD toxicity. Diethylhexylphthalate (DEHP) pretreatment attenuated TCDD-induced Fgf21 expression in mouse liver and white adipose tissue, which may explain a previous report that DEHP pretreatment decreases TCDD-induced wasting. In conclusion, Fgf21 appears to be a target gene of AhR-signaling pathway in mouse and human liver. - Highlights: • TCDD induced Fgf21 expression at both mRNA and protein levels. • Fgf21 induction by TCDD is AhR-dependent. • DEHP attenuated TCDD-induced Fgf21 expression.

  10. Characterizing the role of endothelin-1 in the progression of cardiac hypertrophy in aryl hydrocarbon receptor (AhR) null mice

    SciTech Connect

    Lund, Amie K.; Goens, M. Beth; Nunez, Bethany A.; Walker, Mary K. . E-mail: mkwalker@unm.edu

    2006-04-15

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor characterized to play a role in detection and adaptation to environmental stimuli. Genetic deletion of AhR results in hypertension, and cardiac hypertrophy and fibrosis, associated with elevated plasma angiotensin II (Ang II) and endothelin-1 (ET-1), thus AhR appears to contribute to cardiovascular homeostasis. In these studies, we tested the hypothesis that ET-1 mediates cardiovascular pathology in AhR null mice via ET{sub A} receptor activation. First, we determine the time courses of cardiac hypertrophy, and of plasma and tissue ET-1 expression in AhR wildtype and null mice. AhR null mice exhibited increases in heart-to-body weight ratio and age-related expression of cardiac hypertrophy markers, {beta}-myosin heavy chain ({beta}-MHC), and atrial natriuretic factor (ANF), which were significant at 2 months. Similarly, plasma and tissue ET-1 expression was significantly elevated at 2 months and increased further with age. Second, AhR null mice were treated with ET{sub A} receptor antagonist, BQ-123 (100 nmol/kg/day), for 7, 28, or 58 days and blood pressure, cardiac fibrosis, and cardiac hypertrophy assessed, respectively. BQ-123 for 7 days significantly reduced mean arterial pressure in conscious, catheterized mice. BQ-123 for 28 days significantly reduced the histological appearance of cardiac fibrosis. Treatment for 58 days significantly reduced cardiac mass, assessed by heart weight, echocardiography, and {beta}-MHC and ANF expression; and reduced cardiac fibrosis as determined by osteopontin and collagen I mRNA expression. These findings establish ET-1 and the ET{sub A} receptor as primary determinants of hypertension and cardiac pathology in AhR null mice.

  11. Molecular characterization of the aryl hydrocarbon receptor (AhR) pathway in goldfish (Carassius auratus) exposure to TCDD: the mRNA and protein levels.

    PubMed

    Lu, Ming; Chang, Ziwei; Bae, Min-Ji; Oh, Seung Min; Chung, Kyu-Hyuck; Park, Jang-Su

    2013-08-01

    In bony fish or other aquatic vertebrates, the aryl hydrocarbon receptor (AhR) signaling pathway is initiated by exposure to polycyclic (or/and halogenated) aromatic hydrocarbons (PAHs, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD), which subsequently induces the up-regulated expression of a series of related genes (such as cytochrome P4501A (CYP1A)). However, a lack of applicable protein reagents hinders our further understanding of the AhR signaling pathway, which focuses only on gene-based investigations. The goldfish (Carassius auratus) is an ideal model for a study of environmental pollution in whole-Asian fresh water. Here, three sensitive and specific polyclonal antisera against goldfish AhR1, AhR2, and CYP1A proteins were developed. These antisera not only bound the in-vitro synthesized target proteins, but recognized the real proteins expressed in goldfish tissues, with minimal cross-reactivity to non-specific proteins. Together with the analysis of semi-quantitative RT-PCR and polyclonal-antibody-based sandwich ELISA, we confirmed that goldfish AhRs differed in the expression (mRNA and protein levels) patterns among test tissues. Importantly, the relative abundance of each AhR mRNA levels from the different tissues showed no obvious consistency with their protein levels. After exposure to TCDD, goldfish AhR2 showed a more sensitivity than AhR1, and stimulated CYP1A expression directly, similar with the other reported fish models. Overall, development of these antibodies in this study will allow valuable and versatile investigations to further understand the AhR signaling pathway, and different expression (mRNA and protein) patterns represent the first step in determining the regulatory mechanisms underlying the TCDD-exposed aquatic environment.

  12. Fibroblast growth factor (Fgf) 21 is a novel target gene of the aryl hydrocarbon receptor (AhR).

    PubMed

    Cheng, Xingguo; Vispute, Saurabh G; Liu, Jie; Cheng, Christine; Kharitonenkov, Alexei; Klaassen, Curtis D

    2014-07-01

    The toxic effects of dioxins, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), mainly through activation of the aryl hydrocarbon receptor (AhR) are well documented. Fibroblast growth factor (Fgf) 21 plays critical roles in metabolic adaptation to fasting by increasing lipid oxidation and ketogenesis in the liver. The present study was performed to determine whether activation of the AhR induces Fgf21 expression. In mouse liver, TCDD increased Fgf21 mRNA in both dose- and time-dependent manners. In addition, TCDD markedly increased Fgf21 mRNA expression in cultured mouse and human hepatocytes. Moreover, TCDD increased mRNA (in liver) and protein levels (in both liver and serum) of Fgf21 in wild-type mice, but not in AhR-null mice. Chromatin immunoprecipitation assays showed that TCDD increased AhR protein binding to the Fgf21 promoter (-105/+1 base pair). Fgf21-null mice administered 200μg/kg of TCDD died within 20days, whereas wild-type mice receiving the same treatment were still alive at one month after administration. This indicates that TCDD-induced Fgf21 expression protects against TCDD toxicity. Diethylhexylphthalate (DEHP) pretreatment attenuated TCDD-induced Fgf21 expression in mouse liver and white adipose tissue, which may explain a previous report that DEHP pretreatment decreases TCDD-induced wasting. In conclusion, Fgf21 appears to be a target gene of AhR-signaling pathway in mouse and human liver.

  13. Fibroblast growth factor (Fgf) 21 is a novel target gene of the aryl hydrocarbon receptor (AhR).

    PubMed

    Cheng, Xingguo; Vispute, Saurabh G; Liu, Jie; Cheng, Christine; Kharitonenkov, Alexei; Klaassen, Curtis D

    2014-07-01

    The toxic effects of dioxins, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), mainly through activation of the aryl hydrocarbon receptor (AhR) are well documented. Fibroblast growth factor (Fgf) 21 plays critical roles in metabolic adaptation to fasting by increasing lipid oxidation and ketogenesis in the liver. The present study was performed to determine whether activation of the AhR induces Fgf21 expression. In mouse liver, TCDD increased Fgf21 mRNA in both dose- and time-dependent manners. In addition, TCDD markedly increased Fgf21 mRNA expression in cultured mouse and human hepatocytes. Moreover, TCDD increased mRNA (in liver) and protein levels (in both liver and serum) of Fgf21 in wild-type mice, but not in AhR-null mice. Chromatin immunoprecipitation assays showed that TCDD increased AhR protein binding to the Fgf21 promoter (-105/+1 base pair). Fgf21-null mice administered 200μg/kg of TCDD died within 20days, whereas wild-type mice receiving the same treatment were still alive at one month after administration. This indicates that TCDD-induced Fgf21 expression protects against TCDD toxicity. Diethylhexylphthalate (DEHP) pretreatment attenuated TCDD-induced Fgf21 expression in mouse liver and white adipose tissue, which may explain a previous report that DEHP pretreatment decreases TCDD-induced wasting. In conclusion, Fgf21 appears to be a target gene of AhR-signaling pathway in mouse and human liver. PMID:24769090

  14. DDE and PCB 153 independently induce aryl hydrocarbon receptor (AhR) expression in peripheral blood mononuclear cells.

    PubMed

    Gaspar-Ramírez, Octavio; Pérez-Vázquez, Francisco J; Salgado-Bustamante, Mariana; González-Amaro, Roberto; Hernandez-Castro, Berenice; Pérez-Maldonado, Ivan N

    2015-01-01

    Recent studies have demonstrated that compounds inducing pro-inflammatory cytokines enhance AhR expression. The aim of this study was 2-fold: (1) to determine if two pro-inflammatory compounds, dichlorodiphenyldichloroethylene (DDE) and 2,2',4,4',5,5'-hexa-chlorobiphenyl (PCB 153), independently affect AhR gene expression in peripheral blood mononuclear cells (PBMC); and (2) if affected, to determine whether the mechanism involved was due to AhR activation or to a pro-inflammatory effect of the chemicals. PBMC isolated from healthy individuals were incubated in the presence of DDE (10 µg/ml) and PCB 153 (20 ng/ml) over time and AhR and CYP1A1 expression was assessed with a real-time PCR technique. The results indicated there was over-expression of the AhR mRNA in PBMC when the cells were treated with DDE and PCB 153. No changes in expression levels of CYP1A1 mRNA were found. Importantly, when the cells were exposed to DDE and PCB 153 in the presence of an antagonist of tumor necrosis factor (TNF)-α, the over-expression of AhR was abolished; as expected, the expression of CYP1A1 was unaffected. In conclusion, these studies demonstrated for the first time an increment of AhR expression "in vitro" in PBMC treated with two pro-inflammatory environmental pollutants, DDE and PCB153. Moreover, the over-expression of AhR was dependent of TNFα induced by DDE and PCB 153 and was independent of AhR activation.

  15. Mixed-ligand copper(II) complexes activate aryl hydrocarbon receptor AhR and induce CYP1A genes expression in human hepatocytes and human cell lines.

    PubMed

    Kubešová, Kateřina; Dořičáková, Aneta; Trávníček, Zdeněk; Dvořák, Zdeněk

    2016-07-25

    The effects of four copper(II) mixed-ligand complexes [Cu(qui1)(L)]NO3·H2O (1-3) and [Cu(qui2)(phen)]NO3 (4), where qui1=2-phenyl-3-hydroxy-4(1H)-quinolinone, Hqui2=2-(4-amino-3,5-dichlorophenyl)-N-propyl-3-hydroxy-4(1H)-quinolinone-7-carboxamide, L=1,10-phenanthroline (phen) (1), 5-methyl-1,10-phenanthroline (mphen) (2), bathophenanthroline (bphen) (3), on transcriptional activities of steroid receptors, nuclear receptors and xenoreceptors have been studied. The complexes (1-4) did not influence basal or ligand-inducible activities of glucocorticoid receptor, androgen receptor, thyroid receptor, pregnane X receptor and vitamin D receptor, as revealed by gene reporter assays. The complexes 1 and 2 dose-dependently induced luciferase activity in stable gene reporter AZ-AhR cell line, and this induction was reverted by resveratrol, indicating involvement of aryl hydrocarbon receptor (AhR) in the process. The complexes 1, 2 and 3 induced CYP1A1 mRNA in LS180 cells and CYP1A1/CYP1A2 in human hepatocytes through AhR. Electrophoretic mobility shift assay EMSA showed that the complexes 1 and 2 transformed AhR in its DNA-binding form. Collectively, we demonstrate that the complexes 1 and 2 activate AhR and induce AhR-dependent genes in human hepatocytes and cancer cell lines. In conclusion, the data presented here might be of toxicological importance, regarding the multiple roles of AhR in human physiology and pathophysiology. PMID:27180721

  16. Aryl hydrocarbon receptor (AhR) inducers and estrogen receptor (ER) activities in surface sediments of Three Gorges Reservoir, China evaluated with in vitro cell bioassays.

    PubMed

    Wang, Jingxian; Bovee, Toine F H; Bi, Yonghong; Bernhöft, Silke; Schramm, Karl-Werner

    2014-02-01

    Two types of biological tests were employed for monitoring the toxicological profile of sediment cores in the Three Gorges Reservoir (TGR), China. In the present study, sediments collected in June 2010 from TGR were analyzed for estrogen receptor (ER)- and aryl hydrocarbon receptor (AhR)-mediated activities. The estrogenic activity was assessed using a rapid yeast estrogen bioassay, based on the expression of a green fluorescent reporter protein. Weak anti-estrogenic activity was detected in sediments from an area close to the dam of the reservoir, and weak estrogenic activities ranging from 0.3 to 1 ng 17β-estradiol (E2) equivalents (EQ) g(-1) dry weight sediment (dw) were detected in sediments from the Wanzhou to Guojiaba areas. In the upstream areas Wanzhou and Wushan, sediments demonstrated additive effects in co-administration of 1 nM E2 in the yeast test system, while sediments from the downstream Badong and Guojiaba areas showed estrogenic activities which seemed to be more than additive (synergistic activity). There was an increasing tendency in estrogenic activity from upstream of TGR to downstream, while this tendency terminated and converted into anti-estrogenic activity in the area close to the dam. The AhR activity was detected employing rat hepatoma cell line (H4IIE). EROD activities were found homogenously distributed in sediments in TGR ranging from 200 to 311 pg 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) EQ g(-1) dw for total AhR agonists and from 45 to 76 pg TCDD EQ g(-1) dw for more persistent AhR agonists. The known AhR agonists polycyclic aromatic hydrocarbon, polychlorinated biphenyl, and PCDD/F only explained up to 8 % of the more persistent AhR agonist activity in the samples, which suggests that unidentified AhR-active compounds represented a great proportion of the TCDD EQ in sediments from TGR. These findings of estrogenic potential and dioxin-like activity in TGR sediments provide possible weight-of-evidence of potential

  17. Aryl hydrocarbon receptor (AhR) inducers and estrogen receptor (ER) activities in surface sediments of Three Gorges Reservoir, China evaluated with in vitro cell bioassays.

    PubMed

    Wang, Jingxian; Bovee, Toine F H; Bi, Yonghong; Bernhöft, Silke; Schramm, Karl-Werner

    2014-02-01

    Two types of biological tests were employed for monitoring the toxicological profile of sediment cores in the Three Gorges Reservoir (TGR), China. In the present study, sediments collected in June 2010 from TGR were analyzed for estrogen receptor (ER)- and aryl hydrocarbon receptor (AhR)-mediated activities. The estrogenic activity was assessed using a rapid yeast estrogen bioassay, based on the expression of a green fluorescent reporter protein. Weak anti-estrogenic activity was detected in sediments from an area close to the dam of the reservoir, and weak estrogenic activities ranging from 0.3 to 1 ng 17β-estradiol (E2) equivalents (EQ) g(-1) dry weight sediment (dw) were detected in sediments from the Wanzhou to Guojiaba areas. In the upstream areas Wanzhou and Wushan, sediments demonstrated additive effects in co-administration of 1 nM E2 in the yeast test system, while sediments from the downstream Badong and Guojiaba areas showed estrogenic activities which seemed to be more than additive (synergistic activity). There was an increasing tendency in estrogenic activity from upstream of TGR to downstream, while this tendency terminated and converted into anti-estrogenic activity in the area close to the dam. The AhR activity was detected employing rat hepatoma cell line (H4IIE). EROD activities were found homogenously distributed in sediments in TGR ranging from 200 to 311 pg 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) EQ g(-1) dw for total AhR agonists and from 45 to 76 pg TCDD EQ g(-1) dw for more persistent AhR agonists. The known AhR agonists polycyclic aromatic hydrocarbon, polychlorinated biphenyl, and PCDD/F only explained up to 8 % of the more persistent AhR agonist activity in the samples, which suggests that unidentified AhR-active compounds represented a great proportion of the TCDD EQ in sediments from TGR. These findings of estrogenic potential and dioxin-like activity in TGR sediments provide possible weight-of-evidence of potential

  18. Aromatic hydrocarbons upregulate glyceraldehyde-3-phosphate dehydrogenase and induce changes in actin cytoskeleton. Role of the aryl hydrocarbon receptor (AhR).

    PubMed

    Reyes-Hernández, O D; Mejía-García, A; Sánchez-Ocampo, E M; Castro-Muñozledo, F; Hernández-Muñoz, R; Elizondo, G

    2009-12-21

    Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional enzyme involved in several cellular functions including glycolysis, membrane transport, microtubule assembly, DNA replication and repair, nuclear RNA export, apoptosis, and the detection of nitric oxide stress. Therefore, modifications in the regulatory ability and function of GAPDH may alter cellular homeostasis. We report here that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and beta-naphthoflavone, which are well-known ligands for the aryl hydrocarbon receptor (AhR), increase GAPDH mRNA levels in vivo and in vitro, respectively. These compounds fail to induce GAPDH transcription in an AhR-null mouse model, suggesting that the increase in GAPDH level is dependent upon AhR activation. To analyse the consequences of AhR ligands on GAPDH function, mice were treated with TCDD and the level of liver activity of GAPDH was determined. The results showed that TCDD treatment increased GAPDH activity. On the other hand, treatment of Hepa-1 cells with beta-naphthoflavone leads to an increase in microfilament density when compared to untreated cultures. Collectively, these results suggest that AhR ligands, such as polycyclic hydrocarbons, can modify GAPDH expression and, therefore, have the potential to alter the multiple functions of this enzyme.

  19. Selective Aryl Hydrocarbon Receptor Modulator 3,3'-Diindolylmethane Impairs AhR and ARNT Signaling and Protects Mouse Neuronal Cells Against Hypoxia.

    PubMed

    Rzemieniec, J; Litwa, E; Wnuk, A; Lason, W; Krzeptowski, W; Kajta, M

    2016-10-01

    The neuroprotective potential of 3,3'-diindolylmethane (DIM), which is a selective aryl hydrocarbon receptor modulator, has recently been shown in cellular and animal models of Parkinson's disease and lipopolysaccharide-induced inflammation. However, there are no data concerning the protective capacity and mechanisms of DIM action in neuronal cells exposed to hypoxia. The aim of the present study was to investigate the neuroprotective potential of DIM against the hypoxia-induced damage in mouse hippocampal cells in primary cultures, with a particular focus on DIM interactions with the aryl hydrocarbon receptor (AhR), its nuclear translocator ARNT, and estrogen receptor β (ERβ). In the present study, 18 h of hypoxia induced apoptotic processes, in terms of the mitochondrial membrane potential, activation of caspase-3, and fragmentation of cell nuclei. These effects were accompanied by substantial lactate dehydrogenase release and neuronal cell death. The results of the present study demonstrated strong neuroprotective and anti-apoptotic actions of DIM in hippocampal cells exposed to hypoxia. In addition, DIM decreased the Ahr and Arnt mRNA expression and stimulated Erβ mRNA expression level. DIM-induced mRNA alterations were mirrored by changes in protein levels, except for ERβ, as detected by ELISA, Western blotting, and immunofluorescence labeling. We also demonstrated that DIM decreased the expression of AhR-regulated CYP1A1. Using specific siRNAs, we provided evidence that impairment of AhR and ARNT, but not ERβ plays a key role in the neuroprotective action of DIM against hypoxia-induced cell damage. This study may have implication for identifying new agents that could protect neurons against hypoxia by targeting AhR/ARNT signaling. PMID:26476840

  20. Use of natural AhR ligands as potential therapeutic modalities against inflammatory disorders

    PubMed Central

    Busbee, Philip B; Rouse, Michael; Nagarkatti, Mitzi; Nagarkatti, Prakash S

    2014-01-01

    The aim of this review is to discuss research involving ligands for the aryl hydrocarbon receptor (AhR) and their role in immunomodulation. While activation of the AhR is well known for its ability to regulate the biochemical and toxic effects of environmental chemicals, more recently an exciting discovery has been made indicating that AhR ligation can also regulate T-cell differentiation, specifically through activation of Foxp3+ regulatory T cells (Tregs) and downregulation of the proinflammatory Th17 cells. Such findings have opened new avenues of research on the possibility of targeting the AhR to treat inflammatory and autoimmune diseases. Specifically, this review will discuss the current research involving natural and dietary AhR ligands. In addition, evidence indicating the potential use of these ligands in regulating inflammation in various diseases will be highlighted. The importance of the AhR in immunological processes can be illustrated by expression of this receptor on a majority of immune cell types. In addition, AhR signaling pathways have been reported to influence a number of genes responsible for mediating inflammation and other immune responses. As interest in the AhR and its ligands increases, it seems prudent to consolidate current research on the contributions of these ligands to immune regulation during the course of inflammatory diseases. PMID:23731446

  1. Pityriazepin and other potent AhR ligands isolated from Malassezia furfur yeast

    PubMed Central

    Mexia, Nikitia; Gaitanis, George; Velegraki, Aristea; Soshilov, Anatoly; Denison, Michael S.; Magiatis, Prokopios

    2015-01-01

    Malassezia furfur yeast strains isolated from diseased human skin preferentially biosynthesize indole alkaloids which can be detected in human skin and are highly potent activators of the aryl hydrocarbon receptor (AhR) and AhR-dependent gene expression. Chemical analysis of an EtOAc extract of a M. furfur strain obtained from diseased human skin and grown on L-tryptophan agar revealed several known AhR active tryptophan metabolites along with a previously unidentified compound, pityriazepin. While its structure resembled that of the known alkaloid pityriacitrin, the comprised pyridine ring had been transformed into an azepinone. The indoloazepinone scaffold of pityriazepin is extremely rare in nature and has only been reported once previously. Pityriazepin, like the other isolated compounds, was found to be a potent activator of the AhR-dependent reporter gene assays in recombinant cell lines derived from four different species, although significant species differences in relative potency was observed. The ability of pityriazepin to competitively bind to the AhR and directly stimulate AhR DNA binding classified it as a new naturally-occurring potent AhR agonist. Malassezia furfur produces an expanded collection of extremely potent naturally occurring AhR agonists, which produce their biological effects in a species-specific manner.1 PMID:25721496

  2. Aryl hydrocarbon receptor (AhR) activation during pregnancy, and in adult nulliparous mice, delays the subsequent development of DMBA-induced mammary tumors

    PubMed Central

    Wang, Tao; Gavin, Heather M.; Arlt, Volker M.; Lawrence, B. Paige; Fenton, Suzanne E.; Medina, Daniel; Vorderstrasse, Beth A.

    2010-01-01

    TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin), the prototypic ligand for the aryl hydrocarbon receptor (AhR), promotes tumor formation in some model systems. However with regard to breast cancer, epidemiological and animal studies are inconclusive as to whether exposure increases tumor incidence or may instead be protective. We have previously reported that mice exposed to TCDD during pregnancy have impaired differentiation of mammary tissue, including decreased branching and poor development of lobuloalveolar structures. Because normal pregnancy-induced mammary differentiation may protect against subsequent neoplastic transformation, we hypothesized that TCDD-treated mice would be more susceptible to chemical carcinogenesis after parturition. To test this, mice were treated with TCDD or vehicle during pregnancy. Four weeks later, DMBA (7,12-dimethylbenz[a]anthracene) was administered to induce mammary tumor formation. Contrary to our hypothesis, TCDD-exposed parous mice showed a four-week delay in tumor formation relative to controls, and had a lower tumor incidence throughout the 27-week time course. The same results were obtained in nulliparous mice given TCDD and DMBA on the same schedule. We next addressed whether the delayed tumor incidence was a reflection of decreased tumor initiation, by testing the formation of DMBA-DNA adducts and preneoplastic lesions, induction of cytochrome P450s, and cell proliferation. None of these markers of tumor initiation differed between vehicle- and TCDD-treated animals. The expression of CXCL12 and CXCR4 was also measured to address their possible role in tumorigenesis. Taken together, our results suggest that AhR activation by TCDD slows the promotion of preneoplastic lesions to overt mammary tumors. PMID:20521247

  3. AhR expression and polymorphisms are associated with risk of coronary arterial disease in Chinese population

    PubMed Central

    Huang, Shian; Shui, Xiaorong; He, Yuan; Xue, Yiqiang; Li, Jianwen; Li, Guoming; Lei, Wei; Chen, Can

    2015-01-01

    The aryl hydrocarbon receptor (AhR) mediates the control of environmental toxicity, and modulates the development and pathogenesis of the cardiovascular system. However, little is known about the role of AhR in coronary arterial disease (CAD) susceptibility. We therefore conducted a case-control study in a Chinese population, and assessed the potential association between AhR variants and CAD susceptibility. Compared with the controls, circulating AhR expression was found to be significantly increased in patients with CAD and its subtypes including ST-segment and non-ST-segment elevation myocardial infarction, and stable and unstable angina pectoris. Receiver operating characteristic (ROC) analysis to evaluate the effect of AhR on CAD progression showed it to be a potent biomarker for CAD. Genotype frequencies of AhR rs2066853 differed significantly between CAD and control subjects, while smoking and hyperlipidemia markedly promoted CAD risk relative to the AhR polymorphism. Moreover, a significant difference in AhR variant distribution was observed between the four CAD subtypes with different severities. The expression level and functional polymorphisms of circulating AhR may affect the susceptibility and progression of CAD in Chinese populations. This provides a novel view of the etiology and epidemiology of CAD, and will contribute to the diagnosis and therapy of this severe disease. PMID:25620626

  4. Enhancement of hypoxia-induced gene expression in fish liver by the aryl hydrocarbon receptor (AhR) ligand, benzo[a]pyrene (BaP).

    PubMed

    Yu, Richard Man Kit; Ng, Patrick Kwok Shing; Tan, Tianfeng; Chu, Daniel Ling Ho; Wu, Rudolf Shiu Sun; Kong, Richard Yuen Chong

    2008-11-21

    Fish in polluted coastal habitats commonly suffer simultaneous exposure to both hypoxia and xenobiotics. Although the adaptive molecular responses to each stress have been described, little is known about the interaction between the signaling pathways mediating these responses. Previous studies in mammalian hepatoma cell lines have shown that hypoxia-inducible factor (HIF)- and/or aryl hydrocarbon receptor (AhR)-activated gene expression is suppressed following co-exposure to hypoxia and the hallmark AhR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). However, whether similar crosstalk exists in the non-tumor liver tissues of fish and whether other non-TCDD ligands also play the same inhibitory role in this crosstalk remain unknown. Here, the in vivo hepatic mRNA expression profiles of multiple hypoxia- and AhR-responsive genes (later gene expression=mRNA expression of the gene) were examined in the orange-spotted grouper (Epinephelus coioides) upon single and combined exposures to hypoxia and benzo[a]pyrene (BaP). Combined exposure enhanced hypoxia-induced gene expression but did not significantly alter BaP-induced gene expression. Protein carbonyl content was markedly elevated in fish subjected to combined exposure, indicating accumulation of reactive oxygen species (ROS). Application of diethyldithiocarbamate (DDC) to hypoxia-treated grouper liver explants similarly exaggerated hypoxia-induced gene expression as in the combined stress tissues in vivo. These observations suggest that ROS derived from the combined hypoxia and BaP stress have a role in enhancing hypoxia-induced gene expression.

  5. Induction of human UGT1A1 by bilirubin through AhR dependent pathway.

    PubMed

    Togawa, Hiroshi; Shinkai, Shigeko; Mizutani, Takaharu

    2008-12-01

    UDP-glucuronosyltransferase1A1 (UGT1A1) plays a key role to conjugate bilirubin and preventing jaundice, but there is no report showing the induction of human UGT1A1 (UGT1A1) by bilirubin. In this report, we show findings of the induction of the reporter gene (-3475/+14) of UGT1A1 in HepG2 cells by bilirubin at 50 microM, 100 microM, with human aryl hydrocarbon receptor (hAhR). We confirmed that induction of the reporter gene by bilirubin is dependent on the position of the xenobiotic responsive element (XRE) (-3328/-3319) of UGT1A1, because the XRE deletion UGT1A1 gene did not respond to stimulation by a complex of bilirubin and hAhR. alpha-Naphthoflavone (alpha-NF) of a typical AhR antagonist at 50 microM inhibited induction by bilirubin, suggesting that bilirubin stimulates through binding with hAhR. Meanwhile, bilirubin itself did not stimulate the induction of AhR, because we detected no-elevation of the mRNA level of AhR by RT-PCR. These results indicate that the induction of UGT1A1 by bilirubin-AhR did not depend on the elevation of AhR but on ligand binding. From this result, we considered that high bilirubin in neonates must induce the elevation of UGT1A1 after birth to prevent jaundice, and bilirubin in adults also regulates the level of UGT1A1. This is the first report showing direct induction of UGT1A1 by a bilirubin through AhR pathway. PMID:19356098

  6. Effects of scorched food leachates with or without activated charcoal pretreatment on AhR activation in cultured cells.

    PubMed

    Takahashi, Satoshi; Morita, Koji; Kinoshita, Makoto; Fujimori, Shin; Ishikawa, Toshio

    2015-12-01

    Aryl hydrocarbon receptor (AhR) is a transcription factor activated by xenobiotics, including dioxins and polycyclic aromatic hydrocarbons (PAHs). Although AhR is also activated by some dietary constituents, it has not been completely clarified in what circumstances AhR ligands are ingested in our daily life. Because PAHs are formed by the incomplete combustion of organic materials, we hypothesized that scorched foods might contain and leach out AhR ligands sufficient to stimulate AhR in vitro. To test this hypothesis, scorched foods (bread, cheese, etc.) were mixed vigorously with water, and the supernatants were retrieved as samples. The samples were added to HepG2 cells stably expressing an AhR-responsive reporter gene. Also, expression of CYP1A1, an endogenous AhR-responsive gene, was analyzed by RT-PCR in different cell lines treated with the samples. We further tested whether pretreatment of the samples with activated charcoal would alter their AhR-stimulating activity. All the supernatant samples tested induced AhR-dependent reporter gene activity and CYP1A1 mRNA expression. In some samples, these inductions were inhibited by pretreatment with activated charcoal. Our findings indicate that scorched food leachates stimulate AhR in cultured cells and that activated charcoal adsorbs the AhR-stimulating substances in some leachates. Thus, people who habitually eat scorched foods are exposed to AhR ligands on a regular basis. Further studies are needed to elucidate whether burnt foods actually exert biological effects on our health.

  7. Effects of scorched food leachates with or without activated charcoal pretreatment on AhR activation in cultured cells.

    PubMed

    Takahashi, Satoshi; Morita, Koji; Kinoshita, Makoto; Fujimori, Shin; Ishikawa, Toshio

    2015-12-01

    Aryl hydrocarbon receptor (AhR) is a transcription factor activated by xenobiotics, including dioxins and polycyclic aromatic hydrocarbons (PAHs). Although AhR is also activated by some dietary constituents, it has not been completely clarified in what circumstances AhR ligands are ingested in our daily life. Because PAHs are formed by the incomplete combustion of organic materials, we hypothesized that scorched foods might contain and leach out AhR ligands sufficient to stimulate AhR in vitro. To test this hypothesis, scorched foods (bread, cheese, etc.) were mixed vigorously with water, and the supernatants were retrieved as samples. The samples were added to HepG2 cells stably expressing an AhR-responsive reporter gene. Also, expression of CYP1A1, an endogenous AhR-responsive gene, was analyzed by RT-PCR in different cell lines treated with the samples. We further tested whether pretreatment of the samples with activated charcoal would alter their AhR-stimulating activity. All the supernatant samples tested induced AhR-dependent reporter gene activity and CYP1A1 mRNA expression. In some samples, these inductions were inhibited by pretreatment with activated charcoal. Our findings indicate that scorched food leachates stimulate AhR in cultured cells and that activated charcoal adsorbs the AhR-stimulating substances in some leachates. Thus, people who habitually eat scorched foods are exposed to AhR ligands on a regular basis. Further studies are needed to elucidate whether burnt foods actually exert biological effects on our health. PMID:26558458

  8. Functional and phenotypic effects of AhR activation in inflammatory dendritic cells

    SciTech Connect

    Bankoti, Jaishree; Rase, Ben; Simones, Tom; Shepherd, David M.

    2010-07-15

    Aryl hydrocarbon receptor (AhR) activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces immune suppression. Dendritic cells (DCs) are key antigen presenting cells governing T cell activation and differentiation. However, the consequences of AhR activation in DCs are not fully defined. We hypothesized that AhR activation alters DC differentiation and generates dysfunctional DCs. To test this hypothesis, inflammatory bone marrow-derived DCs (BMDCs) from C57Bl/6 mice were generated in the presence of vehicle or TCDD. TCDD decreased CD11c expression but increased MHC class II, CD86 and CD25 expression on the BMDCs. The effects of TCDD were strictly AhR-dependent but not exclusively DRE-mediated. Similar effects were observed with two natural AhR ligands, 6-formylindolo[3,2-b]carbazole (FICZ) and 2-(1H-Indol-3-ylcarbonyl)-4-thiazolecarboxylic acid (ITE). TCDD increased LPS- and CpG-induced IL-6 and TNF-{alpha} production by BMDCs but decreased their NO production. TCDD decreased CpG-induced IL-12p70 production by BMDCs but did not affect their secretion of IL-10. TCDD downregulated LPS- and CpG-induced NF-kB p65 levels and induced a trend towards upregulation of RelB levels in the BMDCs. AhR activation by TCDD modulated BMDC uptake of both soluble and particulate antigens. Induction of indoleamine-2,3-dioxygenase (IDO) and TGF-{beta}3 has been implicated in the generation of regulatory T cells following AhR activation. TCDD increased IDO1, IDO2 and TGF-{beta}3 mRNA levels in BMDCs as compared to vehicle. Despite the induction of regulatory mediators, TCDD-treated BMDCs failed to suppress antigen-specific T cell activation. Thus, AhR activation can directly alter the differentiation and innate functions of inflammatory DCs without affecting their ability to successfully interact with T cells.

  9. Newspapers and newspaper ink contain agonists for the ah receptor.

    PubMed

    Bohonowych, Jessica E S; Zhao, Bin; Timme-Laragy, Alicia; Jung, Dawoon; Di Giulio, Richard T; Denison, Michael S

    2008-04-01

    Ligand-dependent activation of the aryl hydrocarbon receptor (AhR) pathway leads to a diverse array of biological and toxicological effects. The best-studied ligands for the AhR include polycyclic and halogenated aromatic hydrocarbons, the most potent of which is 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). However, as new AhR ligands are identified and characterized, their structural and physiochemical diversity continues to expand. Our identification of AhR agonists in crude extracts from diverse materials raises questions as to the magnitude and extent of human exposure to AhR ligands through normal daily activities. We have found that solvent extracts of newspapers from countries around the world stimulate the AhR signaling pathway. AhR agonist activity was observed for dimethyl sulfoxide (DMSO), ethanol, and water extracts of printed newspaper, unprinted virgin paper, and black printing ink, where activation of luciferase reporter gene expression was transient, suggesting that the AhR active chemical(s) was metabolically labile. DMSO and ethanol extracts also stimulated AhR transformation and DNA binding, and also competed with [(3)H]TCDD for binding to the AhR. In addition, DMSO extracts of printed newspaper induced cytochrome P450 1A associated 7-ethoxyresorufin-O-deethylase activity in zebrafish embryos in vivo. Although the responsible bioactive chemical(s) remain to be identified, our results demonstrate that newspapers and printing ink contain relatively potent metabolically labile agonists of the AhR. Given the large amount of recycling and reprocessing of newspapers throughout the world, release of these easily extractable AhR agonists into the environment should be examined and their potential effects on aquatic organisms assessed. PMID:18203687

  10. The AhR is involved in the regulation of LoVo cell proliferation through cell cycle-associated proteins.

    PubMed

    Yin, Jiuheng; Sheng, Baifa; Han, Bin; Pu, Aimin; Yang, Kunqiu; Li, Ping; Wang, Qimeng; Xiao, Weidong; Yang, Hua

    2016-05-01

    Some ingredients in foods can activate the aryl hydrocarbon receptor (AhR) and arrest cell proliferation. In this study, we hypothesized that 6-formylindolo [3, 2-b] carbazole (FICZ) arrests the cell cycle in LoVo cells (a colon cancer line) through the AhR. The AhR agonist FICZ and the AhR antagonist CH223191 were used to treat LoVo cells. Real-time PCR and Western blot analyses were performed to detect the expression of the AhR, CYP1A1, CDK4, cyclinD1, cyclin E, CDK2, P27, and pRb. The distribution and activation of the AhR were detected with immunofluorescence. A 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay and flow cytometric analysis were performed to measure cell viability, cell cycle stage, and apoptosis. Our results show that FICZ inhibited LoVo cell proliferation by inducing G1 cell cycle arrest but had no effect on epithelial apoptosis. Further analysis found that FICZ downregulated cyclinD1 and upregulated p27 expression to arrest Rb phosphorylation. The downregulation of cyclinD1 and upregulation of p27 were abolished by co-treatment with CH223191. We conclude that the AhR, when activated by FICZ (an endogenous AhR ligand), can arrest the cell cycle and block LoVo cell proliferation.

  11. A novel AhR ligand, 2AI, protects the retina from environmental stress

    PubMed Central

    Gutierrez, Mark A.; Davis, Sonnet S.; Rosko, Andrew; Nguyen, Steven M.; Mitchell, Kylie P.; Mateen, Samiha; Neves, Joana; Garcia, Thelma Y.; Mooney, Shaun; Perdew, Gary H.; Hubbard, Troy D.; Lamba, Deepak A.; Ramanathan, Arvind

    2016-01-01

    Various retinal degenerative diseases including dry and neovascular age-related macular degeneration (AMD), retinitis pigmentosa, and diabetic retinopathy are associated with the degeneration of the retinal pigmented epithelial (RPE) layer of the retina. This consequently results in the death of rod and cone photoreceptors that they support, structurally and functionally leading to legal or complete blindness. Therefore, developing therapeutic strategies to preserve cellular homeostasis in the RPE would be a favorable asset in the clinic. The aryl hydrocarbon receptor (AhR) is a conserved, environmental ligand-dependent, per ARNT-sim (PAS) domain containing bHLH transcription factor that mediates adaptive response to stress via its downstream transcriptional targets. Using in silico, in vitro and in vivo assays, we identified 2,2′-aminophenyl indole (2AI) as a potent synthetic ligand of AhR that protects RPE cells in vitro from lipid peroxidation cytotoxicity mediated by 4-hydroxynonenal (4HNE) as well as the retina in vivo from light-damage. Additionally, metabolic characterization of this molecule by LC-MS suggests that 2AI alters the lipid metabolism of RPE cells, enhancing the intracellular levels of palmitoleic acid. Finally, we show that, as a downstream effector of 2AI-mediated AhR activation, palmitoleic acid protects RPE cells from 4HNE-mediated stress, and light mediated retinal degeneration in mice. PMID:27364765

  12. Deciphering Dimerization Modes of PAS Domains: Computational and Experimental Analyses of the AhR:ARNT Complex Reveal New Insights Into the Mechanisms of AhR Transformation

    PubMed Central

    Corrada, Dario; Soshilov, Anatoly A.; Denison, Michael S.

    2016-01-01

    The Aryl hydrocarbon Receptor (AhR) is a transcription factor that mediates the biochemical response to xenobiotics and the toxic effects of a number of environmental contaminants, including dioxins. Recently, endogenous regulatory roles for the AhR in normal physiology and development have also been reported, thus extending the interest in understanding its molecular mechanisms of activation. Since dimerization with the AhR Nuclear Translocator (ARNT) protein, occurring through the Helix-Loop-Helix (HLH) and PER-ARNT-SIM (PAS) domains, is needed to convert the AhR into its transcriptionally active form, deciphering the AhR:ARNT dimerization mode would provide insights into the mechanisms of AhR transformation. Here we present homology models of the murine AhR:ARNT PAS domain dimer developed using recently available X-ray structures of other bHLH-PAS protein dimers. Due to the different reciprocal orientation and interaction surfaces in the different template dimers, two alternative models were developed for both the PAS-A and PAS-B dimers and they were characterized by combining a number of computational evaluations. Both well-established hot spot prediction methods and new approaches to analyze individual residue and residue-pairwise contributions to the MM-GBSA binding free energies were adopted to predict residues critical for dimer stabilization. On this basis, a mutagenesis strategy for both the murine AhR and ARNT proteins was designed and ligand-dependent DNA binding ability of the AhR:ARNT heterodimer mutants was evaluated. While functional analysis disfavored the HIF2α:ARNT heterodimer-based PAS-B model, most mutants derived from the CLOCK:BMAL1-based AhR:ARNT dimer models of both the PAS-A and the PAS-B dramatically decreased the levels of DNA binding, suggesting this latter model as the most suitable for describing AhR:ARNT dimerization. These novel results open new research directions focused at elucidating basic molecular mechanisms underlying the

  13. Gene cloning and expression analysis of AhR and CYP4 from Pinctada martensii after exposed to pyrene.

    PubMed

    Du, Junqiao; Liao, Chenghong; Zhou, Hailong; Diao, Xiaoping; Li, Yuhu; Zheng, Pengfei; Wang, Fuqiang

    2015-10-01

    Pyrene, a typical polycyclic aromatic hydrocarbon, is a common pollutant in the marine environment. Polycyclic aromatic hydrocarbons initiate cellular detoxification in an exposed organism via the activation of the aryl hydrocarbon receptor (AhR). Subsequent metabolism of these xenobiotics is mainly by the cytochrome P450 enzymes of the phase I detoxification system. Full-length complementary DNA sequences from the pearl oyster Pinctada martensii (pm) encoding AhR and cytochrome P4 were cloned. The P. martensii AhR complementary DNA sequence constitutes an open reading frame that encodes for 848 amino acids. Sequence analysis indicated PmAhR showed high similarity with its homologues of other bivalve species. The cytochrome P(CYP)4 complementary DNA sequence of P. martensii constitutes an open reading frame that encodes for 489 amino acids. Quantitative real-time analysis detected both PmAhR and PmCYP4 messenger RNA expressions in the mantle, gill, hepatapancreas and adductor muscle of P. martensii exposed to pyrene. The highest transcript-band intensities of PmAhR and PmCYP4 were observed in the gill. Temporal expression of PmAhR and PmCYP4 messenger RNAs induction was observed in gills and increased between 3 and 5 days post exposure; then returned to control level. These results suggest that messenger RNAs of PmAhR and PmCYP4 in pearl oysters might be useful parameters for monitoring marine environment pyrene pollution.

  14. AhR activation underlies the CYP1A autoinduction by A-998679 in rats

    PubMed Central

    Liguori, Michael J.; Lee, Chih-Hung; Liu, Hong; Ciurlionis, Rita; Ditewig, Amy C.; Doktor, Stella; Andracki, Mark E.; Gagne, Gerard D.; Waring, Jeffrey F.; Marsh, Kennan C.; Gopalakrishnan, Murali; Blomme, Eric A. G.; Yang, Yi

    2012-01-01

    Xenobiotic-mediated induction of cytochrome P450 (CYP) drug metabolizing enzymes (DMEs) is frequently encountered in drug discovery and can influence disposition, pharmacokinetic, and toxicity profiles. The CYP1A subfamily of DMEs plays a central role in the biotransformation of several drugs and environmental chemicals. Autoinduction of drugs through CYP3A enzymes is a common mechanism for their enhanced clearance. However, autoinduction via CYP1A is encountered less frequently. In this report, an experimental compound, A-998679 [3-(5-pyridin-3-yl-1,2,4-oxadiazol-3-yl) benzonitrile], was shown to enhance its own clearance via induction of Cyp1a1 and Cyp1a2. Rats were dosed for 5 days with 30, 100, and 200 mg/kg/day A-998679. During the dosing period, the compound's plasma AUC decreased at 30 mg/kg (95%) and 100 mg/kg (80%). Gene expression analysis and immunohistochemistry of the livers showed a large increase in the mRNA and protein levels of Cyp1a, which was involved in the biotransformation of A-998679. Induction of CYP1A was confirmed in primary rat, human, and dog hepatocytes. The compound also weakly inhibited CYP1A2 in human liver microsomes. A-998679 activated the aryl hydrocarbon receptor (AhR) in a luciferase gene reporter assay in HepG2 cells, upregulated expression of genes associated with AhR activation in rat liver and enhanced nuclear migration of AhR in HepG2 cells. Collectively these results demonstrate that A-998679 is an AhR activator that induces Cyp1a1 and Cyp1a2 expression, resulting in an autoinduction phenomenon. The unique properties of A-998679, along with its novel structure distinct from classical polycyclic aromatic hydrocarbons (PAHs), may warrant its further evaluation as a tool compound for use in studies involving AhR biology and CYP1A-related mechanisms of drug metabolism and toxicity. PMID:23112805

  15. Aryl hydrocarbon receptor (AHR)-active pharmaceuticals are selective AHR modulators in MDA-MB-468 and BT474 breast cancer cells.

    PubMed

    Jin, Un-Ho; Lee, Syng-ook; Safe, Stephen

    2012-11-01

    Leflunomide, flutamide, nimodipine, mexiletine, sulindac, tranilast, 4-hydroxytamoxifen, and omeprazole are pharmaceuticals previously characterized as aryl hydrocarbon receptor (AHR) agonists in various cell lines and animal models. In this study, the eight AHR-active pharmaceuticals were investigated in highly aggressive aryl hydrocarbon (Ah)-responsive BT474 and MDA-MB-468 breast cancer cell lines, and their effects on AHR protein, CYP1A1 (protein and mRNA), CYP1B1 (mRNA), and cell migration were determined. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) was used as a positive control. The AHR agonist activities of the pharmaceuticals depended on structure, response, and cell context. Most compounds induced one or more AHR-mediated responses in BT474 cells, whereas in Ah-responsive MDA-MB-468 cells effects of the AHR-active pharmaceuticals were highly variable. 4-Hydroxytamoxifen, mexiletine, and tranilast did not induce CYP1A1 in MDA-MB-468 cells; moreover, in combination with TCDD, mexiletine was a potent AHR antagonist, tranilast was a partial antagonist, and 4-hydroxytamoxifen also exhibited some AHR antagonist activity. Omeprazole and, to a lesser extent, sulindac and leflunomide were full and partial AHR agonists, respectively, in both breast cancer cell lines. These data indicate that the AHR-active pharmaceuticals are selective AHR modulators, and applications of these drugs for targeting the AHR must be confirmed by studies using the most relevant cell context. PMID:22879383

  16. Non-dioxin-like AhR ligands in a mouse peanut allergy model.

    PubMed

    Schulz, Veronica J; Smit, Joost J; Huijgen, Veerle; Bol-Schoenmakers, Marianne; van Roest, Manon; Kruijssen, Laura J W; Fiechter, Daniëlle; Hassing, Ine; Bleumink, Rob; Safe, Stephen; van Duursen, Majorie B M; van den Berg, Martin; Pieters, Raymond H H

    2012-07-01

    Recently, we have shown that AhR activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) suppresses sensitization to peanut at least in part by inducing a functional shift toward CD4(+)CD25(+)Foxp3(+) T cells. Next to TCDD, numerous other AhR ligands have been described. In this study, we investigated the effect of three structurally different non-dioxin-like AhR ligands, e.g., 6-formylindolo[3,2-b]carbazole (FICZ), β-naphthoflavone (β-NF), and 6-methyl-1,3,8-trichlorodibenzofuran (6-MCDF), on peanut sensitization. Female C57BL/6 mice were sensitized by administering peanut extract (PE) by gavage in the presence of cholera toxin. Before and during peanut sensitization, mice were treated with FICZ, β-NF, or 6-MCDF. AhR gene transcription in duodenum and liver was investigated on day 5, even as the effect of these AhR ligands on CD4(+)CD25(+)Foxp3(+) T(reg) cells in spleen and mesenteric lymph nodes (MLNs). Mice treated with TCDD were included as a positive control. Furthermore, the murine reporter cell line H1G1.1c3 (CAFLUX) was used to investigate the possible role of metabolism of TCDD, FICZ, β-NF, and 6-MCDF on AhR activation in vitro. TCDD, but not FICZ, β-NF, and 6-MCDF, suppressed sensitization to peanut (measured by PE-specific IgE, IgG1, IgG2a and PE-induced interleukin (IL)-5, IL-10, IL-13, IL-17a, IL-22, and interferon-γ). In addition, FICZ, β-NF, and 6-MCDF treatments less effectively induced AhR gene transcription (measured by gene expression of AhR, AhRR, CYP1A1, CYP1A2, CYP1B1) compared with TCDD-treated mice. Furthermore, FICZ, β-NF and 6-MCDF did not increase the percentage of CD4(+)CD25(+)Foxp3(+) T(reg) cells in spleen and mesenteric lymph nodes compared with PE-sensitized mice, in contrast to TCDD. Inhibition of metabolism in vitro increased AhR activation. Together, these data shows that TCDD, but not FICZ, β-NF, and 6-MCDF suppresses sensitization to peanut. Differences in metabolism, AhR binding and subsequent gene transcription might

  17. Integration of Genome-Wide Computation DRE Search, AhR ChIP-chip and Gene Expression Analyses of TCDD-Elicited Responses in the Mouse Liver

    PubMed Central

    2011-01-01

    Background The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor (TF) that mediates responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Integration of TCDD-induced genome-wide AhR enrichment, differential gene expression and computational dioxin response element (DRE) analyses further elucidate the hepatic AhR regulatory network. Results Global ChIP-chip and gene expression analyses were performed on hepatic tissue from immature ovariectomized mice orally gavaged with 30 μg/kg TCDD. ChIP-chip analysis identified 14,446 and 974 AhR enriched regions (1% false discovery rate) at 2 and 24 hrs, respectively. Enrichment density was greatest in the proximal promoter, and more specifically, within ± 1.5 kb of a transcriptional start site (TSS). AhR enrichment also occurred distal to a TSS (e.g. intergenic DNA and 3' UTR), extending the potential gene expression regulatory roles of the AhR. Although TF binding site analyses identified over-represented DRE sequences within enriched regions, approximately 50% of all AhR enriched regions lacked a DRE core (5'-GCGTG-3'). Microarray analysis identified 1,896 number of TCDD-responsive genes (|fold change| ≥ 1.5, P1(t) > 0.999). Integrating this gene expression data with our ChIP-chip and DRE analyses only identified 625 differentially expressed genes that involved an AhR interaction at a DRE. Functional annotation analysis of differentially regulated genes associated with AhR enrichment identified overrepresented processes related to fatty acid and lipid metabolism and transport, and xenobiotic metabolism, which are consistent with TCDD-elicited steatosis in the mouse liver. Conclusions Details of the AhR regulatory network have been expanded to include AhR-DNA interactions within intragenic and intergenic genomic regions. Moreover, the AhR can interact with DNA independent of a DRE core suggesting there are alternative mechanisms of AhR-mediated gene regulation. PMID:21762485

  18. Access Path to the Ligand Binding Pocket May Play a Role in Xenobiotics Selection by AhR

    PubMed Central

    Szöllősi, Dániel; Erdei, Áron; Gyimesi, Gergely; Magyar, Csaba; Hegedűs, Tamás

    2016-01-01

    Understanding of multidrug binding at the atomic level would facilitate drug design and strategies to modulate drug metabolism, including drug transport, oxidation, and conjugation. Therefore we explored the mechanism of promiscuous binding of small molecules by studying the ligand binding domain, the PAS-B domain of the aryl hydrocarbon receptor (AhR). Because of the low sequence identities of PAS domains to be used for homology modeling, structural features of the widely employed HIF-2α and a more recent suitable template, CLOCK were compared. These structures were used to build AhR PAS-B homology models. We performed molecular dynamics simulations to characterize dynamic properties of the PAS-B domain and the generated conformational ensembles were employed in in silico docking. In order to understand structural and ligand binding features we compared the stability and dynamics of the promiscuous AhR PAS-B to other PAS domains exhibiting specific interactions or no ligand binding function. Our exhaustive in silico binding studies, in which we dock a wide spectrum of ligand molecules to the conformational ensembles, suggest that ligand specificity and selection may be determined not only by the PAS-B domain itself, but also by other parts of AhR and its protein interacting partners. We propose that ligand binding pocket and access channels leading to the pocket play equally important roles in discrimination of endogenous molecules and xenobiotics. PMID:26727491

  19. Behavioral Rhythmicity of Mice Lacking AhR and Attenuation of Light-induced Phase Shift by 2,3,7,8-Tetracholordibenzo-p-dioxin

    PubMed Central

    Mukai, Motoko; Lin, Tien-Min; Peterson, Richard E.; Cooke, Paul S.; Tischkau, Shelley A.

    2008-01-01

    Transcription factors belonging to the Per/Arnt/Sim (PAS) domain family are highly conserved and many are involved in circadian rhythm regulation. One member of this family, aryl hydrocarbon receptor (AhR), is an orphan receptor whose physiological role is unknown. Recent findings have led to the hypothesis that AhR has a role in circadian rhythm, which is the focus of the present investigation. First, time-of-day dependent mRNA expression of AhR and its signaling target, cytochrome p4501A1 (Cyp1a1) was determined in C57BL/6J mice by quantitative RT-PCR. Circadian expression of AhR and Cyp1a1 was observed both in the suprachiasmatic nucleus (SCN) and liver. Next, the circadian phenotype of mice lacking AhR (AhRKO) was investigated using behavioral monitoring. Intact AhRKO mice had robust circadian rhythmicity with a similar tau under constant conditions compared to wild-type mice, but a significant difference in tau was observed between genotypes in ovariectomized female mice. Time to re-entrainment following 6-h advances or delays of the light/dark cycle was not significantly different between genotypes. However, mice exposed to the AhR agonist 2,3,7,8-tetracholorodibenzo-p-dioxin (TCDD, 1 μg/kg BW) displayed decreased phase shifts in response to light and had altered expression of Per1 and Bmal1. These results suggest that chronic activation of AhR may affect the ability of the circadian timekeeping system to adjust to alterations in environmental lighting by affecting canonical clock genes. Further studies are necessary to decipher the mechanism of how AhR agonists could disrupt light-induced phase shifts. If AhR does have a role in circadian rhythm, it may share redundant roles with other PAS domain proteins and/or the role of AhR may not be exhibited in the behavioral activity rhythm, but could be important elsewhere in the peripheral circadian system. PMID:18487412

  20. Ultraviolet light converts propranolol, a nonselective β-blocker and potential lupus-inducing drug, into a proinflammatory AhR ligand.

    PubMed

    Dorgham, Karim; Amoura, Zahir; Parizot, Christophe; Arnaud, Laurent; Frances, Camille; Pionneau, Cédric; Devilliers, Hervé; Pinto, Sandra; Zoorob, Rima; Miyara, Makoto; Larsen, Martin; Yssel, Hans; Gorochov, Guy; Mathian, Alexis

    2015-11-01

    UV light and some medications are known to trigger lupus erythematosus (LE). A common mechanism underlying the immunopathologic effect, resulting from exposure to these two seemingly unrelated factors, remains unknown. The aryl hydrocarbon receptor (AhR) plays a key role in the regulation of IL-22 production in humans and can be activated by both xenobiotics and naturally occurring photoproducts. A significant expansion of Th17 and Th22 cells was observed in the peripheral blood of active systemic LE (SLE) patients, compared to inactive patients and controls. We also show that propranolol, a potential lupus-inducing drug, induced stronger AhR activation in PBMCs of SLE patients than in those of controls. AhR agonist activity of propranolol was enhanced by UV light exposure. MS analysis of irradiated propranolol revealed the generation of a proinflammatory photoproduct. This compound behaves like the prototypic AhR ligand 6-formylindolo[3,2-b]carbazole, a cutaneous UV light-induced tryptophan metabolite, both promoting IL-22, IL-8, and CCL2 secretion by T-cells and macrophages. Finally, LE patients exhibit signs of cutaneous AhR activation that correlate with lesional expression of the same proinflammatory cytokines, suggesting a role for photometabolites in the induction of skin inflammation. The AhR might therefore represent a target for therapeutic intervention in LE. PMID:26354876

  1. Ultraviolet light converts propranolol, a nonselective β-blocker and potential lupus-inducing drug, into a proinflammatory AhR ligand.

    PubMed

    Dorgham, Karim; Amoura, Zahir; Parizot, Christophe; Arnaud, Laurent; Frances, Camille; Pionneau, Cédric; Devilliers, Hervé; Pinto, Sandra; Zoorob, Rima; Miyara, Makoto; Larsen, Martin; Yssel, Hans; Gorochov, Guy; Mathian, Alexis

    2015-11-01

    UV light and some medications are known to trigger lupus erythematosus (LE). A common mechanism underlying the immunopathologic effect, resulting from exposure to these two seemingly unrelated factors, remains unknown. The aryl hydrocarbon receptor (AhR) plays a key role in the regulation of IL-22 production in humans and can be activated by both xenobiotics and naturally occurring photoproducts. A significant expansion of Th17 and Th22 cells was observed in the peripheral blood of active systemic LE (SLE) patients, compared to inactive patients and controls. We also show that propranolol, a potential lupus-inducing drug, induced stronger AhR activation in PBMCs of SLE patients than in those of controls. AhR agonist activity of propranolol was enhanced by UV light exposure. MS analysis of irradiated propranolol revealed the generation of a proinflammatory photoproduct. This compound behaves like the prototypic AhR ligand 6-formylindolo[3,2-b]carbazole, a cutaneous UV light-induced tryptophan metabolite, both promoting IL-22, IL-8, and CCL2 secretion by T-cells and macrophages. Finally, LE patients exhibit signs of cutaneous AhR activation that correlate with lesional expression of the same proinflammatory cytokines, suggesting a role for photometabolites in the induction of skin inflammation. The AhR might therefore represent a target for therapeutic intervention in LE.

  2. AhR signaling activation disrupts migration and dendritic growth of olfactory interneurons in the developing mouse

    PubMed Central

    Kimura, Eiki; Ding, Yunjie; Tohyama, Chiharu

    2016-01-01

    Perinatal exposure to a low level of dioxin, a ubiquitous environmental pollutant, has been shown to induce abnormalities in learning and memory, emotion, and sociality in laboratory animals later in adulthood. However, how aryl hydrocarbon receptor (AhR) signaling activation disrupts the higher brain function remains unclear. Therefore, we studied the possible effects of excessive activation of AhR signaling on neurodevelopmental processes, such as cellular migration and neurite growth, in mice. To this end, we transfected a constitutively active-AhR plasmid into stem cells in the lateral ventricle by in vivo electroporation on postnatal day 1. Transfection was found to induce tangential migration delay and morphological abnormalities in neuronal precursors in the rostral migratory stream at 6 days post-electroporation (dpe) as well as disrupt radial migration in the olfactory bulb and apical and basal dendritic growth of the olfactory interneurons in the granule cell layer at 13 and 20 dpe. These results suggest that the retarded development of interneurons by the excessive AhR signaling may at least in part explain the dioxin-induced abnormal behavioral alterations previously reported in laboratory animals. PMID:27197834

  3. Characterization of AhR agonists reveals antagonistic activity in European herring gull (Larus argentatus) eggs.

    PubMed

    Muusse, Martine; Christensen, Guttorm; Gomes, Tânia; Kočan, Anton; Langford, Katherine; Tollefsen, Knut Erik; Vaňková, Lenka; Thomas, Kevin V

    2015-05-01

    European herring gull (Larus argentatus) eggs from two Norwegian islands, Musvær in the south east and Reiaren in Northern Norway, were screened for dioxins, furans, and dioxin-like and selected non-dioxin-like polychlorinated biphenyls (PCBs), and subjected to non-target analysis to try to identify the aryl hydrocarbon receptor (AhR) agonists, responsible for elevated levels measured using the dioxin responsive chemically activated luciferase expression (DR-CALUX) assay. Eggs from Musvær contained chemically calculated toxic equivalent (WHO TEQ) levels of between 109 and 483 pg TEQ/g lw, and between 82 and 337 pg TEQ/g lw was determined in eggs from Reiaren. In particular PCB126 contributed highly to the total TEQ (69-82%). In 19 of the 23 samples the calculated WHO TEQ was higher than the TEQCALUX. Using CALUX specific relative effect potencies (REPs), the levels were lower at between 77 and 292 pg/g lw in eggs from Musvær and between 55 and 223 pg/g lw in eggs from Reiaren, which was higher than the TEQCALUX in 16 of the 23 samples. However, the means of the REP values and the TEQCALUX were not significantly different. This suggests the presence of compounds that can elicit antagonist effects, with a low binding affinity to the AhR. Non-target analysis identified the presence of hexachlorobenzene (HCB) (quantified at 9.6-185 pg/g lw) but neither this compound nor high concentrations of PCB126 and non-dioxin-like PCBs could explain the differences between the calculated TEQ or REP values and the TEQCALUX. Even though, for most AhR agonists, the sensitivity of herring gulls is not known, the reported levels can be considered to represent a risk for biological effects in the developing embryo, compared to LC50 values in chicken embryos. For human consumers of herring gull eggs, these eggs contain TEQ levels up to four times higher than the maximum tolerable weekly intake.

  4. Measured and predicted affinities of binding and relative potencies to activate the AhR of PAHs and their alkylated analogues.

    PubMed

    Lee, Sangwoo; Shin, Woong-Hee; Hong, Seongjin; Kang, Habyeong; Jung, Dawoon; Yim, Un Hyuk; Shim, Won Joon; Khim, Jong Seong; Seok, Chaok; Giesy, John P; Choi, Kyungho

    2015-11-01

    Polycyclic aromatic hydrocarbons (PAHs) and their alkylated forms are important components of crude oil. Both groups of PAHs have been reported to cause dioxin-like responses, mediated by aryl hydrocarbon receptor (AhR). Thus, characterization of binding affinity to the AhR of unsubstituted or alkylated PAHs is important to understand the toxicological consequences of oil contamination on ecosystems. We investigated the potencies of major PAHs of crude oil, e.g., chrysene, phenanthrene and dibenzothiophene, and their alkylated forms (n=17) to upregulate expression of AhR-mediated processes by use of the H4IIE-luc transactivation bioassay. In addition, molecular descriptors of different AhR activation potencies among PAHs were investigated by use of computational molecular docking models. Based on responses of the H4IIE-luc in vitro assay, it was shown that potencies of PAHs were determined by alkylation in addition to the number and conformation of rings. Potencies of AhR-mediated processes were generally greater when a chrysene group was substituted, especially in 1-methyl-chrysene. Significant negative correlations were observed between the in vitro dioxin-like potency measured in H4IIE-luc cells and the binding distance estimated from the in silico modeling. The difference in relative potency for AhR activation observed among PAHs and their alkylated forms could be explained by differences among binding distances in the ligand binding domain of the AhR caused by alkylation. The docking model developed in the present study may have utility in predicting risks of environmental contaminants of which toxicities are mediated by AhR binding.

  5. Baicalein induces G1 arrest in oral cancer cells by enhancing the degradation of cyclin D1 and activating AhR to decrease Rb phosphorylation

    SciTech Connect

    Cheng, Ya-Hsin; Li, Lih-Ann; Lin, Pinpin; Cheng, Li-Chuan; Hung, Chein-Hui; Chang, Nai Wen; Lin, Chingju

    2012-09-15

    Baicalein is a flavonoid, known to have anti-inflammatory and anti-cancer effects. As an aryl hydrocarbon receptor (AhR) ligand, baicalein at high concentrations blocks AhR-mediated dioxin toxicity. Because AhR had been reported to play a role in regulating the cell cycle, we suspected that the anti-cancer effect of baicalein is associated with AhR. This study investigated the molecular mechanism involved in the anti-cancer effect of baicalein in oral cancer cells HSC-3, including whether such effect would be AhR-mediated. Results revealed that baicalein inhibited cell proliferation and increased AhR activity in a dose-dependent manner. Cell cycle was arrested at the G1 phase and the expression of CDK4, cyclin D1, and phosphorylated retinoblastoma (pRb) was decreased. When the AhR was suppressed by siRNA, the reduction of pRb was partially reversed, accompanied by a decrease of cell population at G1 phase and an increase at S phase, while the reduction of cyclin D1 and CDK4 did not change. This finding suggests that the baicalein activation of AhR is indeed associated with the reduction of pRb, but is independent of the reduction of cyclin D1 and CDK4. When cells were pre-treated with LiCl, the inhibitor of GSK-3β, the decrease of cyclin D1 was blocked and the reduction of pRb was recovered. The data indicates that in HSC-3 the reduction of pRb is both mediated by baicalein through activation of AhR and facilitation of cyclin D1 degradation, which causes cell cycle arrest at the G1 phase, and results in the inhibition of cell proliferation. -- Highlights: ► Baicalein causes the G1 phase arrest by decreasing Rb phosphorylation. ► Baicalein modulates AhR-mediated cell proliferation. ► Both AhR activation and cyclin D1 degradation results in hypophosphorylation of Rb. ► Baicalein facilitates cyclin D1 degradation by signalling the GSK-3β pathway.

  6. Combined chemical and toxicological long-term monitoring for AhR agonists with SPMD-based virtual organisms in drinking water Danjiangkou Reservoir, China.

    PubMed

    Wang, Jingxian; Song, Guoqiang; Li, Aimin; Henkelmann, Bernhard; Pfister, Gerd; Tong, Anthony Z; Schramm, Karl-Werner

    2014-08-01

    SPMD-based virtual organisms (VOs) were employed for time-integrating, long-term sampling combined biological and chemical analyses for exposure assessment of hydrophobic organic pollutants (HOPs) in a drinking water reservoir, China. The SPMDs were deployed at four and five sites in the Danjiangkou (DJK) reservoir over two periods of 26 and 31 d to sequester the hydrophobic contaminants in water. The chosen bioassay response for the extracts of the SPMDs, the induction of 7-ethoxyresorufin-o-deethylase (EROD) was assayed using a rat hepatoma cell line (H4IIE). The known aryl hydrocarbon receptor (AhR) agonists PAHs and PCBs were analyzed by HRGC/HRMS instrument. The cause-effect relationship between the observed AhR activities and chemical concentrations of detected AhR agonists was examined. The results show that the extracts from the SPMD samples could induce AhR activity significantly, whereas the chemically derived 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) equivalent (TEQcal) was not correlated with the bioassay-derived TCDD equivalent (TEQbio). The known AhR agonists could only account for 2-10% of the observed AhR responses among which the contribution of PCBs could almost be neglected. Unidentified AhR-active compounds represented a greater proportion of the TCDD equivalent (TCDD-EQ) in SPMD samples from DJK. Based on the first assessment, the VO followed by the combination of chemical and biological analyses emerges as a resource efficient water monitoring device in ecotoxicological assessment for toxicologically relevant compounds which are readily available for uptake by resident aquatic biota in drinking water resources. PMID:24548649

  7. Combined chemical and toxicological long-term monitoring for AhR agonists with SPMD-based virtual organisms in drinking water Danjiangkou Reservoir, China.

    PubMed

    Wang, Jingxian; Song, Guoqiang; Li, Aimin; Henkelmann, Bernhard; Pfister, Gerd; Tong, Anthony Z; Schramm, Karl-Werner

    2014-08-01

    SPMD-based virtual organisms (VOs) were employed for time-integrating, long-term sampling combined biological and chemical analyses for exposure assessment of hydrophobic organic pollutants (HOPs) in a drinking water reservoir, China. The SPMDs were deployed at four and five sites in the Danjiangkou (DJK) reservoir over two periods of 26 and 31 d to sequester the hydrophobic contaminants in water. The chosen bioassay response for the extracts of the SPMDs, the induction of 7-ethoxyresorufin-o-deethylase (EROD) was assayed using a rat hepatoma cell line (H4IIE). The known aryl hydrocarbon receptor (AhR) agonists PAHs and PCBs were analyzed by HRGC/HRMS instrument. The cause-effect relationship between the observed AhR activities and chemical concentrations of detected AhR agonists was examined. The results show that the extracts from the SPMD samples could induce AhR activity significantly, whereas the chemically derived 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) equivalent (TEQcal) was not correlated with the bioassay-derived TCDD equivalent (TEQbio). The known AhR agonists could only account for 2-10% of the observed AhR responses among which the contribution of PCBs could almost be neglected. Unidentified AhR-active compounds represented a greater proportion of the TCDD equivalent (TCDD-EQ) in SPMD samples from DJK. Based on the first assessment, the VO followed by the combination of chemical and biological analyses emerges as a resource efficient water monitoring device in ecotoxicological assessment for toxicologically relevant compounds which are readily available for uptake by resident aquatic biota in drinking water resources.

  8. Effects of 4-nitrophenol on expression of the ER-α and AhR signaling pathway-associated genes in the small intestine of rats.

    PubMed

    Tang, Juan; Song, Meiyan; Watanabe, Gen; Nagaoka, Kentaro; Rui, Xiaoli; Li, ChunMei

    2016-09-01

    4-Nitrophenol (PNP) is a persistent organic pollutant that was proven to be an environmental endocrine disruptor. The aim of this study was to evaluate the role of the estrogen receptor-α (ER-α) and aryl hydrocarbon receptor (AhR) signaling pathway in regulating the damage response to PNP in the small intestine of rats. Wistar-Imamichi male rats (21 d) were randomly divided into two groups: the control group and PNP group. Each group had three processes that were gavaged with PNP or vehicle daily: single dose (1 d), repeated dose (3 consecutive days) (3 d), and repeated dose with recovery (3 consecutive days and 3 recovery days) (6 d). The weight of the body, the related viscera, and small intestine were examined. Histological parameters of the small intestine and the quantity of mucus proteins secreted by small goblet cells were determined using HE staining and PAS staining. The mRNA expression of AhR, ER-α, CYP1A1, and GST was measured by real-time qPCR. In addition, we also analyzed the AhR, ER-α, and CYP1A1 expression in the small intestine by immunohistochemical staining. The small intestines histologically changed in the PNP-treated rat and the expression of AhR, CYP1A1, and GST was increased. While ER-α was significantly decreased in the small intestine, simultaneously, when rats were exposed to a longer PNP treatment, the damages disappeared. Our results demonstrate that PNP has an effect on the expression of AhR signaling pathway genes, AhR, CYP1A1, and GST, and ER-α in the rat small intestine.

  9. Disruption of period gene expression alters the inductive effects of dioxin on the AhR signaling pathway in the mouse liver

    SciTech Connect

    Qu Xiaoyu; Metz, Richard P.; Porter, Weston W.; Cassone, Vincent M.; Earnest, David J.

    2009-02-01

    The aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (ARNT) are transcription factors that express Per-Arnt-Sim (PAS) DNA-binding motifs and mediate the metabolism of drugs and environmental toxins in the liver. Because these transcription factors interact with other PAS genes in molecular feedback loops forming the mammalian circadian clockworks, we determined whether targeted disruption or siRNA inhibition of Per1 and Per2 expression alters toxin-mediated regulation of the AhR signaling pathway in the mouse liver and Hepa1c1c7 hepatoma cells in vitro. Treatment with the prototypical Ahr ligand, 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), had inductive effects on the primary targets of AhR signaling, Cyp1A1 and Cyp1B1, in the liver of all animals, but genotype-based differences were evident such that the toxin-mediated induction of Cyp1A1 expression was significantly greater (2-fold) in mice with targeted disruption of Per1 (Per1{sup ldc} and Per1{sup ldc}/Per2{sup ldc}). In vitro experiments yielded similar results demonstrating that siRNA inhibition of Per1 significantly increases the TCDD-induced expression of Cyp1A1 and Cyp1B1 in Hepa1c1c7 cells. Per2 inhibition in siRNA-infected Hepa1c1c7 cells had the opposite effect and significantly decreased both the induction of these p450 genes as well as AhR and Arnt expression in response to TCDD treatment. These findings suggest that Per1 may play a distinctive role in modulating AhR-regulated responses to TCDD in the liver.

  10. The aryl hydrocarbon receptor (AHR) transcription factor regulates megakaryocytic polyploidization

    PubMed Central

    Lindsey, Stephan; T. Papoutsakis, Eleftherios

    2012-01-01

    Summary We propose that the aryl hydrocarbon receptor (AHR) is a novel transcriptional regulator of megakaryopoietic polyploidization. Functional evidence was obtained that AHR impacts in vivo megakaryocytic differentiation and maturation; compared to wild-type mice, AHR-null mice had lower platelet counts, fewer numbers of newly synthesized platelets, increased bleeding times and lower-ploidy megakaryocytes (Mks). AHR mRNA increased 3·6-fold during ex vivo megakaryocytic differentiation, but reduced or remained constant during parallel isogenic granulocytic or erythroid differentiation. We interrogated the role of AHR in megakaryopoiesis using a validated Mk model of megakaryopoiesis, the human megakaryoblastic leukaemia CHRF cell line. Upon CHRF Mk differentiation, AHR mRNA and protein levels increased, AHR protein shifted from the cytoplasm to the nucleus and AHR binding to its consensus DNA binding sequence increased. Protein and mRNA levels of the AHR transcriptional target HES1 also increased. Mk differentiation of CHRF cells where AHR or HES1 was knocked-down using RNAi resulted in lower ploidy distributions and cells that were incapable of reaching ploidy classes ≥16n. AHR knockdown also resulted in increased DNA synthesis of lower ploidy cells, without impacting apoptosis. Together, these data support a role for AHR in Mk polyploidization and in vivo platelet function, and warrant further detailed investigations. PMID:21226706

  11. Effects of artificial sweeteners on the AhR- and GR-dependent CYP1A1 expression in primary human hepatocytes and human cancer cells.

    PubMed

    Kamenickova, Alzbeta; Pecova, Michaela; Bachleda, Petr; Dvorak, Zdenek

    2013-12-01

    Food constituents may cause a phenomenon of food-drug interactions. In the current study, we examined the effects of artificial sweeteners (aspartame, acesulfame, cyclamate, saccharin) on the aryl hydrocarbon receptor (AhR) and glucocorticoid receptor (GR)-dependent expression of CYP1A1 in human hepatocytes, hepatic HepG2 and intestinal LS174T cancer cell lines. Sweeteners were tested in concentrations up to those occurring in non-alcoholic beverages. Basal and ligand-inducible AhR- and GR-dependent reporter gene activation in stably transfected HepG2 and HeLa cells, respectively, were not affected by either of the sweeteners tested after 24h of incubation. The expression of CYP1A1 mRNA and protein in primary cultures of human hepatocytes and in LS174T and HepG2 cells was not induced by any of the tested sweeteners. Overall, aspartame, acesulfame, saccharin and cyclamate had no effects on CYP1A1 expression and transcriptional activities of AhR and GR. These data imply the safety of artificial sweeteners in terms of interference with AhR, GR and CYP1A1.

  12. Preferential induction of the AhR gene battery in HepaRG cells after a single or repeated exposure to heterocyclic aromatic amines

    SciTech Connect

    Dumont, Julie Josse, Rozenn Lambert, Carine Antherieu, Sebastien Laurent, Veronique Loyer, Pascal Robin, Marie-Anne Guillouzo, Andre

    2010-11-15

    2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) are two of the most common heterocyclic aromatic amines (HAA) produced during cooking of meat, fish and poultry. Both HAA produce different tumor profiles in rodents and are suspected to be carcinogenic in humans. In order to better understand the molecular basis of HAA toxicity, we have analyzed gene expression profiles in the metabolically competent human HepaRG cells using pangenomic oligonucleotide microarrays, after either a single (24-h) or a repeated (28-day) exposure to 10 {mu}M PhIP or MeIQx. The most responsive genes to both HAA were downstream targets of the arylhydrocarbon receptor (AhR): CYP1A1 and CYP1A2 after both time points and CYP1B1 and ALDH3A1 after 28 days. Accordingly, CYP1A1/1A2 induction in HAA-treated HepaRG cells was prevented by chemical inhibition or small interference RNA-mediated down-regulation of the AhR. Consistently, HAA induced activity of the CYP1A1 promoter, which contains a consensus AhR-related xenobiotic-responsive element (XRE). In addition, several other genes exhibited both time-dependent and compound-specific expression changes with, however, a smaller magnitude than previously reported for the prototypical AhR target genes. These changes concerned genes mainly related to cell growth and proliferation, apoptosis, and cancer. In conclusion, these results identify the AhR gene battery as the preferential target of PhIP and MeIQx in HepaRG cells and further support the hypothesis that intake of HAA in diet might increase human cancer risk.

  13. TCDD and a putative endogenous AhR ligand, ITE, elicit the same immediate changes in gene expression in mouse lung fibroblasts.

    PubMed

    Henry, Ellen C; Welle, Stephen L; Gasiewicz, Thomas A

    2010-03-01

    The aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor, mediates toxicity of several classes of xenobiotics and also has important physiological roles in differentiation, reproduction, and immunity, although the endogenous ligand(s) mediating these functions is/are as yet unidentified. One candidate endogenous ligand, 2-(1'H-indolo-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE), is a potent AhR agonist in vitro, activates the murine AhR in vivo, but does not induce toxicity. We hypothesized that ITE and the toxic ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), may modify transcription of different sets of genes to account for their different toxicity. To test this hypothesis, primary mouse lung fibroblasts were exposed to 0.5muM ITE, 0.2nM TCDD, or vehicle for 4 h, and total gene expression was evaluated using microarrays. After this short-term and low-dose treatment, several hundred genes were changed significantly, and the response to ITE and TCDD was remarkably similar, both qualitatively and quantitatively. Induced gene sets included the expected battery of AhR-dependent xenobiotic-metabolizing enzymes, as well as several sets that reflect the inflammatory role of lung fibroblasts. Real time quantitative RT-qPCR assay of several selected genes confirmed these microarray data and further suggested that there may be kinetic differences in expression between ligands. These data suggest that ITE and TCDD elicit an analogous change in AhR conformation such that the initial transcription response is the same. Furthermore, if the difference in toxicity between TCDD and ITE is mediated by differences in gene expression, then it is likely that secondary changes enabled by the persistent TCDD, but not by the shorter lived ITE, are responsible.

  14. INSIGHTS FROM AHR AND ARNT GENE KNOCKOUT STUDIES REGARDING RESPONSES TO TCDD AND REGULATION OF NORMAL EMBRYONIC DEVELOPMENT

    EPA Science Inventory

    The aryl hydrocarbon receptor (AhR) and the AhR nuclear translocator (ARNT) are members of the Per-ARNT-Sim (PAS) family of proteins. The AhR binds members of the chemical family that includes dioxins, furans and coplanar polychlorinated biphenyls (PCBs). A ligand-AhR-ARNT comp...

  15. Effects of Developmental Activation of the AhR on CD4+ T-Cell Responses to Influenza Virus Infection in Adult Mice

    PubMed Central

    Boule, Lisbeth A.; Winans, Bethany

    2014-01-01

    Background: Epidemiological and animal studies indicate that maternal exposure to pollutants that bind the aryl hydrocarbon receptor (AhR) correlates with poorer ability to combat respiratory infection and lower antibody levels in the offspring. These observations point to an impact on CD4+ T cells. Yet, the consequence of developmental exposure to AhR ligands on the activation and differentiation of CD4+ T cells has not been directly examined. Objectives: Our goal was to determine whether maternal exposure to an AhR ligand directly alters CD4+ T cell differentiation and function later in life. Methods: C57BL/6 mice were exposed to a prototypical AhR ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), in utero and via suckling. We then measured CD4+ T-cell activation and differentiation into distinct effector populations in adult offspring that were infected with influenza A virus (IAV). Reciprocal adoptive transfers were used to define whether modifications in CD4+ T-cell responses resulted from direct effects of developmental TCDD exposure on CD4+ T cells. Results: Developmental exposure skewed CD4+ T-cell responses to IAV infection. We observed fewer virus-specific, activated CD4+ T cells and a reduced frequency of conventional CD4+ effector-cell subsets. However, there was an increase in regulatory CD4+ T cells. Direct effects of AhR activation on CD4+ T cells resulted in impaired differentiation into conventional effector subsets; this defect was transferred to mice that had not been developmentally exposed to TCDD. Conclusions: Maternal exposure to TCDD resulted in durable changes in the responsive capacity and differentiation of CD4+ T cells in adult C57BL/6 mice. Citation: Boule LA, Winans B, Lawrence BP. 2014. Effects of developmental activation of the AhR on CD4+ T-cell responses to influenza virus infection in adult mice. Environ Health Perspect 122:1201–1208; http://dx.doi.org/10.1289/ehp.1408110 PMID:25051576

  16. Agonistic effect of selected isoflavones on arylhydrocarbon receptor in a novel AZ-AhR transgenic gene reporter human cell line.

    PubMed

    Bialesova, Lucia; Novotna, Aneta; Macejova, Dana; Brtko, Julius; Dvorak, Zdenek

    2015-07-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that controls the expression of a diverse set of genes. Structurally diverse compounds bind to AhR and act as AhR agonists. Well characterised family of natural AhR ligands are isoflavones, which are compounds found predominantly in soy beans or red clover. In this study we have examined agonistic effect of selected isoflavones (genistein, daidzein, biochanin A, formononetin and equol) on AhR in the novel transgenic gene reporter human cell line AZ-AhR, a stably transfected AhR-responsive cell line allowing rapid and sensitive assessment of AhR transcriptional activity. We demonstrated that biochanin A, formononetin and genistein at concentration 10(-4) mol/l exerted agonistic effects on AhR with fold activation of 309- fold, 108-fold and 27-fold, which is about 84.8%, 29.6% and 7.4%, respectively, of the value attained by 2,3,7,8-tetrachlorodibenzo-p-dioxin. Daidzein and equol did not show any significant effects on AhR. PMID:25926549

  17. The Ah receptor regulates growth factor expression in head and neck squamous cell carcinoma cell lines.

    PubMed

    John, Kaarthik; Lahoti, Tejas S; Wagner, Kelly; Hughes, Jarod M; Perdew, Gary H

    2014-10-01

    Previous studies in head and neck squamous cell carcinoma (HNSCC) cell lines have revealed that the Ah receptor (AHR) plays a significant role in mediating the "aggressive" phenotype of these cells, which includes enhanced inflammatory signaling (e.g., IL6) and migratory potential. Here we sought to identify putative novel targets of the AHR associated with enhanced tumor invasiveness. Global gene expression analysis identified a number of genes that are repressed upon treatment of OSC-19 or HN30 cells with an AHR antagonist. Three growth factors were targets of AHR activity; amphiregulin (AREG), epiregulin (EREG), and platelet-derived growth factor A (PDGFA) were repressed by an AHR antagonist and further examined. Quantitative PCR analysis, ELISA, and siRNA-mediated knock down of AHR revealed an attenuation of basal and/or induced levels of expression of these growth factors in two HNSCC lines, following AHR antagonism. In silico analysis revealed that these growth factors possess dioxin-like response elements. Two other AHR ligands, 6-formylindolo[3,2-b]carbazole and benzo(a)pyrene (BP) also elicited similar responses. In conclusion, this study identified AREG, EREG, and PDGFA as growth factor targets of AHR activity associated with metastatic phenotype of HNSCC cells, suggesting that attenuation of AHR activity may be a therapeutic strategy.

  18. Transgenic Overexpression of Aryl Hydrocarbon Receptor Repressor (AhRR) and AhR-Mediated Induction of CYP1A1, Cytokines, and Acute Toxicity

    PubMed Central

    Vogel, Christoph F.A.; Chang, W.L. William; Kado, Sarah; McCulloh, Kelly; Vogel, Helena; Wu, Dalei; Haarmann-Stemmann, Thomas; Yang, GuoXiang; Leung, Patrick S.C.; Matsumura, Fumio; Gershwin, M. Eric

    2016-01-01

    Background: The aryl hydrocarbon receptor repressor (AhRR) is known to repress aryl hydrocarbon receptor (AhR) signaling, but very little is known regarding the role of the AhRR in vivo. Objective: This study tested the role of AhRR in vivo in AhRR overexpressing mice on molecular and toxic end points mediated through a prototypical AhR ligand. Methods: We generated AhRR-transgenic mice (AhRR Tg) based on the genetic background of C57BL/6J wild type (wt) mice. We tested the effect of the prototypical AhR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on the expression of cytochrome P450 (CYP)1A1 and cytokines in various tissues of mice. We next analyzed the infiltration of immune cells in adipose tissue of mice after treatment with TCDD using flow cytometry. Results: AhRR Tg mice express significantly higher levels of AhRR compared to wt mice. Activation of AhR by TCDD caused a significant increase of the inflammatory cytokines Interleukin (IL)-1β, IL-6 and IL-10, and CXCL chemokines in white epididymal adipose tissue from both wt and AhRR Tg mice. However, the expression of IL-1β, CXCL2 and CXCL3 were significantly lower in AhRR Tg versus wt mice following TCDD treatment. Exposure to TCDD caused a rapid accumulation of neutrophils and macrophages in white adipose tissue of wt and AhRR Tg mice. Furthermore we found that male AhRR Tg mice were protected from high-dose TCDD-induced lethality associated with a reduced inflammatory response and liver damage as indicated by lower levels of TCDD-induced alanine aminotransferase and hepatic triglycerides. Females from both wt and AhRR Tg mice were less sensitive than male mice to acute toxicity induced by TCDD. Conclusion: In conclusion, the current study identifies AhRR as a previously uncharacterized regulator of specific inflammatory cytokines, which may protect from acute toxicity induced by TCDD. Citation: Vogel CF, Chang WL, Kado S, McCulloh K, Vogel H, Wu D, Haarmann-Stemmann T, Yang GX, Leung PS, Matsumura F

  19. Promoter analysis of TCDD-inducible genes in a thymic epithelial cell line indicates the potential for cell-specific transcription factor crosstalk in the AhR response

    SciTech Connect

    Frericks, Markus; Burgoon, Lyle D.; Zacharewski, Timothy R.; Esser, Charlotte

    2008-10-15

    Activation of the aryl hydrocarbon receptor (AhR{sup 1}) by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) elicits severe immunosuppression accompanied by thymic atrophy. Previous evidence suggests that TCDD targets both thymocytes and thymic epithelial cells. The AhR induces cell-specific changes in gene transcription via binding to the dioxin response element DRE; however, the underlying specificity-mechanisms, in particular with regard to the role of promoter element context, and possible transcription factor crosstalk remain poorly understood. Global gene expression in the cortical thymic epithelial cell line ET at 2, 4, and 6 h following 5 nM TCDD exposure resulted in differential regulation of 201 genes. JASPAR and TRANSFAC mapped the statistically over-represented promoter elements in the regulated genes to specific transcription factor binding sites, suggesting a regulatory role in AhR signaling. Over-represented elements included the xenobiotic response element XRE, NF{kappa}B-Rel, HRE, PPAR{gamma}, GR, PAX-4 and estrogen receptor binding sites. Co-treatment experiments with TCDD and CoCl{sub 2}, to induce hypoxia, or TCDD and 17-{beta}-estradiol (E2) indicated crosstalk between AhR and Hif or ER, in agreement with other experimental models. The computational identification of TFBS and the demonstration of interaction confirm their interactions with AhR signaling and suggest that the other over-represented elements may also be important in the immunosuppressive effects elicited by TCDD. In conclusion, we demonstrated the importance of promoter element cooperation in the shaping of a cell-specific AhR response. Our findings regarding the transcriptional changes in cortical epithelial cells are congruent with the well-known thymotoxic TCDD-phenotype, and useful in new hypothesis generation of the role of cortical TECs in TCDD toxicity.

  20. TCDD promoted EMT of hFPECs via AhR, which involved the activation of EGFR/ERK signaling.

    PubMed

    Gao, Zhan; Bu, Yongjun; Liu, Xiaozhuan; Wang, Xugang; Zhang, Guofu; Wang, Erhui; Ding, Shibin; Liu, Yongfeng; Shi, Ruling; Li, Qiaoyun; Fu, Jianhong; Yu, Zengli

    2016-05-01

    One critical step of second palatal fusion is the newly formed medial epithelia seam (MES) disintegration, which involves apoptosis, epithelial to mesenchymal transition (EMT), and cell migration. Although the environmental toxicant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) produces cleft palate at high rates, little is known about the effects of TCDD exposure on the fate of palatal epithelial cells. By using primary epithelial cells isolated from human fetal palatal shelves (hFPECs), we show that TCDD increased cell proliferation and EMT, as demonstrated by increased the epithelial markers (E-cadherin and cytokeratin14) and enhanced the mesenchymal markers (vimentin and fibronectin), but had no effect on cell migration and apoptosis. TCDD exposure led to a dose-dependent increase in Slug protein expression. Coimmunoprecipitation revealed that TCDD promoted AhR to form a protein complex with Slug. ChIP assay confirmed that TCDD exposure recruited AhR to the xenobiotic responsive element of Slug promoter. Knockdown of AhR by siRNA remarkably weakened TCDD-induced binding of AhR to the XRE promoter of slug, thereby suppressed TCDD-induced vimentin. Further experiment showed that TCDD stimulated EGFR phosphorylation did not influence the TGFβ3/Smad signaling; whereas TCDD increased phosphorylation of ERK1/2 and p38 with no effect on activation of JNK. By using varieties of inhibitors, we confirmed that TCDD promoted proliferation and EMT of hFPECs via activation of EGFR/ERK pathway. These data make a novel contribution to the molecular mechanism of cleft palate by TCDD. PMID:26971374

  1. Assessment of energetic costs of AhR activation by β-naphthoflavone in rainbow trout (Oncorhynchus mykiss) hepatocytes using metabolic flux analysis

    SciTech Connect

    Nault, Rance; Abdul-Fattah, Hiba; Mironov, Gleb G.; Berezovski, Maxim V.; Moon, Thomas W.

    2013-08-15

    Exposure to environmental contaminants such as activators of the aryl hydrocarbon receptor (AhR) leads to the induction of defense and detoxification mechanisms. While these mechanisms allow organisms to metabolize and excrete at least some of these environmental contaminants, it has been proposed that these mechanisms lead to significant energetic challenges. This study tests the hypothesis that activation of the AhR by the model agonist β-naphthoflavone (βNF) results in increased energetic costs in rainbow trout (Oncorhynchus mykiss) hepatocytes. To address this hypothesis, we employed traditional biochemical approaches to examine energy allocation and metabolism including the adenylate energy charge (AEC), protein synthesis rates, Na{sup +}/K{sup +}-ATPase activity, and enzyme activities. Moreover, we have used for the first time in a fish cell preparation, metabolic flux analysis (MFA) an in silico approach for the estimation of intracellular metabolic fluxes. Exposure of trout hepatocytes to 1 μM βNF for 48 h did not alter hepatocyte AEC, protein synthesis, or Na{sup +}/K{sup +}-ATPase activity but did lead to sparing of glycogen reserves and changes in activities of alanine aminotransferase and citrate synthase suggesting altered metabolism. Conversely, MFA did not identify altered metabolic fluxes, although we do show that the dynamic metabolism of isolated trout hepatocytes poses a significant challenge for this type of approach which should be considered in future studies. - Highlights: • Energetic costs of AhR activation by βNF was examined in rainbow trout hepatocytes. • Metabolic flux analysis was performed on a fish cell preparation for the first time. • Exposure to βNF led to sparing of glycogen reserves and altered enzyme activities. • Adenylate energy charge was maintained despite temporal changes in metabolism.

  2. TOXICITY OF AHR AGONISTS TO FISH EARLY LIFE STAGES

    EPA Science Inventory

    Fish early life stages are exceptionally sensitive to the lethal toxicity of chemicals that act as arylhydrocarbon receptor (AhR) agonists. Toxicity characterizations based on 2,3,7,8-tetrachlorodibenzo-p-dioxin, generally the most potent AhR agonist, support the toxicity equiva...

  3. Complementary PLS and KNN algorithms for improved 3D-QSDAR consensus modeling of AhR binding

    PubMed Central

    2013-01-01

    Multiple validation techniques (Y-scrambling, complete training/test set randomization, determination of the dependence of R2test on the number of randomization cycles, etc.) aimed to improve the reliability of the modeling process were utilized and their effect on the statistical parameters of the models was evaluated. A consensus partial least squares (PLS)-similarity based k-nearest neighbors (KNN) model utilizing 3D-SDAR (three dimensional spectral data-activity relationship) fingerprint descriptors for prediction of the log(1/EC50) values of a dataset of 94 aryl hydrocarbon receptor binders was developed. This consensus model was constructed from a PLS model utilizing 10 ppm x 10 ppm x 0.5 Å bins and 7 latent variables (R2test of 0.617), and a KNN model using 2 ppm x 2 ppm x 0.5 Å bins and 6 neighbors (R2test of 0.622). Compared to individual models, improvement in predictive performance of approximately 10.5% (R2test of 0.685) was observed. Further experiments indicated that this improvement is likely an outcome of the complementarity of the information contained in 3D-SDAR matrices of different granularity. For similarly sized data sets of Aryl hydrocarbon (AhR) binders the consensus KNN and PLS models compare favorably to earlier reports. The ability of 3D-QSDAR (three dimensional quantitative spectral data-activity relationship) to provide structural interpretation was illustrated by a projection of the most frequently occurring bins on the standard coordinate space, thus allowing identification of structural features related to toxicity. PMID:24257141

  4. Methylated pentavalent arsenic metabolites are bifunctional inducers, as they induce cytochrome P450 1A1 and NAD(P)H:quinone oxidoreductase through AhR- and Nrf2-dependent mechanisms.

    PubMed

    Anwar-Mohamed, Anwar; Elshenawy, Osama H; Soshilov, Anatoly A; Denison, Michael S; Chris Le, X; Klotz, Lars-Oliver; El-Kadi, Ayman O S

    2014-02-01

    Activation of the aryl hydrocarbon receptor (AhR) ultimately leads to the induction of the carcinogen-activating enzyme cytochrome P450 1A1 (CYP1A1), and activation of the nuclear factor-erythroid 2 p45-related factor 2 (Nrf2) in addition to the AhR pathway induces the expression of the NADP(H):quinone oxidoreductase (NQO1). Therefore, the aim of this study was to examine the effect of As(III) pentavalent metabolites, MMA(V), DMA(V), and TMA(V), on AhR and Nrf2 activation and on the expression of their prototypical downstream targets CYP1A1 and NQO1, respectively. Our results showed that treatment of HepG2 cells with MMA(V), DMA(V), or TMA(V) in the absence and presence of 2,3,7,8-tetrachlorodibenzo-p-dioxin or sulforaphane significantly induced both CYP1A1 and NQO1 at the mRNA, protein, and catalytic activity levels. Furthermore, these metabolites increased the AhR-dependent XRE-driven and the Nrf2-dependent ARE-driven luciferase reporter activities, which coincided with increased nuclear accumulation of both transcription factors. However, none of these metabolites were shown to be AhR ligands. The induction of CYP1A1 by these metabolites seems to be ligand-independent, possibly through a decrease in HSP90 protein expression levels. The metabolites also increased ROS production, which was significantly higher than that produced by As(III). Upon knockdown of AhR and Nrf2 the MMA(V)-, DMA(V)-, and TMA(V)-mediated induction of both CYP1A1 and NQO1 proteins was significantly decreased. In conclusion, this study demonstrates for the first time that methylated pentavalent arsenic metabolites are bifunctional inducers, as they increase CYP1A1 by activating the AhR/XRE signaling pathway and they increase NQO1 by activating the Nrf2/ARE signaling pathway in addition to the AhR/XRE pathway.

  5. ITE and TCDD differentially regulate the vascular remodeling of rat placenta via the activation of AhR.

    PubMed

    Wu, Yanming; Chen, Xiao; Zhou, Qian; He, Qizhi; Kang, Jiuhong; Zheng, Jing; Wang, Kai; Duan, Tao

    2014-01-01

    Vascular remodeling in the placenta is essential for normal fetal development. The previous studies have demonstrated that in utero exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, an environmental toxicant) induces the intrauterine fetal death in many species via the activation of aryl hydrocarbon receptor (AhR). In the current study, we compared the effects of 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) and TCDD on the vascular remodeling of rat placentas. Pregnant rats on gestational day (GD) 15 were randomly assigned into 5 groups, and were exposed to a single dose of 1.6 and 8.0 mg/kg body weight (bw) ITE, 1.6 and 8.0 µg/kg bw TCDD, or an equivalent volume of the vehicle, respectively. The dams were sacrificed on GD20 and the placental tissues were gathered. The intrauterine fetal death was observed only in 8.0 µg/kg bw TCDD-exposed group and no significant difference was seen in either the placental weight or the fetal weight among all these groups. The immunohistochemical and histological analyses revealed that as compared with the vehicle-control, TCDD, but not ITE, suppressed the placental vascular remodeling, including reduced the ratio of the placental labyrinth zone to the basal zone thickness (at least 0.71 fold of control), inhibited the maternal sinusoids dilation and thickened the trophoblastic septa. However, no marked difference was observed in the density of fetal capillaries in the labyrinth zone among these groups, although significant differences were detected in the expression of angiogenic growth factors between ITE and TCDD-exposed groups, especially Angiopoietin-2 (Ang-2), Endoglin, Interferon-γ (IFN-γ) and placenta growth factor (PIGF). These results suggest ITE and TCDD differentially regulate the vascular remodeling of rat placentas, as well as the expression of angiogenic factors and their receptors, which in turn may alter the blood flow in the late gestation and partially resulted in

  6. Tumorigenic effects of endocrine-disrupting chemicals are alleviated by licorice (Glycyrrhiza glabra) root extract through suppression of AhR expression in mammalian cells.

    PubMed

    Chu, Xiao Ting; de la Cruz, Joseph; Hwang, Seong Gu; Hong, Heeok

    2014-01-01

    Endocrine-disrupting chemicals (EDCs) have been reported to interfere with estrogen signaling. Exposure to these chemicals decreases the immune response and causes a wide range of diseases in animals and humans. Recently, many studies showed that licorice (Glycyrrhiza glabra) root extract (LRE) commonly called "gamcho" in Korea exhibits antioxidative, chemoprotective, and detoxifying properties. This study aimed to investigate the mechanism of action of LRE and to determine if and how LRE can alleviate the toxicity of EDCs. LRE was prepared by vacuum evaporation and freeze-drying after homogenization of licorice root powder that was soaked in 80% ethanol for 72 h. We used 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) as a representative EDC, which is known to induce tumors or cancers; MCF-7 breast cancer cells, used as a tumor model, were treated with TCDD and various concentrations of LRE (0, 50, 100, 200, 400 μg/mL) for 24, 48, and 72 h. As a result, TCDD stimulated MCF-7 cell proliferation, but LRE significantly inhibited TCDD-induced MCF-7 cell proliferation in a dose- and time-dependent manner. The expression of TCDD toxicity-related genes, i.e., aryl hydrocarbon receptor (AhR), AhR nuclear translocator, and cytochrome P450 1A1, was also down-regulated by LRE in a dose-dependent manner. Analysis of cell cycle distribution after treatment of MCF-7 cells with TCDD showed that LRE inhibited the proliferation of MCF-7 cells via G2/M phase arrest. Reverse transcription-polymerase chain reaction and Western blot analysis also revealed that LRE dose-dependently increased the expression of the tumor suppressor genes p53 and p27 and down-regulated the expression of cell cycle-related genes. These data suggest that LRE can mitigate the tumorigenic effects of TCDD in breast cancer cells by suppression of AhR expression and cell cycle arrest. Thus, LRE can be used as a potential toxicity-alleviating agent against EDC-mediated diseases.

  7. Divergent Ah Receptor Ligand Selectivity during Hominin Evolution.

    PubMed

    Hubbard, Troy D; Murray, Iain A; Bisson, William H; Sullivan, Alexis P; Sebastian, Aswathy; Perry, George H; Jablonski, Nina G; Perdew, Gary H

    2016-10-01

    We have identified a fixed nonsynonymous sequence difference between humans (Val381; derived variant) and Neandertals (Ala381; ancestral variant) in the ligand-binding domain of the aryl hydrocarbon receptor (AHR) gene. In an exome sequence analysis of four Neandertal and Denisovan individuals compared with nine modern humans, there are only 90 total nucleotide sites genome-wide for which archaic hominins are fixed for the ancestral nonsynonymous variant and the modern humans are fixed for the derived variant. Of those sites, only 27, including Val381 in the AHR, also have no reported variability in the human dbSNP database, further suggesting that this highly conserved functional variant is a rare event. Functional analysis of the amino acid variant Ala381 within the AHR carried by Neandertals and nonhuman primates indicate enhanced polycyclic aromatic hydrocarbon (PAH) binding, DNA binding capacity, and AHR mediated transcriptional activity compared with the human AHR. Also relative to human AHR, the Neandertal AHR exhibited 150-1000 times greater sensitivity to induction of Cyp1a1 and Cyp1b1 expression by PAHs (e.g., benzo(a)pyrene). The resulting CYP1A1/CYP1B1 enzymes are responsible for PAH first pass metabolism, which can result in the generation of toxic intermediates and perhaps AHR-associated toxicities. In contrast, the human AHR retains the ancestral sensitivity observed in primates to nontoxic endogenous AHR ligands (e.g., indole, indoxyl sulfate). Our findings reveal that a functionally significant change in the AHR occurred uniquely in humans, relative to other primates, that would attenuate the response to many environmental pollutants, including chemicals present in smoke from fire use during cooking. PMID:27486223

  8. Divergent Ah Receptor Ligand Selectivity during Hominin Evolution.

    PubMed

    Hubbard, Troy D; Murray, Iain A; Bisson, William H; Sullivan, Alexis P; Sebastian, Aswathy; Perry, George H; Jablonski, Nina G; Perdew, Gary H

    2016-10-01

    We have identified a fixed nonsynonymous sequence difference between humans (Val381; derived variant) and Neandertals (Ala381; ancestral variant) in the ligand-binding domain of the aryl hydrocarbon receptor (AHR) gene. In an exome sequence analysis of four Neandertal and Denisovan individuals compared with nine modern humans, there are only 90 total nucleotide sites genome-wide for which archaic hominins are fixed for the ancestral nonsynonymous variant and the modern humans are fixed for the derived variant. Of those sites, only 27, including Val381 in the AHR, also have no reported variability in the human dbSNP database, further suggesting that this highly conserved functional variant is a rare event. Functional analysis of the amino acid variant Ala381 within the AHR carried by Neandertals and nonhuman primates indicate enhanced polycyclic aromatic hydrocarbon (PAH) binding, DNA binding capacity, and AHR mediated transcriptional activity compared with the human AHR. Also relative to human AHR, the Neandertal AHR exhibited 150-1000 times greater sensitivity to induction of Cyp1a1 and Cyp1b1 expression by PAHs (e.g., benzo(a)pyrene). The resulting CYP1A1/CYP1B1 enzymes are responsible for PAH first pass metabolism, which can result in the generation of toxic intermediates and perhaps AHR-associated toxicities. In contrast, the human AHR retains the ancestral sensitivity observed in primates to nontoxic endogenous AHR ligands (e.g., indole, indoxyl sulfate). Our findings reveal that a functionally significant change in the AHR occurred uniquely in humans, relative to other primates, that would attenuate the response to many environmental pollutants, including chemicals present in smoke from fire use during cooking.

  9. Exposure to Diesel Exhaust Particle Extracts (DEPe) Impairs Some Polarization Markers and Functions of Human Macrophages through Activation of AhR and Nrf2

    PubMed Central

    Jaguin, Marie; Fardel, Olivier; Lecureur, Valérie

    2015-01-01

    Macrophages (MΦ), well-known to play an important role in immune response, also respond to environmental toxic chemicals such as diesel exhaust particles (DEP). Potential effects of DEPs towards MΦ polarization, a key hall-mark of MΦ physiology, remain however poorly documented. This study was therefore designed to evaluate the effects of a reference DEP extract (DEPe) on human MΦ polarization. Human blood monocytes-derived MΦ were incubated with IFNγ+LPS or IL-4 to obtain M1 and M2 subtypes, respectively; a 24 h exposure of polarizing MΦ to 10 μg/ml DEPe was found to impair expression of some macrophagic M1 and M2 markers, without however overall inhibition of M1 and M2 polarization processes. Notably, DEPe treatment increased the secretion of the M1 marker IL-8 and the M2 marker IL-10 in both MΦ subtypes, whereas it reduced lipopolysaccharide-induced IL-6 and IL-12p40 secretion in M1 MΦ. In M2 MΦ, DEPe exposure led to a reduction of CD200R expression and of CCL17, CCL18 and CCL22 secretion, associated with a lower chemotaxis of CCR4-positive cells. DEPe activated the Nrf2 and AhR pathways and induced expression of their reference target genes such as Hmox-1 and cytochrome P-4501B1 in M1 and M2 MΦ. Nrf2 or AhR silencing through RNA interference prevented DEPe-related down-regulation of IL-6. AhR silencing also inhibited the down-secretion of IL-12p40 and CCL18 in M1- and M2-DEPe-exposed MΦ, respectively. DEPs are therefore likely to alter expression of some M1 and M2 markers in an AhR- and Nrf2-dependent manner; such regulations may contribute to deleterious immune effects of atmospheric DEP. PMID:25710172

  10. Ah receptor ligands in tobacco smoke.

    PubMed

    Löfroth, G; Rannug, A

    1988-08-01

    Tar particulates from cigarette smoke contain compounds with affinity for the Ah receptor. The sidestream activity is larger than that of the mainstream with a ratio of about 5. The compounds causing the affinity appear in the neutral fraction after chemical fractionation excluding basic and acidic components as major contributors to the affinity. The affinity cannot be explained by benzo[a]pyrene and other polycyclic aromatic hydrocarbons but it might be caused by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related compounds and by oxidized tryptophan derivatives.

  11. In vitro re-expression of the aryl hydrocarbon receptor (Ahr) in cultured Ahr-deficient mouse antral follicles partially restores the phenotype to that of cultured wild-type mouse follicles

    PubMed Central

    Ziv-Gal, A; Gao, L.; Karman, B.N.; Flaws, J.A.

    2014-01-01

    Background The aryl hydrocarbon receptor (AHR) mediates the toxic effects of various endocrine disrupting chemicals. In female mice, global deletion of the Ahr (AhrKO) results in slow growth of ovarian antral follicles. No studies, however, have examined whether injection of the Ahr restores the phenotypes of cultured AhrKO ovarian antral follicles to wild-type levels. Methods We developed a system to construct a recombinant adenovirus containing the Ahr to re-express the Ahr in AhrKO granulosa cells and whole antral follicles. We then compared follicle growth and levels of factors in the AHR signaling pathway (Ahr, Ahrr, Cyp1a1, and Cyp1b1) in wild-type, AhrKO, and Ahr re-expressed follicles. Further, we compared the response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in wild-type, AhrKO, and Ahr re-expressed follicles. Results AdAhr injection into AhrKO follicles partially restored their growth pattern to wild-type levels. Further, Ahr re-expressed follicles had significantly higher levels of Ahr, Ahrr, Cyp1a1, and Cyp1b1 compared to wild-type follicles. Upon TCDD treatment, only Cyp1a1 levels were significantly higher in Ahr re-expressed follicles compared to the levels in wild-type follicles. Conclusion Our system of re-expression of the Ahr partially restores follicle growth and transcript levels of factors in the AHR signaling pathway to wild-type levels. PMID:25500125

  12. Plant polyphenols differentially modulate inflammatory responses of human keratinocytes by interfering with activation of transcription factors NF{kappa}B and AhR and EGFR-ERK pathway

    SciTech Connect

    Potapovich, Alla I.; Lulli, Daniela; Fidanza, Paolo; Kostyuk, Vladimir A.; De Luca, Chiara; Pastore, Saveria; Korkina, Liudmila G.

    2011-09-01

    Molecular mechanisms underlying modulation of inflammatory responses in primary human keratinocytes by plant polyphenols (PPs), namely the glycosylated phenylpropanoid verbascoside, the stilbenoid resveratrol and its glycoside polydatin, and the flavonoid quercetin and its glycoside rutin were evaluated. As non-lethal stimuli, the prototypic ligand for epidermal growth factor receptor (EGFR) transforming growth factor alpha (TGFalpha), the combination of tumor necrosis factor (TNFalpha) and interferon (IFNgamma) (T/I), UVA + UVB irradiation, and bacterial lipopolysaccharide (LPS) were used. We demonstrated differential modulation of inflammatory responses in keratinocytes at signal transduction, gene transcription, and protein synthesis levels as a function of PP chemical structure, the pro-inflammatory trigger used, and PP interaction with intracellular detoxifying systems. The PPs remarkably inhibited constitutive, LPS- and T/I-induced but not TGFalpha-induced ERK phosphorylation. They also suppressed NFkappaB activation by LPS and T/I. Verbascoside and quercetin invariably impaired EGFR phosphorylation and UV-associated aryl hydrocarbon receptor (AhR)-mediated signaling, while rutin, polydatin and resveratrol did not affect EGFR phosphorylation and further activated AhR machinery in UV-exposed keratinocytes. In general, PPs down-regulated gene expression of pro-inflammatory cytokines/enzymes, except significant up-regulation of IL-8 observed under stimulation with TGFalpha. Both spontaneous and T/I-induced release of IL-8 and IP-10 was suppressed, although 50 {mu}M resveratrol and polydatin up-regulated IL-8. At this concentration, resveratrol activated both gene expression and de novo synthesis of IL-8 and AhR-mediated mechanisms were involved. We conclude that PPs differentially modulate the inflammatory response of human keratinocytes through distinct signal transduction pathways, including AhR and EGFR. - Graphical abstract: Display Omitted Highlights

  13. Autoradiographic localization of aromatic hydrocarbon receptor (AHR) in rhesus monkey ovary.

    PubMed

    Baldridge, Monika G; Hutz, Reinhold J

    2007-06-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most toxic congener of a large class of manmade pollutants that persist in the environment. TCDD exerts its toxic effects, in part, by binding to its receptor known as the aromatic hydrocarbon receptor (AHR). TCDD is estrogen modulatory and in some systems its receptor associates directly with estrogen receptors via co-activator molecules. TCDD inhibits steroid synthesis in human ovarian granulosa cells and AHR is found in these cells. We have previously shown that AHR is found in whole rhesus monkey ovary, but have yet to establish its location. In the present study, we set out to show that radiolabeled TCDD binds to monkey ovarian follicles and that this binding is receptor mediated. Ovaries from Macaca mulatta were sectioned on a cryostat at 10 micro m; and sections were incubated with either control vehicle, (3)H-TCDD, or (3)H-TCDD plus alpha-naphthoflavone (ANF), a known receptor-blocking agent. Here, we show for the first time specific binding of TCDD to the granulosa cells of antral follicles and other regions of the rhesus monkey ovary. Our data indicate a 60-fold increase in binding with (3)H-TCDD over that of control, and that this binding is reduced to the levels seen in controls with the addition of the competitive antagonist ANF. These findings support the hypothesis that TCDD directly affects primate ovarian function via the AHR.

  14. Identification of ah receptor agonists in soil of E-waste recycling sites from Taizhou area in China.

    PubMed

    Shen, Chaofeng; Huang, Shengbiao; Wang, Zijian; Qiao, Min; Tang, Xianjin; Yu, Chunna; Shi, Dezhi; Zhu, Youfeng; Shi, Jiyan; Chen, Xincai; Setty, Karen; Chen, Yingxu

    2008-01-01

    In recent years, increasing concern has surrounded the consequences of improper electric and electronic waste (e-waste) disposal. In order to mitigate or remediate the potentially severe toxic effects of e-waste recycling on the environment, organisms, and humans, many contaminated sites must first be well-characterized. In this study, soil samples were taken from Taizhou city, one of the largest e-waste disposal centers in China, which was involved in recycling for nearly 30 years. The extracts of the samples were assayed for aryl hydrocarbon receptor (AhR)-mediated ethoxyresorufin-O-deethylase (EROD) induction in the rat hepatoma cell line H4IIE. Some of the target AhR agonists, including polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), polychlorinated biphenyls (PCBs), and polycyclic aromatic hydrocarbons (PAHs), were instrumentally analyzed as well. The cause-effect relationship and dose-response relationship between the chemical concentrations of AhR agonists and observed EROD activity were examined. The results showed that soil extracts could induce AhR activity significantly, and the chemically derived 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) equivalents (TEQcal) were perfectly correlated to bioassay-derived TCDD equivalents (TEQbio; R = 0.96, P < 0.001), which indicated that the known AhR agonists could account for the observed responses. Among different contributors, PCBs accounted for 87.2-98.2% and PCDD/Fs contributed 1.7-11.6% of TEQcal, while the contribution of PAHs could almost be neglected. Under these conditions, a quantitative dose-effect relationship between TEQ(PCB) and EROD activity could be evaluated, suggesting that the observed AhR effect was mainly caused by PCBs. Further source identification by congener profiles analysis showed that the crude dismantling of electric power devices and open burning of electric wires and printed circuit boards may be the main sources of these dioxin-like compounds. This study suggests that

  15. A human intervention study with foods containing natural Ah-receptor agonists does not significantly show AhR-mediated effects as measured in blood cells and urine.

    PubMed

    de Waard, Pim W J; Peijnenburg, Ad A C M; Baykus, Hakan; Aarts, Jac M M J G; Hoogenboom, Ron L A P; van Schooten, Frederik J; de Kok, Theo M C M

    2008-10-22

    Binding and activation of the aryl hydrocarbon receptor (AhR) is thought to be an essential step in the toxicity of the environmental pollutants dioxins and dioxin-like PCBs. However, also a number of natural compounds, referred to as NAhRAs (natural Ah-receptor agonists), which are present in, for example, fruits and vegetables, can bind and activate this receptor. To study their potential effects in humans, we first investigated the effect of the prototypical AhR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on gene expression in ex vivo exposed freshly isolated human lymphocytes, and compared the resulting gene expression profile with those caused by the well-known NAhRA indolo[3,2-b]carbazole (ICZ), originating from cruciferous vegetables, and by a hexane extract of NAhRA-containing grapefruit juice (GJE). Only ICZ induced a gene expression profile similar to TCDD in the lymphocytes, and both significantly up-regulated CYP1B1 and TIPARP (TCDD-inducible poly (ADP-ribose) polymerase) mRNA. Next, we performed a human intervention study with NAhRA-containing cruciferous vegetables and grapefruit juice. The expression of the prototypical AhR-responsive genes CYP1A1, CYP1B1 and NQO1 in whole blood cells and in freshly isolated lymphocytes was not significantly affected. Also enzyme activities of CYP1A2, CYP2A6, N-acetyltransferase 2 (NAT2) and xanthine oxidase (XO), as judged by caffeine metabolites in urine, were unaffected, except for a small down-regulation of NAT2 activity by grapefruit juice. Examination of blood plasma with DR CALUX showed a 12% increased AhR agonist activity 3 and 24 h after consumption of cruciferous vegetables, but did not show a significant effect of grapefruit juice consumption. We conclude that intake of NAhRAs from food may result in minor AhR-related effects measurable in human blood and urine. PMID:18762178

  16. Endogenous and exogenous ligands of aryl hydrocarbon receptor: current state of art.

    PubMed

    Stejskalova, Lucie; Dvorak, Zdenek; Pavek, Petr

    2011-02-01

    Aryl hydrocarbon receptor (AhR) is an important transcriptional regulator of drug metabolizing enzymes that dominantly controls the expression of cytochrome P450 CYP1 family genes and some phase II enzymes. AhR also has many endogenous functions including cell cycle control, immune response, and cell differentiation. In addition, AhR is well-known to be involved in chemically-induced carcinogenesis. AhR is activated by a variety of endogenous and exogenous ligands. While exogenous activation of AhR has deleterious effects on human organism, sustained activation of AhR by endogenous ligands is indispensable for proper cell functions. Therefore, the effects of exogenous and endogenous ligands on AhR resemble the Dr. Jekyll and Mr. Hyde story. The aim of the current paper is to summarize and update the knowledge on exogenous and endogenous AhR ligands. PMID:21395538

  17. Ligand-dependent interactions of the Ah receptor with coactivators in a mammalian two-hybrid assay

    SciTech Connect

    Zhang Shu; Rowlands, Craig; Safe, Stephen

    2008-03-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a high affinity ligand for the aryl hydrocarbon receptor (AhR). In this study, we investigated structure-dependent differences in activation of the AhR by a series of halogenated aromatic hydrocarbons. TCDD, 1,2,3,7,8-pentachlorodibenzo-p-dioxin (PeCDD), 2,3,7,8-tetrachlorodibenzofuran (TCDF), 2,3,4,7,8-pentachlorodibenzofuran (PeCDF), and 3,3',4,4',5-pentachlorobiphenyl (PCB126) induced CYP1A1-dependent activities in HEK293 human embryonic kidney, Panc1 pancreatic cancer, and Hepa1c1c7 mouse hepatoma cell lines. There was a structure-dependent difference in the efficacy of TCDF and PCB126 in HEK293 and Panc1 cells since induced CYP1A1 mRNA levels were lower than observed for the other congeners. A mammalian two-hybrid assay in cells transfected with GAL4-coactivator and AhR-VP16 chimeras was used to investigate structure-dependent interactions of these chimeras in Panc1, HEK293, and Hepa1c1c7 cells. The reporter construct pGAL4-luc contains five tandem GAL4 response elements linked to the luciferase gene and the GAL4-coactivator chimeras express several coactivators including steroid receptor coactivator 1 (SRC-1), SRC-2 and SRC-3, the mediator coactivator TRAP220, coactivator associated arginine methyl transferase 1 (CARM-1), and peroxisome proliferator-activated receptor {gamma} coactivator 1 (PGC-1). Results of the mammalian two-hybrid studies clearly demonstrate that activation of pGAL4-luc in cells transfected with VP-AhR and GAL4-coactivator chimeras is dependent on the structure of the HAH congener, cell context, and coactivator, suggesting that the prototypical HAH congeners used in this study exhibit selective AhR modulator activity.

  18. Indole and Tryptophan Metabolism: Endogenous and Dietary Routes to Ah Receptor Activation

    PubMed Central

    Hubbard, Troy D.; Murray, Iain A.

    2015-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor recognized for its role in xenobiotic metabolism. The physiologic function of AHR has expanded to include roles in immune regulation, organogenesis, mucosal barrier function, and the cell cycle. These functions are likely dependent upon ligand-mediated activation of the receptor. High-affinity ligands of AHR have been classically defined as xenobiotics, such as polychlorinated biphenyls and dioxins. Identification of endogenous AHR ligands is key to understanding the physiologic functions of this enigmatic receptor. Metabolic pathways targeting the amino acid tryptophan and indole can lead to a myriad of metabolites, some of which are AHR ligands. Many of these ligands exhibit species selective preferential binding to AHR. The discovery of specific tryptophan metabolites as AHR ligands may provide insight concerning where AHR is activated in an organism, such as at the site of inflammation and within the intestinal tract. PMID:26041783

  19. Functional interaction of nuclear receptor coactivator 4 with aryl hydrocarbon receptor

    SciTech Connect

    Kollara, Alexandra; Brown, Theodore J. . E-mail: brown@mshri.on.ca

    2006-07-28

    Aryl hydrocarbon receptor (AhR) transcriptional activity is enhanced by interaction with p160 coactivators. We demonstrate here that NcoA4, a nuclear receptor coactivator, interacts with and amplifies AhR action. NcoA4-AhR and NcoA4-ARNT interactions were demonstrated by immunoprecipitation in T47D breast cancer and COS cells and was independent of ligand. Overexpression of NcoA4 enhanced AhR transcriptional activity 3.2-fold in the presence of dioxin, whereas overexpression of a splice variant, NcoA4{beta}, as well as a variant lacking the C-terminal region enhanced AhR transcriptional activity by only 1.6-fold. Enhanced AhR signaling by NcoA4 was independent of the LXXLL and FXXLF motifs or of the activation domain. NcoA4 protein localized to cytoplasm in the absence of dioxin and in both the cytoplasm and nucleus following dioxin treatment. NcoA4-facilitation of AhR activity was abolished by overexpression of androgen receptor, suggesting a potential competition of AhR and androgen receptor for NcoA4. These findings thus demonstrate a functional interaction between NcoA4 and AhR that may alter AhR activity to affect disease development and progression.

  20. Genetic variation at aryl hydrocarbon receptor (AHR) loci in populations of Atlantic killifish (Fundulus heteroclitus) inhabiting polluted and reference habitats

    PubMed Central

    2014-01-01

    Background The non-migratory killifish Fundulus heteroclitus inhabits clean and polluted environments interspersed throughout its range along the Atlantic coast of North America. Several populations of this species have successfully adapted to environments contaminated with toxic aromatic hydrocarbon pollutants such as polychlorinated biphenyls (PCBs). Previous studies suggest that the mechanism of resistance to these and other “dioxin-like compounds” (DLCs) may involve reduced signaling through the aryl hydrocarbon receptor (AHR) pathway. Here we investigated gene diversity and evidence for positive selection at three AHR-related loci (AHR1, AHR2, AHRR) in F. heteroclitus by comparing alleles from seven locations ranging over 600 km along the northeastern US, including extremely polluted and reference estuaries, with a focus on New Bedford Harbor (MA, USA), a PCB Superfund site, and nearby reference sites. Results We identified 98 single nucleotide polymorphisms within three AHR-related loci among all populations, including synonymous and nonsynonymous substitutions. Haplotype distributions were spatially segregated and F-statistics suggested strong population genetic structure at these loci, consistent with previous studies showing strong population genetic structure at other F. heteroclitus loci. Genetic diversity at these three loci was not significantly different in contaminated sites as compared to reference sites. However, for AHR2 the New Bedford Harbor population had significant FST values in comparison to the nearest reference populations. Tests for positive selection revealed ten nonsynonymous polymorphisms in AHR1 and four in AHR2. Four nonsynonymous SNPs in AHR1 and three in AHR2 showed large differences in base frequency between New Bedford Harbor and its reference site. Tests for isolation-by-distance revealed evidence for non-neutral change at the AHR2 locus. Conclusion Together, these data suggest that F. heteroclitus populations in reference

  1. Ah-receptor controlled luciferase expression: A novel species-specific bioassay for Ah-receptor active compounds in environmental matrices

    SciTech Connect

    Murk, A.J.; Aarts, J.M.M.J.G.; Koeman, J.H.; Brouwer, A.; Denison, M.S.

    1995-12-31

    Polyhalogenated aromatic hydrocarbons (PHAHs) such as polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs) and biphenyls (PCBs) are persistent lipophilic compounds that accumulate especially in sediments and in top predators of the aquatic foodchain. PHAHs elicit a number of common toxic responses, which are highly species-specific. The most toxic, planar, PHAHs share a common mechanism of action mediated by the aryl hydrocarbon receptor (AhR). Based on this mechanism, the toxic equivalency factor (TEF) concept has been developed, allowing hazard and risk assessment for mixtures of PHAHs. The TEF-approach assumes additive responses, but also synergistic and antagonistic interactions have been observed. In addition, the often large number of compounds in a mixture, low levels of individual congeners, possible presence of unknown AhR-active substances, and species differences in inducibility, ask for an comprehensive approach in hazard assessment. A number of recombinant cell lines, including Hepa1c1c7 mouse and H411E rat hepatoma cell lines, were developed, showing AhR-mediated firefly (Photinuspyralis) luciferase gene expression. The response by 2,3,7,8-TCDD in the CALUX (chemical activated luciferase expression) assay with these cell lines is dose-dependent, and not subjected to substrate inhibition at higher ligand concentrations. The detection limit for 2,3,7,8-TCDD is below 1 pM (0.2 fmol). The luciferase assay has been successfully applied for monitoring the amount of AhR-active compounds in small aliquots of blood plasma and in both sediment and pore-water samples, of which examples will be presented.

  2. Comparative Analysis of Homology Models of the Ah Receptor Ligand Binding Domain: Verification of Structure-Function Predictions by Site-Directed Mutagenesis of a Non-Functional AHR†

    PubMed Central

    Fraccalvieri, Domenico; Soshilov, Anatoly A.; Karchner, Sibel I.; Franks, Diana G.; Pandini, Alessandro; Bonati, Laura; Hahn, Mark E.; Denison, Michael S.

    2013-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-dependent transcription factor that mediates the biological and toxic effects of a wide variety of structurally diverse chemicals, including the toxic environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). While significant interspecies differences in AHR ligand binding specificity, selectivity and response have been observed, the structural determinants responsible have not been determined and homology models of the AHR ligand-binding domain (LBD) are available for only a few species. Here we describe the development and comparative analysis of homology models of the LBD of sixteen AHRs from twelve mammalian and nonmammalian species and identify the specific residues contained within their ligand binding cavities. The ligand-binding cavity of the fish AHR exhibits differences from mammalian and avian AHRs, suggesting a slightly different TCDD binding mode. Comparison of the internal cavity in the LBD model of zebrafish (zf) AHR2, which binds TCDD with high affinity, to that of zfAHR1a, which does not bind TCDD, revealed that the latter has a dramatically shortened binding cavity due to the side chains of three residues (Tyr296, Thr386, His388) that reduce the internal space available to TCDD. Mutagenesis of two of these residues in zfAhR1a to those present in zfAHR2 (Y296H, T386A) restored the ability of zfAHR1a to bind TCDD and to exhibit TCDD-dependent binding to DNA. These results demonstrate the importance of these two amino acids and highlight the predictive potential of comparative analysis of homology models from diverse species. The availability of these AHR LBD homology models will facilitate in depth comparative studies of AHR ligand binding and ligand-dependent AHR activation and provide a novel avenue to examine species specific differences in AHR responsiveness. PMID:23286227

  3. A novel role for the dioxin receptor in fatty acid metabolism and hepatic steatosis

    PubMed Central

    Lee, Jung Hoon; Wada, Taira; Febbraio, Maria; He, Jinhan; Matsubara, Tsutomu; Lee, Min Jae; Gonzalez, Frank J.; Xie, Wen

    2010-01-01

    Background & Aims The aryl hydrocarbon receptor (AhR) is a PAS domain transcription factor previously known as the “dioxin receptor” or “xenobiotic receptor.” The goal of this study is to determine the endobiotic role of AhR in hepatic steatosis. Methods Wild type, constitutively activated AhR (CA-AhR) transgenic, AhR null (AhR-/-), and fatty acid translocase CD36/FAT null (CD36-/-) mice were used to investigate the role of AhR in steatosis and the involvement of CD36 in the steatotic effect of AhR. The promoters of the mouse and human CD36 genes were cloned and their regulation by AhR was analyzed. Results Activation of AhR induced spontaneous hepatic steatosis characterized by the accumulation of triglycerides. The steatotic effect of AhR is likely due to the combined upregulation of CD36 and fatty acid transport proteins (FATPs), suppression of fatty acid oxidation, inhibition of hepatic export of triglycerides, increase in peripheral fat mobilization, and increased hepatic oxidative stress. Promoter analysis established CD36 as a novel transcriptional target of AhR. Activation of AhR in liver cells induced CD36 gene expression and enhanced fatty acid uptake. The steatotic effect of an AhR agonist was inhibited in CD36-/- mice. Conclusions Our study reveals a novel link between AhR-induced steatosis and the expression of CD36. Industrial or military exposures to dioxin and related compounds have been linked to increased prevalence of fatty liver in humans. Results from this study may help to establish AhR and its target CD36 as novel therapeutic and preventive targets for fatty liver disease. PMID:20303349

  4. AhR activation by 6-formylindolo[3,2-b]carbazole and 2,3,7,8-tetrachlorodibenzo-p-dioxin inhibit the development of mouse intestinal epithelial cells.

    PubMed

    Park, Joo-Hung; Choi, Ah-Jeong; Kim, Soo-Ji; Cheong, Seon-Woo; Jeong, So-Yeon

    2016-04-01

    The intestinal epithelium plays a central role in immune homeostasis in the intestine. AhR, a ligand-activated transcription factor, plays an important role in diverse physiological processes. The intestines are exposed to various exogenous and endogenous AhR ligands. Thus, AhR may regulate the intestinal homeostasis, directly acting on the development of intestinal epithelial cells (IEC). In this study, we demonstrated that 6-formylindolo[3,2-b]carbazole (FICZ) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) inhibited the in vitro development of mouse intestinal organoids. The number of Paneth cells in the small intestine and the depth of crypts of the small and large intestines were reduced in mice administrated with FICZ. Immunohistochemical and flow cytometric assays revealed that AhR was highly expressed in Lgr5(+) stem cells. FICZ inhibited Wnt signaling lowering the level of β-catenin protein. Gene expression analyses demonstrated that FICZ increased expression of Lgr5, Math1, BMP4, and Indian Hedgehog while inhibiting that of Lgr4. PMID:26950395

  5. Expression of genes for AhR and Nrf2 signal pathways in the retina of OXYS rats during the development of retinopathy and melatonin-induced changes in this process.

    PubMed

    Perepechaeva, M L; Stefanova, N A; Grishanova, A Yu

    2014-08-01

    Modulation of oxidative stress is one of the experimental approaches to the therapy of age-related macular degeneration. Melatonin holds much promise in this respect. It was hypothesized that the efficiency of melatonin in age-related macular degeneration is associated with its ability to modulate gene expression for the AhR and Nrf2 signal pathways. Experiments were performed on premature aging OXYS rats, which serve as a reliable model of age-related macular degeneration in humans. We studied the effect of melatonin on gene mRNA for the AhR and Nrf2 signal pathways. Melatonin was shown to decrease the level of mRNA for AhR-dependent genes of CYP1A2 and CYP1B1 cytochromes in the retina, but had no effect on the content of mRNA for Nrf2-dependent genes in OXYS rats.

  6. EGCG protects endothelial cells against PCB 126-induced inflammation through inhibition of AhR and induction of Nrf2-regulated genes

    SciTech Connect

    Han, Sung Gu; Han, Seong-Su; Toborek, Michal; Hennig, Bernhard

    2012-06-01

    Tea flavonoids such as epigallocatechin gallate (EGCG) protect against vascular diseases such as atherosclerosis via their antioxidant and anti-inflammatory functions. Persistent and widespread environmental pollutants, including polychlorinated biphenyls (PCB), can induce oxidative stress and inflammation in vascular endothelial cells. Even though PCBs are no longer produced, they are still detected in human blood and tissues and thus considered a risk for vascular dysfunction. We hypothesized that EGCG can protect endothelial cells against PCB-induced cell damage via its antioxidant and anti-inflammatory properties. To test this hypothesis, primary vascular endothelial cells were pretreated with EGCG, followed by exposure to the coplanar PCB 126. Exposure to PCB 126 significantly increased cytochrome P450 1A1 (Cyp1A1) mRNA and protein expression and superoxide production, events which were significantly attenuated following pretreatment with EGCG. Similarly, EGCG also reduced DNA binding of NF-κB and downstream expression of inflammatory markers such as monocyte chemotactic protein-1 (MCP-1) and vascular cell adhesion protein-1 (VCAM-1) after PCB exposure. Furthermore, EGCG decreased endogenous or base-line levels of Cyp1A1, MCP-1 and VCAM-1 in endothelial cells. Most of all, treatment of EGCG upregulated expression of NF-E2-related factor 2 (Nrf2)-controlled antioxidant genes, including glutathione S transferase (GST) and NAD(P)H:quinone oxidoreductase 1 (NQO1), in a dose-dependent manner. In contrast, silencing of Nrf2 increased Cyp1A1, MCP-1 and VCAM-1 and decreased GST and NQO1 expression, respectively. These data suggest that EGCG can inhibit AhR regulated genes and induce Nrf2-regulated antioxidant enzymes, thus providing protection against PCB-induced inflammatory responses in endothelial cells. -- Highlights: ► PCBs cause endothelial inflammation and subsequent atherosclerosis. ► Nutrition can modulate toxicity by environmental pollutants. ► We

  7. Activation of AhR-mediated toxicity pathway by emerging pollutants polychlorinated diphenyl sulfides

    EPA Science Inventory

    Polychlorinated diphenyl sulfides (PCDPSs) are a group of environmental pollutants for which limited toxicological information is available. This study tested the hypothesis that PCDPSs could activate the mammalian aryl hydrocarbon receptor (AhR) mediated toxicity pathways. Eight...

  8. Transgenic mouse lines expressing rat AH receptor variants - A new animal model for research on AH receptor function and dioxin toxicity mechanisms

    SciTech Connect

    Pohjanvirta, Raimo

    2009-04-15

    Han/Wistar (Kuopio; H/W) rats are exceptionally resistant to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) toxicity mainly because of their mutated aryl hydrocarbon receptor (AHR) gene. In H/W rats, altered splicing of the AHR mRNA generates two AHR proteins: deletion (DEL) and insertion (INS) variants, with the INS isoform being predominantly expressed. To gain further insight into their functional properties, cDNAs of these and rat wild-type (rWT) isoform were transferred into C57BL/6J-derived mice by microinjection. The endogenous mouse AHR was eliminated by selective crossing with Ahr-null mice. A single mouse line was obtained for each of the three constructs. The AHR mRNA levels in tissues were generally close to those of C57BL/6 mice in INS and DEL mice and somewhat higher in rWT mice; in testis, however, all 3 constructs exhibited marked overexpression. The transgenic mouse lines were phenotypically normal except for increased testis weight. Induction of drug-metabolizing enzymes by TCDD occurred similarly to that in C57BL/6 mice, but there tended to be a correlation with AHR concentrations, especially in testis. In contrast to C57BL/6 mice, the transgenics did not display any major gender difference in susceptibility to the acute lethality and hepatotoxicity of TCDD; rWT mice were highly sensitive, DEL mice moderately resistant and INS mice highly resistant. Co-expression of mouse AHR and rWT resulted in augmented sensitivity to TCDD and abolished the natural resistance of female C57BL/6 mice, whereas mice co-expressing mouse AHR and INS were resistant. Thus, these transgenic mouse lines provide a novel promising tool for molecular studies on dioxin toxicity and AHR function.

  9. Association of polymorphisms in AhR, CYP1A1, GSTM1, and GSTT1 genes with levels of DNA damage in peripheral blood lymphocytes among coke-oven workers

    SciTech Connect

    Yongwen Chen; Yun Bai; Jing Yuan; Weihong Chen; Jianya Sun; Hong Wang; Huashan Liang; Liang Guo; Xiaobo Yang; Hao Tan; Yougong Su; Qingyi Wei; Tangchun Wu

    2006-09-15

    Accumulating evidence has shown that both DNA damage caused by the metabolites of polycyclic aromatic hydrocarbons (PAH) and genetic polymorphisms in PAH-metabolic genes contribute to individual susceptibility to PAH-induced carcinogenesis. However, the functional relevance of genetic polymorphisms in PAH-metabolic genes in exposed individuals is still unclear. In this study of 240 coke-oven workers (the exposed group) and 123 non-coke-oven workers (the control group), we genotyped for polymorphisms in the AhR, CYP1A1, GSTM1, and GSTT1 genes by PCR methods, and determined the levels of DNA damage in peripheral blood lymphocytes using the alkaline comet assay. It was found that the ln-transformed Olive tail moment (Olive TM) values in the exposed group were significantly higher than those in the control group. Furthermore, in the exposed group, the Olive TM values in subjects with the AhR Lys{sup 554} variant genotype were higher than those with the AhR Arg{sup 554}/Arg{sup 554} genotype. Similarly, the Olive TM values in the non-coke-oven workers with the CYP1A1 MspI CC + CT genotype were lower than the values of those with the CYP1A1 MspI TT genotype. However, these differences were not evident for GSTM1 and GSTT1. These results suggested that the polymorphism of AhR might modulate the effects of PAHs in the exposed group; however, the underlying molecular mechanisms by which this polymorphism may have affected the levels of PAH-induced DNA damage warrant further investigation.

  10. Aryl Hydrocarbon Receptor Plays Protective Roles against High Fat Diet (HFD)-induced Hepatic Steatosis and the Subsequent Lipotoxicity via Direct Transcriptional Regulation of Socs3 Gene Expression.

    PubMed

    Wada, Taira; Sunaga, Hiroshi; Miyata, Kazuki; Shirasaki, Haruno; Uchiyama, Yuki; Shimba, Shigeki

    2016-03-25

    Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor regulating the expression of genes involved in xenobiotic response. Recent studies have suggested that AhR plays essential roles not only in xenobiotic detoxification but also energy metabolism. Thus, in this study, we studied the roles of AhR in lipid metabolism. Under high fat diet (HFD) challenge, liver-specific AhR knock-out (AhR LKO) mice exhibited severe steatosis, inflammation, and injury in the liver. Gene expression analysis and biochemical study revealed thatde novolipogenesis activity was significantly increased in AhR LKO mice. In contrast, induction of suppressor of cytokine signal 3 (Socs3) expression by HFD was attenuated in the livers of AhR LKO mice. Rescue of theSocs3gene in the liver of AhR LKO mice cancelled the HFD-induced hepatic lipotoxicities. Promoter analysis established Socs3 as novel transcriptional target of AhR. These results indicated that AhR plays a protective role against HFD-induced hepatic steatosis and the subsequent lipotoxicity effects, such as inflammation, and that the mechanism of protection involves the direct transcriptional regulation ofSocs3expression by AhR. PMID:26865635

  11. Ah receptor mediated suppression of the antibody response in mice is primarily dependent on the Ah phenotype of lymphoid tissue.

    PubMed

    Silkworth, J B; Antrim, L A; Sack, G

    1986-12-01

    Halogenated aromatic hydrocarbons act through the aromatic hydrocarbon (Ah) receptor in mice to produce a series of toxic effects of the immune system. The receptor protein is a product of the Ah gene locus. Ah responsive (Ahb/Ahb) mice express a high affinity receptor in both lymphoid and nonlymphoid tissues whereas nonresponsive Ahd/Ahd mice express a poor affinity receptor. To determine the role of the Ah receptor of lymphoid tissue relative to that of nonlymphoid tissue in the induction of immune impairment, bone marrow was used to reconstitute lethally irradiated mice of the same or opposite Ah phenotype. All mice were given 3,3',4,4'-tetrachlorobiphenyl (35 and 350 mumol/kg) ip 2 days before immunization with sheep erythrocytes (SRBC). The immune response to this T dependent antigen and organ weights were determined 5 or 7 days later in normal or chimeric mice, respectively. Monoclonal Lyt 1.1 and Lyt 1.2 antibodies were used to establish the origin of the cells which repopulated the chimeric thymuses. The immune responses of both BALB/cBy (Ahb/Ahb) and the BALB/cBy X DBA/2 hybrid, CByD2F1 (Ahb/Ahd), were significantly suppressed but DBA/2 mice were unaffected. The immune responses of chimeric BALB/cBy----BALB/cBy and BALB/cBy----DBA/2 (donor----recipient) mice were also significantly suppressed and thymic atrophy was observed in both cases. The serum anti-SRBC antibody titers of DBA/2----BALB/cBy chimeras were also significantly decreased although not to the same extent as in BALB/cBy----DBA/2 mice. Chimeric DBA/2----DBA/2 mice were not affected. These results indicate that the sensitivity to Ah receptor mediated suppression of the antibody response is primarily determined by the Ah phenotype of the lymphoid tissue.

  12. The aryl hydrocarbon receptor promotes aging phenotypes across species

    PubMed Central

    Eckers, Anna; Jakob, Sascha; Heiss, Christian; Haarmann-Stemmann, Thomas; Goy, Christine; Brinkmann, Vanessa; Cortese-Krott, Miriam M.; Sansone, Roberto; Esser, Charlotte; Ale-Agha, Niloofar; Altschmied, Joachim; Ventura, Natascia; Haendeler, Judith

    2016-01-01

    The ubiquitously expressed aryl hydrocarbon receptor (AhR) induces drug metabolizing enzymes as well as regulators of cell growth, differentiation and apoptosis. Certain AhR ligands promote atherosclerosis, an age-associated vascular disease. Therefore, we investigated the role of AhR in vascular functionality and aging. We report a lower pulse wave velocity in young and old AhR-deficient mice, indicative of enhanced vessel elasticity. Moreover, endothelial nitric oxide synthase (eNOS) showed increased activity in the aortas of these animals, which was reflected in increased NO production. Ex vivo, AhR activation reduced the migratory capacity of primary human endothelial cells. AhR overexpression as well as treatment with a receptor ligand, impaired eNOS activation and reduced S-NO content. All three are signs of endothelial dysfunction. Furthermore, AhR expression in blood cells of healthy human volunteers positively correlated with vessel stiffness. In the aging model Caenorhabditis elegans, AhR-deficiency resulted in increased mean life span, motility, pharynx pumping and heat shock resistance, suggesting healthier aging. Thus, AhR seems to have a negative impact on vascular and organismal aging. Finally, our data from human subjects suggest that AhR expression levels could serve as an additional, new predictor of vessel aging. PMID:26790370

  13. Induction of AhR-Mediated Gene Transcription by Coffee

    PubMed Central

    Ishikawa, Toshio; Takahashi, Satoshi; Morita, Koji; Okinaga, Hiroko; Teramoto, Tamio

    2014-01-01

    Background Aryl hydrocarbon receptor (AhR) is classically known to be activated by xenobiotics such as dioxins and polycyclic aromatic hydrocarbons (PAHs). Although it has been reported that PAHs are contained in roasted coffee beans, in general coffee beverages are not considered to be AhR activators. We tested whether exposure to coffee would activate AhR in cultured cells. Methods HepG2 cells stably expressing an AhR-responsive reporter gene were treated with coffee samples. Also, expression of CYP1A1, an endogenous AhR-responsive gene, was quantitated by RT-PCR and Western blotting in HepG2, Caco-2, and MCF-7 cells, after treatment with coffee. In order to obtain sensitive and reproducible results, all the experiments were performed with the cells placed in either phosphate-buffered saline (PBS) or pure serum, instead of routinely-used culture medium, whose intrinsic AhR-stimulating activity turned out to be so strong as to interfere with the analyses. Results All the coffee samples tested robustly stimulated AhR-mediated transcription in the reporter gene assays. Of note, to what extent coffee and other AhR agonists activated AhR was different, depending on whether the experiments were done in PBS or serum. CYP1A1 mRNA was induced by coffee, in HepG2, Caco-2, and MCF-7 cells placed in either PBS or serum. CYP1A1 protein expression, which was not detected in these cells incubated in PBS, was also increased by coffee in cells placed in serum. Conclusions By using culture medium-free experimental settings, we have shown that coffee is a strong AhR activator. Our observation may help elucidate as-yet-unrecognized effects of coffee on human health. PMID:25007155

  14. In Vivo Dioxin Favors Interleukin-22 Production by Human CD4+ T Cells in an Aryl Hydrocarbon Receptor (AhR)-Dependent Manner

    PubMed Central

    Brembilla, Nicolò Costantino; Ramirez, Jean-Marie; Chicheportiche, Rachel; Sorg, Olivier; Saurat, Jean-Hilaire; Chizzolini, Carlo

    2011-01-01

    Background The transcription factor aryl hydrocarbon receptor (AhR) mediates the effects of a group of chemicals known as dioxins, ubiquitously present in our environment. However, it is poorly known how the in vivo exposure to these chemicals affects in humans the adaptive immune response. We therefore assessed the functional phenotype of T cells from an individual who developed a severe cutaneous and systemic syndrome after having been exposed to an extremely high dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Methodology/Principal Findings T cells of the TCDD-exposed individual were studied for their capacity to produce cytokines in response to polyclonal and superantigenic stimulation, and for the expression of chemokine receptors involved in skin homing. The supernatants from T cells of the exposed individual contained a substantially increased amount of interleukin (IL)-22 but not of IL-17A, interferon (IFN)-γ or IL-10 when compared to nine healthy controls. In vitro experiments confirmed a direct, AhR-dependent, enhancing effect of TCDD on IL-22 production by CD4+ T cells. The increased production of IL-22 was not dependent on AhR occupancy by residual TCDD molecules, as demonstrated in competition experiments with the specific AhR antagonist CH-223191. In contrast, it was due to an increased frequency of IL-22 single producing cells accompanied by an increased percentage of cells expressing the skin-homing chemokine receptors CCR6 and CCR4, identified through a multiparameter flow cytometry approach. Of interest, the frequency of CD4+CD25hiFoxP3+ T regulatory cells was similar in the TCDD-exposed and healthy individuals. Conclusions/Significance This case strongly supports the contention that human exposure to persistent AhR ligands in vivo induce a long-lasting effect on the human adaptive immune system and specifically polarizes CD4+ T cells to produce IL-22 and not other T cell cytokines with no effect on T regulatory cells. PMID:21525997

  15. Prenatal 3,3',4,4',5-pentachlorobiphenyl exposure modulates induction of rat hepatic CYP 1A1, 1B1, and AhR by 7,12-dimethylbenz[a]anthracene

    SciTech Connect

    Wakui, Shin . E-mail: wakui@azabu-u.ac.jp; Yokoo, Kiyofumi; Takahashi, Hiroyuki; Muto, Tomoko; Suzuki, Yoshihiko; Kanai, Yoshikatsu; Hano, Hiroshi; Furusato, Masakuni; Endou, Hitoshi

    2006-02-01

    We previously reported the finding that prenatal exposure to a relatively low dose of PCB126 increases the rate of DMBA-induced rat mammary carcinoma, while a high dose decreased it. One of the most important factors determining the sensitivity to mammary carcinogenesis is the metabolic stage at administration of the carcinogenic agent. DMBA is a procarcinogen that recruits the host metabolism to yield its ultimate carcinogenic form, and CYP1A1 and CYP1B1 (CYP1) conduct this metabolism. We investigated the hepatic expression of CYP1 and AhR following oral administration of DMBA (100 mg/kg b.w.) (i.g.) to 50-day-old female Sprague-Dawley rats whose dams had been treated (i.g.) with 2.5 ng, 250 ng, 7.5 {mu}g of PCB126/kg or the vehicle on days 13 to 19 post-conception. Real-time quantitative RT-PCR analysis revealed that the prenatal exposure to a relatively low dose of PCB126 (the 250 ng group) prolonged the higher expression of CYP1A1, CYP1B1, and AhR mRNA, while prenatal exposure to a high dose of PCB126 (the 7.5 {mu}g group) prolonged the higher expression of CYP1A1 and AhR mRNA. Western blotting and immunohistochemical analyses were consistent with mRNAs changes. Because DMBA oxidation produces a highly mutagenic metabolite and is finally catalyzed by CYP1B1, a relatively low PCB126 dose might produce the biological character to potentially increase the risk of DMBA-induced mammary carcinoma.

  16. Identification of the Ah-receptor structural determinants for ligand preferences.

    PubMed

    Xing, Yongna; Nukaya, Manabu; Satyshur, Kenneth A; Jiang, Li; Stanevich, Vitali; Korkmaz, Elif Nihal; Burdette, Lisa; Kennedy, Gregory D; Cui, Qiang; Bradfield, Christopher A

    2012-09-01

    The aryl hydrocarbon receptor (AHR) is a transcription factor that responds to diverse ligands and plays a critical role in toxicology, immune function, and cardiovascular physiology. The structural basis of the AHR for ligand promiscuity and preferences is critical for understanding AHR function. Based on the structure of a closely related protein HIF2α, we modeled the AHR ligand binding domain (LBD) bound to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and benzo(a)pyrene (BaP) and identified residues that control ligand preferences by shape and H-bond potential. Mutations to these residues, particularly Q377 and G298, resulted in robust and opposite changes in the potency of TCDD and BaP and up to a 20-fold change in the ratio of TCDD/BaP efficacy. The model also revealed a flexible "belt" structure; molecular dynamic (MD) simulation suggested that the "belt" and several other structural elements in the AHR-LBD are more flexible than HIF2α and likely contribute to ligand promiscuity. Molecular docking of TCDD congeners to a model of human AHR-LBD ranks their binding affinity similar to experimental ranking of their toxicity. Our study reveals key structural basis for prediction of toxicity and understanding the AHR signaling through diverse ligands. PMID:22659362

  17. Ah Receptor Signaling Controls the Expression of Cardiac Development and Homeostasis Genes.

    PubMed

    Carreira, Vinicius S; Fan, Yunxia; Wang, Qing; Zhang, Xiang; Kurita, Hisaka; Ko, Chia-I; Naticchioni, Mindi; Jiang, Min; Koch, Sheryl; Medvedovic, Mario; Xia, Ying; Rubinstein, Jack; Puga, Alvaro

    2015-10-01

    Congenital heart disease (CHD) is the most common congenital abnormality and one of the leading causes of newborn death throughout the world. Despite much emerging scientific information, the precise etiology of this disease remains elusive. Here, we show that the aryl hydrocarbon receptor (AHR) regulates the expression of crucial cardiogenesis genes and that interference with endogenous AHR functions, either by gene ablation or by agonist exposure during early development, causes overlapping structural and functional cardiac abnormalities that lead to altered fetal heart physiology, including higher heart rates, right and left ventricle dilation, higher stroke volume, and reduced ejection fraction. With striking similarity between AHR knockout (Ahr(-/-)) and agonist-exposed wild type (Ahr(+/+)) embryos, in utero disruption of endogenous AHR functions converge into dysregulation of molecular mechanisms needed for attainment and maintenance of cardiac differentiation, including the pivotal signals regulated by the cardiogenic transcription factor NKH2.5, energy balance via oxidative phosphorylation and TCA cycle and global mitochondrial function and homeostasis. Our findings suggest that AHR signaling in the developing mammalian heart is central to the regulation of pathways crucial for cellular metabolism, cardiogenesis, and cardiac function, which are potential targets of environmental factors associated with CHD.

  18. The aryl hydrocarbon receptor controls cyclin O to promote epithelial multiciliogenesis

    PubMed Central

    Villa, Matteo; Crotta, Stefania; Dingwell, Kevin S.; Hirst, Elizabeth M. A.; Gialitakis, Manolis; Ahlfors, Helena; Smith, James C.; Stockinger, Brigitta; Wack, Andreas

    2016-01-01

    Epithelia function as barriers against environmental insults and express the transcription factor aryl hydrocarbon receptor (AhR). However, AhR function in these tissues is unknown. Here we show that AhR regulates multiciliogenesis in both murine airway epithelia and in Xenopus laevis epidermis. In air-exposed airway epithelia, induction of factors required for multiciliogenesis, including cyclin O (Ccno) and Multicilin (Mcidas), is AhR dependent, and air exposure induces AhR binding to the Ccno promoter. Submersion and hypoxic conditions impede AhR-dependent Ccno induction. This is mediated by the persistence of Notch signalling, as Notch blockade renders multiciliogenesis and Ccno induction by AhR independent from air exposure. In contrast to Ccno induction, air exposure does not induce the canonical AhR target cytochrome P450 1a1 (Cyp1a1). Inversely, exposure to AhR ligands induces Cyp1a1 but not Ccno and impeded ciliogenesis. These data indicate that AhR involvement in detoxification of environmental pollutants may impede its physiological role, resulting in respiratory pathology. PMID:27554288

  19. The aryl hydrocarbon receptor controls cyclin O to promote epithelial multiciliogenesis.

    PubMed

    Villa, Matteo; Crotta, Stefania; Dingwell, Kevin S; Hirst, Elizabeth M A; Gialitakis, Manolis; Ahlfors, Helena; Smith, James C; Stockinger, Brigitta; Wack, Andreas

    2016-01-01

    Epithelia function as barriers against environmental insults and express the transcription factor aryl hydrocarbon receptor (AhR). However, AhR function in these tissues is unknown. Here we show that AhR regulates multiciliogenesis in both murine airway epithelia and in Xenopus laevis epidermis. In air-exposed airway epithelia, induction of factors required for multiciliogenesis, including cyclin O (Ccno) and Multicilin (Mcidas), is AhR dependent, and air exposure induces AhR binding to the Ccno promoter. Submersion and hypoxic conditions impede AhR-dependent Ccno induction. This is mediated by the persistence of Notch signalling, as Notch blockade renders multiciliogenesis and Ccno induction by AhR independent from air exposure. In contrast to Ccno induction, air exposure does not induce the canonical AhR target cytochrome P450 1a1 (Cyp1a1). Inversely, exposure to AhR ligands induces Cyp1a1 but not Ccno and impeded ciliogenesis. These data indicate that AhR involvement in detoxification of environmental pollutants may impede its physiological role, resulting in respiratory pathology. PMID:27554288

  20. Dioxin sensitivity-related two critical amino acids of arylhydrocarbon receptor may not correlate with the taxonomy or phylogeny in avian species.

    PubMed

    Fujisawa, Nozomi; Kawai, Yusuke K; Nakayama, Shouta M M; Ikenaka, Yoshinori; Yamamoto, Hideaki; Ishizuka, Mayumi

    2013-12-30

    There are two arylhydrocarbon receptor (AhR) isoforms in birds, AhR1 and AhR2. The varying sensitivity of AhR is reported to be related to two critical amino acids at positions 325 and 381 in the AhR1 ligand-binding domain. In this study, seven avian species whose in vivo dioxin sensitivity was known, and 13 species with no data regarding their in vivo dioxin sensitivity were examined. The two critical amino acids in the ligand-binding domain were investigated in avian species, and the results were compared with the taxonomy or phylogenetic trees for the bird AhR proteins. We found that the two critical amino acids did not correlate with the taxonomy or phylogeny of these proteins, suggesting that dioxin sensitivity was independent of taxonomy.

  1. Dioxin Sensitivity-Related Two Critical Amino Acids of Arylhydrocarbon Receptor May Not Correlate with the Taxonomy or Phylogeny in Avian Species

    PubMed Central

    FUJISAWA, Nozomi; KAWAI, Yusuke K.; NAKAYAMA, Shouta M. M.; IKENAKA, Yoshinori; YAMAMOTO, Hideaki; ISHIZUKA, Mayumi

    2013-01-01

    ABSTRACT There are two arylhydrocarbon receptor (AhR) isoforms in birds, AhR1 and AhR2. The varying sensitivity of AhR is reported to be related to two critical amino acids at positions 325 and 381 in the AhR1 ligand-binding domain. In this study, seven avian species whose in vivo dioxin sensitivity was known, and 13 species with no data regarding their in vivo dioxin sensitivity were examined. The two critical amino acids in the ligand-binding domain were investigated in avian species, and the results were compared with the taxonomy or phylogenetic trees for the bird AhR proteins. We found that the two critical amino acids did not correlate with the taxonomy or phylogeny of these proteins, suggesting that dioxin sensitivity was independent of taxonomy. PMID:23912877

  2. Benzo[ghi]perylene activates the AHR pathway to exert biological effects on the NL-20 human bronchial cell line.

    PubMed

    Zaragoza-Ojeda, Montserrat; Eguía-Aguilar, Pilar; Perezpeña-Díazconti, Mario; Arenas-Huertero, Francisco

    2016-08-10

    Polycyclic aromatic hydrocarbons (PAH) are produced by incomplete combustion of organic material. In the Mexico City atmosphere, the most abundant PAH is benzo[ghi]perylene (BghiP), a gasoline combustion marker. At present, there are no reports of the effects of BghiP on human bronchial cells, so the aim of the study was to evaluate the effects in vitro of BghiP on the NL-20 cell line. Results showed that BghiP induced the formation of small vesicles throughout the cytoplasm, with absence of nuclear fragmentation. At 48h exposition, damage in cell membrane increased significantly at 1.24μg/mL of BghiP (p<0.05). Immunocytochemistry revealed that BghiP provokes nuclear translocation of AhR receptor, which indicates that this compound can induce transcription of genes via receptor binding (AhR pathway activation). BghiP induced a two-fold increase (p<0.05) in the expression of AhR and CYP4B1 (a lung-specific pathway effector). In the presence of the receptor antagonist CH-223191, the loss of viability, the nuclear translocation and the overexpression of genes decreased, though this did not prevent the formation of vesicles. BghiP induced oxidative stress and in presence of the receptor antagonist this increased significantly. In conclusion, BghiP can activate the overexpression of AhR and CYP4B1, and the effects are abated by the AhR receptor antagonist. This is the first report to prove that BghiP utilizes the AhR pathway to exert its toxic effects on the NL-20 human bronchial cell line . PMID:27234499

  3. Evodiamine as a novel antagonist of aryl hydrocarbon receptor

    SciTech Connect

    Yu, Hui; Tu, Yongjiu; Zhang, Chun; Fan, Xia; Wang, Xi; Wang, Zhanli; Liang, Huaping

    2010-11-05

    Research highlights: {yields} Evodiamine interacted with the AhR. {yields} Evodiamine inhibited the specific binding of [{sup 3}H]-TCDD to the AhR. {yields} Evodiamine acts as an antagonist of the AhR. -- Abstract: Evodiamine, the major bioactive alkaloid isolated from Wu-Chu-Yu, has been shown to interact with a wide variety of proteins and modify their expression and activities. In this study, we investigated the interaction between evodiamine and the aryl hydrocarbon receptor (AhR). Molecular modeling results revealed that evodiamine directly interacted with the AhR. Cytosolic receptor binding assay also provided the evidence that evodiamine could interact with the AhR with the K{sub i} value of 28.4 {+-} 4.9 nM. In addition, we observed that evodiamine suppressed the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induced nuclear translocation of the AhR and the expression of CYP1A1 dose-dependently. These results suggested that evodiamine was able to bind to the AhR as ligand and exhibit antagonistic effects.

  4. Balancing intestinal and systemic inflammation through cell type-specific expression of the aryl hydrocarbon receptor repressor

    PubMed Central

    Brandstätter, Olga; Schanz, Oliver; Vorac, Julia; König, Jessica; Mori, Tetsushi; Maruyama, Toru; Korkowski, Markus; Haarmann-Stemmann, Thomas; von Smolinski, Dorthe; Schultze, Joachim L.; Abel, Josef; Esser, Charlotte; Takeyama, Haruko; Weighardt, Heike; Förster, Irmgard

    2016-01-01

    As a sensor of polyaromatic chemicals the aryl hydrocarbon receptor (AhR) exerts an important role in immune regulation besides its requirement for xenobiotic metabolism. Transcriptional activation of AhR target genes is counterregulated by the AhR repressor (AhRR) but the exact function of the AhRR in vivo is currently unknown. We here show that the AhRR is predominantly expressed in immune cells of the skin and intestine, different from other AhR target genes. Whereas AhRR antagonizes the anti-inflammatory function of the AhR in the context of systemic endotoxin shock, AhR and AhRR act in concert to dampen intestinal inflammation. Specifically, AhRR contributes to the maintenance of colonic intraepithelial lymphocytes and prevents excessive IL-1β production and Th17/Tc17 differentiation. In contrast, the AhRR enhances IFN-γ-production by effector T cells in the inflamed gut. Our findings highlight the physiologic importance of cell-type specific balancing of AhR/AhRR expression in response to microbial, nutritional and other environmental stimuli. PMID:27184933

  5. Balancing intestinal and systemic inflammation through cell type-specific expression of the aryl hydrocarbon receptor repressor.

    PubMed

    Brandstätter, Olga; Schanz, Oliver; Vorac, Julia; König, Jessica; Mori, Tetsushi; Maruyama, Toru; Korkowski, Markus; Haarmann-Stemmann, Thomas; von Smolinski, Dorthe; Schultze, Joachim L; Abel, Josef; Esser, Charlotte; Takeyama, Haruko; Weighardt, Heike; Förster, Irmgard

    2016-05-17

    As a sensor of polyaromatic chemicals the aryl hydrocarbon receptor (AhR) exerts an important role in immune regulation besides its requirement for xenobiotic metabolism. Transcriptional activation of AhR target genes is counterregulated by the AhR repressor (AhRR) but the exact function of the AhRR in vivo is currently unknown. We here show that the AhRR is predominantly expressed in immune cells of the skin and intestine, different from other AhR target genes. Whereas AhRR antagonizes the anti-inflammatory function of the AhR in the context of systemic endotoxin shock, AhR and AhRR act in concert to dampen intestinal inflammation. Specifically, AhRR contributes to the maintenance of colonic intraepithelial lymphocytes and prevents excessive IL-1β production and Th17/Tc17 differentiation. In contrast, the AhRR enhances IFN-γ-production by effector T cells in the inflamed gut. Our findings highlight the physiologic importance of cell-type specific balancing of AhR/AhRR expression in response to microbial, nutritional and other environmental stimuli.

  6. Balancing intestinal and systemic inflammation through cell type-specific expression of the aryl hydrocarbon receptor repressor.

    PubMed

    Brandstätter, Olga; Schanz, Oliver; Vorac, Julia; König, Jessica; Mori, Tetsushi; Maruyama, Toru; Korkowski, Markus; Haarmann-Stemmann, Thomas; von Smolinski, Dorthe; Schultze, Joachim L; Abel, Josef; Esser, Charlotte; Takeyama, Haruko; Weighardt, Heike; Förster, Irmgard

    2016-01-01

    As a sensor of polyaromatic chemicals the aryl hydrocarbon receptor (AhR) exerts an important role in immune regulation besides its requirement for xenobiotic metabolism. Transcriptional activation of AhR target genes is counterregulated by the AhR repressor (AhRR) but the exact function of the AhRR in vivo is currently unknown. We here show that the AhRR is predominantly expressed in immune cells of the skin and intestine, different from other AhR target genes. Whereas AhRR antagonizes the anti-inflammatory function of the AhR in the context of systemic endotoxin shock, AhR and AhRR act in concert to dampen intestinal inflammation. Specifically, AhRR contributes to the maintenance of colonic intraepithelial lymphocytes and prevents excessive IL-1β production and Th17/Tc17 differentiation. In contrast, the AhRR enhances IFN-γ-production by effector T cells in the inflamed gut. Our findings highlight the physiologic importance of cell-type specific balancing of AhR/AhRR expression in response to microbial, nutritional and other environmental stimuli. PMID:27184933

  7. The aryl hydrocarbon receptor suppresses cigarette-smoke-induced oxidative stress in association with dioxin response element (DRE)-independent regulation of sulfiredoxin 1.

    PubMed

    Sarill, Miles; Zago, Michela; Sheridan, Jared A; Nair, Parameswaran; Matthews, Jason; Gomez, Alvin; Roussel, Lucie; Rousseau, Simon; Hamid, Qutayba; Eidelman, David H; Baglole, Carolyn J

    2015-12-01

    The aryl hydrocarbon receptor (AhR) is a ubiquitously expressed receptor/transcription factor that mediates toxicological responses of environmental contaminants such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Emerging evidence indicates that the AhR suppresses apoptosis and proliferation independent of exogenous ligands, including suppression of apoptosis by cigarette smoke, a key risk factor for chronic obstructive pulmonary disease (COPD). As cigarette smoke is a potent inducer of oxidative stress, a feature that may contribute to the development of COPD, we hypothesized that the AhR prevents smoke-induced apoptosis by regulating oxidative stress. Utilizing primary lung fibroblasts derived from AhR(+/+) and AhR(-/-) mice as well as A549 human lung adenocarcinoma cells deficient in AhR expression (A549-AhR(ko)), we first show that AhR(-/-) fibroblasts and A549-AhR(ko) epithelial cells have a significant increase in cigarette smoke extract (CSE)-induced oxidative stress compared to wild type. CSE induced a significant increase in the mRNA expression of key antioxidant genes, including Nqo1 and Srxn1, predominantly in AhR(+/+) fibroblasts, with significantly less induction in AhR(-/-) cells. The induction of Srxn1, but not Nqo1, was independent of dioxin-response element (DRE) binding as AhR(DBD/DBD) lung fibroblasts, which express an AhR incapable of binding the DRE, increased Srxn1 to a degree similar to wild-type cells in response to CSE. There was no difference in Nrf2 expression or activation based on AhR expression. Lung fibroblasts derived from COPD subjects have significantly less AhR protein expression compared with both never-smokers (Normal) and smokers (At Risk). Consequently, COPD-derived fibroblasts were less robust in their induction of both Nqo1 and Srxn1 mRNA after exposure to CSE, which also failed to activate the AhR in the COPD fibroblasts. Taken together, these results support a new role for the AhR in regulating antioxidant defense in lung

  8. Lactobacillus bulgaricus OLL1181 activates the aryl hydrocarbon receptor pathway and inhibits colitis.

    PubMed

    Takamura, Takeyuki; Harama, Daisuke; Fukumoto, Suguru; Nakamura, Yuki; Shimokawa, Naomi; Ishimaru, Kayoko; Ikegami, Shuji; Makino, Seiya; Kitamura, Masanori; Nakao, Atsuhito

    2011-10-01

    Increasing evidence suggests that the aryl hydrocarbon receptor (AhR) pathway has an important role in the regulation of inflammatory responses. Most recently, we have shown that the activation of the AhR pathway by a potent AhR agonist inhibits the development of dextran sodium sulfate (DSS)-induced colitis, a model of human ulcerative colitis, by the induction of prostaglandin E2 (PGE2) in the large intestine. Because several strains of probiotic lactic acid bacteria have been reported to inhibit DSS-induced colitis by unidentified mechanisms, we hypothesized that particular strains of lactic acid bacterium might have the potential to activate the AhR pathway, thereby inhibiting DSS-induced colitis. This study investigated whether there are specific lactic acid bacterial strains that can activate the AhR pathway, and if so, whether this AhR-activating potential is associated with suppression of DSS-induced colitis. By using AhR signaling reporter cells, we found that Lactobacillus bulgaricus OLL1181 had the potential to activate the AhR pathway. OLL1181 also induced the mRNA expression of cytochrome P450 family 1A1 (CYP1A1), a target gene of the AhR pathway, in human colon cells, which was inhibited by the addition of an AhR antagonist, α-naphthoflavon (αNF). In addition, mice treated orally with OLL1181 showed an increase in CYP1A1 mRNA expression in the large intestine and amelioration of DSS-induced colitis. Thus, OLL1181 can induce activation of the intestinal AhR pathway and inhibit DSS-induced colitis in mice. This strain of lactic acid bacterium has therefore the potential to activate the AhR pathway, which may be able to suppress colitis.

  9. The Aryl Hydrocarbon Receptor: A Key Bridging Molecule of External and Internal Chemical Signals

    PubMed Central

    Tian, Jijing; Feng, Yu; Fu, Hualing; Xie, Heidi Qunhui; Jiang, Joy Xiaosong; Zhao, Bin

    2015-01-01

    The aryl hydrocarbon receptor (AhR) is a highly evolutionary conserved, ligand-activated transcription factor that is best known to mediate the toxicities of dioxins and dioxin-like compounds. Phenotype of AhR-null mice, together with the recent discovery of a variety of endogenous and plant-derived ligands, point to the integral roles of AhR in normal cell physiology, in addition to its roles in sensing the environmental chemicals. Here, we summarize the current knowledge about AhR signaling pathways, its ligands and AhR-mediated effects on cell specialization, host defense and detoxification. AhR-mediated health effects particularly in liver, immune, and nervous systems, as well as in tumorgenesis are discussed. Dioxin-initiated embryotoxicity and immunosuppressive effects in fish and birds are reviewed. Recent data demonstrate that AhR is a convergence point of multiple signaling pathways that inform the cell of its external and internal environments. As such, AhR pathway is a promising potential target for therapeutics targeting nervous, liver, and autoimmune diseases through AhR ligand-mediated interventions and other perturbations of AhR signaling. Additionally, using available laboratory data obtained on animal models, AhR-centered adverse outcome pathway analysis is useful in reexamining known and potential adverse outcomes of specific or mixed compounds on wildlife. PMID:26079192

  10. The Dioxin Receptor Regulates the Constitutive Expression of the Vav3 Proto-Oncogene and Modulates Cell Shape and Adhesion

    PubMed Central

    Carvajal-Gonzalez, Jose M.; Mulero-Navarro, Sonia; Roman, Angel Carlos; Sauzeau, Vincent; Merino, Jaime M.; Bustelo, Xose R.

    2009-01-01

    The dioxin receptor (AhR) modulates cell plasticity and migration, although the signaling involved remains unknown. Here, we report a mechanism that integrates AhR into these cytoskeleton-related functions. Immortalized and mouse embryonic fibroblasts lacking AhR (AhR−/−) had increased cell area due to spread cytoplasms that reverted to wild-type morphology upon AhR re-expression. The AhR-null phenotype included increased F-actin stress fibers, depolarized focal adhesions, and enhanced spreading and adhesion. The cytoskeleton alterations of AhR−/− cells were due to down-regulation of constitutive Vav3 expression, a guanosine diphosphate/guanosine triphosphate exchange factor for Rho/Rac GTPases and a novel transcriptional target of AhR. AhR was recruited to the vav3 promoter and maintained constitutive mRNA expression in a ligand-independent manner. Consistently, AhR−/− fibroblasts had reduced Rac1 activity and increased activation of the RhoA/Rho kinase (Rock) pathway. Pharmacological inhibition of Rac1 shifted AhR+/+ fibroblasts to the null phenotype, whereas Rock inhibition changed AhR-null cells to the AhR+/+ morphology. Knockdown of vav3 transcripts by small interfering RNA induced cytoskeleton defects and changes in adhesion and spreading mimicking those of AhR-null cells. Moreover, vav3−/− MEFs, as AhR−/− mouse embryonic fibroblasts, had increased cell area and enhanced stress fibers. By modulating Vav3-dependent signaling, AhR could regulate cell shape, adhesion, and migration under physiological conditions and, perhaps, in certain pathological states. PMID:19158396

  11. Regulatory crosstalk and interference between the xenobiotic and hypoxia sensing pathways at the AhR-ARNT-HIF1α signaling node

    PubMed Central

    Vorrink, Sabine U.; Domann, Frederick E.

    2014-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates many of the responses to toxic environmental chemicals such as TCDD or dioxin-like PCBs. To regulate gene expression, the AhR requires its binding partner, the aryl hydrocarbon receptor nuclear translocator (ARNT). ARNT is also required by the hypoxia-inducible factor-1α (HIF-1α), a crucial regulator of responses to conditions of reduced oxygen. The important role of ARNT in both the AhR and HIF-1α signaling pathways establishes a meaningful foundation for a possible crosstalk between these two vitally important signaling pathways. This crosstalk might lead to interference between the two signaling pathways and thus might play a role in the variety of cellular responses after exposure to AhR ligands and reduced oxygen availability. This review focuses on studies that have analyzed the effect of low oxygen environments and hypoxiamimetic agents on AhR signaling and conversely, the effect of AhR ligands, with a special emphasis on PCBs, on HIF-1α signaling. We highlight studies that assess the role of ARNT, elucidate the mechanism of the crosstalk, and discuss the physiological implications for exposure to AhR-inducing compounds in the context of hypoxia. PMID:24824450

  12. The aryl hydrocarbon receptor regulates an essential transcriptional element in the immunoglobulin heavy chain gene.

    PubMed

    Wourms, Michael J; Sulentic, Courtney E W

    2015-05-01

    Ig heavy chain (Igh) transcription involves several regulatory elements including the 3'Igh regulatory region (3'IghRR). 3'IghRR activity is modulated by several transcription factors, including NF-κB and AP-1 and potentially the aryl hydrocarbon receptor (AhR). The prototypical AhR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) inhibits antibody secretion and 3'IghRR activity. However, the exact mechanism is unknown and TCDD can modulate NF-κB and AP-1 in an AhR-independent manner. To determine if the AhR is a significant regulator of the 3'IghRR, we utilized a mouse B-cell line that stably expresses a 3'IghRR-regulated transgene and either an AhR antagonist or shRNA targeting the AhR. Disruption of the AhR pathway reversed TCDD-induced suppression of the 3'IghRR-regulated transgene and of endogenous Ig demonstrating a biologically significant effect of the AhR on 3'IghRR activation. Altered human 3'IGHRR activity by AhR ligands, which include dietary, environmental, and pharmaceutical chemicals, may have significant implications to human diseases previously associated with the 3'IGHRR.

  13. Zebrafish Cardiotoxicity: The Effects of CYP1A Inhibition and AHR2 Knockdown Following Exposure to Weak Aryl Hydrocarbon Receptor Agonists

    PubMed Central

    Clark, Bryan William; Van Tiem Garner, Lindsey; Di Giulio, Richard Thomas

    2014-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that mediates many of the toxic effects of dioxin-like compounds (DLCs) and some polycyclic aromatic hydrocarbons (PAHs). Strong AHR agonists, such as certain polychlorinated biphenyls and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), cause severe cardiac teratogenesis in fish embryos. Moderately strong AHR agonists, such as benzo[a]pyrene and β-naphthoflavone, have been shown to cause similar cardiotoxic effects when coupled with a cytochrome P450 1A (CYP1A) inhibitor, such as fluoranthene (FL). We sought to determine if weak AHR agonists, when combined with a CYP1A inhibitor (FL) or CYP1A morpholino gene knockdown, are capable of causing cardiac deformities similar to moderately strong AHR agonists (Wassenberg and Di Giulio 2004; Wassenberg and Di Giulio 2004; Billiard, Timme-Laragy et al. 2006; Van Tiem and Di Giulio 2011). The weak AHR agonists included the following: carbaryl, phenanthrene, 2-methylindole, 3-methylindole, indigo, and indirubin. The results showed a complex pattern of cardiotoxic response to weak agonist inhibitor exposure and morpholino-knockdown. Danio rerio (zebrafish) embryos were first exposed to weak AHR agonists at equimolar concentrations. The agonists were assessed for their relative potency as inducers of CYP1 enzyme activity, measured by the ethoxyresorufin-o-deethylase (EROD) assay, and cardiac deformities. Carbaryl, 2-methylindole, and 3-methylindole induced the highest CYP1A activity in zebrafish. Experiments were then conducted to determine the individual cardiotoxicity of each compound. Next, zebrafish were co-exposed to each agonist (at concentrations below those determined to be cardiotoxic) and FL in combination to assess if CYP1A inhibition could induce cardiac deformities. Carbaryl, 2-methylindole, 3-methylindole, and phenanthrene significantly increased pericardial edema relative to controls when combined with FL. To further evaluate the

  14. Common Commercial and Consumer Products Contain Activators of the Aryl Hydrocarbon (Dioxin) Receptor

    PubMed Central

    Zhao, Bin; Bohonowych, Jessica E. S.; Timme-Laragy, Alicia; Jung, Dawoon; Affatato, Alessandra A.; Rice, Robert H.; Di Giulio, Richard T.; Denison, Michael S.

    2013-01-01

    Activation of the Ah receptor (AhR) by halogenated aromatic hydrocarbons (HAHs), such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin), can produce a wide variety of toxic and biological effects. While recent studies have shown that the AhR can bind and be activated by structurally diverse chemicals, how widespread of these AhR agonists are in environmental, biological and synthetic materials remains to be determined. Using AhR-based assays, we demonstrate the presence of potent AhR agonists in a variety of common commercial and consumer items. Solvent extracts of paper, rubber and plastic products contain chemicals that can bind to and stimulate AhR DNA binding and/or AhR-dependent gene expression in hepatic cytosol, cultured cell lines, human epidermis and zebrafish embryos. In contrast to TCDD and other persistent dioxin-like HAHs, activation of AhR-dependent gene expression by these extracts was transient, suggesting that the agonists are metabolically labile. Solvent extracts of rubber products produce AhR-dependent developmental toxicity in zebrafish in vivo, and inhibition of expression of the metabolic enzyme CYP1A, significantly increased their toxic potency. Although the identity of the responsible AhR-active chemicals and their toxicological impact remain to be determined, our data demonstrate that AhR active chemicals are widely distributed in everyday products. PMID:23441220

  15. The Aryl Hydrocarbon Receptor Modulates Production of Cytokines and Reactive Oxygen Species and Development of Myocarditis during Trypanosoma cruzi Infection.

    PubMed

    Barroso, Andréia; Gualdrón-López, Melisa; Esper, Lísia; Brant, Fátima; Araújo, Ronan R S; Carneiro, Matheus B H; Ávila, Thiago V; Souza, Danielle G; Vieira, Leda Q; Rachid, Milene A; Tanowitz, Herbert B; Teixeira, Mauro M; Machado, Fabiana S

    2016-10-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor involved in controlling several aspects of immune responses, including the activation and differentiation of specific T cell subsets and antigen-presenting cells, thought to be relevant in the context of experimental Trypanosoma cruzi infection. The relevance of AhR for the outcome of T. cruzi infection is not known and was investigated here. We infected wild-type (WT) mice and AhR knockout (AhR KO) mice with T. cruzi (Y strain) and determined levels of parasitemia, myocardial inflammation and fibrosis, expression of AhR/cytokines/suppressor of cytokine signaling (SOCS) (spleen/heart), and production of nitric oxide (NO), reactive oxygen species (ROS), and peroxynitrite (ONOO(-)) (spleen). AhR expression was increased in the heart of infected WT mice. Infected AhR KO mice displayed significantly reduced parasitemia, inflammation, and fibrosis of the myocardium. This was associated with an anticipated increased immune response characterized by increased levels of inflammatory cytokines and reduced expression of SOCS2 and SOCS3 in the heart. In vitro, AhR deficiency caused impairment in parasite replication and decreased levels of ROS production. In conclusion, AhR influences the development of murine Chagas disease by modulating ROS production and regulating the expression of key physiological regulators of inflammation, SOCS1 to -3, associated with the production of cytokines during experimental T. cruzi infection.

  16. Leflunomide Induces Pulmonary and Hepatic CYP1A Enzymes via Aryl Hydrocarbon Receptor.

    PubMed

    Patel, Ananddeep; Zhang, Shaojie; Paramahamsa, Maturu; Jiang, Weiwu; Wang, Lihua; Moorthy, Bhagavatula; Shivanna, Binoy

    2015-12-01

    Emerging evidence indicates that the aryl hydrocarbon receptor (AhR) plays a crucial role in normal physiologic homeostasis. Additionally, aberrant AhR signaling leads to several pathologic states in the lung and liver. Activation of AhR transcriptionally induces phase I (CYP1A) detoxifying enzymes. Although the effects of the classic AhR ligands such as 3-methylcholanthrene and dioxins on phase 1 enzymes are well studied in rodent lung, liver, and other organs, the toxicity profiles limit their use as therapeutic agents in humans. Hence, there is a need to identify and investigate nontoxic AhR ligands not only to understand the AhR biology but also to develop the AhR as a clinically relevant therapeutic target. Leflunomide is a Food and Drug Administration-approved drug in humans that is known to have AhR agonist activity in vitro. Whether it activates AhR and induces phase 1 enzymes in vivo is unknown. Therefore, we tested the hypothesis that leflunomide will induce pulmonary and hepatic CYP1A enzymes in C57BL/6J wild-type mice, but not in AhR-null mice. We performed real-time reverse-transcription polymerase chain reaction analyses for CYP1A1/2 mRNA expression, western blot assays for CYP1A1/2 protein expression, and ethoxyresorufinO-deethylase assay for CYP1A1 catalytic activity. Leflunomide increased CYP1A1/A2 mRNA, protein, and enzymatic activities in wild-type mice. In contrast, leflunomide failed to increase pulmonary and hepatic CYP1A enzymes in AhR-null mice. In conclusion, we provide evidence that leflunomide induces pulmonary and hepatic CYP1A enzymes via the AhR.

  17. Aryl-hydrocarbon receptor activity modulates prolactin expression in the pituitary.

    PubMed

    Moran, Tyler B; Brannick, Katherine E; Raetzman, Lori T

    2012-11-15

    Pituitary tumors account for 15% of intracranial neoplasms, however the extent to which environmental toxicants contribute to the proliferation and hormone expression of pituitary cells is unknown. Aryl-hydrocarbon receptor (AhR) interacting protein (AIP) loss of function mutations cause somatotrope and lactotrope adenomas in humans. AIP sequesters AhR and inhibits its transcriptional function. Because of the link between AIP and pituitary tumors, we hypothesize that exposure to dioxins, potent exogenous ligands for AhR that are persistent in the environment, may predispose to pituitary dysfunction through activation of AhR. In the present study, we examined the effect of AhR activation on proliferation and endogenous pituitary hormone expression in the GH3 rat somatolactotrope tumor cell line and the effect of loss of AhR action in knockout mice. GH3 cells respond to nM doses of the reversible AhR agonist β-naphthoflavone with a robust induction of Cyp1a1. Although mRNA levels of the anti-proliferative signaling cytokine TGFbeta1 are suppressed upon β-naphthoflavone treatment, we did not observe an alteration in cell proliferation. AhR activation with β-naphthoflavone suppresses Ahr expression and impairs expression of prolactin (PRL), but not growth hormone (GH) mRNA in GH3 cells. In mice, loss of Ahr similarly leads to a reduction in Prl mRNA at P3, while Gh is unaffected. Additionally, there is a significant reduction in pituitary hormones Lhb and Fshb in the absence of Ahr. Overall, these results demonstrate that AhR is important for pituitary hormone expression and suggest that environmental dioxins can exert endocrine disrupting effects at the pituitary.

  18. An endogenous aryl hydrocarbon receptor ligand inhibits proliferation and migration of human ovarian cancer cells.

    PubMed

    Wang, Kai; Li, Yan; Jiang, Yi-Zhou; Dai, Cai-Feng; Patankar, Manish S; Song, Jia-Sheng; Zheng, Jing

    2013-10-28

    The aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor mediates many biological processes. Herein, we investigated if 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE, an endogenous AhR ligand) regulated proliferation and migration of human ovarian cancer cells via AhR. We found that AhR was widely present in many histotypes of ovarian cancer tissues. ITE suppressed OVCAR-3 cell proliferation and SKOV-3 cell migration in vitro, which were blocked by AhR knockdown. ITE also suppressed OVCAR-3 cell growth in mice. These data suggest that the ITE might potentially be used for therapeutic intervention for at least a subset of human ovarian cancer.

  19. The aryl hydrocarbon receptor in barrier organ physiology, immunology, and toxicology.

    PubMed

    Esser, Charlotte; Rannug, Agneta

    2015-01-01

    The aryl hydrocarbon receptor (AhR) is an evolutionarily old transcription factor belonging to the Per-ARNT-Sim-basic helix-loop-helix protein family. AhR translocates into the nucleus upon binding of various small molecules into the pocket of its single-ligand binding domain. AhR binding to both xenobiotic and endogenous ligands results in highly cell-specific transcriptome changes and in changes in cellular functions. We discuss here the role of AhR for immune cells of the barrier organs: skin, gut, and lung. Both adaptive and innate immune cells require AhR signaling at critical checkpoints. We also discuss the current two prevailing views-namely, 1) AhR as a promiscuous sensor for small chemicals and 2) a role for AhR as a balancing factor for cell differentiation and function, which is controlled by levels of endogenous high-affinity ligands. AhR signaling is considered a promising drug and preventive target, particularly for cancer, inflammatory, and autoimmune diseases. Therefore, understanding its biology is of great importance.

  20. Hypoxia perturbs aryl hydrocarbon receptor signaling and CYP1A1 expression induced by PCB 126 in human skin and liver-derived cell lines

    SciTech Connect

    Vorrink, Sabine U.; Severson, Paul L.; Kulak, Mikhail V.; Futscher, Bernard W.; Domann, Frederick E.

    2014-02-01

    The aryl hydrocarbon receptor (AhR) is an important mediator of toxic responses after exposure to xenobiotics including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and dioxin-like polychlorinated biphenyls (PCBs). Activation of AhR responsive genes requires AhR dimerization with the aryl hydrocarbon receptor nuclear translocator (ARNT), a heterodimeric partner also shared by the hypoxia-inducible factor-1α (HIF-1α) protein. TCDD-stimulated AhR transcriptional activity can be influenced by hypoxia; however, it less well known whether hypoxia interferes with AhR transcriptional transactivation in the context of PCB-mediated AhR activation in human cells. Elucidation of this interaction is important in liver hepatocytes which extensively metabolize ingested PCBs and experience varying degrees of oxygen tension during normal physiologic function. This study was designed to assess the effect of hypoxia on AhR transcriptional responses after exposure to 3,3′,4,4′,5-pentachlorobiphenyl (PCB 126). Exposure to 1% O{sub 2} prior to PCB 126 treatment significantly inhibited CYP1A1 mRNA and protein expression in human HepG2 and HaCaT cells. CYP1A1 transcriptional activation was significantly decreased upon PCB 126 stimulation under conditions of hypoxia. Additionally, hypoxia pre-treatment reduced PCB 126 induced AhR binding to CYP1 target gene promoters. Importantly, ARNT overexpression rescued cells from the inhibitory effect of hypoxia on XRE-luciferase reporter activity. Therefore, the mechanism of interference of the signaling crosstalk between the AhR and hypoxia pathways appears to be at least in part dependent on ARNT availability. Our results show that AhR activation and CYP1A1 expression induced by PCB 126 were significantly inhibited by hypoxia and hypoxia might therefore play an important role in PCB metabolism and toxicity. - Highlights: • Significant crosstalk exists between AhR and HIF-1α signaling. • Hypoxia perturbs PCB 126 induced AhR function and

  1. ARYL HYDROCARBON RECEPTOR DEFICIENCY PROTECTS MICE FROM DIET-INDUCED ADIPOSITY AND METABOLIC DISORDERS THROUGH INCREASED ENERGY EXPENDITURE

    PubMed Central

    Zhang, Zhi-Ming; Jaeger, Cassie D.; Krager, Stacey L.; Bottum, Kathleen M.; Liu, Jianghua; Liao, Duan-Fang; Tischkau, Shelley A.

    2015-01-01

    BACKGROUND/OBJECTIVES Epidemics of obesity and diabetes are escalating. High-calorie/high-fat food is a major cause for these global health issues, but molecular mechanisms underlying high-fat, diet-induced obesity are still not well understood. The aryl hydrocarbon receptor (AhR), a transcription factor that acts as a xenobiotic sensor, mediates environmental toxicant-induced obesity, insulin resistance and development of diabetes. AhR also influences lipid metabolism and diet-induced obesity. The effects of AhR deficiency on diet-induced obesity, hepatic steatosis and insulin resistance were examined. METHODS : Male wild type (WT), AhR null (AhR−/−) and AhR heterozygote (AhR+/−) mice were fed a normal chow diet (NCD, 10% kcal from fat) or a high-fat diet (HFD, 60% kcal from fat) for up to 14 weeks. Adiposity, adipose and liver morphology, insulin signaling, metabolic parameters and gene profiles were assessed. RESULTS AhR deficiency protected against HFD-induced obesity, hepatic steatosis, insulin resistance and inflammation. Moreover, AhR deficiency preserved insulin signaling in major metabolic tissues. These protective effects result from a higher energy expenditure in AhR-deficient mice compared to WT. Levels of transcript for both the thermogenic gene, uncoupling protein 1 (Ucp1), in brown adipose tissue and mitochondrial β-oxidation genes in muscle were significantly higher in AhR−/− and AhR+/− mice compared to WT. CONCLUSIONS This work documents a physiologically relevant function for AhR in regulation of body weight, hepatic fat deposition, insulin sensitivity and energy expenditure under HFD exposure, suggesting that AhR signaling may be developed as a potential therapeutic target for treatment of obesity and metabolic disorders. PMID:25907315

  2. Endogenous ligands of the aryl hydrocarbon receptor regulate lung dendritic cell function.

    PubMed

    Thatcher, Thomas H; Williams, Marc A; Pollock, Stephen J; McCarthy, Claire E; Lacy, Shannon H; Phipps, Richard P; Sime, Patricia J

    2016-01-01

    The aryl hydrocarbon receptor (AhR) is a transcription factor that has been extensively studied as a regulator of toxicant metabolism. However, recent evidence indicates that the AhR also plays an important role in immunity. We hypothesized that the AhR is a novel, immune regulator of T helper type 2 (Th2) -mediated allergic airway disease. Here, we report that AhR-deficient mice develop increased allergic responses to the model allergen ovalbumin (OVA), which are driven in part by increased dendritic cell (DC) functional activation. AhR knockout (AhR(-/-) ) mice sensitized and challenged with OVA develop an increased inflammatory response in the lung compared with wild-type controls, with greater numbers of inflammatory eosinophils and neutrophils, greater T-cell proliferation, greater production of Th2 cytokines, and higher levels of OVA-specific IgE and IgG1. Lung DCs from AhR(-/-) mice stimulated antigen-specific proliferation and Th2 cytokine production by naive T cells in vitro. Additionally, AhR(-/-) DCs produced higher levels of tumour necrosis factor-α and interleukin-6, which promote Th2 differentiation, and expressed higher cell surface levels of stimulatory MHC Class II and CD86 molecules. Overall, loss of the AhR was associated with enhanced T-cell activation by pulmonary DCs and heightened pro-inflammatory allergic responses. This suggests that endogenous AhR ligands are involved in the normal regulation of Th2-mediated immunity in the lung via a DC-dependent mechanism. Therefore, the AhR may represent an important target for therapeutic intervention in allergic airways inflammation.

  3. Differences in activation of aryl hydrocarbon receptors of white sturgeon relative to lake sturgeon are predicted by identities of key amino acids in the ligand binding domain.

    PubMed

    Doering, Jon A; Farmahin, Reza; Wiseman, Steve; Beitel, Shawn C; Kennedy, Sean W; Giesy, John P; Hecker, Markus

    2015-04-01

    Dioxin-like compounds (DLCs) are pollutants of global environmental concern. DLCs elicit their adverse outcomes through activation of the aryl hydrocarbon receptor (AhR). However, there is limited understanding of the mechanisms that result in differences in sensitivity to DLCs among different species of fishes. Understanding these mechanisms is critical for protection of the diversity of fishes exposed to DLCs, including endangered species. This study investigated specific mechanisms that drive responses of two endangered fishes, white sturgeon (Acipenser transmontanus) and lake sturgeon (Acipenser fulvescens) to DLCs. It determined whether differences in sensitivity to activation of AhRs (AhR1 and AhR2) can be predicted based on identities of key amino acids in the ligand binding domain (LBD). White sturgeon were 3- to 30-fold more sensitive than lake sturgeon to exposure to 5 different DLCs based on activation of AhR2. There were no differences in sensitivity between white sturgeon and lake sturgeon based on activation of AhR1. Adverse outcomes as a result of exposure to DLCs have been shown to be mediated through activation of AhR2, but not AhR1, in all fishes studied to date. This indicates that white sturgeon are likely to have greater sensitivity in vivo relative to lake sturgeon. Homology modeling and in silico mutagenesis suggests that differences in sensitivity to activation of AhR2 result from differences in key amino acids at position 388 in the LBD of AhR2 of white sturgeon (Ala-388) and lake sturgeon (Thr-388). This indicates that identities of key amino acids in the LBD of AhR2 could be predictive of both in vitro activation by DLCs and in vivo sensitivity to DLCs in these, and potentially other, fishes.

  4. The Aryl Hydrocarbon Receptor Complex and the Control of Gene Expression

    PubMed Central

    Beischlag, Timothy V.; Morales, J. Luis; Hollingshead, Brett D.; Perdew, Gary H.

    2008-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that controls the expression of a diverse set of genes. The toxicity of the potent AhR ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin is almost exclusively mediated through this receptor. However, the key alterations in gene expression that mediate toxicity are poorly understood. It has been established through characterization of AhR-null mice that the AhR has a required physiological function, yet how endogenous mediators regulate this orphan receptor remains to be established. A picture as to how the AhR/ARNT heterodimer actually mediates gene transcription is starting to emerge. The AhR/ARNT complex can alter transcription both by binding to its cognate response element and through tethering to other transcription factors. In addition, many of the coregulatory proteins necessary for AhR-mediated transcription have been identified. Cross talk between the estrogen receptor and the AhR at the promoter of target genes appears to be an important mode of regulation. Inflammatory signaling pathways and the AhR also appear to be another important site of cross talk at the level of transcription. A major focus of this review is to highlight experimental efforts to characterize nonclassical mechanisms of AhR-mediated modulation of gene transcription. PMID:18540824

  5. Role of the Aryl Hydrocarbon Receptor in Colon Neoplasia

    PubMed Central

    Xie, Guofeng; Raufman, Jean-Pierre

    2015-01-01

    For both men and women, colorectal cancer (CRC) is the second leading cause of cancer death in the United States, primarily as a consequence of limited therapies for metastatic disease. The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor with diverse functions in detoxification of xenobiotics, inflammatory responses, and tissue homeostasis. Emerging evidence indicates that AhR also plays an important role in regulating intestinal cell proliferation and tumorigenesis. Here, we review both the pro- and anti-carcinogenic properties of AhR signaling and its potential role as a therapeutic target in CRC. PMID:26264025

  6. Aryl‐hydrocarbon receptor activity modulates prolactin expression in the pituitary

    SciTech Connect

    Moran, Tyler B.; Brannick, Katherine E.; Raetzman, Lori T.

    2012-11-15

    Pituitary tumors account for 15% of intracranial neoplasms, however the extent to which environmental toxicants contribute to the proliferation and hormone expression of pituitary cells is unknown. Aryl-hydrocarbon receptor (AhR) interacting protein (AIP) loss of function mutations cause somatotrope and lactotrope adenomas in humans. AIP sequesters AhR and inhibits its transcriptional function. Because of the link between AIP and pituitary tumors, we hypothesize that exposure to dioxins, potent exogenous ligands for AhR that are persistent in the environment, may predispose to pituitary dysfunction through activation of AhR. In the present study, we examined the effect of AhR activation on proliferation and endogenous pituitary hormone expression in the GH3 rat somatolactotrope tumor cell line and the effect of loss of AhR action in knockout mice. GH3 cells respond to nM doses of the reversible AhR agonist β-naphthoflavone with a robust induction of Cyp1a1. Although mRNA levels of the anti-proliferative signaling cytokine TGFbeta1 are suppressed upon β-naphthoflavone treatment, we did not observe an alteration in cell proliferation. AhR activation with β-naphthoflavone suppresses Ahr expression and impairs expression of prolactin (PRL), but not growth hormone (GH) mRNA in GH3 cells. In mice, loss of Ahr similarly leads to a reduction in Prl mRNA at P3, while Gh is unaffected. Additionally, there is a significant reduction in pituitary hormones Lhb and Fshb in the absence of Ahr. Overall, these results demonstrate that AhR is important for pituitary hormone expression and suggest that environmental dioxins can exert endocrine disrupting effects at the pituitary. -- Highlights: ► AhR signaling suppresses Prl mRNA expression. ► AhR signaling does not influence pituitary proliferation in culture. ► AhR is necessary for Prl, Lhb and Fshb expression at postnatal day 3.

  7. Studies on the role of the Ah receptor in hexachloro-benzene-induced porphyria

    SciTech Connect

    Hahn, M.E.

    1987-01-01

    Many of the effects of hexachlorobenzene (HCB) resemble those of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), whose effects are initiated by its binding to the AH receptor, the regulatory gene product of the Ah locus. I investigated the ability of HCB to interact with the AH receptor and the involvement of this protein in HCB-induced porphyria. The induction of two cytochrome P450 isozymes regulated by the Ah locus was also examined in light of their possible role in the pathogenesis of HCB- and TCDD-induced porphyria. HCB competitively inhibited the in vitro specific binding of ({sup 3}H)-TCDD to the rat hepatic Ah receptor (K{sub I} = 2.1 {mu}M) without affecting the solubility of ({sup 3}H)TCDD. Following the administration of HCB to rats, the number of ({sup 3}H)TCDD specific binding sites was reduced by up to 40%. HCB induced cytochromes P450b, P450e, P450c, and P450d, confirming that it is a mixed-type P450 inducer. The presence of porphyria in mice was assessed by measuring urinary and hepatic porphyrins and hepatic uroporphyrinogen decarboxylase activity.

  8. Responses of mixtures of polyhalogenated aromatic compounds or single compounds in the CALUX-assay a novel species-specific bioassay for Ah-receptor active compounds

    SciTech Connect

    Murk, A.J.; Aarts, J.M.M.J.G.; Jonas, A.; Brouwer, A.; Denison, M.S.

    1995-12-31

    Polyhalogenated aromatic hydrocarbons (PHAHs) elicit a number of common toxic responses, including reproductive toxicity, teratogenicity, impairment of immune responses, alterations in vitamin A and thyroid hormone metabolism and carcinogenesis. The toxic effects however are highly dependent on the animal species used, The most toxic PHAHs are approximate isostereomeres of 2,3,7,8 tetrachlorinated dibenzo-p-dioxin (TCDD) and share a common mechanism of action mediated by the aryl hydrocarbon receptor (AhR). Based on the common receptor mediated mechanism, the toxic equivalency factor concept was developed, in which the potency of each individual congener is expressed relative to TCDD, thus allowing hazard and risk assessment for mixtures of PHAHs. A number of recombinant cell lines were developed, including hepalclc7 mouse and H4IIE rat hepatoma cell lines, with AhR-mediated firefly (Photinus pyralis) luciferase gene expression. The response in this so-called CALUX (chemical activated luciferase expression) assay is additive for polychlorinated dibenzofurans (PCDFs) and PCDDS, but for polychlorinated biphenyls (PCBs) both synergistic and antagonistic interactions have been demonstrated, which are partially species-dependent. Also some structurally related compounds, like polybrominated diphenyl ether, pentachlorinated phenol, benzo(a)pyrene, pyrene, tetrachlorobenzyltoluene (Ugilec 141) and mixtures of polychlorinated terphenyls have been tested in the CALUX assay. The responses of these compounds were sometimes agonistic, but also antagonistic and synergistic effects on the TCDO response were observed.

  9. Beta-naphthoflavone causes an AhR-independent inhibition of invasion and intracellular multiplication of Listeria monocytogenes in murine hepatocytes

    PubMed Central

    Shi, Lewis Zhichang; Czuprynski, Charles J.

    2009-01-01

    We recently reported a heretofore unknown role for the aryl hydrocarbon receptor in host resistance to listeriosis in mice. Hepatocytes are an important site for Listeria monocytogenes multiplication in vivo. In this study, we investigated whether activation of AhR in TIB73 murine embryonic hepatocytes affects the ingestion and intracellular multiplication of L. monocytogenes. Treatment of TIB73 cells with the AhR agonist β-naphthoflavone (BNF) significantly inhibited the ingestion and intracellular growth of L. monocytogenes. The inhibitory effects of BNF were dose-dependent and correlated with up-regulation of CYP1A1. Surprisingly, pretreatment with AhR antagonists (3′-MNF or α-naphthoflavone) or knocking-down of AhR with siRNA did not abolish the inhibitory effects of BNF. Moreover, the inhibitory effects of BNF on invasion and intracellular growth of L. monocytogenes by BNF were observed in AhR-deficient (CRL-2710), or ARNT-dysfunctional (CRL-2717) Hepa cells. We also observed similar inhibitory effects of BNF treatment using primary hepatocytes recovered from AhR+/- or AhR-/- mice. Moreover, the prototypic AhR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) did not inhibit the invasion and intracellular growth of L. monocytogenes in TIB73 cells. Mechanistic studies demonstrated that ROS, but not TNF-α or iNOS, plays an important role in mediating BNF-induced inhibition. In conclusion, BNF caused an AhR-independent inhibition of ingestion and intracellular multiplication of L. monocytogenes in murine hepatocytes, mediated in part by production of ROS. PMID:19715752

  10. Aryl hydrocarbon receptor-driven signals inhibit collagen synthesis in the gut.

    PubMed

    Monteleone, Ivan; Zorzi, Francesca; Marafini, Irene; Di Fusco, Davide; Dinallo, Vincenzo; Caruso, Roberta; Izzo, Roberta; Franzè, Eleonora; Colantoni, Alfredo; Pallone, Francesco; Monteleone, Giovanni

    2016-04-01

    Fibrostrictures (FS) are a major complication of Crohn's disease (CD). Pathogenesis of FS is not fully understood, but activation of fibroblasts and excessive collagen deposition are crucial in the development of FS. Here, we investigated the role of aryl hydrocarbon receptor (AhR) in intestinal fibrosis. AhR RNA and protein expression were evaluated in intestinal fibroblasts of CD patients and controls. CD fibroblasts were stimulated with TGF-β1 or TNF-α in the presence or absence of the AhR activator Ficz, an AhR antagonist CH223191, or a specific AhR-silencing RNA. In CD fibroblasts, TGF-β1 and TNF-α increased Col1A1, Col3A1 and α-SMA transcripts and collagen secretion and this effect was reduced by Ficz and upregulated by CH22319. TGF-β1 or TNF-α induced activation of p38 and ERK1/2 MAP kinases was decreased by Ficz and increased by CH223191. The inhibitory effect of Ficz on Map kinase activation and collagen induction was abolished by AhR silencing. To assess the role of AhR in vivo, mice with trinitrobenzene-sulfonic-acid induced colonic fibrosis were given Ficz or CH223191. Mice given either Ficz or CH223191 produced less or more collagen respectively as compared with control mice. Our results indicate that AhR is a negative regulator of profibrotic signals in the gut. PMID:26786786

  11. Laquinimod arrests experimental autoimmune encephalomyelitis by activating the aryl hydrocarbon receptor

    PubMed Central

    Kaye, Joel; Piryatinsky, Victor; Birnberg, Tal; Hingaly, Tal; Raymond, Emanuel; Kashi, Rina; Amit-Romach, Einat; Caballero, Ignacio S.; Towfic, Fadi; Ator, Mark A.; Rubinstein, Efrat; Laifenfeld, Daphna; Orbach, Aric; Shinar, Doron; Marantz, Yael; Grossman, Iris; Knappertz, Volker; Hayden, Michael R.; Laufer, Ralph

    2016-01-01

    Laquinimod is an oral drug currently being evaluated for the treatment of relapsing, remitting, and primary progressive multiple sclerosis and Huntington’s disease. Laquinimod exerts beneficial activities on both the peripheral immune system and the CNS with distinctive changes in CNS resident cell populations, especially astrocytes and microglia. Analysis of genome-wide expression data revealed activation of the aryl hydrocarbon receptor (AhR) pathway in laquinimod-treated mice. The AhR pathway modulates the differentiation and function of several cell populations, many of which play an important role in neuroinflammation. We therefore tested the consequences of AhR activation in myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE) using AhR knockout mice. We demonstrate that the pronounced effect of laquinimod on clinical score, CNS inflammation, and demyelination in EAE was abolished in AhR−/− mice. Furthermore, using bone marrow chimeras we show that deletion of AhR in the immune system fully abrogates, whereas deletion within the CNS partially abrogates the effect of laquinimod in EAE. These data strongly support the idea that AhR is necessary for the efficacy of laquinimod in EAE and that laquinimod may represent a first-in-class drug targeting AhR for the treatment of multiple sclerosis and other neurodegenerative diseases. PMID:27671624

  12. Zinc finger transcription factor Slug is a novel target gene of aryl hydrocarbon receptor

    SciTech Connect

    Ikuta, Togo; Kawajiri, Kaname . E-mail: kawajiri@cancer-c.pref.saitama.jp

    2006-11-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor. We previously showed that AhR localizes predominantly in the cytoplasm under high cell densities of a keratinocytes cell line, HaCaT, but accumulates in the nucleus at low cell densities. In the current report, we show that the Slug, which is a member of the snail/slug family of zinc finger transcriptional repressors critical for induction of epithelial-mesenchymal transitions (EMT), is activated transcriptionally in accordance with nuclear accumulation of AhR. By reporter assay of the promoter of the Slug gene, gel shift and chromatin immunoprecipitation analyses showed AhR directly binds to xenobiotic responsive element 5 at - 0.7 kb of the gene. AhR-targeted gene silencing by small interfering RNA duplexes led to the abolishment of not only CYP1A1 but also Slug induction by 3-methycholanthrene. The Slug was co-localized to the AhR at the wound margins of HaCaT cells, where apparent nuclear distribution of AhR and Slug was observed. The induced Slug was associated with reduction of an epithelial marker of cytokeratin-18 and with an increase in the mesenchymal marker, fibronectin. Taken together, these findings suggest that AhR participated in Slug induction, which, in turn, regulates cellular physiology including cell adhesion and migration.

  13. Hypoxia perturbs aryl hydrocarbon receptor signaling and CYP1A1 expression induced by PCB 126 in human skin and liver-derived cell lines

    PubMed Central

    Vorrink, Sabine U.; Severson, Paul L.; Kulak, Mikhail V.; Futscher, Bernard W.; Domann, Frederick E.

    2014-01-01

    The aryl hydrocarbon receptor (AhR) is an important mediator of toxic responses after exposure to xenobiotics including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and dioxin-like polychlorinated biphenyls (PCBs). Activation of AhR responsive genes requires AhR dimerization with the aryl hydrocarbon receptor nuclear translocator (ARNT), a heterodimeric partner also shared by the hypoxia-inducible factor-1α (HIF-1α) protein. TCDD-stimulated AhR transcriptional activity can be influenced by hypoxia; however, it less well known whether hypoxia interferes with AhR transcriptional transactivation in the context of PCB-mediated AhR activation in human cells. Elucidation of this interaction is important in liver hepatocytes which extensively metabolize ingested PCBs and experience varying degrees of oxygen tension during normal physiologic function. This study was designed to assess the effect of hypoxia on AhR transcriptional responses after exposure to 3,3',4,4',5-pentachlorobiphenyl (PCB 126). Exposure to 1% O2 prior to PCB 126 treatment significantly inhibited CYP1A1 mRNA and protein expression in human HepG2 and HaCaT cells. CYP1A1 transcriptional activation was significantly decreased upon PCB 126 stimulation under conditions of hypoxia. Additionally, hypoxia pre-treatment reduced PCB 126 induced AhR binding to CYP1 target gene promoters. Importantly, ARNT overexpression rescued cells from the inhibitory effect of hypoxia on XRE-luciferase reporter activity. Therefore, the mechanism of interference of the signaling crosstalk between the AhR and hypoxia pathways appears to be at least in part dependent on ARNT availability. Our results show that AhR activation and CYP1A1 expression induced by PCB 126 were significantly inhibited by hypoxia and hypoxia might therefore play an important role in PCB metabolism and toxicity. PMID:24355420

  14. Interaction of aryl hydrocarbon receptor and NF-κB subunit RelB in breast cancer is associated with interleukin-8 overexpression.

    PubMed

    Vogel, Christoph Franz Adam; Li, Wen; Wu, Dalei; Miller, Jamie K; Sweeney, Colleen; Lazennec, Gwendal; Fujisawa, Yasuko; Matsumura, Fumio

    2011-08-01

    The aryl hydrocarbon receptor (AhR) has been best known for its role in mediating the toxicity of dioxin. Here we show that AhR overexpression is found among estrogen receptor (ER)α-negative human breast tumors and that its overexpression is positively correlated to that of the NF-κB subunit RelB and Interleukin (IL)-8. Increased DNA binding activity of the AhR and RelB is coupled to IL-8 overexpression in primary breast cancer tissue, which was also supported by in situ hybridization. Activation of AhR in vitro by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induced IL-8 expression in MDA-MB 436 and MCF-7 cells in an AhR and RelB dependent manner. Consistently, downregulation of RelB or AhR by small interfering RNAs (siRNA) decreased the level of IL-8 but increased expression of ERα in vitro in MCF-7 cells. Our results strongly suggest that RelB and AhR have a critical role in the regulation of IL-8 and reveal a supportive role of RelB and AhR in the anti-apoptotic response in human breast cancer cells. AhR and RelB may present a novel therapeutic target for inflammatory driven breast carcinogenesis and tumor progression. Overexpression of pro-survival factors AhR and RelB may explain the process of the development of environmentally-induced type of breast cancers.

  15. Aryl hydrocarbon receptor-dependent regulation of miR-196a expression controls lung fibroblast apoptosis but not proliferation

    SciTech Connect

    Hecht, Emelia; Zago, Michela; Sarill, Miles; Rico de Souza, Angela; Gomez, Alvin; Matthews, Jason; Hamid, Qutayba; Eidelman, David H.; Baglole, Carolyn J.

    2014-11-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor implicated in the regulation of apoptosis and proliferation. Although activation of the AhR by xenobiotics such as dioxin inhibits the cell cycle and control apoptosis, paradoxically, AhR expression also promotes cell proliferation and survival independent of exogenous ligands. The microRNA (miRNA) miR-196a has also emerged as a regulator of proliferation and apoptosis but a relationship between the AhR and miR-196a is not known. Therefore, we hypothesized that AhR-dependent regulation of endogenous miR-196a expression would promote cell survival and proliferation. Utilizing lung fibroblasts from AhR deficient (AhR{sup −/−}) and wild-type (AhR{sup +/+}) mice, we show that there is ligand-independent regulation of miRNA, including low miR-196a in AhR{sup −/−} cells. Validation by qRT-PCR revealed a significant decrease in basal expression of miR-196a in AhR{sup −/−} compared to AhR{sup +/+} cells. Exposure to AhR agonists benzo[a]pyrene (B[a]P) and FICZ as well as AhR antagonist CH-223191 decreased miR-196a expression in AhR{sup +/+} fibroblasts concomitant with decreased AhR protein levels. There was increased proliferation only in AhR{sup +/+} lung fibroblasts in response to serum, corresponding to a decrease in p27{sup KIP1} protein, a cyclin-dependent kinase inhibitor. Increasing the cellular levels of miR-196a had no effect on proliferation or expression of p27{sup KIP1} in AhR{sup −/−} fibroblasts but attenuated cigarette smoke-induced apoptosis. This study provides the first evidence that AhR expression is essential for the physiological regulation of cellular miRNA levels- including miR-196a. Future experiments designed to elucidate the functional relationship between the AhR and miR-196a may delineate additional novel ligand-independent roles for the AhR. - Highlights: • The AhR controls proliferation and apoptosis in lung cells. • The AhR regulates the

  16. The aryl hydrocarbon receptor mediates raloxifene-induced apoptosis in estrogen receptor-negative hepatoma and breast cancer cells

    PubMed Central

    O'Donnell, E F; Koch, D C; Bisson, W H; Jang, H S; Kolluri, S K

    2014-01-01

    Identification of new molecular targets for the treatment of breast cancer is an important clinical goal, especially for triple-negative breast cancer, which is refractory to existing targeted treatments. The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor known primarily as the mediator of dioxin toxicity. However, the AhR can also inhibit cellular proliferation in a ligand-dependent manner and act as a tumor suppressor in mice, and thus may be a potential anticancer target. To investigate the AhR as an anticancer target, we conducted a small molecule screen to discover novel AhR ligands with anticancer properties. We identified raloxifene, a selective estrogen receptor (ER) modulator currently used in the clinic for prevention of ER-positive breast cancer and osteoporosis in post-menopausal women, as an AhR activator. Raloxifene directly bound the AhR and induced apoptosis in ER-negative mouse and human hepatoma cells in an AhR-dependent manner, indicating that the AhR is a molecular target of raloxifene and mediates raloxifene-induced apoptosis in the absence of ER. Raloxifene selectively induced apoptosis of triple-negative MDA-MB-231 breast cancer cells compared with non-transformed mammary epithelial cells via the AhR. Combined with recent data showing that raloxifene inhibits triple-negative breast cancer xenografts in vivo (Int J Oncol. 43(3):785-92, 2013), our results support the possibility of repurposing of raloxifene as an AhR-targeted therapeutic for triple-negative breast cancer patients. To this end, we also evaluated the role of AhR expression on survival of patients diagnosed with breast cancer. We found that higher expression of the AhR is significantly associated with increased overall survival and distant metastasis-free survival in both hormone-dependent (ER-positive) and hormone-independent (ER and progesterone receptor (PR)-negative) breast cancers. Together, our data strongly support the possibility of using the AhR

  17. Activation of the aryl hydrocarbon receptor by carbaryl: Computational evidence of the ability of carbaryl to assume a planar conformation.

    PubMed

    Casado, Susana; Alonso, Mercedes; Herradón, Bernardo; Tarazona, José V; Navas, José

    2006-12-01

    It has been accepted that aryl hydrocarbon receptor (AhR) ligands are compounds with two or more aromatic rings in a coplanar conformation. Although general agreement exists that carbaryl is able to activate the AhR, it has been proposed that such activation could occur through alternative pathways without ligand binding. This idea was supported by studies showing a planar conformation of carbaryl as unlikely. The objective of the present work was to clarify the process of AhR activation by carbaryl. In rat H4IIE cells permanently transfected with a luciferase gene under the indirect control of AhR, incubation with carbaryl led to an increase of luminescence. Ligand binding to the AhR was studied by means of a cell-free in vitro system in which the activation of AhR can occur only by ligand binding. In this system, exposure to carbaryl also led to activation of AhR. These results were similar to those obtained with the AhR model ligand beta-naphthoflavone, although this compound exhibited higher potency than carbaryl in both assays. By means of computational modeling (molecular mechanics and quantum chemical calculations), the structural characteristics and electrostatic properties of carbaryl were described in detail, and it was observed that the substituent at C-1 and the naphthyl ring were not coplanar. Assuming that carbaryl would interact with the AhR through a hydrogen bond, this interaction was studied computationally using hydrogen fluoride as a model H-bond donor. Under this situation, the stabilization energy of the carbaryl molecule would permit it to adopt a planar conformation. These results are in accordance with the mechanism traditionally accepted for AhR activation: Binding of ligands in a planar conformation.

  18. Linking Ah receptor mediated effects of sediments and impacts on fish to key pollutants in the Yangtze Three Gorges Reservoir, China - A comprehensive perspective.

    PubMed

    Floehr, Tilman; Scholz-Starke, Björn; Xiao, Hongxia; Hercht, Hendrik; Wu, Lingling; Hou, Junli; Schmidt-Posthaus, Heike; Segner, Helmut; Kammann, Ulrike; Yuan, Xingzhong; Roß-Nickoll, Martina; Schäffer, Andreas; Hollert, Henner

    2015-12-15

    The Three Gorges Reservoir (TGR), created in consequence of the Yangtze River's impoundment by the Three Gorges Dam, faces numerous anthropogenic impacts that challenge its unique ecosystem. Organic pollutants, particularly aryl hydrocarbon receptor (AhR) agonists, have been widely detected in the Yangtze River, but only little research was yet done on AhR-mediated activities. Hence, in order to assess effects of organic pollution, with particular focus on AhR-mediated activities, several sites in the TGR area were examined applying the "triad approach". It combines chemical analysis, in vitro, in vivo and in situ investigations to a holistic assessment. Sediments and the benthic fish species Pelteobagrus vachellii were sampled in 2011/2012, respectively, to identify relevant endpoints. Sediment was tested in vitro with the ethoxyresorufin-O-deethylase (EROD) induction assay, and in vivo with the Fish Embryo Toxicity Test and Sediment Contact Assay with Danio rerio. Activities of phase I (EROD) and phase II (glutathione-S-transferase) biotransformation enzymes, pollutant metabolites and histopathological alterations were studied in situ in P. vachellii. EROD induction was tested in vitro and in situ to evaluate possible relationships. Two sites, near Chongqing and Kaixian city, were identified as regional hot-spots and further investigated in 2013. The sediments induced in the in vitro/in vivo bioassays AhR-mediated activities and embryotoxic/teratogenic effects - particularly on the cardiovascular system. These endpoints could be significantly correlated to each other and respective chemical data. However, particle-bound pollutants showed only low bioavailability. The in situ investigations suggested a rather poor condition of P. vachellii, with histopathological alterations in liver and excretory kidney. Fish from Chongqing city exhibited significant hepatic EROD induction and obvious parasitic infestations. The polycyclic aromatic hydrocarbon (PAH) metabolite 1

  19. Linking Ah receptor mediated effects of sediments and impacts on fish to key pollutants in the Yangtze Three Gorges Reservoir, China - A comprehensive perspective.

    PubMed

    Floehr, Tilman; Scholz-Starke, Björn; Xiao, Hongxia; Hercht, Hendrik; Wu, Lingling; Hou, Junli; Schmidt-Posthaus, Heike; Segner, Helmut; Kammann, Ulrike; Yuan, Xingzhong; Roß-Nickoll, Martina; Schäffer, Andreas; Hollert, Henner

    2015-12-15

    The Three Gorges Reservoir (TGR), created in consequence of the Yangtze River's impoundment by the Three Gorges Dam, faces numerous anthropogenic impacts that challenge its unique ecosystem. Organic pollutants, particularly aryl hydrocarbon receptor (AhR) agonists, have been widely detected in the Yangtze River, but only little research was yet done on AhR-mediated activities. Hence, in order to assess effects of organic pollution, with particular focus on AhR-mediated activities, several sites in the TGR area were examined applying the "triad approach". It combines chemical analysis, in vitro, in vivo and in situ investigations to a holistic assessment. Sediments and the benthic fish species Pelteobagrus vachellii were sampled in 2011/2012, respectively, to identify relevant endpoints. Sediment was tested in vitro with the ethoxyresorufin-O-deethylase (EROD) induction assay, and in vivo with the Fish Embryo Toxicity Test and Sediment Contact Assay with Danio rerio. Activities of phase I (EROD) and phase II (glutathione-S-transferase) biotransformation enzymes, pollutant metabolites and histopathological alterations were studied in situ in P. vachellii. EROD induction was tested in vitro and in situ to evaluate possible relationships. Two sites, near Chongqing and Kaixian city, were identified as regional hot-spots and further investigated in 2013. The sediments induced in the in vitro/in vivo bioassays AhR-mediated activities and embryotoxic/teratogenic effects - particularly on the cardiovascular system. These endpoints could be significantly correlated to each other and respective chemical data. However, particle-bound pollutants showed only low bioavailability. The in situ investigations suggested a rather poor condition of P. vachellii, with histopathological alterations in liver and excretory kidney. Fish from Chongqing city exhibited significant hepatic EROD induction and obvious parasitic infestations. The polycyclic aromatic hydrocarbon (PAH) metabolite 1

  20. Disruption of Ah Receptor Signaling during Mouse Development Leads to Abnormal Cardiac Structure and Function in the Adult

    PubMed Central

    Carreira, Vinicius S.; Fan, Yunxia; Kurita, Hisaka; Wang, Qin; Ko, Chia-I; Naticchioni, Mindi; Jiang, Min; Koch, Sheryl; Zhang, Xiang; Biesiada, Jacek; Medvedovic, Mario; Xia, Ying; Rubinstein, Jack; Puga, Alvaro

    2015-01-01

    The Developmental Origins of Health and Disease (DOHaD) Theory proposes that the environment encountered during fetal life and infancy permanently shapes tissue physiology and homeostasis such that damage resulting from maternal stress, poor nutrition or exposure to environmental agents may be at the heart of adult onset disease. Interference with endogenous developmental functions of the aryl hydrocarbon receptor (AHR), either by gene ablation or by exposure in utero to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a potent AHR ligand, causes structural, molecular and functional cardiac abnormalities and altered heart physiology in mouse embryos. To test if embryonic effects progress into an adult phenotype, we investigated whether Ahr ablation or TCDD exposure in utero resulted in cardiac abnormalities in adult mice long after removal of the agent. Ten-months old adult Ahr-/- and in utero TCDD-exposed Ahr+/+ mice showed sexually dimorphic abnormal cardiovascular phenotypes characterized by echocardiographic findings of hypertrophy, ventricular dilation and increased heart weight, resting heart rate and systolic and mean blood pressure, and decreased exercise tolerance. Underlying these effects, genes in signaling networks related to cardiac hypertrophy and mitochondrial function were differentially expressed. Cardiac dysfunction in mouse embryos resulting from AHR signaling disruption seems to progress into abnormal cardiac structure and function that predispose adults to cardiac disease, but while embryonic dysfunction is equally robust in males and females, the adult abnormalities are more prevalent in females, with the highest severity in Ahr-/- females. The findings reported here underscore the conclusion that AHR signaling in the developing heart is one potential target of environmental factors associated with cardiovascular disease. PMID:26555816

  1. Regulation of estrogen sulfotransferase expression by confluence of MCF10A breast epithelial cells: role of the aryl hydrocarbon receptor.

    PubMed

    Fu, Jiaqi; Fang, Hailin; Paulsen, Michelle; Ljungman, Mats; Kocarek, Thomas A; Runge-Morris, Melissa

    2011-11-01

    Estrogen sulfotransferase (SULT1E1) catalyzes the sulfonation of estrogens, which limits estrogen mitogenicity. We recently reported that SULT1E1 expression is low in preconfluent MCF10A human breast epithelial cells but increases when the cells become confluent. Pulse-chase labeling experiments with 5-bromouridine demonstrated that the confluence-mediated increase in SULT1E1 expression was due to increased mRNA synthesis. Because aryl hydrocarbon receptor (AhR) activation has been shown to suppress SULT1E1 expression and loss of cell-cell contact has been shown to activate the AhR in other cell types, we tested whether the confluence-associated changes in SULT1E1 expression were mediated by the AhR. Relative to confluent MCF10A cells, preconfluent cells had higher levels of CYP1A1 mRNA and greater activation of an AhR-responsive luciferase reporter, demonstrating that the AhR was active in the preconfluent cells. AhR and aryl hydrocarbon receptor nuclear translocator mRNA and protein levels were also higher in preconfluent than in confluent cultures. Treatment of preconfluent cells with the AhR antagonist, 3'-methoxy-4'-nitroflavone (MNF), or AhR knockdown significantly increased SULT1E1 expression. MCF10A cells stably transfected with a luciferase reporter containing ∼7 kilobases of the SULT1E1 5'-flanking region showed both MNF- and confluence-inducible luciferase expression. Preconfluent cells transiently transfected with the reporter showed both MNF treatment- and AhR knockdown-mediated luciferase induction, but mutation of a computationally predicted dioxin response element (DRE) at nucleotide (nt) -3476 did not attenuate these effects. These results demonstrate that SULT1E1 expression in MCF10A cells is transcriptionally regulated by confluence through a suppressive action of the AhR, which is not mediated through a DRE at nt -3476.

  2. Aryl hydrocarbon receptor protects lung adenocarcinoma cells against cigarette sidestream smoke particulates-induced oxidative stress

    SciTech Connect

    Cheng, Ya-Hsin; Huang, Su-Chin; Lin, Chun-Ju; Cheng, Li-Chuan; Li, Lih-Ann

    2012-03-15

    Environmental cigarette smoke has been suggested to promote lung adenocarcinoma progression through aryl hydrocarbon receptor (AhR)-signaled metabolism. However, whether AhR facilitates metabolic activation or detoxification in exposed adenocarcinoma cells remains ambiguous. To address this question, we have modified the expression level of AhR in two human lung adenocarcinoma cell lines and examined their response to an extract of cigarette sidestream smoke particulates (CSSP). We found that overexpression of AhR in the CL1-5 cell line reduced CSSP-induced ROS production and oxidative DNA damage, whereas knockdown of AhR expression increased ROS level in CSSP-exposed H1355 cells. Oxidative stress sensor Nrf2 and its target gene NQO1 were insensitive to AhR expression level and CSSP treatment in human lung adenocarcinoma cells. In contrast, induction of AhR expression concurrently increased mRNA expression of xenobiotic-metabolizing genes CYP1B1, UGT1A8, and UGT1A10 in a ligand-independent manner. It appeared that AhR accelerated xenobiotic clearing and diminished associated oxidative stress by coordinate regulation of a set of phase I and II metabolizing genes. However, the AhR-signaled protection could not shield cells from constant oxidative stress. Prolonged exposure to high concentrations of CSSP induced G0/G1 cell cycle arrest via the p53–p21–Rb1 signaling pathway. Despite no effect on DNA repair rate, AhR facilitated the recovery of cells from growth arrest when CSSP exposure ended. AhR-overexpressing lung adenocarcinoma cells exhibited an increased anchorage-dependent and independent proliferation when recovery from exposure. In summary, our data demonstrated that AhR protected lung adenocarcinoma cells against CSSP-induced oxidative stress and promoted post-exposure clonogenicity. -- Highlights: ► AhR expression level influences cigarette sidestream smoke-induced ROS production. ► AhR reduces oxidative stress by coordinate regulation of

  3. Rats fed soy protein isolate (SPI) have impaired hepatic CYP1A1 induction by polycyclic aromatic hydrocarbons as a result of interference with aryl hydrocarbon receptor signaling.

    PubMed

    Singhal, Rohit; Badger, Thomas M; Ronis, Martin J

    2008-03-01

    Consumption of soy diets has been found to reduce cancer incidence in animals and is associated with reduced cancer risk in humans. Previously, we have demonstrated that female Sprague-Dawley rats fed purified AIN-93G diets with soy protein isolate (SPI) as the sole protein source had reduced CYP1A1 induction and basal aryl hydrocarbon receptor (AhR) levels relative to those fed the same diet containing casein (CAS). In the present study, the molecular mechanisms underlying reduced AhR expression have been studied. The SPI-effect on AhR was not observed after feeding diets containing the purified soy isoflavones genistein or daidzein. Rat hepatoma FGC-4 cells were treated with the serum obtained from rats fed CAS- or SPI-containing diets. Reduced AhR levels (P<0.05) were observed after 24 h exposure to SPI-serum without any changes in the overall expression of chaperone proteins--HSP90 and XAP2. SPI-serum-stimulated AhR degradation was inhibited by treating the cells with the proteasome inhibitor, MG132, and was observed to be preceded by ubiquitination of the receptor. A reduced association of XAP2 with the immunoprecipitated AhR complex was observed. SPI-serum-mediated AhR degradation was preceded by nuclear translocation of the receptor. However, the translocated receptor was found to be unable to heterodimerize with ARNT or to bind to XRE elements on the CYP1A1 enhancer. These data suggest that feeding SPI-containing diets antagonizes AhR signaling by a novel mechanism which differs from those established for known AhR antagonists.

  4. Rats fed soy protein isolate (SPI) have impaired hepatic CYP1A1 induction by polycyclic aromatic hydrocarbons as a result of interference with aryl hydrocarbon receptor signaling

    SciTech Connect

    Singhal, Rohit; Badger, Thomas M.; Ronis, Martin J.

    2008-03-01

    Consumption of soy diets has been found to reduce cancer incidence in animals and is associated with reduced cancer risk in humans. Previously, we have demonstrated that female Sprague-Dawley rats fed purified AIN-93G diets with soy protein isolate (SPI) as the sole protein source had reduced CYP1A1 induction and basal aryl hydrocarbon receptor (AhR) levels relative to those fed the same diet containing casein (CAS). In the present study, the molecular mechanisms underlying reduced AhR expression have been studied. The SPI-effect on AhR was not observed after feeding diets containing the purified soy isoflavones genistein or daidzein. Rat hepatoma FGC-4 cells were treated with the serum obtained from rats fed CAS- or SPI-containing diets. Reduced AhR levels (P < 0.05) were observed after 24 h exposure to SPI-serum without any changes in the overall expression of chaperone proteins-HSP90 and XAP2. SPI-serum-stimulated AhR degradation was inhibited by treating the cells with the proteasome inhibitor, MG132, and was observed to be preceded by ubiquitination of the receptor. A reduced association of XAP2 with the immunoprecipitated AhR complex was observed. SPI-serum-mediated AhR degradation was preceded by nuclear translocation of the receptor. However, the translocated receptor was found to be unable to heterodimerize with ARNT or to bind to XRE elements on the CYP1A1 enhancer. These data suggest that feeding SPI-containing diets antagonizes AhR signaling by a novel mechanism which differs from those established for known AhR antagonists.

  5. Aryl hydrocarbon receptor expression and activity in cerebellar granule neuroblasts: implications for development and dioxin neurotoxicity.

    PubMed

    Williamson, Mary A; Gasiewicz, Thomas A; Opanashuk, Lisa A

    2005-02-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent teratogen that produces neurobehavioral abnormalities associated with both cognitive and locomotor systems, yet the precise regional and cellular targets of developmental neurotoxicity remain largely unknown. Most, if not all, TCDD-induced pathology is mediated via binding to the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor that belongs to the basic helix-loop-helix/Per-Arnt-Sim (bHLH/PAS) superfamily. Upon ligand binding, AhR translocates to the nucleus, dimerizes with the AhR nuclear translocator protein (Arnt), and regulates transcription by interaction with dioxin-response elements (DREs) in target genes, most notably specific cytochrome P450 (CYP) family members. To assess whether developing cerebellar granule neuroblasts are potential direct targets for TCDD toxicity, AhR expression and transcriptional activity were examined. AhR and Arnt proteins were present in mouse cerebellum from birth throughout postnatal development. AhR protein levels peaked between postnatal day (PND) 3-10, a critical period for granule neuroblast growth and maturation. Transcriptionally active AhR was detected in immature cerebellar granule cells in a transgenic dioxin-responsive lacZ mouse model after acute TCDD exposure. AhR and Arnt were also expressed in cerebellar granule neuroblast cultures. AhR localized to the nucleus in granule cells 15 min after TCDD treatment. TCCD elicited time-dependent and concentration-dependent increases in CYP1A1 and 1B1 mRNA and protein levels. Moreover, TCDD treatment reduced both thymidine incorporation and granule neuroblast survival in a concentration-dependent manner. These data suggest that (1) granule neuroblasts are direct targets for developmental AhR-mediated TCDD neurotoxicity and (2) TCDD exposure may disrupt granule cell neurogenesis.

  6. Detecting Polychlorinated Biphenyls by Ah Receptor and Fluorescence Quantitative PCR with Exonuclease

    NASA Astrophysics Data System (ADS)

    Zhao, Xiaoxiang; Zhuang, Huisheng

    2010-11-01

    Tetrachlorobiphenyls as ligands were cultivated with goldfish, Ah receptors were extracted from the liver of goldfish and purified by hydroxyapatite. The complex of TCB ligands-receptors were analyzed by Surface Plasmon Resonance. DNA probes were amplified by PCR using Primers F1 and F2 with the DNA recognition site of responsive enhancer. DNA probes bound to the complex were not digested by exonuclease. The DNA that bound to the complex was quantified by real time PCR. A standard curve with TCB concentration to Ct values was obtained in the range of 10-12mol/L to 10-8 mol/L, according to TCB concentration in samples. The detection limit of the assay was below 10-12mol/L of TCB. Compared with HPLC, this assay is much more sensitive. These results suggest that fluorescence quantitative PCR with exonuclease by Ah receptors fits for detection of trace PCB.

  7. Activation of aryl hydrocarbon receptor signaling by extracts of teak and other wood dusts.

    PubMed

    Wilson, Mark J; Sabbioni, Gabriele; Rando, Roy; Miller, Charles A

    2015-12-01

    Wood dusts, as a group, are categorized as known human carcinogens, but the risks of exposure to specific types of wood dusts and the carcinogenic chemicals they contain are not well studied. Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is linked to the carcinogenic action of specific classes of chemicals. Here we examined whether chemicals in various wood dusts had the potential to activate AhR signaling as a potential toxic mechanism of action. We found that methanol extracts of teak, walnut, mahogany, and poplar dusts contained a wide range of AhR ligand activity, whereas extracts of oak, pine, and other softwoods did not contain appreciable activity. Teak dust extract, being particularly potent, was subjected to chemical analysis. The 2-methylanthraquinone (2-MAQ) accounted for the AhR ligand activity and was present at an average concentration of 0.27 parts per hundred in teak dust. Pure 2-MAQ potently induced AhR signaling (EC50 115 nM), confirming that this was the active ligand. Aqueous extracts of teak dust made using yeast or mammalian cell culture medium also contained robust AhR activity, suggesting the 2-MAQ ligand is soluble at bioactive concentrations in physiologically relevant fluids. The high concentration and potency of 2-MAQ in teak wood suggest it may mediate toxic effects through activation of AhR signaling in exposed wood workers.

  8. Decreased expression of the aryl hydrocarbon receptor in ocular Behcet's disease.

    PubMed

    Wang, Chaokui; Ye, Zi; Kijlstra, Aize; Zhou, Yan; Yang, Peizeng

    2014-01-01

    Recent studies show that the aryl hydrocarbon receptor (AhR) is involved in immune responses. AhR is activated following interaction with its ligands, such as 6-formylindolo[3,2-b]carbazole (FICZ) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE). In this study, we investigated the role of AhR activation by its endogenous ligands in the pathogenesis of ocular Behcet's disease (BD). The expression of AhR was significantly decreased in active BD patients as compared to inactive BD patients and normal controls. Both FICZ and ITE inhibited Th1 and Th17 polarization and induced the expression of IL-22 by PBMCs and by CD4(+)T cells in active BD patients and normal controls. Stimulation of purified CD4(+)T cells with FICZ or ITE caused a decreased expression of RORC, IL-17, IL-23R, and CCR6 and an increased phosphorylation of STAT3 and STAT5. The present study suggests that a decreased AhR expression is associated with disease activity in BD patients. The activation of AhR by either FICZ or ITE was able to inhibit Th1 and Th17 cell polarization. Further studies are needed to investigate whether modulation of AhR might be used in the treatment of BD.

  9. TGF-β1 signaling plays a dominant role in the crosstalk between TGF-β1 and the aryl hydrocarbon receptor ligand in prostate epithelial cells.

    PubMed

    Staršíchová, Andrea; Hrubá, Eva; Slabáková, Eva; Pernicová, Zuzana; Procházková, Jiřina; Pěnčíková, Kateřina; Seda, Václav; Kabátková, Markéta; Vondráček, Jan; Kozubík, Alois; Machala, Miroslav; Souček, Karel

    2012-08-01

    Crosstalk between the aryl hydrocarbon receptor (AhR) and transforming growth factor-β1 (TGF-β1) signaling has been observed in various experimental models. However, both molecular mechanism underlying this crosstalk and tissue-specific context of this interaction are still only partially understood. In a model of human non-tumorigenic prostate epithelial cells BPH-1, derived from the benign prostatic hyperplasia, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) persistently activates the AhR signaling pathway and induces expression of xenobiotic metabolizing enzymes, such as CYP1A1 or CYP1B1. Here we demonstrate that TGF-β1 suppresses the AhR-mediated gene expression through multiple mechanisms, involving inhibition of AhR expression and down-regulation of nuclear AhR, via a SMAD4-dependent pathway. In contrast, TCDD-induced AhR signaling does not affect either TGF-β1-regulated gene expression or epithelial-to-mesenchymal transition. These observations suggest that, in the context of prostate epithelium, TGF-β1 signaling plays a dominant role in the crosstalk with AhR signaling pathway. Given the importance of TGF-β1 signaling in regulation of prostate epithelial tissue homeostasis, as well as the recently revealed role of AhR in prostate development and tumorigenesis, the above findings contribute to our understanding of the mechanisms underlying the crosstalk between the two signaling pathways in the prostate-specific context.

  10. Water exposure assessment of aryl hydrocarbon receptor agonists in Three Gorges Reservoir, China using SPMD-based virtual organisms.

    PubMed

    Wang, Jingxian; Bernhöft, Silke; Pfister, Gerd; Schramm, Karl-Werner

    2014-10-15

    SPMD-based virtual organisms (VOs) were deployed at five to eight sites in the Three Gorges Reservoir (TGR), China for five periods in 2008, 2009 and 2011. The water exposure of aryl hydrocarbon receptor (AhR) agonists was assessed by the VOs. The chosen bioassay response for the extracts of the VOs, the induction of 7-ethoxyresorufin-O-deethylase (EROD) was assayed using a rat hepatoma cell line (H4IIE). The results show that the extracts from the VOs could induce AhR activity significantly, whereas the chemically derived 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) equivalent (TEQcal) accounted for <11% of the observed AhR responses (TEQbio). Unidentified AhR-active compounds represented a greater proportion of the TCDD equivalent in VOs from TGR. High TEQbio value in diluted extract and low TEQbio in concentrated extract of the same sample was observed suggesting potential non-additive effects in the mixture. The levels of AhR agonists in VOs from upstream TGR were in general higher than those from downstream reservoir, indicating urbanization effect on AhR agonist pollution. The temporal variation showed that levels of AhR agonists in 2009 and 2011 were higher than those in 2008, and the potential non-additive effects in the area close to the dam were also obviously higher in 2009 and 2011 than in 2008, indicating big changes in the composition of pollutants in the area after water level reached a maximum of 175 m. Although the aqueous concentration of AhR agonists of 0.8-4.8 pg TCDDL(-1) in TGR was not alarming, the tendency of accumulating high concentration of AhR agonists in VO lipid and existence of possible synergism or antagonism in the water may exhibit a potential hazard to local biota being exposed to AhR agonists.

  11. Water exposure assessment of aryl hydrocarbon receptor agonists in Three Gorges Reservoir, China using SPMD-based virtual organisms.

    PubMed

    Wang, Jingxian; Bernhöft, Silke; Pfister, Gerd; Schramm, Karl-Werner

    2014-10-15

    SPMD-based virtual organisms (VOs) were deployed at five to eight sites in the Three Gorges Reservoir (TGR), China for five periods in 2008, 2009 and 2011. The water exposure of aryl hydrocarbon receptor (AhR) agonists was assessed by the VOs. The chosen bioassay response for the extracts of the VOs, the induction of 7-ethoxyresorufin-O-deethylase (EROD) was assayed using a rat hepatoma cell line (H4IIE). The results show that the extracts from the VOs could induce AhR activity significantly, whereas the chemically derived 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) equivalent (TEQcal) accounted for <11% of the observed AhR responses (TEQbio). Unidentified AhR-active compounds represented a greater proportion of the TCDD equivalent in VOs from TGR. High TEQbio value in diluted extract and low TEQbio in concentrated extract of the same sample was observed suggesting potential non-additive effects in the mixture. The levels of AhR agonists in VOs from upstream TGR were in general higher than those from downstream reservoir, indicating urbanization effect on AhR agonist pollution. The temporal variation showed that levels of AhR agonists in 2009 and 2011 were higher than those in 2008, and the potential non-additive effects in the area close to the dam were also obviously higher in 2009 and 2011 than in 2008, indicating big changes in the composition of pollutants in the area after water level reached a maximum of 175 m. Although the aqueous concentration of AhR agonists of 0.8-4.8 pg TCDDL(-1) in TGR was not alarming, the tendency of accumulating high concentration of AhR agonists in VO lipid and existence of possible synergism or antagonism in the water may exhibit a potential hazard to local biota being exposed to AhR agonists. PMID:25058931

  12. Relationship between Ah receptor-mediated polychlorinated biphenyl (PCB)-induced humoral immunosuppression and thymic atrophy.

    PubMed

    Silkworth, J B; Antrim, L

    1985-12-01

    Thymic atrophy and humoral immunosuppression by certain polychlorinated biphenyls is associated with the aromatic hydrocarbon (Ah) receptor in mice. We examined the relationship between these two toxic effects. 3,3',4,4'-Tetrachlorobiphenyl (TCB), which causes immunosuppression and thymic atrophy, and 2,3,3',4,4',5-hexachlorobiphenyl, which causes immunosuppression without thymic atrophy, were administered i.p. to C57BL/6 mice at 0, 35 and 350 mumol/kg b.wt. 2 days before i.v. immunization with 10 micrograms of Escherichia coli lipopolysaccharide. Both congeners caused significant suppression of the day 4 anti-lipopolysaccharide plaque-forming cell response/spleen (less than or equal to 46% of control). TCB (350 mumol/kg) was also administered 2 days before either a primary or secondary i.p. immunization with sheep erythrocytes. TCB treatment before primary immunization had no effects on the day 5 secondary response, whereas treatment before the secondary immunization significantly inhibited both day 5 immunoglobulin M and immunoglobulin G plaque-forming cells (less than 10 and less than 2% of control, respectively) and decreased serum antibody. TCB administered either 8 or 2 days before or 2 or 4 days after immunization with sheep erythrocytes demonstrated that significant suppression of both plaque-forming cells and serum antibody could occur without thymic atrophy. Immunity was most impaired when TCB was given 2 days before immunization. These results demonstrate that thymic atrophy does not always accompany the severe immunosuppression caused by Ah receptor ligands and suggests that it may not be a sensitive measure of Ah receptor-mediated immunosuppression. The data also suggests that differentiation of B lymphocytes into antibody producing cells is impaired during Ah receptor-mediated gene activation.

  13. Ah Receptor–Mediated Suppression of Liver Regeneration through NC-XRE–Driven p21Cip1 Expression

    PubMed Central

    Jackson, Daniel P.; Li, Hui; Mitchell, Kristen A.; Joshi, Aditya D.

    2014-01-01

    Previous studies in hepatocyte-derived cell lines and the whole liver established that the aryl hydrocarbon receptor (AhR) can disrupt G1-phase cell cycle progression following exposure to persistent AhR agonists, such as TCDD (dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin). Growth arrest was attributed to inhibition of G1-phase cyclin-dependent kinase 2 (CDK2) activity. The present study examined the effect of TCDD exposure on liver regeneration following 70% partial hepatectomy in mice lacking the Cip/Kip inhibitors p21Cip1 or p27Kip1 responsible for regulating CDK2 activity. Assessment of the regenerative process in wild-type, p21Cip1 knockout, and p27Kip1 knockout mice confirmed that TCDD-induced inhibition of liver regeneration is entirely dependent on p21Cip1 expression. Compared with wild-type mice, the absence of p21Cip1 expression completely abrogated the TCDD inhibition, and accelerated hepatocyte progression through G1 phase during the regenerative process. Analysis of the transcriptional response determined that increased p21Cip1 expression during liver regeneration involved an AhR-dependent mechanism. Chromatin immunoprecipitation studies revealed that p21Cip1 induction required AhR binding to the newly characterized nonconsensus xenobiotic response element, in conjunction with the tumor suppressor protein Kruppel-like factor 6 functioning as an AhR binding partner. The evidence also suggests that AhR functionality following partial hepatectomy is dependent on a p21Cip1-regulated signaling process, intimately linking AhR biology to the G1-phase cell cycle program. PMID:24431146

  14. A Possible Role of Aryl Hydrocarbon Receptor in Spontaneous Preterm Birth

    PubMed Central

    Li, Yan; Wang, Kai; Zou, Qing-Yun; Zhou, Chi; Magness, Ronald R.; Zheng, Jing

    2015-01-01

    Preterm birth (PTB) is defined as birth before 37 weeks of gestation and is a leading cause of neonatal mortality and morbidity. To date, the etiology of spontaneous PTB (sPTB) remains unclear; however, intrauterine bacterial infection-induced inflammation is considered to be one of the major triggers. Aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor. Upon activation, AhR signaling mediates many biological processes. AhR is abundantly expressed in human placentas, primarily in trophoblasts, and several fetal organs and tissues. The activation of AhR signaling can modulate inflammatory responses via promoting production of pro-inflammatory cytokines by the placenta and fetal membranes. These cytokines could enhance expression and/or activity of cyclooxygenase-2 (COX2) in human trophoblasts and amniotic epithelia, which in turn stimulate synthesis and release of prostaglandins (PGs; e.g., PGE2 and PGF2α). Given the discovery of a number of natural and endogenous AhR ligands in human, we hypothesize that in a subset of patients with high AhR expression in placentas and fetal membranes, repeated exposure to these AhR ligands hyperactivates AhR, inducing hyperactivation of the cytokines/COX2/PGs pathway, resulting in myometrial contractions, ultimately leading to sPTB. We further hypothesize that hyperactivation of this AhR pathway can induce sPTB either directly or in synergy with the bacterial infection. Proof of this hypothesis may provide a novel mechanism underlying sPTB. PMID:25697115

  15. Different regulation of aryl hydrocarbon receptor-regulated genes in response to dioxin in undifferentiated and neuronally differentiated human neuroblastoma SH-SY5Y cells.

    PubMed

    Imran, Saima; Ferretti, Patrizia; Vrzal, Radim

    2015-01-01

    Some environmental pollutants derived from industrial processes have been suggested to be responsible for neurological impairment in children, especially in heavily polluted areas. Since these compounds are usually activators of aryl hydrocarbon receptor (AhR), it would be important to better understand the molecular pathways downstream of AhR leading to neural deficits. To this purpose, appropriate in vitro human neural model is much needed. Here we have investigated whether undifferentiated and neuronally differentiated human neuroblastoma cells, SH-SY5Y cells, can provide a suitable model for monitoring AhR activity induced by environmental pollutants, focusing on 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD), a known activator of AhR. Further characterization of differentiated SH-SY5Y showed an increase in AhRR (aryl hydrocarbon receptor repressor), no change in ARNT1 (AhR nuclear translocator 1), and a decrease in ARNT2 expression with differentiation; in contrast, AhR was undetectable in both undifferentiated and differentiated cells. Nonetheless, treatment of parental as well as differentiated SH-SY5Y cells with TCDD resulted in the induction of AhR-regulated genes, CYP1A1 and CYP1B1; AhRR expression was also affected, but to a much smaller extent. These results indicate that undifferentiated SH-SY5Y are less sensitive to TCDD than neuronally differentiated ones, suggesting a higher resistance of the undifferentiated tumor cells to toxic insults. They also suggest that TCDD in these cells may not act via direct activation of AhR that is undetectable in SH-SY5Y as well as in differentiated neurons. Hence, these cells do not provide an appropriate model for studying ligand-mediated activation of AhR.

  16. Hepatic microsomal cytochrome p450s and chlorinated hydrocarbons in largha and ribbon seals from Hokkaido, Japan: differential response of seal species to Ah receptor agonist exposure.

    PubMed

    Chiba, Issei; Sakakibara, Akihito; Iwata, T Hisato; Ishizuka, Mayumi; Tanabe, Shinsuke; Akahori, Fumiaki; Kazusaka, Akio; Fujita, Shoichi

    2002-04-01

    From 16 largha seals (Phoca largha) and 15 ribbon seals (Phoca fasciata) in the coastal waters of Hokkaido, Japan, blubber chlorinated hydrocarbon (CHC) levels and hepatic cytochrome P450 (CYP) catalytic activities and their immunochemically detected protein content levels were measured. Concentrations of DDTs (2,2-bis(4-chlorophenyl)-1,1-dichloroethylene,p,p'-DDE; 2,2-bis(4-chlorophenyl)-1,1-dichloroethane, p,p'-DDD; dichlorodiphenyltrichloroethane, p,p'-DDT), polychlorinated biphenyl congeners (PCBs), and chlordane compounds (oxychlordane, chlordanes, and nonachlors) in both species were in the range of 290 to 5,300, 420 to 4,000, and 130 to 1,500 ng/g lipid weight, respectively. Aryl hydrocarbon receptor (AhR) agonists, non-ortho (IUPAC 77 and 126) and mono-ortho (IUPAC 105, 118, and 156) coplanar PCB congeners, were also detected, and the 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD) toxic equivalents (TEQs) were 4.9 to 120 pg TEQ/g lipid weight. Cross-reactive proteins with polyclonal antibodies against rat CYP1A1 and CYP3A2 were notably detected in seal liver microsomes. Interestingly, a polyclonal antibody against rat CYP2B1 recognized proteins only at trace levels. In largha seals, both levels of alkoxyresorufin- (methoxy-, ethoxy-, pentoxy-, and benzyloxyresorufin) O-dealkylase (AROD) activities and proteins detected by polyclonal antibodies against rat CYP1A1 were significantly correlated with the concentrations of individual coplanar PCB congeners, total TEQs, and total PCBs. Threshold concentrations for TEQs in blubber of the largha seal to induce hepatic CYP1A protein and EROD activity were estimated to be 8.5 and 19 pg TEQ/g fat weight, respectively. In ribbon seals, similar correlations were not detected, although the TEQ levels were not significantly lower than those in largha seals. These results suggest that AROD activity and CYP1A1 protein in the liver of the largha seal could be a biomarker for the exposure to AhR agonists such as coplanar PCB

  17. Monitoring of xenobiotic ligands for human estrogen receptor and aryl hydrocarbon receptor in industrial wastewater effluents.

    PubMed

    Chou, Pei-Hsin; Liu, Tong-Cun; Lin, Yi-Ling

    2014-07-30

    Industrial wastewater contains a variety of toxic substances, which may severely contaminate the aquatic environment if discharged without adequate treatment. In this study, effluents from a thin film transistor liquid crystal display wastewater treatment plant and the receiving water were analyzed by bioassays and liquid chromatography-tandem mass spectrometry to investigate the presence of estrogenic compounds, aryl hydrocarbon receptor (AhR) agonists, and genotoxicants. Xenobiotic AhR agonists were frequently detected and, in particular, strong AhR agonist activity and genotoxicity were found in the suspended solids of the aeration tank outflow. The high AhR agonist activity in the final effluent (FE) and the downstream river water suggested that the treatment plant failed to remove the wastewater-related AhR agonists. In contrast, although significant estrogenic potency could be detected in raw wastewater or effluents from different treatment processes, the FE and the receiving river water exhibited no or weak estrogenicity. Instrumental analysis showed that bisphenol A was often detected in water samples. However, the investigated estrogenic compounds could only account for a small portion of the estrogenicity in the collected samples. Therefore, further investigation is necessary to identify the major estrogenic compounds and AhR agonist contaminants in the wastewater effluents.

  18. Aryl hydrocarbon receptor mediates benzene-induced hematotoxicity.

    PubMed

    Yoon, Byung-Il; Hirabayashi, Yoko; Kawasaki, Yasushi; Kodama, Yukio; Kaneko, Toyozo; Kanno, Jun; Kim, Dae-Yong; Fujii-Kuriyama, Yoshiaki; Inoue, Tohru

    2002-11-01

    Benzene can induce hematotoxicity and leukemia in humans and mice. Since a review of the literature shows that the CYP2E1 knockout mouse is not known to possess any benzene toxicity, the metabolism of benzene by CYP2E1 in the liver is regarded to be prerequisite for its cytotoxicity and genotoxicity, although the mechanism is not fully understood yet. Because it was found some years ago that benzene was also a substrate for CYP1A1, we investigated the involvement of the aryl hydrocarbon receptor (AhR) in benzene hematotoxicity using AhR wild-type (AhR(+/+)), heterozygous (AhR(+/-)), and homozygous (AhR(-/-)) male mice. Interestingly, following a 2-week inhalation of 300 ppm benzene (a potent dose for leukemogenicity), no hematotoxicity was induced in AhR(-/-) mice. Further, there were no changes in cellularity of peripheral blood and bone marrow (BM), nor in levels of granulocyte-macrophage colony-forming units in BM. This lack of hematotoxicity was associated with the lack of p21 overexpression, which was regularly seen in the wild-type mice following benzene inhalation. Combined treatment with two major benzene metabolites, phenol and hydroquinone, induced hemopoietic toxicity, although it was not known whether this happened due to a surprising lack of expression of CYP2E1 by AhR knockout, or due to a lack of other AhR-mediated CYP enzymes, including 1A1 (i.e., a possible alternative pathway of benzene metabolism). The former possibility, evaluated in the present study, failed to show a significant relationship between AhR and the expression of CYP2E1. Furthermore, a subsequent evaluation of AhR expression after benzene inhalation tended to show higher but less significant expression in the liver, and none in the BM, compared with sham control. Although this study failed to identify the more likely of the above-mentioned two possibilities, the study using AhR knockout mice on benzene inhalation presents the unique possibility that the benzene toxicity may be

  19. Naturally-Occurring Marine Brominated Indoles are Aryl Hydrocarbon Receptor Ligands/Agonists

    PubMed Central

    DeGroot, Danica E.; Franks, Diana G.; Higa, Tatsuo; Tanaka, Junichi; Hahn, Mark E.; Denison, Michael S.

    2015-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that mediates the toxic and biological effects of structurally diverse chemicals, including the environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). As part of a larger effort to identify the full spectrum of chemicals that can bind to and activate the AhR, we have examined the ability of several naturally-occurring marine-derived brominated indoles and brominated (methylthio)indoles (collectively referred to as “brominated indoles”) to bind to the AhR and stimulate AhR-dependent gene expression. Incubation of mouse, rat and guinea pig recombinant cell lines containing a stably transfected AhR-responsive luciferase reporter gene with eight brominated indoles revealed that all compounds stimulated luciferase reporter gene activity, although some species-specific differences were observed. All compounds induced significantly more luciferase activity when incubated with cells for 4 h as compared to 24 h, demonstrating that these compounds are transient activators of the AhR signaling pathway. Three of the brominated indoles induced CYP1A1 mRNA in human HepG2 cells in vitro and Cyp1a mRNA in zebrafish embryos in vivo. The identification of the brominated indoles as direct ligands and activators/agonists of the AhR was confirmed by their ability to compete with [3H]TCDD for binding to the AhR and to stimulate AhR transformation and DNA binding in vitro. Taken together, these marine-derived brominated indoles are members of a new class of naturally-occurring AhR agonists. PMID:26001051

  20. In vitro and in silico evaluation of transactivation potencies of avian AHR1 and AHR2 by endogenous ligands: Implications for the physiological role of avian AHR2.

    PubMed

    Kim, In-Sung; Hwang, Ji-Hee; Hirano, Masashi; Iwata, Hisato; Kim, Eun-Young

    2016-09-01

    Aryl hydrocarbon receptor (AHR) is well conserved from invertebrates to vertebrates, and it mediates the toxic effects of exogenous ligands, including dioxins. Recent studies reported that AHRs activated by endogenous ligands play critical roles in mammalian physiological homeostasis. Avian species possess at least two AHR isoforms (AHR1 and AHR2), which exhibit species- and isoform-specific transactivation potencies to exogenous ligands, whereas mammals possess a single AHR. To delineate the profiles and roles of endogenous ligands for avian AHR isoforms, we investigated in vitro transactivation potencies of avian AHRs (AHR1 and AHR2 from the jungle crow, Corvus macrorhynchos; common cormorant, Phalacrocorax carbo; and black-footed albatross, Phoebastria nigripes) treated with the endogenous tryptophan metabolites 6-formylindolo [3,2-b] carbazole (FICZ), l-kynurenine (l-Kyn), kynurenic acid (KYNA), and indoxyl sulfate (IS). Furthermore, we analyzed the binding mode of these ligands to each avian AHR isoform by in silico docking simulations. The EC50 of FICZ (0.009-0.032nM) was similar regardless of the species or isoform of AHR. The estimated in silico binding mode of FICZ to AHRs was well conserved in both isoforms. The transactivation potencies of avian AHRs to other tryptophan metabolites were 10(5)-10(7) fold lower than those for FICZ, and EC50 values varied in a species- and isoform-specific manner. This was consistent with poor conservation of the binding mode of l-Kyn, KYNA, and IS predicted in in silico docking simulations. Our results suggest that in avian species, FICZ is the most potent endogenous AHR ligand, and that AHR1 and AHR2 are physiologically functional. PMID:27060260

  1. Kynurenine, by activating aryl hydrocarbon receptor, decreases erythropoietin and increases hepcidin production in HepG2 cells: A new mechanism for anemia of inflammation.

    PubMed

    Eleftheriadis, Theodoros; Pissas, Georgios; Antoniadi, Georgia; Liakopoulos, Vassilios; Stefanidis, Ioannis

    2016-01-01

    It is known that inadequate erythropoietin (EPO) production contributes to the pathogenesis of anemia of inflammation, although the exact molecular mechanism is unknown. Aryl hydrocarbon receptor (AhR) may compete with hypoxia-inducible factor 2α (HIF-2α), the master regulator of EPO production, for binding with HIF-1β. The effect of kynurenine, an endogenous AhR activator that increases in inflammation, on EPO and hepcidin production was evaluated. HepG2 cells were treated with the hypoxia mimetic CoCl2, kynurenine, the AhR inhibitor CH223191, and combinations of these. EPO and hepcidin production was measured with enzyme-linked immunosorbent assay. HIF-2α and CYP1A1 levels, a transcriptional target of AhR, were assessed by Western blotting. CoCl2 increased EPO production and decreased hepcidin and CYP1A1. Kynurenine exerted the opposite effects. Wherever CH223191 was added, the inhibitor overcorrected kynurenine-induced alterations in both the presence and the absence of CoCl2. Also, treatment with CH223191 alone increased EPO and decreased hepcidin, indicating that there is a degree of constitutive AhR activation, possibly by other endogenous AhR activators. In conclusion, kynurenine, by competing with HIF-2α, may contribute to anemia of inflammation by decreasing EPO and increasing hepcidin production. The fact that inactivation of AhR alone induces EPO makes this transcription factor a potential therapeutic target in situations that require increased EPO.

  2. Expression of the Aryl Hydrocarbon Receptor in Growth Plate Cartilage and the Impact of Its Local Modulation on Longitudinal Bone Growth

    PubMed Central

    Cedervall, Therése; Lind, Pia Monica; Sävendahl, Lars

    2015-01-01

    Although dioxin has been reported to impair bone growth in both humans and animals, the underlying mechanisms have not been clarified. We conducted this study to rule out if dioxin may directly target the growth plate, via local modulation of the aryl hydrocarbon receptor (AhR). Initial studies in rare tissue samples of the human growth plate confirmed that the AhR protein is widely expressed in growth plate cartilage. To explore the local role of the AhR, mechanistic studies were performed in a well-established model of cultured fetal rat metatarsal bones. The longitudinal growth of these bones was monitored while being exposed to AhR modulators. The AhR agonist, 2,3,7,8-tetrachlorodibenzo-p-dioxin, did not affect bone growth at any concentrations tested (1 pM–10 nM). In contrast, the AhR antagonist, alpha-naphthoflavone, suppressed bone growth and increased chondrocyte apoptosis, although only at a high, potentially cytotoxic concentration (50 µM). We conclude that although the AhR is widely expressed in the growth plate, bone growth is not modulated when locally activated, and therefore, dioxin-induced growth failure is likely mediated through systemic rather than local actions. PMID:25867478

  3. Inhibition of the aryl hydrocarbon receptor prevents Western diet-induced obesity. Model for AHR activation by kynurenine via oxidized-LDL, TLR2/4, TGFβ, and IDO1.

    PubMed

    Moyer, Benjamin J; Rojas, Itzel Y; Kerley-Hamilton, Joanna S; Hazlett, Haley F; Nemani, Krishnamurthy V; Trask, Heidi W; West, Rachel J; Lupien, Leslie E; Collins, Alan J; Ringelberg, Carol S; Gimi, Barjor; Kinlaw, William B; Tomlinson, Craig R

    2016-06-01

    Obesity is an increasingly urgent global problem, yet, little is known about its causes and less is known how obesity can be effectively treated. We showed previously that the aryl hydrocarbon receptor (AHR) plays a role in the regulation of body mass in mice fed Western diet. The AHR is a ligand-activated nuclear receptor that regulates genes involved in a number of biological pathways, including xenobiotic metabolism and T cell polarization. This study was an investigation into whether inhibition of the AHR prevents Western diet-based obesity. Male C57Bl/6J mice were fed control and Western diets with and without the AHR antagonist α-naphthoflavone or CH-223191, and a mouse hepatocyte cell line was used to delineate relevant cellular pathways. Studies are presented showing that the AHR antagonists α-naphthoflavone and CH-223191 significantly reduce obesity and adiposity and ameliorates liver steatosis in male C57Bl/6J mice fed a Western diet. Mice deficient in the tryptophan metabolizing enzyme indoleamine 2,3-dioxygenase 1 (IDO1) were also resistant to obesity. Using an AHR-directed, luciferase-expressing mouse hepatocyte cell line, we show that the transforming growth factor β1 (TGFβ1) signaling pathway via PI3K and NF-κB and the toll-like receptor 2/4 (TLR2/4) signaling pathway stimulated by oxidized low-density lipoproteins via NF-κB, each induce luciferase expression; however, TLR2/4 signaling was significantly reduced by inhibition of IDO1. At physiological levels, kynurenine but not kynurenic acid (both tryptophan metabolites and known AHR agonists) activated AHR-directed luciferase expression. We propose a hepatocyte-based model, in which kynurenine production is increased by enhanced IDO1 activity stimulated by TGFβ1 and TLR2/4 signaling, via PI3K and NF-κB, to perpetuate a cycle of AHR activation to cause obesity; and inhibition of the AHR, in turn, blocks the cycle's output to prevent obesity. The AHR with its broad ligand binding specificity

  4. Norisoboldine, an Anti-Arthritis Alkaloid Isolated from Radix Linderae, Attenuates Osteoclast Differentiation and Inflammatory Bone Erosion in an Aryl Hydrocarbon Receptor-Dependent Manner.

    PubMed

    Wei, Zhi-feng; Lv, Qi; Xia, Ying; Yue, Meng-fan; Shi, Can; Xia, Yu-feng; Chou, Gui-xin; Wang, Zheng-tao; Dai, Yue

    2015-01-01

    Norisoboldine (NOR), the primary isoquinoline alkaloid constituent of the root of Lindera aggregata, has previously been demonstrated to attenuate osteoclast (OC) differentiation. Accumulative evidence has shown that aryl hydrocarbon receptor (AhR) plays an important role in regulating the differentiation of various cells, and multiple isoquinoline alkaloids can modulate AhR. In the present study, we explored the role of NOR in the AhR signaling pathway. These data showed that the combination of AhR antagonist resveratrol (Res) or α-naphthoflavone (α-NF) nearly reversed the inhibition of OC differentiation through NOR. NOR could stably bind to AhR, up-regulate the nuclear translocation of AhR, and enhance the accumulation of the AhR-ARNT complex, AhR-mediated reporter gene activity and CYP1A1 expression in RAW 264.7 cells, suggesting that NOR might be an agonist of AhR. Moreover, NOR inhibited the nuclear translocation of NF-κB-p65, resulting in the evident accumulation of the AhR-NF-κB-p65 complex, which could be markedly inhibited through either Res or α-NF. Although NOR only slightly affected the expression of HIF-1α, NOR markedly reduced VEGF mRNA expression and ARNT-HIF-1α complex accumulation. In vivo studies indicated that NOR decreased the number of OCs and ameliorated the bone erosion in the joints of rats with collagen-induced arthritis, accompanied by the up-regulation of CYP1A1 and the down-regulation of VEGF mRNA expression in the synovium of rats. A combination of α-NF nearly completely reversed the effects of NOR. In conclusion, NOR attenuated OC differentiation and bone erosion through the activation of AhR and the subsequent inhibition of both NF-κB and HIF pathways.

  5. Biology and function of the aryl hydrocarbon receptor: report of an international and interdisciplinary conference.

    PubMed

    Esser, Charlotte

    2012-08-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor present in many cells. The AhR links environmental chemical stimuli with adaptive responses, such as detoxification, cellular homoeostasis or immune responses. Furthermore, novel roles of AhR in physiological and genetic functions are being discovered. This is a report of a recent meeting in Düsseldorf. The meeting highlighted that AhR research has moved from its focus on toxic effects of dioxins and other environmental pollutants to its biological roles. For instance, it was recently discovered that AhR-responsive elements in retrotransposons contribute to the functional structure of the genome. Other exciting new reports concerned the way plant-derived compounds in our diet are necessary for a fully functioning immune system of the gut. Also, human brain tumours use the AhR system to gain growth advantages. Other aspects covered were neurotoxicology, the circadian rhythm, or the breadth of the adaptive and innate immune system (hematopoietic stem cells, dendritic cells, T cells, mast cells). Finally, the meeting dealt with the discovery of new xenobiotic and natural ligands and their use in translational medicine, or cancer biology and AhR.

  6. Growth suppression of Leydig TM3 cells mediated by aryl hydrocarbon receptor

    SciTech Connect

    Iseki, Minoru; Ikuta, Togo; Kobayashi, Tetsuya; Kawajiri, Kaname . E-mail: kawajiri@cancer-c.pref.saitama.jp

    2005-06-17

    Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin induces developmental toxicity in reproductive organs. To elucidate the function of AhR, we generated stable transformants of TM3 cells overexpressing wild-type aryl hydrocarbon receptor (AhR) or its mutants which carried mutations in nuclear localization signal or nuclear export signal. In the presence of 3-methylcholanthrene (MC), proliferation of the cells transfected with wild-type AhR was completely suppressed, whereas cells expressing AhR mutants proliferated in a manner equivalent to control TM3 cells, suggesting AhR-dependent growth inhibition. The suppression was associated with up-regulation of cyclin-dependent kinase inhibitor p21{sup Cip1}, which was abolished by pretreatment with actinomycin D. A p38 MAPK specific inhibitor, SB203580, blocked the increase of p21{sup Cip1} mRNA in response to MC. Treatment with indigo, another AhR ligand, failed to increase of p21{sup Cip1} mRNA, although up-regulation of mRNA for CYP1A1 was observed. These data suggest AhR in Leydig cells mediates growth inhibition by inducing p21{sup Cip1}.

  7. Aryl hydrocarbon receptor-dependent stanniocalcin 2 induction by cinnabarinic acid provides cytoprotection against endoplasmic reticulum and oxidative stress.

    PubMed

    Joshi, Aditya D; Carter, Dwayne E; Harper, Tod A; Elferink, Cornelis J

    2015-04-01

    The aryl hydrocarbon receptor (AhR) is a cytosolic ligand-activated transcription factor historically known for its role in xenobiotic metabolism. Although AhR activity has previously been shown to play a cytoprotective role against intrinsic apoptotic stimuli, the underlying mechanism by which AhR confers cytoprotection against apoptosis is largely unknown. Here, we demonstrate that activation of AhR by the tryptophan catabolite cinnabarinic acid (CA) directly upregulates expression of stanniocalcin 2 (Stc2) to elicit cytoprotection against apoptosis induced by endoplasmic reticulum stress and oxidative stress. Chromatin immunoprecipitation studies demonstrated that CA treatment induces direct AhR binding to a region of the Stc2 promoter containing multiple xenobiotic response elements. Using isolated primary hepatocytes from AhR wild-type (AhR floxed) and liver-specific AhR conditional knockout mice, we showed that pretreatment with CA conferred cytoprotection against hydrogen peroxide (H(2)O(2))-, thapsigargin-, and ethanol-induced apoptosis in an AhR-dependent manner. Furthermore, suppressing Stc2 expression using RNA interference confirmed that the cytoprotective properties of CA against H(2)O(2), thapsigargin, and ethanol injury were absolutely dependent on Stc2. Immunochemistry revealed the presence of Stc2 in the endoplasmic reticulum and on the cell surface, consistent with Stc2 secretion and autocrine and/or paracrine signaling. Finally, in vivo data using a mouse model of acute alcohol hepatotoxicity demonstrated that CA provided cytoprotection against ethanol-induced apoptosis, hepatic microvesicular steatosis, and liver injury. Collectively, our data uncovered a novel mechanism for AhR-mediated cytoprotection in the liver that is dependent on CA-induced Stc2 activity. PMID:25672339

  8. Role of Aryl Hydrocarbon Receptor in Circadian Clock Disruption and Metabolic Dysfunction.

    PubMed

    Jaeger, Cassie; Tischkau, Shelley A

    2016-01-01

    The prevalence of metabolic syndrome, a clustering of three or more risk factors that include abdominal obesity, increased blood pressure, and high levels of glucose, triglycerides, and high-density lipoproteins, has reached dangerous and costly levels worldwide. Increases in morbidity and mortality result from a combination of factors that promote altered glucose metabolism, insulin resistance, and metabolic dysfunction. Although diet and exercise are commonly touted as important determinants in the development of metabolic dysfunction, other environmental factors, including circadian clock disruption and activation of the aryl hydrocarbon receptor (AhR) by dietary or other environmental sources, must also be considered. AhR binds a range of ligands, which prompts protein-protein interactions with other Per-Arnt-Sim (PAS)-domain-containing proteins and subsequent transcriptional activity. This review focuses on the reciprocal crosstalk between the activated AhR and the molecular circadian clock. AhR exhibits a rhythmic expression and time-dependent sensitivity to activation by AhR agonists. Conversely, AhR activation influences the amplitude and phase of expression of circadian clock genes, hormones, and the behavioral responses of the clock system to changes in environmental illumination. Both the clock and AhR status and activation play significant and underappreciated roles in metabolic homeostasis. This review highlights the state of knowledge regarding how AhR may act together with the circadian clock to influence energy metabolism. Understanding the variety of AhR-dependent mechanisms, including its interactions with the circadian timing system that promote metabolic dysfunction, reveals new targets of interest for maintenance of healthy metabolism. PMID:27559298

  9. Role of Aryl Hydrocarbon Receptor in Circadian Clock Disruption and Metabolic Dysfunction

    PubMed Central

    Jaeger, Cassie; Tischkau, Shelley A.

    2016-01-01

    The prevalence of metabolic syndrome, a clustering of three or more risk factors that include abdominal obesity, increased blood pressure, and high levels of glucose, triglycerides, and high-density lipoproteins, has reached dangerous and costly levels worldwide. Increases in morbidity and mortality result from a combination of factors that promote altered glucose metabolism, insulin resistance, and metabolic dysfunction. Although diet and exercise are commonly touted as important determinants in the development of metabolic dysfunction, other environmental factors, including circadian clock disruption and activation of the aryl hydrocarbon receptor (AhR) by dietary or other environmental sources, must also be considered. AhR binds a range of ligands, which prompts protein–protein interactions with other Per-Arnt-Sim (PAS)-domain-containing proteins and subsequent transcriptional activity. This review focuses on the reciprocal crosstalk between the activated AhR and the molecular circadian clock. AhR exhibits a rhythmic expression and time-dependent sensitivity to activation by AhR agonists. Conversely, AhR activation influences the amplitude and phase of expression of circadian clock genes, hormones, and the behavioral responses of the clock system to changes in environmental illumination. Both the clock and AhR status and activation play significant and underappreciated roles in metabolic homeostasis. This review highlights the state of knowledge regarding how AhR may act together with the circadian clock to influence energy metabolism. Understanding the variety of AhR-dependent mechanisms, including its interactions with the circadian timing system that promote metabolic dysfunction, reveals new targets of interest for maintenance of healthy metabolism. PMID:27559298

  10. Persistent Binding of Ligands to the Aryl Hydrocarbon Receptor

    PubMed Central

    Bohonowych, Jessica E.; Denison, Michael S.

    2010-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates many of the biological and toxic effects of halogenated aromatic hydrocarbons (HAHs), polycyclic aromatic hydrocarbons (PAHs), and other structurally diverse ligands. While HAHs are several orders of magnitude more potent in producing AhR-dependent biochemical effects than PAHs or other AhR agonists, only the HAHs have been observed to produce AhR-dependent toxicity in vivo. Here we have characterized the dissociation of a prototypical HAH ligand ([3H] 2,3,7,8-tetrachlorodibenzo-p-dioxin [TCDD]) and PAH-like ligand ([3H] β-naphthoflavone [βNF]) from the guinea pig, hamster, mouse, and rat hepatic cytosolic AhR in order to elucidate the relationship between the apparent ligand-binding affinities and the divergent potency of these chemicals. Both compounds dissociated very slowly from the AhR with the amount of specific binding remaining at 96 h ranging from 53% to 70% for [3H]TCDD and 26% to 85% for [3H] βNF, depending upon the species examined. The rate of ligand dissociation was unaffected by protein concentration or incubation temperature. Preincubation of cytosol with 2,3,7,8-tetrachlorodibenzofuran, carbaryl, or primaquine, prior to the addition of [3H]TCDD, shifted the apparent IC50 of these compounds as competitive AhR ligands by ∼10- to 50-fold. Our results support the need for reassessment of previous AhR ligand-binding affinity calculations and competitive binding analysis since these measurements are not carried out at equilibrium binding conditions. Our studies suggest that AhR binding affinity/occupancy has little effect on the observed differences in the persistence of gene expression by HAHs and PAHs. PMID:17431010

  11. New Trends in Aryl Hydrocarbon Receptor Biology

    PubMed Central

    Mulero-Navarro, Sonia; Fernandez-Salguero, Pedro M.

    2016-01-01

    Traditionally considered as a critical intermediate in the toxic and carcinogenic response to dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin, TCDD), the Aryl hydrocarbon/Dioxin receptor (AhR) has proven to be also an important regulator of cell physiology and organ homeostasis. AhR has become an interesting and actual area of research mainly boosted by a significant number of recent studies analyzing its contribution to the proper functioning of the immune, hepatic, cardiovascular, vascular and reproductive systems. At the cellular level, AhR establishes functional interactions with signaling pathways governing cell proliferation and cell cycle, cell morphology, cell adhesion and cell migration. Two exciting new aspects in AhR biology deal with its implication in the control of cell differentiation and its more than likely involvement in cell pluripotency and stemness. In fact, it is possible that AhR could help modulate the balance between differentiation and pluripotency in normal and transformed tumor cells. At the molecular level, AhR regulates an increasingly large array of physiologically relevant genes either by traditional transcription-dependent mechanisms or by unforeseen processes involving genomic insulators, chromatin dynamics and the transcription of mobile genetic elements. AhR is also closely related to epigenetics, not only from the point of view of target gene expression but also with respect to its own regulation by promoter methylation. It is reasonable to consider that deregulation of these many functions could have a causative role, or at least contribute to, human disease. Consequently, several laboratories have proposed that AhR could be a valuable tool as diagnostic marker and/or therapeutic target in human pathologies. An additional point of interest is the possibility of regulating AhR activity by endogenous non-toxic low weight molecules agonist or antagonist molecules that could be present or included in the diet. In this review, we will

  12. Implication of cytochrome P-450 1A isoforms and the AH receptor in the genotoxicity of coal-tar fume condensate and bitumen fume condensates.

    PubMed

    Genevois, C; Pfohl-Leszkowicz, A; Boillot, K; Brandt, H; Castegnaro, M

    1998-06-01

    During the hot application of bitumen- or coal-tar-containing materials, fumes are emitted that contain polycyclic aromatic compounds. Although workers' exposure to these fumes is low, it might lead to health problems. No study has reported the metabolic pathways involved in the genotoxicity of coal tar or bitumen fume condensates (CTFC, BFCs). We have therefore studied the DNA adducts formed by incubation of CTFC or BFCs with liver microsomes from several type of mice and with yeast microsomes expressing individual human CYP enzymes. Our results demonstrates that: (1) the aryl hydrocarbon receptor (AHR) plays an important role in the biotransformation of BFCs and to a lesser extent of CTFC; (2) for CTFC, both cytochrome P450 (CYP) 1A isoforms are involved in the formation of genotoxic compounds, and the reactive metabolites formed via CYP 1A1, are substrates for epoxide hydrolase (mEH); (3) for BFCs, the genotoxicity is partially dependent upon CYP 1A1 and the reactive metabolites are not substrates for mEH; (4) CYP 1A isoforms are not exclusively responsible for the genotoxicity of the CTFC and BFCs as other CYPs and also enzymes of the [AH] gene battery, may play an important role. PMID:21781875

  13. Implication of cytochrome P-450 1A isoforms and the AH receptor in the genotoxicity of coal-tar fume condensate and bitumen fume condensates.

    PubMed

    Genevois, C; Pfohl-Leszkowicz, A; Boillot, K; Brandt, H; Castegnaro, M

    1998-06-01

    During the hot application of bitumen- or coal-tar-containing materials, fumes are emitted that contain polycyclic aromatic compounds. Although workers' exposure to these fumes is low, it might lead to health problems. No study has reported the metabolic pathways involved in the genotoxicity of coal tar or bitumen fume condensates (CTFC, BFCs). We have therefore studied the DNA adducts formed by incubation of CTFC or BFCs with liver microsomes from several type of mice and with yeast microsomes expressing individual human CYP enzymes. Our results demonstrates that: (1) the aryl hydrocarbon receptor (AHR) plays an important role in the biotransformation of BFCs and to a lesser extent of CTFC; (2) for CTFC, both cytochrome P450 (CYP) 1A isoforms are involved in the formation of genotoxic compounds, and the reactive metabolites formed via CYP 1A1, are substrates for epoxide hydrolase (mEH); (3) for BFCs, the genotoxicity is partially dependent upon CYP 1A1 and the reactive metabolites are not substrates for mEH; (4) CYP 1A isoforms are not exclusively responsible for the genotoxicity of the CTFC and BFCs as other CYPs and also enzymes of the [AH] gene battery, may play an important role.

  14. Activation of aryl hydrocarbon receptor mediates suppression of hypoxia-inducible factor-dependent erythropoietin expression by indoxyl sulfate.

    PubMed

    Asai, Hirobumi; Hirata, Junya; Hirano, Ayumi; Hirai, Kazuya; Seki, Sayaka; Watanabe-Akanuma, Mie

    2016-01-15

    Indoxyl sulfate (IS) is a representative uremic toxin that accumulates in the blood of patients with chronic kidney disease (CKD). In addition to the involvement in the progression of CKD, a recent report indicates that IS suppresses hypoxia-inducible factor (HIF)-dependent erythropoietin (EPO) production, suggesting that IS may also contribute to the progression of renal anemia. In this report, we provide evidence that aryl hydrocarbon receptor (AhR) mediates IS-induced suppression of HIF activation and subsequent EPO production. In HepG2 cells, IS at concentrations similar to the blood levels in CKD patients suppressed hypoxia- or cobalt chloride-induced EPO mRNA expression and transcriptional activation of HIF. IS also induced AhR activation, and AhR blockade resulted in abolishment of IS-induced suppression of HIF activation. The HIF transcription factor is a heterodimeric complex composed of HIF-α subunits (HIF-1α and HIF-2α) and AhR nuclear translocator (ARNT). IS suppressed nuclear accumulation of the HIF-α-ARNT complex accompanied by an increase of the AhR-ARNT complex in the nucleus, implying the involvement of interactions among AhR, HIF-α, and ARNT in the suppression mechanism. In rats, oral administration of indole, a metabolic precursor of IS, inhibited bleeding-induced elevation of renal EPO mRNA expression and plasma EPO concentration and strongly induced AhR activation in the liver and renal cortex tissues. Collectively, this study is the first to elucidate the detailed mechanism by which AhR plays an indispensable role in the suppression of HIF activation by IS. Hence, IS-induced activation of AhR may be a potential therapeutic target for treating renal anemia. PMID:26561638

  15. Activation of estrogen receptor signaling by the dioxin-like aryl hydrocarbon receptor agonist, 3,3',4,4',5-Pentachlorobiphenyl (PCB126) in salmon in vitro system

    SciTech Connect

    Mortensen, Anne Skjetne; Arukwe, Augustine

    2008-03-01

    Available toxicological evidence indicates that environmental contaminants with strong affinity to the aryl hydrocarbon receptor (AhR) have anti-estrogenic properties in both mammalian and non-mammalian in vivo and in vitro studies. The primary objective of the present study was to investigate the interactions between the AhR and estrogen receptor (ER) in salmon in vitro system. Two separate experiments were performed and gene expression patterns were analyzed using real-time PCR, while protein analysis was done by immunoblotting. Firstly, salmon primary hepatocytes were exposed to the dioxin-like PCB126 at 1, 10 and 50 pM and ER agonist nonylphenol (NP) at 5 and 10 {mu}M, singly or in combination. Our data showed increased levels of ER-mediated gene expression (vitellogenin: Vtg, zona radiata protein: Zr-protein, ER{alpha}, ER{beta} and vigilin) as well as increased cellular ER{alpha} protein levels after treatment with NP and PCB126, singly or in combination. PCB126 treatment alone produced, as expected, increased transcription of AhR nuclear translocator (Arnt), CYP1A1 and AhR repressor (AhRR) mRNA, and these responses were reduced in the presence of NP concentrations. PCB126 exposure alone did not produce significant effect on AhR2{alpha} mRNA but increased (at 1 and 50 pM) and decreased (at 10 pM) AhR2{beta} mRNA below control level. For AhR2{delta} and AhR2{gamma} isotypes, PCB126 (at 1 pM) produced significant decreases (total inhibition for AhR2{gamma}) of mRNA levels but was indifferent at 10 and 50 pM, compared to control. NP exposure alone produced concentration-dependent significant decrease of AhR2{beta} mRNA. In contrast, while 5 {mu}M NP produced an indifferent effect on AhR2{delta} and AhR2{gamma}, 10 {mu}M NP produced significant decrease (total inhibition for AhR2{gamma}) and the presence of NP produced apparent PCB126 concentration-specific modulation of all AhR isotypes. A second experiment was performed to evaluate the involvement of ER

  16. An in vitro rainbow trout cell bioassay for AhR-mediated toxins

    SciTech Connect

    Richter, C.A.; Giesy, J.P.; Denison, M.S.

    1995-12-31

    The toxicity of PCBs, dioxins, and other halogenated aromatic hydrocarbons (HAHS) at environmentally relevant concentrations is in large part mediated through the aromatic hydrocarbon receptor (AhR). Bioassays which measure the activity of genes regulated by the receptor provide an integrative measure of the total AhR-mediated toxicity of a sample. The authors have recently developed and characterized a bioassay using recombinant rainbow trout hepatoma cells containing the firefly luciferase reporter gene under the regulation of the AhR. The cell line is designated Remodulated Lightning Trout (RLT). The RLT bioassay is relevant to fish, and is useful as a rapid screening device, a guide for chemical analysis, and a tool for studies of the AhR mechanism. The responses of the RLT cell line to various PCB congeners are similar to responses of in vivo fish bioassays. The authors now report on the responses of the bioassay to dioxins, dibenzofurans, and other related compounds as compared to in vivo fish bioassays. The authors will also report on the utility of the RLT bioassay in measuring the total TEQ of complex mixtures.

  17. Sinomenine induces the generation of intestinal Treg cells and attenuates arthritis via activation of aryl hydrocarbon receptor.

    PubMed

    Tong, Bei; Yuan, Xusheng; Dou, Yannong; Wu, Xin; Wang, Yuhui; Xia, Yufeng; Dai, Yue

    2016-10-01

    Sinomenine (SIN), an anti-arthritis drug, has previously been proven to exert immunomodulatory activity in rats by inducing intestinal regulatory T-cells (Treg cells). Here, we assessed the effect of SIN on the generation and function of Treg cells in autoimmune arthritis, and the underlying mechanisms in view of aryl hydrocarbon receptor (AhR). The proportions of Treg cells and IL-17-producing T-cells (Th17 cells) differentiated from naive T-cells were analyzed by flow cytometric analysis. The AhR agonistic effect of SIN was tested by analyzing the activation of downstream signaling pathways and target genes. The dependence of intestinal Treg cell induction and arthritis alleviation by SIN on AhR activation was confirmed in a mouse collagen-induced arthritis (CIA) model. SIN promoted the differentiation and function of intestinal Treg cells in vitro. It induced the expression and activity of AhR target gene, promoted AhR/Hsp90 dissociation and AhR nuclear translocation, induced XRE reporter activity, and facilitated AhR/XRE binding in vitro, displaying the potential to be an agonist of AhR. In CIA mice, SIN induced the generation of intestinal Treg cells, and facilitated the immunosuppressive function of these Treg cells as shown by an adoptive transfer test. In addition, the induction of intestinal Treg cells and the anti-arthritic effect of SIN in CIA mice could be largely diminished by the AhR antagonist resveratrol. SIN attenuates arthritis by promoting the generation and function of Treg cells in an AhR-dependent manner. PMID:27617398

  18. Skatole (3-Methylindole) Is a Partial Aryl Hydrocarbon Receptor Agonist and Induces CYP1A1/2 and CYP1B1 Expression in Primary Human Hepatocytes.

    PubMed

    Rasmussen, Martin Krøyer; Balaguer, Patrick; Ekstrand, Bo; Daujat-Chavanieu, Martine; Gerbal-Chaloin, Sabine

    2016-01-01

    Skatole (3-methylindole) is a product of bacterial fermentation of tryptophan in the intestine. A significant amount of skatole can also be inhaled during cigarette smoking. Skatole is a pulmonary toxin that induces the expression of aryl hydrocarbon receptor (AhR) regulated genes, such as cytochrome P450 1A1 (CYP1A1), in human bronchial cells. The liver has a high metabolic capacity for skatole and is the first organ encountered by the absorbed skatole; however, the effect of skatole in the liver is unknown. Therefore, we investigated the impact of skatole on hepatic AhR activity and AhR-regulated gene expression. Using reporter gene assays, we showed that skatole activates AhR and that this is accompanied by an increase of CYP1A1, CYP1A2 and CYP1B1 expression in HepG2-C3 and primary human hepatocytes. Specific AhR antagonists and siRNA-mediated AhR silencing demonstrated that skatole-induced CYP1A1 expression is dependent on AhR activation. The effect of skatole was reduced by blocking intrinsic cytochrome P450 activity and indole-3-carbinole, a known skatole metabolite, was a more potent inducer than skatole. Finally, skatole could reduce TCDD-induced CYP1A1 expression, suggesting that skatole is a partial AhR agonist. In conclusion, our findings suggest that skatole and its metabolites affect liver homeostasis by modulating the AhR pathway. PMID:27138278

  19. Skatole (3-Methylindole) Is a Partial Aryl Hydrocarbon Receptor Agonist and Induces CYP1A1/2 and CYP1B1 Expression in Primary Human Hepatocytes

    PubMed Central

    Balaguer, Patrick; Ekstrand, Bo; Daujat-Chavanieu, Martine; Gerbal-Chaloin, Sabine

    2016-01-01

    Skatole (3-methylindole) is a product of bacterial fermentation of tryptophan in the intestine. A significant amount of skatole can also be inhaled during cigarette smoking. Skatole is a pulmonary toxin that induces the expression of aryl hydrocarbon receptor (AhR) regulated genes, such as cytochrome P450 1A1 (CYP1A1), in human bronchial cells. The liver has a high metabolic capacity for skatole and is the first organ encountered by the absorbed skatole; however, the effect of skatole in the liver is unknown. Therefore, we investigated the impact of skatole on hepatic AhR activity and AhR-regulated gene expression. Using reporter gene assays, we showed that skatole activates AhR and that this is accompanied by an increase of CYP1A1, CYP1A2 and CYP1B1 expression in HepG2-C3 and primary human hepatocytes. Specific AhR antagonists and siRNA-mediated AhR silencing demonstrated that skatole-induced CYP1A1 expression is dependent on AhR activation. The effect of skatole was reduced by blocking intrinsic cytochrome P450 activity and indole-3-carbinole, a known skatole metabolite, was a more potent inducer than skatole. Finally, skatole could reduce TCDD-induced CYP1A1 expression, suggesting that skatole is a partial AhR agonist. In conclusion, our findings suggest that skatole and its metabolites affect liver homeostasis by modulating the AhR pathway. PMID:27138278

  20. The novel Aryl hydrocarbon receptor inhibitor biseugenol inhibits gastric tumor growth and peritoneal dissemination

    PubMed Central

    Lai, De-Wei; Karlsson, Anna Isabella; Wang, Keh-Bin; Chen, Yi-Ching; Shen, Chin-Chang; Wu, Sheng-Mao; Liu, Chia-Yu; Tien, Hsing-Ru; Peng, Yen-Chun; Jan, Yee-Jee; Chao, Te-Hsin; Lan, Keng-Hsin; Arbiser, Jack L.; Sheu, Meei-Ling

    2014-01-01

    Biseugenol (Eug) is known to antiproliferative of cancer cells; however, to date, the antiperitoneal dissemination effects have not been studied in any mouse cancer model. In this study, Aryl hydrocarbon receptor (AhR) expression was associated with lymph node and distant metastasis in patients with gastric cancer and was correlated with clinicolpathological pattern. We evaluated the antiperitoneal dissemination potential of knockdown AhR and Biseugenol in cancer mouse model and assessed mesenchymal characteristics. Our results demonstrate that tumor growth, peritoneal dissemination and peritoneum or organ metastasis implanted MKN45 cells were significantly decreased in shAhR and Biseugenol-treated mice and that endoplasmic reticulum (ER) stress was caused. Biseugenol-exposure tumors showed acquired epithelial features such as phosphorylation of E-cadherin, cytokeratin-18 and loss mesenchymal signature Snail, but not vimentin regulation. Snail expression, through AhR activation, is an epithelial-to-mesenchymal transition (EMT) determinant. Moreover, Biseugenol enhanced Calpain-10 (Calp-10) and AhR interaction resulted in Snail downregulation. The effect of shCalpain-10 in cancer cells was associated with inactivation of AhR/Snail promoter binding activity. Inhibition of Calpain-10 in gastric cancer cells by short hairpin RNA or pharmacological inhibitor was found to effectively reduced growth ability and vessel density in vivo. Importantly, knockdown of AhR completed abrogated peritoneal dissemination. Herein, Biseugenol targeting ER stress provokes Calpain-10 activity, sequentially induces reversal of EMT and apoptosis via AhR may involve the paralleling processes. Taken together, these data suggest that Calpain-10 activation and AhR inhibition by Biseugenol impedes both gastric tumor growth and peritoneal dissemination by inducing ER stress and inhibiting EMT. PMID:25226618

  1. Genetic association of aromatic hydrocarbon receptor (AHR) and cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) polymorphisms with dioxin blood concentrations among pregnant Japanese women.

    PubMed

    Kobayashi, Sumitaka; Sata, Fumihiro; Sasaki, Seiko; Ban, Susumu; Miyashita, Chihiro; Okada, Emiko; Limpar, Mariko; Yoshioka, Eiji; Kajiwara, Jumboku; Todaka, Takashi; Saijo, Yasuaki; Kishi, Reiko

    2013-06-01

    Dioxins are metabolized by cytochrome P450, family 1 (CYP1) via the aromatic hydrocarbon receptor (AHR). We determined whether different blood dioxin concentrations are associated with polymorphisms in AHR (dbSNP ID: rs2066853), AHR repressor (AHRR; rs2292596), CYP1 subfamily A polypeptide 1 (CYP1A1; rs4646903 and rs1048963), CYP1 subfamily A polypeptide 2 (CYP1A2; rs762551), and CYP1 subfamily B polypeptide 1 (CYP1B1; rs1056836) in pregnant Japanese women. These six polymorphisms were detected in 421 healthy pregnant Japanese women. Differences in dioxin exposure concentrations in maternal blood among the genotypes were investigated. Comparisons among the GG, GA, and AA genotypes of AHR showed a significant difference (genotype model: P=0.016 for the mono-ortho polychlorinated biphenyl concentrations and toxicity equivalence quantities [TEQs]). Second, we found a significant association with the dominant genotype model ([TT+TC] vs. CC: P=0.048 for the polychlorinated dibenzo-p-dioxin TEQs; P=0.035 for polychlorinated dibenzofuran TEQs) of CYP1A1 (rs4646903). No significant differences were found among blood dioxin concentrations and polymorphisms in AHRR, CYP1A1 (rs1048963), CYP1A2, and CYP1B1. Thus, polymorphisms in AHR and CYP1A1 (rs4646903) were associated with maternal dioxin concentrations. However, differences in blood dioxin concentrations were relatively low. PMID:23528250

  2. Hematopoietic stem and progenitor cell activation during chronic dermatitis provoked by constitutively active aryl-hydrocarbon receptor driven by Keratin 14 promoter.

    PubMed

    Murakami, Shohei; Yamamoto, Masayuki; Motohashi, Hozumi

    2014-03-01

    Polycyclic aromatic hydrocarbons (PAHs) activate aryl-hydrocarbon receptor (AhR). Because PAHs are known as a risk factor for allergic diseases, PAH-induced AhR activation is expected to be involved in the development of the pathology. We previously generated transgenic mice expressing a constitutively active AhR (AhR-CA) under the control of Keratin 14 (K14) promoter (AhR-CA mouse). The mice develop chronic dermatitis with immune imbalance toward Th2 predominance, indicating that the AhR activation driven by K14 promoter provokes allergic response. Because hematopoietic cells actively participate in the development of allergic inflammation, it is important to understand the hematopoietic status under allergic conditions. To clarify how the K14 promoter-driven AhR activation influences hematopoiesis, we analyzed bone marrow and spleen of AhR-CA mice. We verified that AhR-CA was expressed in keratinocytes and thymic epithelial cells but not in hematopoietic cells. The AhR-CA mice with full-blown dermatitis exhibited leukocytosis and skewed differentiation of hematopoietic progenitor cells toward granulocyte-monocyte lineages. They also showed a significant expansion of short-term hematopoietic stem cells and multipotent progenitors and a subtle reduction in long-term hematopoietic stem cells (LT-HSCs). Their spleens were enlarged and abundantly accumulated hematopoietic stem and progenitor cells. AhR-CA mice at the early stage of dermatitis did not show leukocytosis or splenomegaly but exhibited the granulocyte-monocyte skewing and the reduction in LT-HSCs. Thus, AhR activation driven by K14 promoter already alters the hematopoietic differentiation and reduces LT-HSCs at the initial stage of dermatitis development. These results suggest that nonhematopoietic exposure to PAHs triggers allergic response and concomitantly affects hematopoiesis.

  3. A new cross-talk between the Aryl hydrocarbon receptor and RelB, a member of the NF-κB family

    PubMed Central

    Vogel, Christoph F. A.; Matsumura, Fumio

    2009-01-01

    The discovery of the new crosstalk between the aryl hydrocarbon receptor (AhR) and the NF-κB subunit RelB may extend our understanding of the biological functions of the AhR and at the same time raises a number of questions, which will be addressed in this review. The characteristics of this interaction differ from that of AhR with RelA in that the latter appears to be mostly negative unlike the collaborative interactions of AhR/RelB. The AhR/RelB dimer is capable of binding to DNA response elements including the dioxin response element (DRE) as well as NF-κB binding sites supporting the activation of target genes of the AhR as well as NF-κB pathway. Further studies show that AhR/RelB complexes can be found not only in lymphoid cells but also in a human hepatoma cell line (HepG2) or breast cancer cell line (MDA-MB-231). RelB has been implicated in carcinogenesis of breast cancer for instance and RelB is known to be a critical factor for the function and differentiation of dendritic cells; interestingly the participation of AhR in both processes has been suggested recently, which offers the great potential to expand the scope of the physiological roles of the AhR. There is evidence indicating that RelB may serve as a pro-survival factor, including its ability to promote “inflammation resolution” besides the association of RelB with inflammatory disorders. Based on such information, a hypothesis has been proposed in this review that AhR together with RelB functions as a coordinator of inflammatory responses. PMID:18955032

  4. Identification of interacting proteins with aryl hydrocarbon receptor in scallop Chlamys farreri by yeast two hybrid screening.

    PubMed

    Cai, Yuefeng; Pan, Luqing; Miao, Jingjing; Liu, Tong

    2016-11-01

    The aryl hydrocarbon receptor (AhR) belongs to the basic-helix-loop helix (bHLH) Per-Arnt-Sim (PAS) family of transcription factors. AhR has been known primarily for its role in the regulation of several drug and xenobiotic metabolizing enzymes, as well as the mediation of the toxicity of certain xenobiotics, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Although the AhR is well-studied as a mediator of the toxicity of certain xenobiotics in marine bivalves, the normal physiological function remains unknown. In order to explore the function of the AhR, the bait protein expression plasmid pGBKT7-CfAhR and the cDNA library of gill from Chlamys farreri were constructed. By yeast two hybrid system, after multiple screening with the high screening rate medium, rotary verification, sequencing and bioinformatics analysis, the interactions of the CfAhR with receptor for activated protein kinase C 1 (RACK1), thyroid peroxidase-like protein (TPO), Toll-like receptor 4(TLR 4), androglobin-like, store-operated Ca(2+) entry (SocE), ADP/ATP carrier protein, cytochrome b, thioesterase, actin, ferritin subunit 1, poly-ubiquitin, short-chain collagen C4-like and one hypothetical protein in gill cells were identified. This study suggests that the CfAhR played fundamental roles in immune system homeostasis, oxidative stress response, and in grow and development of C. farreri. The elucidation of these protein interactions is of much importance both in understanding the normal physiological function of AhR, and as potential targets for further research on protein function in AhR interactions. PMID:27497785

  5. Identification of interacting proteins with aryl hydrocarbon receptor in scallop Chlamys farreri by yeast two hybrid screening.

    PubMed

    Cai, Yuefeng; Pan, Luqing; Miao, Jingjing; Liu, Tong

    2016-11-01

    The aryl hydrocarbon receptor (AhR) belongs to the basic-helix-loop helix (bHLH) Per-Arnt-Sim (PAS) family of transcription factors. AhR has been known primarily for its role in the regulation of several drug and xenobiotic metabolizing enzymes, as well as the mediation of the toxicity of certain xenobiotics, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Although the AhR is well-studied as a mediator of the toxicity of certain xenobiotics in marine bivalves, the normal physiological function remains unknown. In order to explore the function of the AhR, the bait protein expression plasmid pGBKT7-CfAhR and the cDNA library of gill from Chlamys farreri were constructed. By yeast two hybrid system, after multiple screening with the high screening rate medium, rotary verification, sequencing and bioinformatics analysis, the interactions of the CfAhR with receptor for activated protein kinase C 1 (RACK1), thyroid peroxidase-like protein (TPO), Toll-like receptor 4(TLR 4), androglobin-like, store-operated Ca(2+) entry (SocE), ADP/ATP carrier protein, cytochrome b, thioesterase, actin, ferritin subunit 1, poly-ubiquitin, short-chain collagen C4-like and one hypothetical protein in gill cells were identified. This study suggests that the CfAhR played fundamental roles in immune system homeostasis, oxidative stress response, and in grow and development of C. farreri. The elucidation of these protein interactions is of much importance both in understanding the normal physiological function of AhR, and as potential targets for further research on protein function in AhR interactions.

  6. A potential endogenous ligand for the aryl hydrocarbon receptor has potent agonist activity in vitro and in vivo.

    PubMed

    Henry, E C; Bemis, J C; Henry, O; Kende, A S; Gasiewicz, T A

    2006-06-01

    The aryl hydrocarbon receptor (AhR) is best known as a mediator of toxicity of a diverse family of xenobiotic chemicals such as dioxins and PCBs. However, many naturally occurring compounds also activate AhR. One such compound, 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE), was isolated from tissue and found to be potent in preliminary tests [J. Song, M. Clagett-Dame, R.E. Peterson, M.E. Hahn, W.M. Westler, R.R. Sicinski, H.F. DeLuca, Proc. Natl. Acad. Sci. USA 99 (2002) 14694-14699]. We have synthesized ITE and [(3)H]ITE and further evaluated its AhR activity in several in vitro and in vivo assays in comparison with the toxic ligand, TCDD. AhR in Hepa1c1c7 cell cytosol bound [(3)H]ITE with high affinity and the AhR.ITE complex formed in vitro bound dioxin response element (DRE) oligonucleotide as potently as TCDD.AhR. In cells treated with ITE, nuclear translocation of AhR, and induction of CYP1A1 protein and of a DRE-dependent luciferase reporter gene were observed. ITE administered to pregnant DRE-LacZ transgenic mice activated fetal AhR, observed as X-gal staining in the same sites as in TCDD-treated mice. However, unlike TCDD, ITE did not induce cleft palate or hydronephrosis. TCDD but not ITE induced thymic atrophy in young adult mice, but both ITE and TCDD caused similar loss of cells and alterations of cell profiles in cultured fetal thymi. These data demonstrate that ITE is a potent AhR agonist in cell extracts, cultured cells, and intact animals, but does not cause the toxicity associated with the more stable xenobiotic ligand, TCDD.

  7. Cobaltous chloride and hypoxia inhibit aryl hydrocarbon receptor-mediated responses in breast cancer cells

    SciTech Connect

    Khan, Shaheen; Liu Shengxi; Stoner, Matthew; Safe, Stephen

    2007-08-15

    The aryl hydrocarbon receptor (AhR) is expressed in estrogen receptor (ER)-positive ZR-75 breast cancer cells. Treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces CYP1A1 protein and mRNA levels and also activates inhibitory AhR-ER{alpha} crosstalk associated with hormone-induced reporter gene expression. In ZR-75 cells grown under hypoxia, induction of these AhR-mediated responses by TCDD was significantly inhibited. This was not accompanied by decreased nuclear AhR levels or decreased interaction of the AhR complex with the CYP1A1 gene promoter as determined in a chromatin immunoprecipitation assay. Hypoxia-induced loss of Ah-responsiveness was not associated with induction of hypoxia-inducible factor-1{alpha} or other factors that sequester the AhR nuclear translocation (Arnt) protein, and overexpression of Arnt under hypoxia did not restore Ah-responsiveness. The p65 subunit of NF{kappa}B which inhibits AhR-mediated transactivation was not induced by hypoxia and was primarily cytosolic in ZR-75 cells grown under hypoxic and normoxic conditions. In ZR-75 cells maintained under hypoxic conditions for 24 h, BRCA1 (an enhancer of AhR-mediated transactivation in breast cancer cells) was significantly decreased and this contributed to loss of Ah-responsiveness. In cells grown under hypoxia for 6 h, BRCA1 was not decreased, but induction of CYP1A1 by TCDD was significantly decreased. Cotreatment of ZR-75 cells with TCDD plus the protein synthesis inhibitor cycloheximide for 6 h enhanced CYP1A1 expression in cells grown under hypoxia and normoxia. These results suggest that hypoxia rapidly induces protein(s) that inhibit Ah-responsiveness and these may be similar to constitutively expressed inhibitors of Ah-responsiveness (under normoxia) that are also inhibited by cycloheximide.

  8. An introduction to the molecular basics of aryl hydrocarbon receptor biology.

    PubMed

    Abel, Josef; Haarmann-Stemmann, Thomas

    2010-11-01

    Depending on their chemical structure and properties, environmental chemicals and other xenobiotics that enter the cell can affect cellular function by either nonselective binding to cellular macromolecules or by interference with cellular receptors, which would initiate a more defined cell biological response. One of these intracellular chemosensor molecules is the aryl hydrocarbon receptor (AhR), a transcription factor of the bHLH/PAS family that is known to mediate the biochemical and toxic effects of dioxins, polyaromatic hydrocarbons and related compounds. Numerous investigations have revealed that the AhR is not only a master regulator of drug metabolism activated by anthropogenic chemicals, but is also triggered by natural and endogenous ligands and can influence cell biological endpoints such as growth and differentiation. Cutting-edge research has identified new intriguing functions of the AhR, such as during proteasomal degradation of steroid hormone receptors, the cellular UVB stress response and the differentiation of certain T-cell subsets. In this review we provide both a survey of the fundamental basics of AhR biology and an insight into new functional aspects of AhR signaling to further stimulate research on this intriguing transcription factor at the interface between toxicology, cell biology and immunology.

  9. AHR-11797: a novel benzodiazepine antagonist

    SciTech Connect

    Johnson, D.N.; Kilpatrick, B.F.; Hannaman, P.K.

    1986-03-01

    AHR-11797(5,6-dihydro-6-methyl-1-phenyl-/sup 3/H-pyrrolo(3,2,1-ij)quinazolin-3-one) displaced /sup 3/H-flunitrazepam (IC/sub 50/ = 82 nM) and /sup 3/H-Ro 15-1877 (IC/sub 50/ = 104 nM) from rat brain synaptosomes. AHR-11797 did not protect mice from seizures induced by maximal electroshock or subcutaneous Metrazol (scMET), nor did it induce seizures in doses up to the lethal dose. However, at 31.6 mg/kg, IP, it significantly increased the anticonvulsant ED/sub 50/ of chlordiazepoxide (CDPX) from 1.9 to 31.6 mg/kg, IP. With 56.7 mg/kg, IP, of AHR-11797, CDPX was inactive in doses up to 100 mg/kg, IP. AHR-11797 did not significantly increase punished responding in the Geller and Seifter conflict procedure, but it did attenuate the effects of diazepam. Although the compound is without anticonvulsant or anxiolytic activity, it did have muscle relaxant properties. AHR-11797 blocked morphine-induced Straub tail in mice (ED/sub 50/ = 31 mg/kg, IP) and it selectively suppressed the polysnaptic linguomandibular reflex in barbiturate-anesthetized cats. The apparent muscle relaxant activity of AHR-11797 suggests that different receptor sites are involved for muscle relaxant vs. anxiolytic/anticonvulsant activities of the benzodiazepines.

  10. Molecular docking, molecular dynamics simulation, and structure-based 3D-QSAR studies on the aryl hydrocarbon receptor agonistic activity of hydroxylated polychlorinated biphenyls.

    PubMed

    Cao, Fu; Li, Xiaolin; Ye, Li; Xie, Yuwei; Wang, Xiaoxiang; Shi, Wei; Qian, Xiangping; Zhu, Yongliang; Yu, Hongxia

    2013-09-01

    The binding interactions between hydroxylated polychlorinated biphenyls (HO-PCBs) and the aryl hydrocarbon receptor (AhR) are suspected of causing toxic effects. To understand the binding mode between HO-PCBs and AhR, and to explore the structural characteristics that influence the AhR agonistic activities of HO-PCBs, the combination of molecular docking, three-dimensional quantitative structure-activity relationship (3D-QSAR), and molecular dynamics (MD) simulations was performed. Using molecular docking, the HO-PCBs were docked into the binding pocket of AhR, which was generated by homology modeling. Comparative molecular similarity index analysis (CoMSIA) models were subsequently developed from three different alignment rules. The optimum 3D-QSAR model showed good predictive ability (q(2)=0.583, R(2)=0.913) and good mechanism interpretability. The statistical reliability of the CoMSIA model was also validated. In addition, molecular docking and MD simulations were applied to explore the binding modes between the ligands and AhR. The results obtained from this study may lead to a better understanding of the interaction mechanism between HO-PCBs and AhR.

  11. Exactly the Same but Different: Promiscuity and Diversity in the Molecular Mechanisms of Action of the Aryl Hydrocarbon (Dioxin) Receptor

    PubMed Central

    Denison, Michael S.; Soshilov, Anatoly A.; He, Guochun; DeGroot, Danica E.; Zhao, Bin

    2011-01-01

    The Ah receptor (AhR) is a ligand-dependent transcription factor that mediates a wide range of biological and toxicological effects that result from exposure to a structurally diverse variety of synthetic and naturally occurring chemicals. Although the overall mechanism of action of the AhR has been extensively studied and involves a classical nuclear receptor mechanism of action (i.e., ligand-dependent nuclear localization, protein heterodimerization, binding of liganded receptor as a protein complex to its specific DNA recognition sequence and activation of gene expression), details of the exact molecular events that result in most AhR-dependent biochemical, physiological, and toxicological effects are generally lacking. Ongoing research efforts continue to describe an ever-expanding list of ligand-, species-, and tissue-specific spectrum of AhR-dependent biological and toxicological effects that seemingly add even more complexity to the mechanism. However, at the same time, these studies are also identifying and characterizing new pathways and molecular mechanisms by which the AhR exerts its actions and plays key modulatory roles in both endogenous developmental and physiological pathways and response to exogenous chemicals. Here we provide an overview of the classical and nonclassical mechanisms that can contribute to the differential sensitivity and diversity in responses observed in humans and other species following ligand-dependent activation of the AhR signal transduction pathway. PMID:21908767

  12. Recruitment of CREB1 and Histone Deacetylase 2 (HDAC2) to the Mouse Ltbp-1 Promoter Regulates its Constitutive Expression in a Dioxin Receptor-dependent Manner

    PubMed Central

    Gomez-Duran, Aurea; Ballestar, Esteban; Carvajal-Gonzalez, Jose M.; Marlowe, Jennifer L.; Puga, Alvaro; Esteller, Manel; Fernandez-Salguero, Pedro M.

    2010-01-01

    Latent TGFβ-binding protein 1 (LTBP-1) is a key regulator of TGFβ targeting and activation in the extracellular matrix. LTBP-1 is recognized as a major docking molecule to localize, and possibly to activate, TGFβ in the extracellular matrix. Despite this relevant function, the molecular mechanisms regulating Ltbp-1 transcription remain largely unknown. Previous results from our laboratory revealed that mouse embryonic fibroblasts (MEF) lacking dioxin receptor (AhR) had increased Ltbp-1 mRNA expression and elevated TGFβ activity, suggesting that AhR repressed Ltbp-1 transcription. Here, we have cloned the mouse Ltbp-1 gene promoter and analysed its mechanism of transcriptional repression by AhR. Reporter gene assays, AhR over-expression and site-directed mutagenesis showed that basal Ltbp-1 transcription is AhR-dependent. Chromatin immunoprecipitation (ChIP) and RNA interference (RNAi) revealed that AhR regulates Ltbp-1 transcription by a mechanism involving recruitment of co-activators such as CREB1 and co-repressors such as HDAC2 to the Ltbp-1 promoter. In AhR-expressing (AhR+/+) MEF cells, the recruitment of HDAC1, 2 and 4 correlated with decreased K8H4 acetylation and impaired binding of pCREBSer133 to the Ltbp-1 promoter, likely maintaining a constitutive repressed state. AhR−/− MEF cells had the opposite pattern of HDACs and pCREB1Ser133 binding to Ltbp-1 promoter, and therefore, over-expressed Ltbp-1 mRNA. In agreement, siRNA for HDAC2 increased Ltbp-1 expression and K8H4 acetylation in AhR+/+ but not in AhR−/− MEF cells. We suggest that HDAC2 binding keeps Ltbp-1 promoter repressed in AhR+/+ MEF cells, whereas in AhR-null MEF cells the absence of HDAC2 and the binding of pCREBSer133 allow Ltbp-1 transcription. Thus, epigenetics can contribute to constitutive Ltbp-1 repression by a mechanism requiring AhR activity. PMID:18508077

  13. Aryl hydrocarbon receptor SNP -130 C/T associates with dioxins susceptibility through regulating its receptor activity and downstream effectors including interleukin 24.

    PubMed

    Liu, Ge; Asanoma, Kazuo; Takao, Tomoka; Tsukimori, Kiyomi; Uchi, Hiroshi; Furue, Masutaka; Kato, Kiyoko; Wake, Norio

    2015-01-22

    Dioxins are persistent environmental pollutants that cause multiple adverse health effects in humans, mainly through binding to the ligand-activated transcription factor, aryl hydrocarbon receptor (AhR). Genetic variation in AhR may modulate the susceptibility to dioxins. In this study, we aimed to evaluate the effects of the single nucleotide polymorphism (SNP) -130 C/T in the AhR promoter on dioxin-inducible gene transcription, and to investigate interleukin-24 (IL-24) and interleukin-1β (IL-1β) as proxies for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure. Using primary human chorionic stromal cells, we found that cells with the TT genotype showed higher AhR mRNA and protein levels than did those of the CC genotype. Microarray was carried out to analyze the gene expression profiles of cells (CC and TT genotype) after exposing the cells to TCDD. Several genes associated with human disorders were more highly up-regulated in cells of the TT genotype. Higher up-regulation of IL-24 and IL-1β mRNA in cells with the TT genotype was observed. Furthermore, blood samples from 64 Yusho patients who were accidentally exposed to high concentrations of dioxins were analyzed for the genotype, dioxins concentrations and serum levels of IL-24 and IL-1β. We observed higher serum IL-24 levels and lower serum IL-1β levels in Yusho patients with the TT genotype than in those with the CC genotype. AhR SNP -130 C/T affects serum IL-24 and IL-1β levels, independently of serum dioxins concentrations in Yusho patients. Our observations demonstrate that SNP -130 C/T modulates AhR expression and expression levels of IL-24 and IL-1β, and suggest an association of AhR SNP -130 C/T with the susceptibility to dioxins.

  14. Genetic and pharmacological analysis identifies a physiological role for the AHR in epidermal differentiation

    PubMed Central

    van den Bogaard, Ellen; Podolsky, Michael; Smits, Jos; Cui, Xiao; John, Christian; Gowda, Krishne; Desai, Dhimant; Amin, Shantu; Schalkwijk, Joost; Perdew, Gary H.

    2015-01-01

    Stimulation of the aryl hydrocarbon receptor (AHR) by xenobiotics is known to affect epidermal differentiation and skin barrier formation. The physiological role of endogenous AHR signaling in keratinocyte differentiation is not known. We used murine and human skin models to address the hypothesis that AHR activation is required for normal keratinocyte differentiation. Using transcriptome analysis of Ahr-/- and Ahr+/+ murine keratinocytes, we found significant enrichment of differentially expressed genes linked to epidermal differentiation. Primary Ahr-/- keratinocytes showed a significant reduction in terminal differentiation gene and protein expression, similar to Ahr+/+ keratinocytes treated with AHR antagonists GNF351 and CH223191, or the selective AHR modulator (SAhRM), SGA360. In vitro keratinocyte differentiation led to increased AHR levels and subsequent nuclear translocation, followed by induced CYP1A1 gene expression. Monolayer cultured primary human keratinocytes treated with AHR antagonists also showed an impaired terminal differentiation program. Inactivation of AHR activity during human skin equivalent development severely impaired epidermal stratification, terminal differentiation protein expression and stratum corneum formation. As disturbed epidermal differentiation is a main feature of many skin diseases, pharmacological agents targeting AHR signaling or future identification of endogenous keratinocyte-derived AHR ligands should be considered as potential new drugs in dermatology. PMID:25602157

  15. Aryl hydrocarbon receptor is necessary to protect fetal human pulmonary microvascular endothelial cells against hyperoxic injury: Mechanistic roles of antioxidant enzymes and RelB

    SciTech Connect

    Zhang, Shaojie; Patel, Ananddeep; Chu, Chun; Jiang, Weiwu; Wang, Lihua; Welty, Stephen E.; Moorthy, Bhagavatula; Shivanna, Binoy

    2015-07-15

    Hyperoxia contributes to the development of bronchopulmonary dysplasia (BPD) in premature infants. Activation of the aryl hydrocarbon receptor (AhR) protects adult and newborn mice against hyperoxic lung injury by mediating increases in the expression of phase I (cytochrome P450 (CYP) 1A) and phase II (NADP(H) quinone oxidoreductase (NQO1)) antioxidant enzymes (AOE). AhR positively regulates the expression of RelB, a component of the nuclear factor-kappaB (NF-κB) protein that contributes to anti-inflammatory processes in adult animals. Whether AhR regulates the expression of AOE and RelB, and protects fetal primary human lung cells against hyperoxic injury is unknown. Therefore, we tested the hypothesis that AhR-deficient fetal human pulmonary microvascular endothelial cells (HPMEC) will have decreased RelB activation and AOE, which will in turn predispose them to increased oxidative stress, inflammation, and cell death compared to AhR-sufficient HPMEC upon exposure to hyperoxia. AhR-deficient HPMEC showed increased hyperoxia-induced reactive oxygen species (ROS) generation, cleavage of poly(ADP-ribose) polymerase (PARP), and cell death compared to AhR-sufficient HPMEC. Additionally, AhR-deficient cell culture supernatants displayed increased macrophage inflammatory protein 1α and 1β, indicating a heightened inflammatory state. Interestingly, loss of AhR was associated with a significantly attenuated CYP1A1, NQO1, superoxide dismutase 1(SOD1), and nuclear RelB protein expression. These findings support the hypothesis that decreased RelB activation and AOE in AhR-deficient cells is associated with increased hyperoxic injury compared to AhR-sufficient cells. - Highlights: • AhR deficiency potentiates oxygen toxicity in human fetal lung cells. • Deficient AhR signaling increases hyperoxia-induced cell death. • AhR deficiency increases hyperoxia-induced ROS generation and inflammation. • Anti-oxidant enzyme levels are attenuated in AhR-deficient lung cells

  16. AHR signaling in prostate growth, morphogenesis, and disease

    PubMed Central

    Vezina, Chad M.; Lin, Tien-Min; Peterson, Richard E.

    2010-01-01

    Most evidence of aryl hydrocarbon receptor (AHR) signaling in prostate growth, morphogenesis, and disease stems from research using 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to pharmacologically activate the AHR at various stages of development. This review discusses effects of TCDD on prostate morphogenesis and highlights interactions between AHR and other signaling pathways during normal and aberrant prostate growth. Although AHR signaling modulates estrogen and androgen signaling in other tissues, crosstalk between these steroid hormone receptors and AHR signaling cannot account for actions of TCDD on prostate morphogenesis. Instead, the AHR appears to act within a cooperative framework of developmental signals to regulate timing and patterning of prostate growth. Inappropriate activation of AHR signaling as a result of early life TCDD exposure disrupts the balance of these signals, impairs prostate morphogenesis, and has an imprinting effect on the developing prostate that predisposes to prostate disease in adulthood. Mechanisms of AHR signaling in prostate growth and disease are only beginning to be unraveled and recent studies have revealed its interactions with WNT5A, retinoic acid, fibroblast growth factor 10, and vascular endothelial growth factor signaling pathways. PMID:18977204

  17. The aryl hydrocarbon receptor is required for normal gonadotropin responsiveness in the mouse ovary

    SciTech Connect

    Barnett, Kimberly R.; Tomic, Dragana; Gupta, Rupesh K.; Babus, Janice K.; Roby, Katherine F.; Terranova, Paul F.; Flaws, Jodi A.

    2007-08-15

    The aryl hydrocarbon receptor (AHR) mediates the toxicity of a variety of environmental chemicals. Although little is known about the physiological role of the AHR, studies suggest that it plays an important role in regulating ovulation because Ahr deficient (AhRKO) mice have a reduced number of ovulations compared to wild-type (WT) mice. The reasons for the reduced ability of AhRKO mice to ovulate are unknown. Normal ovulation, however, requires estrous cyclicity, appropriate luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels, and LH and FSH responsiveness. Thus, the purpose of this study was to test the hypothesis that Ahr deletion regulates ovulation by altering cyclicity, FSH and LH levels, follicle-stimulating hormone receptor (Fshr) and luteinizing hormone receptor (Lhcgr) levels and/or gonadotropin responsiveness. The data indicate that AhRKO and WT mice have similar levels of FSH and LH, but AhRKO mice have reduced Fshr and Lhcgr mRNA levels compared to WT mice. Furthermore, AhRKO ovaries contain fewer corpora lutea compared to WT ovaries after 5 IU equine chorionic gonadotropin (eCG) treatment. Lastly, both AhRKO and WT mice ovulate a similar number of eggs in response to 5 IU human chorionic gonadotropin (hCG), but AhRKO mice ovulate fewer eggs than WT mice in response to 2.5 IU and 1.25 IU hCG. Collectively, these data indicate that AhRKO follicles have a reduced capacity to ovulate compared to WT follicles and that this is due to reduced responsiveness to gonadotropins. Thus, in addition to mediating toxicity of environmental chemicals, the Ahr is required for normal ovulation.

  18. Developmental toxicity of 4-ring polycyclic aromatic hydrocarbons in zebrafish is differentially dependent on AH receptor isoforms and hepatic cytochrome P4501A metabolism

    SciTech Connect

    Incardona, John P. . E-mail: john.incardona@noaa.gov; Day, Heather L.; Collier, Tracy K.; Scholz, Nathaniel L.

    2006-12-15

    Polycyclic aromatic hydrocarbons (PAHs) derived from fossil fuels are ubiquitous contaminants and occur in aquatic habitats as highly variable and complex mixtures of compounds containing 2 to 6 rings. For aquatic species, PAHs are generally accepted as acting through either of two modes of action: (1) 'dioxin-like' toxicity mediated by activation of the aryl hydrocarbon receptor (AHR), which controls a battery of genes involved in PAH metabolism, such as cytochrome P4501A (CYP1A) and (2) 'nonpolar narcosis', in which tissue uptake is dependent solely on hydrophobicity and toxicity is mediated through non-specific partitioning into lipid bilayers. As part of a systematic analysis of mechanisms of PAH developmental toxicity in zebrafish, we show here that three tetracyclic PAHs (pyrene, chrysene, and benz[a]anthracene) activate the AHR pathway tissue-specifically to induce distinct patterns of CYP1A expression. Using morpholino knockdown of ahr1a, ahr2, and cyp1a, we show that distinct embryolarval syndromes induced by exposure to two of these compounds are differentially dependent on tissue-specific activation of AHR isoforms or metabolism by CYP1A. Exposure of embryos with and without circulation (silent heart morphants) resulted in dramatically different patterns of CYP1A induction, with circulation required to deliver some compounds to internal tissues. Therefore, biological effects of PAHs cannot be predicted simply by quantitative measures of AHR activity or a compound's hydrophobicity. These results indicate that current models of PAH toxicity in fish are greatly oversimplified and that individual PAHs are pharmacologically active compounds with distinct and specific cellular targets.

  19. Characterization of the Antiallergic Drugs 3-[2-(2-Phenylethyl) benzoimidazole-4-yl]-3-hydroxypropanoic Acid and Ethyl 3-Hydroxy-3-[2-(2-phenylethyl)benzoimidazol-4-yl]propanoate as Full Aryl Hydrocarbon Receptor Agonists

    PubMed Central

    Morales, José Luis; Krzeminski, Jacek; Amin, Shantu; Perdew, Gary H.

    2008-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates most of the toxic effects of numerous chlorinated (e.g., TCDD) and nonchlorinated polycyclic aromatic compounds (e.g., benzo[a]pyrene). Studies in AhR null mice suggested that this receptor may also play a role in the modulation of immune responses. Recently, two drugs, namely, M50354 and M50367 (ethyl ester derivative of M50354), were described as AhR ligands with high efficacy toward reducing atopic allergic symptoms in an AhR-dependent manner by skewing T helper cell differentiation toward a TH1 phenotype [Negishi et al. (2005) J. Immunol. 175 (11), 7348–7356]. Surprisingly, these drugs were shown to have minimal activity toward inducing classical dioxin responsive element-driven AhR-mediated CYP1A1 transcription. We synthesized and reevaluated the ability of these drugs to regulate AhR activity. In contrast to previously published data, both M50354 and M50367 were found to be potent inducers of several AhR target genes, namely, CYP1A1, CYP1B1, and UGT1A2. M50367 was a more effective agonist than M50354, perhaps accounting for its higher bioavailability in vivo. However, M50354 was capable of displacing an AhR-specific radioligand more effectively than M50367. This is consistent with M50354 being the active metabolite of M50367. In conclusion, two selective inhibitors of TH2 differentiation are full AhR agonists. PMID:18179178

  20. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) exposure of normal human dermal fibroblasts results in AhR-dependent and -independent changes in gene expression

    SciTech Connect

    Akintobi, A.M.; Villano, C.M.; White, L.A. . E-mail: lawhite@aesop.rutgers.edu

    2007-04-01

    Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) results in a variety of lesions in mammals including severe skin lesions. The majority of TCDD's biological effects are mediated through activation of the aryl hydrocarbon receptor (AhR). We have chosen to examine the effect of TCDD and the AhR pathway on dermal fibroblasts because this cell type plays an integral role in skin homeostasis through the production of cytokines and other factors that regulate epidermal proliferation and differentiation. Our data show that normal human dermal fibroblasts (NHDFs) are responsive to TCDD, as demonstrated by induction of cytochrome p450 1B1 (CYP1B1) expression. Further, our data demonstrate that TCDD treatment of NHDFs results in significant (75-90%) decrease in expression of Id-1 and Id-3, proteins that are involved in regulation of cell proliferation and differentiation. The Id (Inhibitor of DNA binding) proteins are transcriptional inhibitors that function by forming inactive heterodimers with other HLH proteins. TCDD-repression of Id-1 and -3 is independent of de novo protein synthesis; co-treatment with cycloheximide has no effect on TCDD inhibition of Id-1 and Id-3. Co-treatment with the AhR antagonist {alpha}-naphthoflavone also does not block inhibition of Id-1 and Id-3 by TCDD, suggesting that TCDD inhibition of Id-1 and Id-3 is, at least in part, mediated independently of the AhR pathway. Our data also show that TCDD inhibits expression of the cell cycle regulatory gene p16{sup ink4a}, which is often linked to Id expression. TCDD-induced reduction of p16{sup ink4a} expression is also independent of protein synthesis and the AhR pathway.

  1. Upregulation of ABCG2 by romidepsin via the aryl hydrocarbon receptor pathway.

    PubMed

    To, Kenneth K W; Robey, Robert; Zhan, Zhirong; Bangiolo, Lois; Bates, Susan E

    2011-04-01

    Histone deacetylase inhibitors (HDACI) are promising anticancer agents and their use in combination with conventional anticancer drugs is currently under investigation. We previously reported cell line-specific upregulation of ABCG2, a multidrug resistance transporter shown to control oral bioavailability and CNS penetration, by the HDACI romidepsin, although the precise mechanism in a particular cell line remains to be determined. The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that can be activated by numerous environmental contaminants and has been shown to be a client protein of heat shock protein 90 (Hsp90). A xenobiotic response element was defined in the ABCG2 promoter and was shown to mediate AhR signaling. Activated AhR was found to be associated with the ABCG2 promoter only in cell line models that respond to romidepsin with ABCG2 upregulation. Our data suggest that romidepsin acetylated Hsp70 and inhibited the chaperone function of Hsp90, thereby allowing the dissociation of AhR from Hsp90. The dissociation of AhR from Hsp90 may be a prerequisite for the differential upregulation of ABCG2 by romidepsin. Increasing our understanding of the mechanism(s) governing differential upregulation of ABCG2 in response to romidepsin could provide an insight into strategies needed to tackle resistance to HDACIs in cancer therapeutics.

  2. The aryl hydrocarbon receptor ligand ITE inhibits TGFβ1-induced human myofibroblast differentiation.

    PubMed

    Lehmann, Geniece M; Xi, Xia; Kulkarni, Ajit A; Olsen, Keith C; Pollock, Stephen J; Baglole, Carolyn J; Gupta, Shikha; Casey, Ann E; Huxlin, Krystel R; Sime, Patricia J; Feldon, Steven E; Phipps, Richard P

    2011-04-01

    Fibrosis can occur in any human tissue when the normal wound healing response is amplified. Such amplification results in fibroblast proliferation, myofibroblast differentiation, and excessive extracellular matrix deposition. Occurrence of these sequelae in organs such as the eye or lung can result in severe consequences to health. Unfortunately, medical treatment of fibrosis is limited by a lack of safe and effective therapies. These therapies may be developed by identifying agents that inhibit critical steps in fibrotic progression; one such step is myofibroblast differentiation triggered by transforming growth factor-β1 (TGFβ1). In this study, we demonstrate that TGFβ1-induced myofibroblast differentiation is blocked in human fibroblasts by a candidate endogenous aryl hydrocarbon receptor (AhR) ligand 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE). Our data show that ITE disrupts TGFβ1 signaling by inhibiting the nuclear translocation of Smad2/3/4. Although ITE functions as an AhR agonist, and biologically persistent AhR agonists, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin, cause severe toxic effects, ITE exhibits no toxicity. Interestingly, ITE effectively inhibits TGFβ1-driven myofibroblast differentiation in AhR(-/-) fibroblasts: Its ability to inhibit TGFβ1 signaling is AhR independent. As supported by the results of this study, the small molecule ITE inhibits myofibroblast differentiation and may be useful clinically as an antiscarring agent.

  3. Aryl hydrocarbon receptor facilitates DNA strand breaks and 8-oxo-2'-deoxyguanosine formation by the aldo-keto reductase product benzo[a]pyrene-7,8-dione.

    PubMed

    Park, Jong-Heum; Mangal, Dipti; Frey, Alexander J; Harvey, Ronald G; Blair, Ian A; Penning, Trevor M

    2009-10-23

    Polycyclic aromatic hydrocarbon (PAH) o-quinones produced by aldo-keto reductases are ligands for the aryl hydrocarbon receptor (AhR) (Burczynski, M. E., and Penning, T. M. (2000) Cancer Res. 60, 908-915). They induce oxidative DNA lesions (reactive oxygen species-mediated DNA strand breaks and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dGuo) formation) in human lung cells. We tested whether the AhR enhances PAH o-quinone-mediated oxidative DNA damage by translocating these ligands to the nucleus. Using the single cell gel electrophoresis (comet) assay to detect DNA strand breaks in murine hepatoma Hepa1c1c7 cells and its AhR- and aryl hydrocarbon receptor nuclear translocator-deficient variants, benzo[a]pyrene-7,8-dione (B[a]P-7,8-dione) produced fewer DNA strand breaks in AhR-deficient cells compared with aryl hydrocarbon receptor nuclear translocator-deficient and wild type Hepa1c1c7 cells. Decreased DNA strand breaks were also observed in human bronchoalveolar H358 cells in which the AhR was silenced by siRNA. The antioxidant alpha-tocopherol and the iron chelator/antioxidant desferal decreased the formation of B[a]P-7,8-dione-mediated DNA strand breaks indicating that they were reactive oxygen species-dependent. By coupling the comet assay to 8-oxoguanine glycosylase (hOGG1), which excises 8-oxo-Gua, strand breaks dependent upon this lesion were measured. hOGG1 treatment produced more DNA single strand breaks in B[a]P-7,8-dione-treated Hepa cells and H358 cells than in its absence. The levels of hOGG1-dependent DNA strand breaks mediated by B[a]P-7,8-dione were lower in AhR-deficient Hepa and AhR knockdown H358 cells. The AhR antagonist alpha-naphthoflavone also attenuated B[a]P-7,8-dione-mediated DNA strand breaks. The decrease in 8-oxo-dGuo levels in AhR-deficient Hepa cells and AhR knockdown H358 cells was validated by immunoaffinity capture stable isotope dilution ([(15)N(5)]8-oxo-dGuo) liquid chromatography-electrospray ionization/multiple reaction

  4. Mono-Substituted Isopropylated Triaryl Phosphate, a Major Component of Firemaster 550, is an AHR Agonist that Exhibits AHR-Independent Cardiotoxicity in Zebrafish

    PubMed Central

    Gerlach, Cory V.; Das, Siba R.; Volz, David C.; Bisson, William H.; Kolluri, Siva K.; Tanguay, Robert L.

    2014-01-01

    Firemaster 550 (FM550) is an additive flame retardant mixture used within polyurethane foam and is increasingly found in house dust and the environment due to leaching. Despite the widespread use of FM550, very few studies have investigated the potential toxicity of its ingredients during early vertebrate development. In the current study, we sought to specifically investigate mono-substituted isopropylated triaryl phosphate (mITP), a component comprising approximately 32% of FM550, which has been shown to cause cardiotoxicity during zebrafish embryogenesis. Previous research showed that developmental defects are rescued using an aryl hydrocarbon receptor (AHR) antagonist (CH223191), suggesting that mITP-induced toxicity was AHR-dependent. As zebrafish have three known AHR isoforms, we used a functional AHR2 knockout line along with AHR1A-and AHR1B-specific morpholinos to determine which AHR isoform, if any, mediates mITP-induced cardiotoxicity. As in silico structural homology modeling predicted that mITP may bind favorably to both AHR2 and AHR1B isoforms, we evaluated AHR involvement in vivo by measuring CYP1A mRNA and protein expression following exposure to mITP in the presence or absence of CH223191 or AHR-specific morpholinos. Based on these studies, we found that mITP interacts with both AHR2 and AHR1B isoforms to induce CYP1A expression. However, while CH223191 blocked mITP-induced CYP1A induction and cardiotoxicity, knockdown of all three AHR isoforms failed to block mITP-induced cardiotoxicity in the absence of detectable CYP1A induction. Overall, these results suggest that, while mITP is an AHR agonist, mITP causes AHR-independent cardiotoxicity through a pathway that is also antagonized by CH223191. PMID:24865613

  5. The aryl hydrocarbon receptor-activating effect of uremic toxins from tryptophan metabolism: a new concept to understand cardiovascular complications of chronic kidney disease.

    PubMed

    Sallée, Marion; Dou, Laetitia; Cerini, Claire; Poitevin, Stéphane; Brunet, Philippe; Burtey, Stéphane

    2014-03-04

    Patients with chronic kidney disease (CKD) have a higher risk of cardiovascular diseases and suffer from accelerated atherosclerosis. CKD patients are permanently exposed to uremic toxins, making them good candidates as pathogenic agents. We focus here on uremic toxins from tryptophan metabolism because of their potential involvement in cardiovascular toxicity: indolic uremic toxins (indoxyl sulfate, indole-3 acetic acid, and indoxyl-β-d-glucuronide) and uremic toxins from the kynurenine pathway (kynurenine, kynurenic acid, anthranilic acid, 3-hydroxykynurenine, 3-hydroxyanthranilic acid, and quinolinic acid). Uremic toxins derived from tryptophan are endogenous ligands of the transcription factor aryl hydrocarbon receptor (AhR). AhR, also known as the dioxin receptor, interacts with various regulatory and signaling proteins, including protein kinases and phosphatases, and Nuclear Factor-Kappa-B. AhR activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin and some polychlorinated biphenyls is associated with an increase in cardiovascular disease in humans and in mice. In addition, this AhR activation mediates cardiotoxicity, vascular inflammation, and a procoagulant and prooxidant phenotype of vascular cells. Uremic toxins derived from tryptophan have prooxidant, proinflammatory, procoagulant, and pro-apoptotic effects on cells involved in the cardiovascular system, and some of them are related with cardiovascular complications in CKD. We discuss here how the cardiovascular effects of these uremic toxins could be mediated by AhR activation, in a "dioxin-like" effect.

  6. Aryl hydrocarbon receptor knock-out exacerbates choroidal neovascularization via multiple pathogenic pathways.

    PubMed

    Choudhary, Mayur; Kazmin, Dmitri; Hu, Peng; Thomas, Russell S; McDonnell, Donald P; Malek, Goldis

    2015-01-01

    The aryl hydrocarbon receptor (AhR) is a heterodimeric transcriptional regulator with pleiotropic functions in xenobiotic metabolism and detoxification, vascular development and cancer. Herein, we report a previously undescribed role for the AhR signalling pathway in the pathogenesis of the wet, neovascular subtype of age-related macular degeneration (AMD), the leading cause of vision loss in the elderly in the Western world. Comparative analysis of gene expression profiles of aged AhR(-/-) and wild-type (wt) mice, using high-throughput RNA sequencing, revealed differential modulation of genes belonging to several AMD-related pathogenic pathways, including inflammation, angiogenesis and extracellular matrix regulation. To investigate AhR regulation of these pathways in wet AMD, we experimentally induced choroidal neovascular lesions in AhR(-/-) mice and found that they measured significantly larger in area and volume compared to age-matched wt mice. Furthermore, these lesions displayed a higher number of ionized calcium-binding adaptor molecule 1-positive (Iba1(+) ) microglial cells and a greater amount of collagen type IV deposition, events also seen in human wet AMD pathology specimens. Consistent with our in vivo observations, AhR knock-down was sufficient to increase choroidal endothelial cell migration and tube formation in vitro. Moreover, AhR knock-down caused an increase in collagen type IV production and secretion in both retinal pigment epithelial (RPE) and choroidal endothelial cell cultures, increased expression of angiogenic and inflammatory molecules, including vascular endothelial growth factor A (VEGFA) and chemokine (C-C motif) ligand 2 (CCL2) in RPE cells, and increased expression of secreted phosphoprotein 1 (SPP1) and transforming growth factor-β1 (TGFβ1) in choroidal endothelial cells. Collectively, our findings identify AhR as a regulator of multiple pathogenic pathways in experimentally induced choroidal neovascularization, findings that

  7. Diphenylarsinic acid, a chemical warfare-related neurotoxicant, promotes liver carcinogenesis via activation of aryl hydrocarbon receptor signaling and consequent induction of oxidative DAN damage in rats

    SciTech Connect

    Wei, Min; Yamada, Takanori; Yamano, Shotaro; Kato, Minoru; Kakehashi, Anna; Fujioka, Masaki; Tago, Yoshiyuki; Kitano, Mistuaki; Wanibuchi, Hideki

    2013-11-15

    Diphenylarsinic acid (DPAA), a chemical warfare-related neurotoxic organic arsenical, is present in the groundwater and soil in some regions of Japan due to illegal dumping after World War II. Inorganic arsenic is carcinogenic in humans and its organic arsenic metabolites are carcinogenic in animal studies, raising serious concerns about the carcinogenicity of DPAA. However, the carcinogenic potential of DPAA has not yet been evaluated. In the present study we found that DPAA significantly enhanced the development of diethylnitrosamine-induced preneoplastic lesions in the liver in a medium-term rat liver carcinogenesis assay. Evaluation of the expression of cytochrome P450 (CYP) enzymes in the liver revealed that DPAA induced the expression of CYP1B1, but not any other CYP1, CYP2, or CYP3 enzymes, suggesting that CYP1B1 might be the enzyme responsible for the metabolic activation of DPAA. We also found increased oxidative DNA damage, possibly due to elevated CYP1B1 expression. Induction of CYP1B1 has generally been linked with the activation of AhR, and we found that DPAA activates the aryl hydrocarbon receptor (AhR). Importantly, the promotion effect of DPAA was observed only at a dose that activated the AhR, suggesting that activation of AhR and consequent induction of AhR target genes and oxidative DNA damage plays a vital role in the promotion effects of DPAA. The present study provides, for the first time, evidence regarding the carcinogenicity of DPAA and indicates the necessity of comprehensive evaluation of its carcinogenic potential using long-term carcinogenicity studies. - Highlights: • DPAA, an environmental neurotoxicant, promotes liver carcinogenesis in rats. • DPAA is an activator of AhR signaling pathway. • DPAA promoted oxidative DNA damage in rat livers. • AhR target gene CYP 1B1 might be involved in the metabolism of DPAA.

  8. Interactions of polybrominated diphenyl ethers with the aryl hydrocarbon receptor pathway.

    PubMed

    Peters, A K; Nijmeijer, S; Gradin, K; Backlund, M; Bergman, A; Poellinger, L; Denison, M S; Van den Berg, M

    2006-07-01

    Polybrominated diphenyl ethers (PBDEs) are brominated flame retardants that have been in use as additives in various consumer products. Structural similarities of PBDEs with other polyhalogenated aromatic hydrocarbons that show affinity for the aryl hydrocarbon receptor (AhR), such as some polychlorinated biphenyls, raised concerns about their possible dioxin-like properties. We studied the ability of environmentally relevant PBDEs (BDE-47, -99, -100, -153, -154, and -183) and the "planar" congener BDE-77 to bind and/or activate the AhR in stably transfected rodent hepatoma cell lines with an AhR-responsive enhanced green fluorescent protein (AhR-EGFP) reporter gene (H1G1.1c3 mouse and H4G1.1c2 rat hepatoma). 7-Ethoxyresorufin-O-deethylation (EROD) was used as a marker for CYP1A1 activity. Dose- and bromination-specific inhibition of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced responses was measured by their ability to inhibit the induction of AhR-EGFP expression and EROD activity. Individual exposure to these PBDEs did not result in any increase in induction of AhR-EGFP or CYP1A1 activity. The lower brominated PBDEs showed the strongest inhibitory effect on TCDD-induced activities in both cell lines. While the highest brominated PBDE tested, BDE-183, inhibited EROD activity, it did not affect the induction of AhR-EGFP expression. Similar findings were observed after exposing stably transfected human hepatoma (xenobiotic response element [XRE]-HepG2) cells to these PBDEs, resulting in a small but statically significant agonistic effect on XRE-driven luciferase activity. Co-exposure with TCDD resulted again in antagonistic effects, confirming that the inhibitory effect of these PBDEs on TCDD-induced responses was not only due to direct interaction at receptor level but also at DNA-binding level. This antagonism was confirmed for BDE-99 in HepG2 cells transiently transfected with a Gal4-AhR construct and the corresponding Gal4-Luc reporter gene. In addition, a

  9. Interactions of polybrominated diphenyl ethers with the aryl hydrocarbon receptor pathway.

    PubMed

    Peters, A K; Nijmeijer, S; Gradin, K; Backlund, M; Bergman, A; Poellinger, L; Denison, M S; Van den Berg, M

    2006-07-01

    Polybrominated diphenyl ethers (PBDEs) are brominated flame retardants that have been in use as additives in various consumer products. Structural similarities of PBDEs with other polyhalogenated aromatic hydrocarbons that show affinity for the aryl hydrocarbon receptor (AhR), such as some polychlorinated biphenyls, raised concerns about their possible dioxin-like properties. We studied the ability of environmentally relevant PBDEs (BDE-47, -99, -100, -153, -154, and -183) and the "planar" congener BDE-77 to bind and/or activate the AhR in stably transfected rodent hepatoma cell lines with an AhR-responsive enhanced green fluorescent protein (AhR-EGFP) reporter gene (H1G1.1c3 mouse and H4G1.1c2 rat hepatoma). 7-Ethoxyresorufin-O-deethylation (EROD) was used as a marker for CYP1A1 activity. Dose- and bromination-specific inhibition of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced responses was measured by their ability to inhibit the induction of AhR-EGFP expression and EROD activity. Individual exposure to these PBDEs did not result in any increase in induction of AhR-EGFP or CYP1A1 activity. The lower brominated PBDEs showed the strongest inhibitory effect on TCDD-induced activities in both cell lines. While the highest brominated PBDE tested, BDE-183, inhibited EROD activity, it did not affect the induction of AhR-EGFP expression. Similar findings were observed after exposing stably transfected human hepatoma (xenobiotic response element [XRE]-HepG2) cells to these PBDEs, resulting in a small but statically significant agonistic effect on XRE-driven luciferase activity. Co-exposure with TCDD resulted again in antagonistic effects, confirming that the inhibitory effect of these PBDEs on TCDD-induced responses was not only due to direct interaction at receptor level but also at DNA-binding level. This antagonism was confirmed for BDE-99 in HepG2 cells transiently transfected with a Gal4-AhR construct and the corresponding Gal4-Luc reporter gene. In addition, a

  10. Analysis of the AHR gene proximal promoter GGGGC-repeat polymorphism in lung, breast, and colon cancer.

    PubMed

    Spink, Barbara C; Bloom, Michael S; Wu, Susan; Sell, Stewart; Schneider, Erasmus; Ding, Xinxin; Spink, David C

    2015-01-01

    The aryl hydrocarbon receptor (AhR) regulates expression of numerous genes, including those of the CYP1 gene family. With the goal of determining factors that control AHR gene expression, our studies are focused on the role of the short tandem repeat polymorphism, (GGGGC)n, located in the proximal promoter of the human AHR gene. When luciferase constructs containing varying GGGGC repeats were transfected into cancer cell lines derived from the lung, colon, and breast, the number of GGGGC repeats affected AHR promoter activity. The number of GGGGC repeats was determined in DNA from 327 humans and from 38 samples representing 5 species of non-human primates. In chimpanzees and 3 species of macaques, only (GGGGC)2 alleles were observed; however, in western gorilla, (GGGGC)n alleles with n=2, 4, 5, 6, 7, and 8 were identified. In all human populations examined, the frequency of (GGGGC)n was n=4>5≫2, 6. When frequencies of the (GGGGC)n alleles in DNA from patients with lung, colon, or breast cancer were evaluated, the occurrence of (GGGGC)2 was found to be 8-fold more frequent among lung cancer patients in comparison with its incidence in the general population, as represented by New York State neonates. Analysis of matched tumor and non-tumor DNA samples from the same individuals provided no evidence of microsatellite instability. These studies indicate that the (GGGGC)n short tandem repeats are inherited, and that the (GGGGC)2 allele in the AHR proximal promoter region should be further investigated with regard to its potential association with lung cancer susceptibility.

  11. Aryl hydrocarbon receptor-dependent induction of the IgA receptor FcαRI by the environmental contaminant benzo(a)pyrene in human macrophages.

    PubMed

    Pinel-Marie, Marie-Laure; Louarn, Laetitia; Desmots, Sophie; Fardel, Olivier; Sparfel, Lydie

    2011-11-28

    Polycyclic aromatic hydrocarbons (PAHs), such as benzo(a)pyrene (BaP), are widely distributed toxic environmental contaminants well known to regulate gene expression through activation of the aryl hydrocarbon receptor (AhR). In the present study, we demonstrated that the IgA receptor FcαRI/CD89 constitutes a molecular target for PAHs. Indeed, in vitro exposure to BaP markedly increased mRNA and protein expression of FcαRI in primary human macrophages; intratracheal instillation of BaP to rats also enhanced mRNA expression of FcαRI in alveolar macrophages. BaP concomitantly increased activity of the previously uncharacterized -1734 to -42 fragment of the FcaRI promoter that we subcloned in a luciferase reporter vector. Three-methylcholanthrene, a PAH known to activate AhR like BaP, induced FcαRI expression, in contrast to benzo(e)pyrene, a PAH known to poorly interact with AhR. Moreover, FcαRI induction in BaP-exposed human macrophages was fully prevented by down-regulating AhR expression through small interference RNA transfection. In addition, BaP increased nuclear protein binding to a consensus AhR-related xenobiotic-responsive element found in the FcαRI gene promoter, as revealed by electrophoretic mobility shift assay. Overall, these data highlight an AhR-dependent up-regulation of FcαRI in response to BaP, which may contribute to the deleterious effects of environmental PAHs toward the immune/inflammatory response and which also likely emphasizes the role played by AhR in the regulation of genes involved in immunity and inflammation.

  12. Differential suppression of the aryl hydrocarbon receptor nuclear translocator-dependent function by an aryl hydrocarbon receptor PAS-A-derived inhibitory molecule

    PubMed Central

    Xie, Jinghang; Huang, Xin; Park, Miki S.; Pham, Hang M.; Chan, William K.

    2014-01-01

    The aryl hydrocarbon receptor (AhR) heterodimerizes with the aryl hydrocarbon receptor nuclear translocator (Arnt) for transcriptional regulation. We generated three N-terminal deletion constructs of the human AhR of 12–24 KDa in size – namely D1, D2, and D3 – to suppress the Arnt function. We observed that all three deletions interact with the human Arnt with similar affinities. D2, which contains part of the AhR PAS-A domain and interacts with the PAS-A domain of Arnt, inhibits the formation of the AhR gel shift complex. D2 suppresses the 3-methylcholanthrene-induced, dioxin response element (DRE)-driven luciferase activity in Hep3B cells and exogenous Arnt reverses this D2 suppression. D2 suppresses the induction of CYP1A1 at both the message and protein levels in Hep3B cells; however, the CYP1B1 induction is not affected. D2 suppresses the recruitment of Arnt to the cyp1a1 promoter but not to the cyp1b1 promoter, partly because the AhR/Arnt heterodimer binds better to the cyp1b1 DRE than to the cyp1a1 DRE. Interestingly, D2 has no effect on the cobalt chloride-induced, hypoxia inducible factor-1 (HIF-1)-dependent expression of vegf, aldolase c, and ldh-a messages. Our data reveal that the flanking sequences of the DRE contribute to the binding affinity of the AhR/Arnt heterodimer to its endogenous enhancers and the function of AhR and HIF-1 can be differentially suppressed by the D2 inhibitory molecule. PMID:24486526

  13. Molecular evidence for the existence of an aryl hydrocarbon receptor pathway in scallops Chlamys farreri.

    PubMed

    Cai, Yuefeng; Pan, Luqing; Miao, Jingjing

    2016-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that controls the expression of a diverse set of genes. In this study we cloned full-length cDNAs encoding an AhR homologue (designated CfAhR, Accession number: FJ588640) from scallop Chlamys farreri. The CfAhR sequence was constituted by an open reading frame (ORF) of 2466bp encoding 821 amino acids. The predicted molecular weight was 93.0kDa. The CfAhR showed a high conservation of the residues and domains essential to the function of AhR, including basic helix-loop-helix (bHLH) and Per-ARNT-Sim (PAS) domains. Phylogenetic analysis demonstrated that it was clustered within the invertebrate AhR branch. CfAhR expression was detected in gill, digestive gland, ovary, spermary, mantle and adductor, and the highest transcription level was observed in gill. Recombinant plasmid CfAhR-pET32a (designated rCfAhR) was successfully expressed in Escherichia coli BL21. To investigate the molecular detoxification mechanism of benzo(a)pyrene (BaP) detoxification-related genes (AhR; aryl hydrocarbon receptor nuclear translocator, ARNT; heat shock protein 90, HSP90; cytochrome P450 1A1, CYP1A1; glutathione S-transferase pi, GST-pi and P glycoprotein, Pgp) in C. farreri gill, real-time quantitative PCR analysis revealed that the mRNA expression level of CfAhR, xenobiotic-metabolizing enzymes and efflux transporters was induced by BaP and was sensitive to BaP exposure time and concentration, suggesting that BaP influenced the expression of a putative AhR/ARNT signaling pathway in scallops. Our results support the possibility that CfAhR genes are early molecular indicators of BaP through a putative CYP signaling pathway in marine bivalves.

  14. Developing a qPCR method to quantify AhR-PCP-DNA complex for detection of environmental trace-level PCP.

    PubMed

    Zhao, Xiaoxiang; Pang, Xiaoqian; Chaisuwan, Nuanapa

    2011-07-01

    Pentachlorophenol (PCP), a widely-used aseptic or biocide, is known as an environmental toxicant involved in endocrine disruption even at a trace level. In order to reliably and efficiently quantify environmental trace-quantity PCP, this study developed a novel PCP detection method using the aryl hydrocarbon receptor (AhR) and fluorescence quantitative PCR (qPCR). DNA probe with AhR binding sites was synthesized by PCR before added into AhR-PCP complex. After AhR-PCP-DNA complex was digested with exonuclease, copy number of DNA probe was determined using fluorescence qPCR. To calculate PCP concentration in samples, a standard curve (PCP concentration versus Ct value) was constructed and the detection range was 10(-13) to 10(-9) M. PCP detection limit was 0.0089 ppt for the AhR-PCP-DNA complex assay and 8.8780 ppm for high performance liquid chromatography, demonstrating that the method developed in this study is more sensitive. These results suggest that AhR-PCP-DNA complex method may be successfully applicable in detection and quantification of environmental trace-level PCP. PMID:21503612

  15. Analysis of the AHR gene proximal promoter GGGGC-repeat polymorphism in lung, breast, and colon cancer

    SciTech Connect

    Spink, Barbara C.; Bloom, Michael S.; Wu, Susan; Sell, Stewart; Schneider, Erasmus; Ding, Xinxin; Spink, David C.

    2015-01-01

    The aryl hydrocarbon receptor (AhR) regulates expression of numerous genes, including those of the CYP1 gene family. With the goal of determining factors that control AHR gene expression, our studies are focused on the role of the short tandem repeat polymorphism, (GGGGC){sub n}, located in the proximal promoter of the human AHR gene. When luciferase constructs containing varying GGGGC repeats were transfected into cancer cell lines derived from the lung, colon, and breast, the number of GGGGC repeats affected AHR promoter activity. The number of GGGGC repeats was determined in DNA from 327 humans and from 38 samples representing 5 species of non-human primates. In chimpanzees and 3 species of macaques, only (GGGGC){sub 2} alleles were observed; however, in western gorilla, (GGGGC){sub n} alleles with n = 2, 4, 5, 6, 7, and 8 were identified. In all human populations examined, the frequency of (GGGGC){sub n} was n = 4 > 5 ≫ 2, 6. When frequencies of the (GGGGC){sub n} alleles in DNA from patients with lung, colon, or breast cancer were evaluated, the occurrence of (GGGGC){sub 2} was found to be 8-fold more frequent among lung cancer patients in comparison with its incidence in the general population, as represented by New York State neonates. Analysis of matched tumor and non-tumor DNA samples from the same individuals provided no evidence of microsatellite instability. These studies indicate that the (GGGGC){sub n} short tandem repeats are inherited, and that the (GGGGC){sub 2} allele in the AHR proximal promoter region should be further investigated with regard to its potential association with lung cancer susceptibility. - Highlights: • The AHR proximal promoter contains a polymorphism, (GGGGC){sub n}, where n = 4 > 5 ≫ 2, 6 • Matched tumor and non-tumor DNA did not show (GGGGC){sub n} microsatellite instability • AHR promoter activity of a construct with (GGGGC){sub 2} was lower than that of (GGGGC){sub 4} • The frequency of (GGGGC){sub 2} in lung

  16. PATHOGENESIS OF ARYL HYDROCARBON RECEPTOR-MEDIATED DEVELOPMENT OF LYMPHOMA IS ASSOCIATED WITH INCREASED CYCLOOXYGENSE-2 EXPRESSION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Epidemiological studies indicate that exposure to environmental pollutants like pesticides and dioxins lead to the pathogenesis of lymphoma and leukemia. Here we show that activation of the Aryl-hydrocarbon-Receptor (AhR) by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) results in loss of programmed ce...

  17. Quantitative High-Throughput Screening and Confirmation Studies for Identification of Compounds that Activate the Aryl Hydrocarbon Receptor Pathway (SETAC)

    EPA Science Inventory

    The aryl hydrocarbon receptor (AhR) is a transcription factor that mediates adaptive responses to known environmental pollutants, such as aromatic hydrocarbons, through regulation of Phase I and II xenobiotic metabolizing enzymes as well as important growth and differentiation pa...

  18. Source-Related Effects of Wastewater on Transcription Factor (AhR, CAR and PXR)-Mediated Induction of Gene Expression in Cultured Rat Hepatocytes and Their Association with the Prevalence of Antimicrobial-Resistant Escherichia coli

    PubMed Central

    Guruge, Keerthi S.; Yamanaka, Noriko; Sonobe, Miyuki; Fujizono, Wataru; Yoshioka, Miyako; Akiba, Masato; Yamamoto, Takehisa; Joshua, Derrick I.; Balakrishna, Keshava; Yamashita, Nobuyoshi; Kannan, Kurunthachalam; Tsutsui, Toshiyuki

    2015-01-01

    Extracts of wastewater collected from 4 sewage treatment plants (STPs) receiving effluents from different sources in South India were investigated for their levels of transcription factor-mediated gene induction in primary cultured rat hepatocytes. In addition, the relation between gene induction levels and the prevalence of antimicrobial-resistant Escherichia coli (E. coli) in wastewater was examined. STP-3, which treats only hospital wastewater, exhibited significantly greater induction potency of all 6 drug metabolizing cytochrome P450 (CYP) genes examined, CYP1A1, 1A2, 1B1, 2B15, 3A1, and 3A2, whereas the wastewater at STP-1, which exclusively receives domestic sewage, showed significantly diminished levels of induction of 3 CYP genes when compared to the levels of CYP induction at STP-2, which receives mixed wastewater. Samples collected during the monsoon season showed a significantly altered gene induction capacity compared to that of samples from the pre-monsoon period. The data suggest that the toxicity of wastewater in STPs was not significantly diminished during the treatment process. The chemical-gene interaction data predicted that a vast number of chemicals present in the wastewater would stimulate the genes studied in the rat hepatocytes. The multivariable logistic regression analysis demonstrated that the prevalence of isolates resistant to cefotaxime, imipenem and streptomycin was significantly correlated with the levels of induction of at least three CYP-isozymes in STP wastewater. In addition, the resistance of isolates in treatment plants was not altered by the treatment steps, whereas the sampling season did have an impact on the resistance to specific antimicrobials. The identification of receptor-mediated gene regulation capacities offers important data not limited to the (synergistic) physiological role of chemicals in biological systems but may provide new insight into the link between the effects of known/unknown drugs and prevalence of

  19. AHR promoter variant modulates its transcription and downstream effectors by allele-specific AHR-SP1 interaction functioning as a genetic marker for vitiligo.

    PubMed

    Wang, Xiaowen; Li, Kai; Liu, Ling; Shi, Qiong; Song, Pu; Jian, Zhe; Guo, Sen; Wang, Gang; Li, Chunying; Gao, Tianwen

    2015-09-15

    Vitiligo is an acquired depigmentation disorder largely caused by defective melanocyte- or autoimmunity-induced melanocyte destruction. The aryl hydrocarbon receptor (AHR) is essential for melanocyte homeostasis and immune process, and abnormal AHR was observed in vitiligo. We previously identified the T allele of AHR -129C > T variant as a protective factor against vitiligo. However, biological characterization underlying such effects is not fully certain, further validation by mechanistic research is warranted and was conducted in the present study. We showed that -129T allele promoted AHR transcriptional activity through facilitating its interaction with SP1 transcription factor (SP1) compared with -129C allele. We subsequently found reduced peripheral AHR and SP1 transcript expressions in vitiligo and a negative correlation of AHR level with disease duration. We also investigated AHR-related cytokines and observed increased serum TNF-α concentration and diminished serum levels of IL-10 and TGF-β1 in vitiligo. Further genetic analysis showed that -129T carriers possessed higher levels of AHR and IL-10 than -129C carriers. Therefore, our study indicates that the modulation of AHR transcription by a promoter variant has a profound influence on vitiligo, not only advancing our understanding on AHR function but also providing novel insight into the pathogenesis of degenerative or autoimmune diseases including vitiligo.

  20. AHR promoter variant modulates its transcription and downstream effectors by allele-specific AHR-SP1 interaction functioning as a genetic marker for vitiligo

    PubMed Central

    Wang, Xiaowen; Li, Kai; Liu, Ling; Shi, Qiong; Song, Pu; Jian, Zhe; Guo, Sen; Wang, Gang; Li, Chunying; Gao, Tianwen

    2015-01-01

    Vitiligo is an acquired depigmentation disorder largely caused by defective melanocyte- or autoimmunity-induced melanocyte destruction. The aryl hydrocarbon receptor (AHR) is essential for melanocyte homeostasis and immune process, and abnormal AHR was observed in vitiligo. We previously identified the T allele of AHR −129C > T variant as a protective factor against vitiligo. However, biological characterization underlying such effects is not fully certain, further validation by mechanistic research is warranted and was conducted in the present study. We showed that −129T allele promoted AHR transcriptional activity through facilitating its interaction with SP1 transcription factor (SP1) compared with −129C allele. We subsequently found reduced peripheral AHR and SP1 transcript expressions in vitiligo and a negative correlation of AHR level with disease duration. We also investigated AHR-related cytokines and observed increased serum TNF-α concentration and diminished serum levels of IL-10 and TGF-β1 in vitiligo. Further genetic analysis showed that -129T carriers possessed higher levels of AHR and IL-10 than −129C carriers. Therefore, our study indicates that the modulation of AHR transcription by a promoter variant has a profound influence on vitiligo, not only advancing our understanding on AHR function but also providing novel insight into the pathogenesis of degenerative or autoimmune diseases including vitiligo. PMID:26370050

  1. The immunomodulatory effect of laquinimod in CNS autoimmunity is mediated by the aryl hydrocarbon receptor.

    PubMed

    Berg, Johannes; Mahmoudjanlou, Yasaman; Duscha, Alexander; Massa, Megan G; Thöne, Jan; Esser, Charlotte; Gold, Ralf; Haghikia, Aiden

    2016-09-15

    Though several functional properties of laquinimod have been identified, our understanding of the underlying mechanisms is still incomplete. Since the compound elicits similar immunomodulatory effects to ligands of the aryl hydrocarbon receptor (AhR), we compared the efficacy of laquinimod in experimental autoimmune encephalomyelitis (EAE)-afflicted wild-type and AhR-deficient mice. Laquinimod failed to ameliorate clinical symptoms and leukocyte infiltration in AhR-deficient mice; however, treatment exerted neuroprotection by elevation of brain-derived neurotrophic factor (BDNF) independent of genetic profile. Thus, our data identify the AhR pathway in these mutant mice as crucial for the immunomodulatory, but not neuroprotective, efficacy of laquinimod in EAE.

  2. The immunomodulatory effect of laquinimod in CNS autoimmunity is mediated by the aryl hydrocarbon receptor.

    PubMed

    Berg, Johannes; Mahmoudjanlou, Yasaman; Duscha, Alexander; Massa, Megan G; Thöne, Jan; Esser, Charlotte; Gold, Ralf; Haghikia, Aiden

    2016-09-15

    Though several functional properties of laquinimod have been identified, our understanding of the underlying mechanisms is still incomplete. Since the compound elicits similar immunomodulatory effects to ligands of the aryl hydrocarbon receptor (AhR), we compared the efficacy of laquinimod in experimental autoimmune encephalomyelitis (EAE)-afflicted wild-type and AhR-deficient mice. Laquinimod failed to ameliorate clinical symptoms and leukocyte infiltration in AhR-deficient mice; however, treatment exerted neuroprotection by elevation of brain-derived neurotrophic factor (BDNF) independent of genetic profile. Thus, our data identify the AhR pathway in these mutant mice as crucial for the immunomodulatory, but not neuroprotective, efficacy of laquinimod in EAE. PMID:27609269

  3. Microbiome-Derived Tryptophan Metabolites and Their Aryl Hydrocarbon Receptor-Dependent Agonist and Antagonist Activities

    PubMed Central

    Jin, Un-Ho; Lee, Syng-Ook; Sridharan, Gautham; Lee, Kyongbum; Davidson, Laurie A.; Jayaraman, Arul; Chapkin, Robert S.; Alaniz, Robert

    2014-01-01

    The tryptophan metabolites indole, indole-3-acetate, and tryptamine were identified in mouse cecal extracts and fecal pellets by mass spectrometry. The aryl hydrocarbon receptor (AHR) agonist and antagonist activities of these microbiota-derived compounds were investigated in CaCo-2 intestinal cells as a model for understanding their interactions with colonic tissue, which is highly aryl hydrocarbon (Ah)–responsive. Activation of Ah-responsive genes demonstrated that tryptamine and indole 3-acetate were AHR agonists, whereas indole was an AHR antagonist that inhibited TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin)–induced CYP1A1 expression. In contrast, the tryptophan metabolites exhibited minimal anti-inflammatory activities, whereas TCDD decreased phorbol ester-induced CXCR4 [chemokine (C-X-C motif) receptor 4] gene expression, and this response was AHR dependent. These results demonstrate that the tryptophan metabolites indole, tryptamine, and indole-3-acetate modulate AHR-mediated responses in CaCo-2 cells, and concentrations of indole that exhibit AHR antagonist activity (100–250 μM) are detected in the intestinal microbiome. PMID:24563545

  4. Effects of currently used pesticides and their mixtures on the function of thyroid hormone and aryl hydrocarbon receptor in cell culture.

    PubMed

    Ghisari, Mandana; Long, Manhai; Tabbo, Agnese; Bonefeld-Jørgensen, Eva Cecilie

    2015-05-01

    Evidence suggest that exposure to pesticides can interfere with the endocrine system by multiple mechanisms. The endocrine disrupting potential of currently used pesticides in Denmark was analyzed as single compounds and in an equimolar mixture of 5 selected pesticides. The pesticides were previously analyzed for effects on the function of estrogen and androgen receptors, the aromatase enzyme and steroidogenesis in vitro. In this study, the effect on thyroid hormone (TH) function and aryl hydrocarbon receptor (AhR) transactivity was assessed using GH3 cell proliferation assay (T-screen) and AhR responsive luciferase reporter gene bioassay, respectively. Thirteen pesticides were analyzed as follows: 2-methyl-4-chlorophenoxyacetic acid, terbuthylazine, iodosulfuron-methyl-sodium, mesosulfuron-methyl, metsulfuron-methyl, chlormequat chloride, bitertanol, propiconazole, prothioconazole, mancozeb and its metabolite ethylene thiourea, cypermethrin, tau-fluvalinate, and malathion (currently banned in DK). In the T-screen, prothioconazole, malathion, tau-fluvalinate, cypermethrin, terbuthylazine and mancozeb significantly stimulated and bitertanol and propiconazole slightly reduced the GH3 cell proliferation. In the presence of triiodothyronine (T3), prothioconazole, tau-fluvalinate, propiconazole, cypermethrin and bitertanol significantly antagonized the T3-induced GH3 cell proliferation. Eleven of the tested pesticides agonized the AhR function, and bitertanol and prothioconazole inhibited the basal AhR activity. Bitertanol, propiconazole, prothioconazole and cypermethrin antagonized the TCDD-induced AhR transactivation at the highest tested concentration. The 5-component mixture had inducing effect but the combined effect could not be predicted due to the presence of bitertanol eliciting inhibitory effect. Upon removal of bitertanol from the mixture, the remaining four pesticides acted additively. In conclusion, our data suggest that pesticides currently used in Denmark

  5. Effects of currently used pesticides and their mixtures on the function of thyroid hormone and aryl hydrocarbon receptor in cell culture.

    PubMed

    Ghisari, Mandana; Long, Manhai; Tabbo, Agnese; Bonefeld-Jørgensen, Eva Cecilie

    2015-05-01

    Evidence suggest that exposure to pesticides can interfere with the endocrine system by multiple mechanisms. The endocrine disrupting potential of currently used pesticides in Denmark was analyzed as single compounds and in an equimolar mixture of 5 selected pesticides. The pesticides were previously analyzed for effects on the function of estrogen and androgen receptors, the aromatase enzyme and steroidogenesis in vitro. In this study, the effect on thyroid hormone (TH) function and aryl hydrocarbon receptor (AhR) transactivity was assessed using GH3 cell proliferation assay (T-screen) and AhR responsive luciferase reporter gene bioassay, respectively. Thirteen pesticides were analyzed as follows: 2-methyl-4-chlorophenoxyacetic acid, terbuthylazine, iodosulfuron-methyl-sodium, mesosulfuron-methyl, metsulfuron-methyl, chlormequat chloride, bitertanol, propiconazole, prothioconazole, mancozeb and its metabolite ethylene thiourea, cypermethrin, tau-fluvalinate, and malathion (currently banned in DK). In the T-screen, prothioconazole, malathion, tau-fluvalinate, cypermethrin, terbuthylazine and mancozeb significantly stimulated and bitertanol and propiconazole slightly reduced the GH3 cell proliferation. In the presence of triiodothyronine (T3), prothioconazole, tau-fluvalinate, propiconazole, cypermethrin and bitertanol significantly antagonized the T3-induced GH3 cell proliferation. Eleven of the tested pesticides agonized the AhR function, and bitertanol and prothioconazole inhibited the basal AhR activity. Bitertanol, propiconazole, prothioconazole and cypermethrin antagonized the TCDD-induced AhR transactivation at the highest tested concentration. The 5-component mixture had inducing effect but the combined effect could not be predicted due to the presence of bitertanol eliciting inhibitory effect. Upon removal of bitertanol from the mixture, the remaining four pesticides acted additively. In conclusion, our data suggest that pesticides currently used in Denmark

  6. Effects of currently used pesticides and their mixtures on the function of thyroid hormone and aryl hydrocarbon receptor in cell culture

    SciTech Connect

    Ghisari, Mandana; Long, Manhai; Tabbo, Agnese; Bonefeld-Jørgensen, Eva Cecilie

    2015-05-01

    Evidence suggest that exposure to pesticides can interfere with the endocrine system by multiple mechanisms. The endocrine disrupting potential of currently used pesticides in Denmark was analyzed as single compounds and in an equimolar mixture of 5 selected pesticides. The pesticides were previously analyzed for effects on the function of estrogen and androgen receptors, the aromatase enzyme and steroidogenesis in vitro. In this study, the effect on thyroid hormone (TH) function and aryl hydrocarbon receptor (AhR) transactivity was assessed using GH3 cell proliferation assay (T-screen) and AhR responsive luciferase reporter gene bioassay, respectively. Thirteen pesticides were analyzed as follows: 2-methyl-4-chlorophenoxyacetic acid, terbuthylazine, iodosulfuron-methyl-sodium, mesosulfuron-methyl, metsulfuron-methyl, chlormequat chloride, bitertanol, propiconazole, prothioconazole, mancozeb and its metabolite ethylene thiourea, cypermethrin, tau-fluvalinate, and malathion (currently banned in DK). In the T-screen, prothioconazole, malathion, tau-fluvalinate, cypermethrin, terbuthylazine and mancozeb significantly stimulated and bitertanol and propiconazole slightly reduced the GH3 cell proliferation. In the presence of triiodothyronine (T3), prothioconazole, tau-fluvalinate, propiconazole, cypermethrin and bitertanol significantly antagonized the T3-induced GH3 cell proliferation. Eleven of the tested pesticides agonized the AhR function, and bitertanol and prothioconazole inhibited the basal AhR activity. Bitertanol, propiconazole, prothioconazole and cypermethrin antagonized the TCDD-induced AhR transactivation at the highest tested concentration. The 5-component mixture had inducing effect but the combined effect could not be predicted due to the presence of bitertanol eliciting inhibitory effect. Upon removal of bitertanol from the mixture, the remaining four pesticides acted additively. In conclusion, our data suggest that pesticides currently used in Denmark

  7. Binding of aromatic amines to the rat hepatic Ah receptor in vitro and in vivo and the 8S and 4S estrogen receptor of rat uterus and rat liver

    SciTech Connect

    Cikryt, P.; Kaiser, T.; Gottlicher, M. )

    1990-08-01

    Studies on structurally related aromatic amines with different carcinogenic properties have shown that 2-acetylaminofluorene (2-AAF) and 2-acetylaminophenanthrene (AAP) inhibit the binding of 2,3,7,8-tetrachlorodibenzo-p-dioxin to the Ah receptor in vitro. The apparent inhibitor constants (K{sub i}) are 2.3 {mu}M for 2-AAF and 2.7 {mu}M for AAP. In contrast, 4-acetylaminofluorene, an isomer of 2-AAF, and trans-4-acetylaminostilbene do not bind to the rat hepatic cytosolic Ah receptor. Pretreating female Wistar rats with 2-AAF or AAP leads to the induction of the P-450 isoenzymes that are under the control of the Ah receptor. Ornithine decarboxylase activity is induced by all aromatic amines tested irrespective of their Ah receptor affinity. The aromatic amines used as model compounds do not inhibit the binding of 17-{beta}-estradiol to the 8S and 4S estrogen receptor of rat uterus or rat liver in a competition assay analyzed using sucrose density gradient centrifugation. On the other hand, the aromatic amines bind to varying extents to another estrogen-binding protein of rat liver whose function and identity is still unknown. The study demonstrates that structurally related aromatic amines in their unmetabolized form interact differentially with a cellular target protein, the Ah receptor, in vitro as well as in vivo. However, a relationship between these effects and the postulated promoting properties of 2-AAF remains to be established.

  8. Metformin inhibits 7,12-dimethylbenz[a]anthracene-induced breast carcinogenesis and adduct formation in human breast cells by inhibiting the cytochrome P4501A1/aryl hydrocarbon receptor signaling pathway

    SciTech Connect

    Maayah, Zaid H.; Ghebeh, Hazem; Alhaider, Abdulqader A.; El-Kadi, Ayman O.S.; Soshilov, Anatoly A.; Denison, Michael S.; Ansari, Mushtaq Ahmad; Korashy, Hesham M.

    2015-04-15

    Recent studies have established that metformin (MET), an oral anti-diabetic drug, possesses antioxidant activity and is effective against different types of cancer in several carcinogen-induced animal models and cell lines. However, whether MET can protect against breast cancer has not been reported before. Therefore, the overall objectives of the present study are to elucidate the potential chemopreventive effect of MET in non-cancerous human breast MCF10A cells and explore the underlying mechanism involved, specifically the role of cytochrome P4501A1 (CYP1A1)/aryl hydrocarbon receptor (AhR) pathway. Transformation of the MCF10A cells into initiated breast cancer cells with DNA adduct formation was conducted using 7,12-dimethylbenz[a]anthracene (DMBA), an AhR ligand. The chemopreventive effect of MET against DMBA-induced breast carcinogenesis was evidenced by the capability of MET to restore the induction of the mRNA levels of basic excision repair genes, 8-oxoguanine DNA glycosylase (OGG1) and apurinic/apyrimidinic endonuclease1 (APE1), and the level of 8-hydroxy-2-deoxyguanosine (8-OHdG). Interestingly, the inhibition of DMBA-induced DNA adduct formation was associated with proportional decrease in CYP1A1 and in NAD(P)H:quinone oxidoreductase 1 (NQO1) gene expression. Mechanistically, the involvements of AhR and nuclear factor erythroid 2-related factor-2 (Nrf2) in the MET-mediated inhibition of DMBA-induced CYP1A1 and NQO1 gene expression were evidenced by the ability of MET to inhibit DMBA-induced xenobiotic responsive element and antioxidant responsive element luciferase reporter gene expression which suggests an AhR- and Nrf2-dependent transcriptional control. However, the inability of MET to bind to AhR suggests that MET is not an AhR ligand. In conclusion, the present work shows a strong evidence that MET inhibits the DMBA-mediated carcinogenicity and adduct formation by inhibiting the expression of CYP1A1 through an AhR ligand-independent mechanism

  9. The androgenic anabolic steroid tetrahydrogestrinone produces dioxin-like effects via the aryl hydrocarbon receptor.

    PubMed

    Moon, Hyo Youl; Kim, Sun-Hee; Ryu, Sung Ho; Suh, Pann-Ghill

    2012-10-01

    For a long time, athletes have used androgenic anabolic steroids (AASs) in an inappropriate and veiled manner with the aim of improving exercise performance or for cosmetic purposes. Abuse of AASs triggers adverse effects such as hepatocarcinogenesis, heart attacks, and aggressive behavior. However, AAS-induced toxicity is not completely understood at the molecular level. In the present study, we showed, by performing a dioxin response element (DRE)-luciferase reporter gene assay, that tetrahydrogestrinone (THG), a popular and potent androgen receptor agonist, has dioxin-like effects. In addition, we showed that THG increased cytochrome P-450 1A1 (CYP1A1) mRNA and protein levels, and enzyme activity. The gene encoding CYP1A1 is involved in phase 1 xenobiotic metabolism and a target gene of the aryl hydrocarbon receptor (AhR). Using the AhR antagonist CH-223191, we also examined whether the effects of THG on DRE activation depended on AhR. Our results suggest that synthetic anabolic steroids may have dioxin-like side effects that can disturb endocrine systems and may cause other side effects including cancer through AhR.

  10. Environmental Ligands of the Aryl Hydrocarbon Receptor and Their Effects in Models of Adult Liver Progenitor Cells

    PubMed Central

    Vondráček, Jan; Machala, Miroslav

    2016-01-01

    The toxicity of environmental and dietary ligands of the aryl hydrocarbon receptor (AhR) in mature liver parenchymal cells is well appreciated, while considerably less attention has been paid to their impact on cell populations exhibiting phenotypic features of liver progenitor cells. Here, we discuss the results suggesting that the consequences of the AhR activation in the cellular models derived from bipotent liver progenitors could markedly differ from those in hepatocytes. In contact-inhibited liver progenitor cells, the AhR agonists induce a range of effects potentially linked with tumor promotion. They can stimulate cell cycle progression/proliferation and deregulate cell-to-cell communication, which is associated with downregulation of proteins forming gap junctions, adherens junctions, and desmosomes (such as connexin 43, E-cadherin, β-catenin, and plakoglobin), as well as with reduced cell adhesion and inhibition of intercellular communication. At the same time, toxic AhR ligands may affect the activity of the signaling pathways contributing to regulation of liver progenitor cell activation and/or differentiation, such as downregulation of Wnt/β-catenin and TGF-β signaling, or upregulation of transcriptional targets of YAP/TAZ, the effectors of Hippo signaling pathway. These data illustrate the need to better understand the potential role of liver progenitors in the AhR-mediated liver carcinogenesis and tumor promotion. PMID:27274734

  11. Environmental Ligands of the Aryl Hydrocarbon Receptor and Their Effects in Models of Adult Liver Progenitor Cells.

    PubMed

    Vondráček, Jan; Machala, Miroslav

    2016-01-01

    The toxicity of environmental and dietary ligands of the aryl hydrocarbon receptor (AhR) in mature liver parenchymal cells is well appreciated, while considerably less attention has been paid to their impact on cell populations exhibiting phenotypic features of liver progenitor cells. Here, we discuss the results suggesting that the consequences of the AhR activation in the cellular models derived from bipotent liver progenitors could markedly differ from those in hepatocytes. In contact-inhibited liver progenitor cells, the AhR agonists induce a range of effects potentially linked with tumor promotion. They can stimulate cell cycle progression/proliferation and deregulate cell-to-cell communication, which is associated with downregulation of proteins forming gap junctions, adherens junctions, and desmosomes (such as connexin 43, E-cadherin, β-catenin, and plakoglobin), as well as with reduced cell adhesion and inhibition of intercellular communication. At the same time, toxic AhR ligands may affect the activity of the signaling pathways contributing to regulation of liver progenitor cell activation and/or differentiation, such as downregulation of Wnt/β-catenin and TGF-β signaling, or upregulation of transcriptional targets of YAP/TAZ, the effectors of Hippo signaling pathway. These data illustrate the need to better understand the potential role of liver progenitors in the AhR-mediated liver carcinogenesis and tumor promotion.

  12. Aryl hydrocarbon receptor knock-out exacerbates choroidal neovascularization via multiple pathogenic pathways

    PubMed Central

    Choudhary, Mayur; Kazmin, Dmitri; Hu, Peng; Thomas, Russell S; McDonnell, Donald P; Malek, Goldis

    2015-01-01

    The aryl hydrocarbon receptor (AhR) is a heterodimeric transcriptional regulator with pleiotropic functions in xenobiotic metabolism and detoxification, vascular development and cancer. Herein, we report a previously undescribed role for the AhR signalling pathway in the pathogenesis of the wet, neovascular subtype of age-related macular degeneration (AMD), the leading cause of vision loss in the elderly in the Western world. Comparative analysis of gene expression profiles of aged AhR−/− and wild-type (wt) mice, using high-throughput RNA sequencing, revealed differential modulation of genes belonging to several AMD-related pathogenic pathways, including inflammation, angiogenesis and extracellular matrix regulation. To investigate AhR regulation of these pathways in wet AMD, we experimentally induced choroidal neovascular lesions in AhR−/− mice and found that they measured significantly larger in area and volume compared to age-matched wt mice. Furthermore, these lesions displayed a higher number of ionized calcium-binding adaptor molecule 1-positive (Iba1+) microglial cells and a greater amount of collagen type IV deposition, events also seen in human wet AMD pathology specimens. Consistent with our in vivo observations, AhR knock-down was sufficient to increase choroidal endothelial cell migration and tube formation in vitro. Moreover, AhR knock-down caused an increase in collagen type IV production and secretion in both retinal pigment epithelial (RPE) and choroidal endothelial cell cultures, increased expression of angiogenic and inflammatory molecules, including vascular endothelial growth factor A (VEGFA) and chemokine (C–C motif) ligand 2 (CCL2) in RPE cells, and increased expression of secreted phosphoprotein 1 (SPP1) and transforming growth factor-β1 (TGFβ1) in choroidal endothelial cells. Collectively, our findings identify AhR as a regulator of multiple pathogenic pathways in experimentally induced choroidal neovascularization, findings that

  13. Aryl Hydrocarbon Receptor Ligand 5F 203 Induces Oxidative Stress That Triggers DNA Damage in Human Breast Cancer Cells.

    PubMed

    McLean, Lancelot S; Watkins, Cheri N; Campbell, Petreena; Zylstra, Dain; Rowland, Leah; Amis, Louisa H; Scott, Lia; Babb, Crystal E; Livingston, W Joel; Darwanto, Agus; Davis, Willie L; Senthil, Maheswari; Sowers, Lawrence C; Brantley, Eileen

    2015-05-18

    Breast tumors often show profound sensitivity to exogenous oxidative stress. Investigational agent 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203) induces aryl hydrocarbon receptor (AhR)-mediated DNA damage in certain breast cancer cells. Since AhR agonists often elevate intracellular oxidative stress, we hypothesize that 5F 203 increases reactive oxygen species (ROS) to induce DNA damage, which thwarts breast cancer cell growth. We found that 5F 203 induced single-strand break formation. 5F 203 enhanced oxidative DNA damage that was specific to breast cancer cells sensitive to its cytotoxic actions, as it did not increase oxidative DNA damage or ROS formation in nontumorigenic MCF-10A breast epithelial cells. In contrast, AhR agonist and procarcinogen benzo[a]pyrene and its metabolite, 1,6-benzo[a]pyrene quinone, induced oxidative DNA damage and ROS formation, respectively, in MCF-10A cells. In sensitive breast cancer cells, 5F 203 activated ROS-responsive kinases: c-Jun-N-terminal kinase (JNK) and p38 mitogen activated protein kinase (p38). AhR antagonists (alpha-naphthoflavone, CH223191) or antioxidants (N-acetyl-l-cysteine, EUK-134) attenuated 5F 203-mediated JNK and p38 activation, depending on the cell type. Pharmacological inhibition of AhR, JNK, or p38 attenuated 5F 203-mediated increases in intracellular ROS, apoptosis, and single-strand break formation. 5F 203 induced the expression of cytoglobin, an oxidative stress-responsive gene and a putative tumor suppressor, which was diminished with AhR, JNK, or p38 inhibition. Additionally, 5F 203-mediated increases in ROS production and cytoglobin were suppressed in AHR100 cells (AhR ligand-unresponsive MCF-7 breast cancer cells). Our data demonstrate 5F 203 induces ROS-mediated DNA damage at least in part via AhR, JNK, or p38 activation and modulates the expression of oxidative stress-responsive genes such as cytoglobin to confer its anticancer action.

  14. Aryl Hydrocarbon Receptor Ligand 5F 203 Induces Oxidative Stress That Triggers DNA Damage in Human Breast Cancer Cells

    PubMed Central

    McLean, Lancelot S.; Watkins, Cheri N.; Campbell, Petreena; Zylstra, Dain; Rowland, Leah; Amis, Louisa H.; Scott, Lia; Babb, Crystal E.; Livingston, W. Joel; Darwanto, Agus; Davis, Willie L.; Senthil, Maheswari; Sowers, Lawrence C.; Brantley, Eileen

    2015-01-01

    Breast tumors often show profound sensitivity to exogenous oxidative stress. Investigational agent 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203) induces aryl hydrocarbon receptor (AhR)-mediated DNA damage in certain breast cancer cells. Since AhR agonists often elevate intracellular oxidative stress, we hypothesize that 5F 203 increases reactive oxygen species (ROS) to induce DNA damage, which thwarts breast cancer cell growth. We found that 5F 203 induced single-strand break formation. 5F 203 enhanced oxidative DNA damage that was specific to breast cancer cells sensitive to its cytotoxic actions, as it did not increase oxidative DNA damage or ROS formation in nontumorigenic MCF-10A breast epithelial cells. In contrast, AhR agonist and procarcinogen benzo[a]pyrene and its metabolite, 1,6-benzo[a]pyrene quinone, induced oxidative DNA damage and ROS formation, respectively, in MCF-10A cells. In sensitive breast cancer cells, 5F 203 activated ROS-responsive kinases: c-Jun-N-terminal kinase (JNK) and p38 mitogen activated protein kinase (p38). AhR antagonists (alpha-naphthoflavone, CH223191) or antioxidants (N-acetyl-l-cysteine, EUK-134) attenuated 5F 203-mediated JNK and p38 activation, depending on the cell type. Pharmacological inhibition of AhR, JNK, or p38 attenuated 5F 203-mediated increases in intracellular ROS, apoptosis, and single-strand break formation. 5F 203 induced the expression of cytoglobin, an oxidative stress-responsive gene and a putative tumor suppressor, which was diminished with AhR, JNK, or p38 inhibition. Additionally, 5F 203-mediated increases in ROS production and cytoglobin were suppressed in AHR100 cells (AhR ligand-unresponsive MCF-7 breast cancer cells). Our data demonstrate 5F 203 induces ROS-mediated DNA damage at least in part via AhR, JNK, or p38 activation and modulates the expression of oxidative stress-responsive genes such as cytoglobin to confer its anticancer action. PMID:25781201

  15. EXPRESSION OF AHR AND ARNT MRNA IN CULTURED HUMAN ENDOMETRIAL EXPLANTS EXPOSED TO TCDD

    EPA Science Inventory

    Expression of AhR and ARNT mRNA in cultured human endometrial explants exposed to TCDD.

    Pitt JA, Feng L, Abbott BD, Schmid J, Batt RE, Costich TG, Koury ST, Bofinger DP.

    Curriculum in Toxicology, University of North Carolina, Chapel Hill, NC 27599, USA.

    Endom...

  16. Docking-based three-dimensional quantitative structure-activity relationship (3D-QSAR) predicts binding affinities to aryl hydrocarbon receptor for polychlorinated dibenzodioxins, dibenzofurans, and biphenyls.

    PubMed

    Yuan, Jintao; Pu, Yuepu; Yin, Lihong

    2013-07-01

    Polychlorinated dibenzodioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and polychlorinated biphenyls (PCBs) cause toxic effects after binding to an intracellular cytosolic receptor called the aryl hydrocarbon receptor (AhR). Thymic atrophy, weight loss, immunotoxicity, acute lethality, and induction of cytochrome P4501A1 have all been correlated with the binding affinity to AhR. To study the key molecular features for determining binding affinity to AhR, a homology model of AhR ligand-binding domains was developed, a molecular docking approach was employed to obtain docking-based conformations of all molecules in the whole set, and 3-dimensional quantitative structure-activity relationship (3D-QSAR) methodology, namely, comparative molecular field analysis (CoMFA), was applied. A partial least square analysis was performed, and QSAR models were generated for a training set of 59 compounds. The generated QSAR model showed good internal and external statistical reliability, and in a comparison with other reported CoMFA models using different alignment methods, the docking-based CoMFA model showed some advantages.

  17. Cardiac Myocyte-Specific AHR Activation Phenocopies TCDD-Induced Toxicity in Zebrafish

    PubMed Central

    Lanham, Kevin A.; Plavicki, Jessica; Peterson, Richard E.; Heideman, Warren

    2014-01-01

    Exposure of zebrafish embryos to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) activates the zebrafish aryl hydrocarbon receptor 2 (AHR) to produce developmental and cardiovascular toxicity. AHR is found in the heart; however, AHR activation by TCDD is not confined to the heart and occurs throughout the organism. In order to understand the cause of cardiotoxicity, we constructed a constitutively active AHR (caAHR) based on the zebrafish AHR2 and expressed it specifically in cardiomyocytes. We show that AHR activation within the cardiomyocytes can account for the heart failure induced by TCDD. Expression of the caAHR within the heart produced cardiac malformations, loss of circulation, and pericardial edema. The heart-specific activation of AHR reproduced several other well-characterized endpoints of TCDD toxicity outside of the cardiovascular system, including defects in swim bladder and craniofacial development. This work identifies a single cellular site of TCDD action, the myocardial cell, that can account for the severe cardiovascular collapse observed following early life stage exposure to TCDD, and contributes to other forms of toxicity. PMID:25037585

  18. Role of the Aryl Hydrocarbon Receptor in the Immune Response Profile and Development of Pathology during Plasmodium berghei Anka Infection

    PubMed Central

    Brant, Fatima; Miranda, Aline S.; Esper, Lisia; Rodrigues, David Henrique; Kangussu, Lucas Miranda; Bonaventura, Daniella; Soriani, Frederico Marianetti; Pinho, Vanessa; Souza, Danielle G.; Rachid, Milene Alvarenga; Weiss, Louis M.; Tanowitz, Herbert B.; Teixeira, Mauro Martins; Teixeira, Antônio Lucio

    2014-01-01

    Infection with Plasmodium falciparum may result in severe disease affecting various organs, including liver, spleen, and brain, resulting in high morbidity and mortality. Plasmodium berghei Anka infection of mice recapitulates many features of severe human malaria. The aryl hydrocarbon receptor (AhR) is an intracellular receptor activated by ligands important in the modulation of the inflammatory response. We found that AhR-knockout (KO) mice infected with P. berghei Anka displayed increased parasitemia, earlier mortality, enhanced leukocyte-endothelial cell interactions in the brain microvasculature, and increased inflammation in brain (interleukin-17 [IL-17] and IL-6) and liver (gamma interferon [IFN-γ] and tumor necrosis factor alpha [TNF-α]) compared to infected wild-type (WT) mice. Infected AhR-KO mice also displayed a reduction in cytokines required for host resistance, including TNF-α, IL-1β, and IFN-γ, in the brain and spleen. Infection of AhR-KO mice resulted in an increase in T regulatory cells and transforming growth factor β, IL-6, and IL-17 in the brain. AhR modulated the basal expression of SOCS3 in spleen and brain, and P. berghei Anka infection resulted in enhanced expression of SOCS3 in brain, which was absent in infected AhR-KO mice. These data suggest that AhR-mediated control of SOCS3 expression is probably involved in the phenotype seen in infected AhR-KO mice. This is, to our knowledge, the first demonstration of a role for AhR in the pathogenesis of malaria. PMID:24818665

  19. Hexachlorobenzene modulates the crosstalk between the aryl hydrocarbon receptor and transforming growth factor-β1 signaling, enhancing human breast cancer cell migration and invasion.

    PubMed

    Miret, Noelia; Pontillo, Carolina; Ventura, Clara; Carozzo, Alejandro; Chiappini, Florencia; Kleiman de Pisarev, Diana; Fernández, Natalia; Cocca, Claudia; Randi, Andrea

    2016-07-29

    Given the number of women affected by breast cancer, considerable interest has been raised in understanding the relationships between environmental chemicals and disease onset. Hexachlorobenzene (HCB) is a dioxin-like compound that is widely distributed in the environment and is a weak ligand of the aryl hydrocarbon receptor (AhR). We previously demonstrated that HCB acts as an endocrine disruptor capable of stimulating cell proliferation, migration, invasion, and metastasis in different breast cancer models. In addition, increasing evidence indicates that transforming growth factor-β1 (TGF-β1) can contribute to tumor maintenance and progression. In this context, this work investigated the effect of HCB (0.005, 0.05, 0.5, and 5μM) on TGF-β1 signaling and AhR/TGF-β1 crosstalk in the human breast cancer cell line MDA-MB-231 and analyzed whether TGF-β1 pathways are involved in HCB-induced cell migration and invasion. RT-qPCR results indicated that HCB reduces AhR mRNA expression through TGF-β1 signaling but enhances TGF-β1 mRNA levels involving AhR signaling. Western blot analysis demonstrated that HCB could increase TGF-β1 protein levels and activation, as well as Smad3, JNK, and p38 phosphorylation. In addition, low and high doses of HCB were determined to exert differential effects on AhR protein levels, localization, and activation, with a high dose (5μM) inducing AhR nuclear translocation and AhR-dependent CYP1A1 expression. These findings also revealed that c-Src and AhR are involved in HCB-mediated activation of Smad3. HCB enhances cell migration (scratch motility assay) and invasion (Transwell assay) through the Smad, JNK, and p38 pathways, while ERK1/2 is only involved in HCB-induced cell migration. These results demonstrate that HCB modulates the crosstalk between AhR and TGF-β1 and consequently exacerbates a pro-migratory phenotype in MDA-MB-231 cells, which contributes to a high degree of malignancy. Taken together, our findings help to

  20. Imaging the Met Receptor Tyrosine Kinase (Met) and Assessing Tumor Responses to a Met Tyrosine Kinase Inhibitor in Human Xenograft Mouse Models with a [99mTc] (AH-113018) or Cy 5** (AH-112543) Labeled Peptide.

    PubMed

    Jagoda, Elaine M; Bhattacharyya, Sibaprasad; Kalen, Joseph; Riffle, Lisa; Leeder, Avrum; Histed, Stephanie; Williams, Mark; Wong, Karen J; Xu, Biying; Szajek, Lawrence P; Elbuluk, Osama; Cecchi, Fabiola; Raffensperger, Kristen; Golla, Meghana; Bottaro, Donald P; Choyke, Peter

    2015-01-01

    Developing an imaging agent targeting the hepatocyte growth factor receptor protein (Met) status of cancerous lesions would aid in the diagnosis and monitoring of Met-targeted tyrosine kinase inhibitors (TKIs). A peptide targeting Met labeled with [(99m)Tc] had high affinity in vitro (Kd = 3.3 nM) and detected relative changes in Met in human cancer cell lines. In vivo [(99m)Tc]-Met peptide (AH-113018) was retained in Met-expressing tumors, and high-expressing Met tumors (MKN-45) were easily visualized and quantitated using single-photon emission computed tomography or optical imaging. In further studies, MKN-45 mouse xenografts treated with PHA 665752 (Met TKI) or vehicle were monitored weekly for tumor responses by [(99m)Tc]-Met peptide imaging and measurement of tumor volumes. Tumor uptake of [(99m)Tc]-Met peptide was significantly decreased as early as 1 week after PHA 665752 treatment, corresponding to decreases in tumor volumes. These results were comparable to Cy5**-Met peptide (AH-112543) fluorescence imaging using the same treatment model. [(99m)Tc] or Cy5**-Met peptide tumor uptake was further validated by histologic (necrosis, apoptosis) and immunoassay (total Met, p Met, and plasma shed Met) assessments in imaged and nonimaged cohorts. These data suggest that [(99m)Tc] or Cy5**-Met peptide imaging may have clinical diagnostic, prognostic, and therapeutic monitoring applications. PMID:26461980

  1. Harman induces CYP1A1 enzyme through an aryl hydrocarbon receptor mechanism

    SciTech Connect

    El Gendy, Mohamed A.M.; El-Kadi, Ayman O.S.

    2010-11-15

    Harman is a common compound in several foods, plants and beverages. Numerous studies have demonstrated its mutagenic, co-mutagenic and carcinogenic effects; however, the exact mechanism has not been fully identified. Aryl hydrocarbon receptor (AhR) is a transcription factor regulating the expression of the carcinogen-activating enzyme; cytochrome P450 1A1 (CYP1A1). In the present study, we examined the ability of harman to induce AhR-mediated signal transduction in human and rat hepatoma cells; HepG2 and H4IIE cells. Our results showed that harman significantly induced CYP1A1 mRNA in a time- and concentration-dependent manner. Similarly, harman significantly induced CYP1A1 at protein and activity levels in a concentration-dependent manner. Moreover, the AhR antagonist, resveratrol, inhibited the increase in CYP1A1 activity by harman. The RNA polymerase inhibitor, actinomycin D, completely abolished the CYP1A1 mRNA induction by harman, indicating a transcriptional activation. The role of AhR in CYP1A1 induction by harman was confirmed by using siRNA specific for human AhR. The ability of harman to induce CYP1A1 was strongly correlated with its ability to stimulate AhR-dependent luciferase activity and electrophoretic mobility shift assay. At post-transcriptional and post-translational levels, harman did not affect the stability of CYP1A1 at the mRNA and the protein levels, excluding other mechanisms participating in the obtained effects. We concluded that harman can directly induce CYP1A1 gene expression in an AhR-dependent manner and may represent a novel mechanism by which harman promotes mutagenicity, co-mutagenicity and carcinogenicity.

  2. Aryl hydrocarbon receptor activation in lactotropes and gonadotropes interferes with estradiol-dependent and -independent preprolactin, glycoprotein alpha and luteinizing hormone beta gene expression.

    PubMed

    Cao, Jinyan; Patisaul, Heather B; Petersen, Sandra L

    2011-02-20

    Arylhydrocarbon receptor (Ahr) activation by 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD) interferes with female reproductive functions, but there is little information on the specific targets of TCDD in the hypothalamic-pituitary-gonadal (HPG) axis. In these studies, we found that TCDD upregulated known AhR target genes, cytochrome p450 1a1 (Cyp1a1), Cyp1a2 and Cyp1b1 in the rat pituitary gland. Moreover, 75% of pituitary lactotropes and 45% of gonadotropes contained Ahr mRNA, and most Ahr-containing cells were estrogen receptor 1 (Esr1)-positive. TCDD abrogated estradiol (E(2))-induced prolactin (Prl) expression in vivo and in vitro; conversely, E(2) blocked TCDD upregulation of luteinizing hormone beta (Lhb) and glycoprotein hormone alpha polypeptide (Cga) expression. TCDD had no effect on levels of Ahr mRNA, but upregulated Esr1 mRNA. E(2) independently repressed Ahr and Esr1 expression and blocked TCDD upregulation of Esr1. Thus, complex interactions between Ahr and Esr alter Prl and luteinizing hormone (LH) synthesis by direct actions in lactotropes and gonadotropes. These findings provide important insights into how TCDD disrupts female reproductive functions.

  3. Aryl Hydrocarbon Receptor Activation in Lactotropes and Gonadotropes Interferes with Estradiol-Dependent and -Independent Preprolactin, Glycoprotein Alpha and Luteinizing Hormone Beta Gene Expression

    PubMed Central

    Cao, JinYan; Patisaul, Heather B.; Petersen, Sandra L.

    2011-01-01

    Arylhydrocarbon receptor (Ahr) activation by 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD) interferes with female reproductive functions, but there is little information on the specific targets of TCDD in the hypothalamic-pituitary-gonadal (HPG) axis. In these studies, we found that TCDD upregulated known AhR target genes, cytochrome p450 1a1 (Cyp1a1), Cyp1a2 and Cyp1b1 in the rat pituitary gland. Moreover, 75% of pituitary lactotropes and 45% of gonadotropes contained Ahr mRNA, and most Ahr-containing cells were estrogen receptor 1 (Esr1)-positive. TCDD abrogated estradiol (E2)-induced prolactin (Prl) expression in vivo and in vitro; conversely, E2 blocked TCDD upregulation of luteinizing hormone beta (Lhb) and glycoprotein hormone alpha polypeptide (Cga) expression. TCDD had no effect on levels of Ahr mRNA, but upregulated Esr1 mRNA. E2 independently repressed Ahr and Esr1 expression and blocked TCDD upregulation of Esr1. Thus, complex interactions between Ahr and Esr alter Prl and luteinizing hormone (LH) synthesis by direct actions in lactotropes and gonadotropes. These findings provide important insights into how TCDD disrupts female reproductive functions. PMID:21187122

  4. Unique Expression of Angiotensin Type-2 Receptor in Sex-Specific Distribution of Myelinated Ah-Type Baroreceptor Neuron Contributing to Sex-Dimorphic Neurocontrol of Circulation.

    PubMed

    Liu, Yang; Zhou, Jia-Ying; Zhou, Yu-Hong; Wu, Di; He, Jian-Li; Han, Li-Min; Liang, Xiao-Bo; Wang, Lu-Qi; Lu, Xiao-Long; Chen, Hanying; Qiao, Guo-Fen; Shou, Weinian; Li, Bai-Yan

    2016-04-01

    This study aims to understand the special expression patterns of angiotensin-II receptor (AT1R and AT2R) in nodose ganglia and nucleus of tractus solitary of baroreflex afferent pathway and their contribution in sex difference of neurocontrol of blood pressure regulation. In this regard, action potentials were recorded in baroreceptor neurons (BRNs) using whole-cell patch techniques; mRNA and protein expression of AT1R and AT2R in nodose ganglia and nucleus of tractus solitary were evaluated using real time-polymerase chain reaction, Western blot, and immunohistochemistry at both tissue and single-cell levels. The in vivo effects of 17β-estradiol on blood pressure and AT2R expression were also tested. The data showed that AT2R, rather than AT1R, expression was higher in female than age-matched male rats. Moreover, AT2R was downregulated in ovariectomized rats, which was restored by the administration of 17β-estradiol. Single-cell real time-polymerase chain reaction data indicated that AT2R was uniquely expressed in Ah-type BRNs. Functional study showed that long-term administration of 17β-estradiol significantly alleviated the blood pressure increase in ovariectomized rats. Electrophysiological recordings showed that angiotensin-II treatment increased the neuroexcitability more in Ah- than C-type BRNs, whereas no such effect was observed in A-types. In addition, angiotensin-II treatment prolonged action potential duration, which was not further changed by iberiotoxin. The density of angiotensin-II-sensitive K(+) currents recorded in Ah-types was equivalent with iberiotoxin-sensitive component. In summary, the unique, sex- and afferent-specific expression of AT2R was identified in Ah-type BRNs, and AT2R-mediated KCa1.1 inhibition in Ah-type BRNs may exert great impacts on baroreflex afferent function and blood pressure regulation in females. PMID:26883269

  5. Absence of aryl hydrocarbon receptors increases endogenous kynurenic acid levels and protects mouse brain against excitotoxic insult and oxidative stress.

    PubMed

    García-Lara, Lucia; Pérez-Severiano, Francisca; González-Esquivel, Dinora; Elizondo, Guillermo; Segovia, José

    2015-09-01

    L-kynurenine (Kyn) is a key element of tryptophan metabolism; it is enzymatically converted by kynurenine aminotransferase II (KAT II) to kynurenic acid (KYNA), which acts as an antagonist to the NMDA receptor-glycine site. Kyn is also an endogenous ligand of the aryl hydrocarbon receptor (AhR), a transcription factor that regulates the expression of a diverse set of genes. KYNA levels are reduced in several regions of the brain of Huntington's disease (HD) patients. The present work uses an AhR-null mouse and age-matched wild-type mice to determine the effect of the absence of AhR on KYNA availability. We found that, in AhR-null mice, there is an increase of KYNA levels in specific brain areas associated with higher expression of KAT II. Moreover, we induced an excitotoxic insult by intrastriatal administration of quinolinic acid, a biochemical model of HD, in both AhR-null and wild-type mice to evaluate the neurological damage as well as the oxidative stress caused by the lesion. The present work demonstrates that, in specific brain regions of AhR-null mice, the levels of KYNA are increased and that this induces a neuroprotective effect against neurotoxic insults. Moreover, AhR-null mice also show improved motor performance in the rotarod test, indicating a constitutive protection of striatal tissue.

  6. Aryl hydrocarbon receptor signaling mediates expression of indoleamine 2,3-dioxygenase.

    PubMed

    Vogel, Christoph F A; Goth, Samuel R; Dong, Bin; Pessah, Isaac N; Matsumura, Fumio

    2008-10-24

    Aryl hydrocarbon receptor (AhR) activation by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) leads to immune suppression associated with the induction of regulatory T cells (T(reg)) expressing the transcription factor Foxp3. The immunological mechanisms of suppression are not well understood however dendritic cells (DC) are considered a key target for AhR-mediated immune suppression. Here we show that activation of AhR by TCDD induces DC indoleamine 2,3-dioxygenase 1 (IDO1) and indoleamine 2,3-dioxygenase-like protein (IDO2). Induction of IDO1 and IDO2 was also found in lung and spleen associated with an increase of the T(reg) marker Foxp3 in spleen of TCDD-treated C57BL/6 mice, which is suppressed by inhibition of IDO. These data indicate that AhR-activation is an important signaling pathway for IDO expression and suggest a critical role of IDO in the mechanism leading to the generation of T(reg) that mediates the immune suppression through activation of AhR. PMID:18694728

  7. Microarray analysis of the AHR system: Tissue-specific flexibility in signal and target genes

    SciTech Connect

    Frericks, Markus; Meissner, Marc; Esser, Charlotte . E-mail: chesser@uni-duesseldorf.de

    2007-05-01

    Data mining published microarray experiments require that expression profiles are directly comparable. We performed linear global normalization on the data of 1967 Affymetrix U74av2 microarrays, i.e. the transcriptomes of > 100 murine tissues or cell types. The mathematical transformation effectively nullifies inter-experimental or inter-laboratory differences between microarrays. The correctness of expression values was validated by quantitative RT-PCR. Using the database we analyze components of the aryl hydrocarbon receptor (AHR) signaling pathway in various tissues. We identified lineage and differentiation specific variant expression of AHR, ARNT, and HIF1{alpha} in the T-cell lineage and high expression of CYP1A1 in immature B cells and dendritic cells. Performing co-expression analysis we found unorthodox expression of the AHR in the absence of ARNT, particularly in stem cell populations, and can reject the hypothesis that ARNT2 takes over and is highly expressed when ARNT expression is low or absent. Furthermore the AHR shows no co-expression with any other transcript present on the chip. Analysis of differential gene expression under 308 conditions revealed 53 conditions under which the AHR is regulated, numerous conditions under which an intrinsic AHR action is modified as well as conditions activating the AHR even in the absence of known AHR ligands. Thus meta-analysis of published expression profiles is a powerful tool to gain novel insights into known and unknown systems.

  8. Activation of the aryl hydrocarbon receptor affects activation and function of human monocyte-derived dendritic cells.

    PubMed

    Wang, C; Ye, Z; Kijlstra, A; Zhou, Y; Yang, P

    2014-08-01

    Aryl hydrocarbon receptor (AhR) is well known for mediating the toxic effects of dioxin-containing pollutants, but has also been shown to be involved in the natural regulation of the immune response. In this study, we investigated the effect of AhR activation by its endogenous ligands 6-formylindolo[3,2-b]carbazole (FICZ) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) on the differentiation, maturation and function of monocyte-derived DCs in Behçet's disease (BD) patients. In this study, we showed that AhR activation by FICZ and ITE down-regulated the expression of co-stimulatory molecules including human leucocyte antigen D-related (HLA-DR), CD80 and CD86, while it had no effect on the expression of CD83 and CD40 on DCs derived from BD patients and normal controls. Lipopolysaccharide (LPS)-treated dendritic cells (DCs) from active BD patients showed a higher level of interleukin (IL)-1β, IL-6, IL-23 and tumour necrosis factor (TNF)-α production. FICZ or ITE significantly inhibited the production of IL-1β, IL-6, IL-23 and TNF-α, but induced IL-10 production by DCs derived from active BD patients and normal controls. FICZ or ITE-treated DCs significantly inhibited the T helper type 17 (Th17) and Th1 cell response. Activation of AhR either by FICZ or ITE inhibits DC differentiation, maturation and function. Further studies are needed to investigate whether manipulation of the AhR pathway may be used to treat BD or other autoimmune diseases.

  9. A constitutive active MAPK/ERK pathway due to BRAFV600E positively regulates AHR pathway in PTC

    PubMed Central

    Regazzo, Daniela; Bertazza, Loris; Galuppini, Francesca; Guzzardo, Vincenza; Jaffrain-Rea, Marie Lise; Vianello, Federica; Ciato, Denis; Ceccato, Filippo; Watutantrige-Fernando, Sara; Bisognin, Andrea; Bortoluzzi, Stefania; Pennelli, Gianmaria; Boscaro, Marco; Scaroni, Carla; Mian, Caterina

    2015-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor mediating the toxicity and tumor-promoting properties of dioxin. AHR has been reported to be overexpressed and constitutively active in a variety of solid tumors, but few data are currently available concerning its role in thyroid cancer. In this study we quantitatively explored a series of 51 paired-normal and papillary thyroid carcinoma (PTC) tissues for AHR-related genes. We identified an increased AHR expression/activity in PTC, independently from its nuclear dimerization partner and repressor but strictly related to a constitutive active MAPK/ERK pathway. The AHR up-regulation followed by an increased expression of AHR target genes was confirmed by a meta-analysis of published microarray data, suggesting a ligand-independent active AHR pathway in PTC. In-vitro studies using a PTC-derived cell line (BCPAP) and HEK293 cells showed that BRAFV600E may directly modulate AHR localization, induce AHR expression and activity in an exogenous ligand-independent manner. The AHR pathway might represent a potential novel therapeutic target for PTC in the clinical practice. PMID:26392334

  10. Aryl hydrocarbon receptor signaling regulates NF-κB RelB activation during dendritic-cell differentiation

    PubMed Central

    Vogel, Christoph F A; Wu, Dalei; Goth, Samuel R; Baek, Jaeeun; Lollies, Anna; Domhardt, Rowena; Grindel, Annemarie; Pessah, Isaac N

    2013-01-01

    How the aryl hydrocarbon receptor (AhR) regulates dendritic-cell (DC) differentiation is unknown. We show that activation of AhR by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) caused enhanced differentiation from immature DCs (IDCs) to mature DCs (MDCs) in the bone-marrow-derived DCs (BMDC) from B6 wild-type mice but not in the BMDCs from AhR-null mice as indicated by the expression of CD11c and class II major histocompatibility complex (MHC). Enhanced maturation of BMDCs was associated with elevated levels of CD86 and an increased AhR-dependent nuclear accumulation of nuclear factor-kappa-light-chain enhancer of activated B cell (NF-κB) member RelB in BMDCs. The expression of interleukin (IL) 10 and chemokine DC-CK1 was suppressed, whereas that of CXCL2, CXCL3 and IL-22 was significantly increased in AhR-activated BMDCs. Furthermore, TCDD induced expression of the regulatory enzymes indoleamine 2,3-dioxygenase (IDO1) and indoleamine 2,3-dioxygenase-like 1 (IDO2). Increased expression of IDO2 was associated with coexpression of the cell-surface marker CCR6. Interestingly, mRNA expression of the chemokine receptor CCR6 was drastically decreased in AhR-null IDCs and MDCs. Overall, these data demonstrate that AhR modifies the maturation of BMDCs associated with the induction of the regulatory enzyme IDO and altered expression of cytokine, chemokines and DC-specific surface markers and receptors. PMID:23999131

  11. Aryl hydrocarbon receptor signaling regulates NF-κB RelB activation during dendritic-cell differentiation.

    PubMed

    Vogel, Christoph F A; Wu, Dalei; Goth, Samuel R; Baek, Jaeeun; Lollies, Anna; Domhardt, Rowena; Grindel, Annemarie; Pessah, Isaac N

    2013-10-01

    How the aryl hydrocarbon receptor (AhR) regulates dendritic-cell (DC) differentiation is unknown. We show that activation of AhR by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) caused enhanced differentiation from immature DCs (IDCs) to mature DCs (MDCs) in the bone-marrow-derived DCs (BMDC) from B6 wild-type mice but not in the BMDCs from AhR-null mice as indicated by the expression of CD11c and class II major histocompatibility complex (MHC). Enhanced maturation of BMDCs was associated with elevated levels of CD86 and an increased AhR-dependent nuclear accumulation of nuclear factor-kappa-light-chain enhancer of activated B cell (NF-κB) member RelB in BMDCs. The expression of interleukin (IL) 10 and chemokine DC-CK1 was suppressed, whereas that of CXCL2, CXCL3 and IL-22 was significantly increased in AhR-activated BMDCs. Furthermore, TCDD induced expression of the regulatory enzymes indoleamine 2,3-dioxygenase (IDO1) and indoleamine 2,3-dioxygenase-like 1 (IDO2). Increased expression of IDO2 was associated with coexpression of the cell-surface marker CCR6. Interestingly, mRNA expression of the chemokine receptor CCR6 was drastically decreased in AhR-null IDCs and MDCs. Overall, these data demonstrate that AhR modifies the maturation of BMDCs associated with the induction of the regulatory enzyme IDO and altered expression of cytokine, chemokines and DC-specific surface markers and receptors. PMID:23999131

  12. Combination effects of AHR agonists and Wnt/β-catenin modulators in zebrafish embryos: Implications for physiological and toxicological AHR functions

    SciTech Connect

    Wincent, Emma; Stegeman, John J.; Jönsson, Maria E.

    2015-04-15

    Wnt/β-catenin signaling regulates essential biological functions and acts in developmental toxicity of some chemicals. The aryl hydrocarbon receptor (AHR) is well-known to mediate developmental toxicity of persistent dioxin-like compounds (DLCs). Recent studies indicate a crosstalk between β-catenin and the AHR in some tissues. However the nature of this crosstalk in embryos is poorly known. We observed that zebrafish embryos exposed to the β-catenin inhibitor XAV939 display effects phenocopying those of the dioxin-like 3,3′,4,4′,5-pentachlorobiphenyl (PCB126). This led us to investigate the AHR interaction with β-catenin during development and ask whether developmental toxicity of DLCs involves antagonism of β-catenin signaling. We examined phenotypes and transcriptional responses in zebrafish embryos exposed to XAV939 or to a β-catenin activator, 1-azakenpaullone, alone or with AHR agonists, either PCB126 or 6-formylindolo[3,2-b]carbazole (FICZ). Alone 1-azakenpaullone and XAV939 both were embryo-toxic, and we found that in the presence of FICZ, the toxicity of 1-azakenpaullone decreased while the toxicity of XAV939 increased. This rescue of 1-azakenpaullone effects occurred in the time window of Ahr2-mediated toxicity and was reversed by morpholino-oligonucleotide knockdown of Ahr2. Regarding PCB126, addition of either 1-azakenpaullone or XAV939 led to lower mortality than with PCB126 alone but surviving embryos showed severe edemas. 1-Azakenpaullone induced transcription of β-catenin-associated genes, while PCB126 and FICZ blocked this induction. The data indicate a stage-dependent antagonism of β-catenin by Ahr2 in zebrafish embryos. We propose that the AHR has a physiological role in regulating β-catenin during development, and that this is one point of intersection linking toxicological and physiological AHR-governed processes.

  13. Combination effects of AHR agonists and Wnt/β-catenin modulators in zebrafish embryos: implications for physiological and toxicological AHR functions

    PubMed Central

    Wincent, Emma; Stegeman, John J.; Jönsson, Maria E.

    2016-01-01

    Wnt/β-catenin signaling regulates essential biological functions and acts in developmental toxicity of some chemicals. The aryl hydrocarbon receptor (AHR) is well-known to mediate developmental toxicity of persistent dioxin-like compounds (DLCs). Recent studies indicate a crosstalk between β-catenin and the AHR in some tissues. However the nature of this crosstalk in embryos is poorly known. We observed that zebrafish embryos exposed to the β-catenin inhibitor XAV939 display effects phenocopying those of the dioxin-like 3,3′,4,4′,5-pentachlorobiphenyl (PCB126). This led us to investigate AHR interaction with β-catenin during development and ask whether developmental toxicity of DLCs involves antagonism of β-catenin signaling. We examined phenotypes and transcriptional responses in zebrafish embryos exposed to XAV939 or to a β-catenin activator, 1-azakenpaullone, alone or with AHR agonists, either PCB126 or 6-formylindolo[3,2-b]carbazole (FICZ). Alone 1-azakenpaullone and XAV939 both were embryo-toxic, and we found that in presence of FICZ, the toxicity of 1-azakenpaullone decreased while the toxicity of XAV939 increased. This rescue of 1-azakenpaullone effects occurred in the time window of Ahr2-mediated toxicity and was reversed by morpholine-oligonucleotide knockdown of Ahr2. Regarding PCB126, addition of either 1-azakenpaullone or XAV939 led to lower mortality than with PCB126 alone but surviving embryos showed severe edemas. 1-Azakenpaullone induced transcription of β-catenin-associated genes, while PCB126 and FICZ blocked this induction. The data indicate a stage-dependent antagonism of β-catenin by Ahr2 in zebrafish embryos. We propose that the AHR has a physiological role in regulating β-catenin during development, and that this is one point of intersection linking toxicological and physiological AHR-governed processes. PMID:25711857

  14. Aryl hydrocarbon receptor expression is associated with a family history of upper gastrointestinal tract cancer in a high-risk population exposed to aromatic hydrocarbons

    SciTech Connect

    Roth, M.J.; Wei, W.Q.; Baer, J.; Abnet, C.C.; Wang, G.Q.; Sternberg, L.R.; Warner, A.C.; Johnson, L.L.; Lu, N.; Giffen, C.A.; Dawsey, S.M.; Qiao, Y.L.; Cherry, J.

    2009-09-15

    Polycyclic aromatic hydrocarbon (PAH) exposure is a risk factor for esophageal squamous cell carcinoma, and PAHs are ligands of the aryl hydrocarbon receptor (AhR). This study measured the expression of AhR and related genes in frozen esophageal cell samples from patients exposed to different levels of indoor air pollution, who did or did not have high-grade squamous dysplasia and who did or did not have a family history of upper gastrointestinal tract (UGI) cancer. 147 samples were evaluated, including 23 (16%) from patients with high-grade dysplasia and 48 (33%) from patients without dysplasia who heated their homes with coal, without a chimney (a 'high' indoor air pollution group), and 27 (18%) from patients with high-grade dysplasia and 49 (33%) from patients without dysplasia who did not heat their homes at all (a 'low' indoor air pollution group). Sixty-four (44%) had a family history of UGI cancer. RNA was extracted and quantitative PCR analysis was done. AhR gene expression was detectable in 85 (58%) of the samples and was >9-fold higher in those with a family history of UGI cancer (median expression (interquartile range), -1,964 (-18,000, -610) versus -18,000 (-18,000, -1036); P = 0.02, Wilcoxon rank-sum test). Heating status, dysplasia category, age, gender, and smoking were not associated with AhR expression (linear regression; all P values {ge} 0.1). AhR expression was higher in patients with a family history of UGI cancer. Such individuals may be more susceptible to the deleterious effects of PAH exposure, including PAH-induced cancer.

  15. Tetrandrine ameliorates collagen-induced arthritis in mice by restoring the balance between Th17 and Treg cells via the aryl hydrocarbon receptor.

    PubMed

    Yuan, Xusheng; Tong, Bei; Dou, Yannong; Wu, Xin; Wei, Zhifeng; Dai, Yue

    2016-02-01

    Tetrandrine is an alkaloid constituent of the root of Stephania tetrandra S. Moore. The long-term clinical uses of tetrandrine for treatments of rheumatalgia and arthralgia as well as the inhibition of rat adjuvant-induced arthritis imply that tetrandrine may have therapeutic potential in rheumatoid arthritis (RA). Here, we explored its anti-RA mechanism in collagen-induced arthritis (CIA) in relation to the balance between T helper (Th) 17 cells and regulatory T (Treg) cells. DBA/1 mice were immunized with chicken type II collagen and were orally administered tetrandrine for 14 consecutive days. Then, the mice were sacrificed, their joints were removed for histological analysis, and spleens and mesenteric lymph nodes (MLNs) were removed to examine the Th17 and Treg cells. Tetrandrine markedly alleviated the severity of arthritis, reduced the serum levels of pro-inflammatory cytokines, and restored the Th17/Treg balance, as demonstrated by the serum levels of their related cytokines (IL-17 and IL-10) and the proportion of each cell type. Tetrandrine inhibited Th17 cell differentiation and induced Treg cell differentiation in vitro . Notably, aryl hydrocarbon receptor (AhR) was proven to play a crucial role in tetrandrine-mediated T cell differentiation. The correlation between AhR activation, regulation of Th17/Treg and amelioration of arthritis by tetrandrine was verified in the CIA mice. Moreover, tetrandrine might be a ligand of AhR because it facilitated the expression of the AhR target gene cytochrome P450 1A1 (CYP1A1) and the activation of its downstream signaling pathways. Taken together, tetrandrine exerts its anti-arthritis efficacy by restoring Th17/Treg balance via AhR. PMID:26640276

  16. Diphenylarsinic acid, a chemical warfare-related neurotoxicant, promotes liver carcinogenesis via activation of aryl hydrocarbon receptor signaling and consequent induction of oxidative DNA damage in rats.

    PubMed

    Wei, Min; Yamada, Takanori; Yamano, Shotaro; Kato, Minoru; Kakehashi, Anna; Fujioka, Masaki; Tago, Yoshiyuki; Kitano, Mistuaki; Wanibuchi, Hideki

    2013-11-15

    Diphenylarsinic acid (DPAA), a chemical warfare-related neurotoxic organic arsenical, is present in the groundwater and soil in some regions of Japan due to illegal dumping after World War II. Inorganic arsenic is carcinogenic in humans and its organic arsenic metabolites are carcinogenic in animal studies, raising serious concerns about the carcinogenicity of DPAA. However, the carcinogenic potential of DPAA has not yet been evaluated. In the present study we found that DPAA significantly enhanced the development of diethylnitrosamine-induced preneoplastic lesions in the liver in a medium-term rat liver carcinogenesis assay. Evaluation of the expression of cytochrome P450 (CYP) enzymes in the liver revealed that DPAA induced the expression of CYP1B1, but not any other CYP1, CYP2, or CYP3 enzymes, suggesting that CYP1B1 might be the enzyme responsible for the metabolic activation of DPAA. We also found increased oxidative DNA damage, possibly due to elevated CYP1B1 expression. Induction of CYP1B1 has generally been linked with the activation of AhR, and we found that DPAA activates the aryl hydrocarbon receptor (AhR). Importantly, the promotion effect of DPAA was observed only at a dose that activated the AhR, suggesting that activation of AhR and consequent induction of AhR target genes and oxidative DNA damage plays a vital role in the promotion effects of DPAA. The present study provides, for the first time, evidence regarding the carcinogenicity of DPAA and indicates the necessity of comprehensive evaluation of its carcinogenic potential using long-term carcinogenicity studies.

  17. Tetrandrine ameliorates collagen-induced arthritis in mice by restoring the balance between Th17 and Treg cells via the aryl hydrocarbon receptor.

    PubMed

    Yuan, Xusheng; Tong, Bei; Dou, Yannong; Wu, Xin; Wei, Zhifeng; Dai, Yue

    2016-02-01

    Tetrandrine is an alkaloid constituent of the root of Stephania tetrandra S. Moore. The long-term clinical uses of tetrandrine for treatments of rheumatalgia and arthralgia as well as the inhibition of rat adjuvant-induced arthritis imply that tetrandrine may have therapeutic potential in rheumatoid arthritis (RA). Here, we explored its anti-RA mechanism in collagen-induced arthritis (CIA) in relation to the balance between T helper (Th) 17 cells and regulatory T (Treg) cells. DBA/1 mice were immunized with chicken type II collagen and were orally administered tetrandrine for 14 consecutive days. Then, the mice were sacrificed, their joints were removed for histological analysis, and spleens and mesenteric lymph nodes (MLNs) were removed to examine the Th17 and Treg cells. Tetrandrine markedly alleviated the severity of arthritis, reduced the serum levels of pro-inflammatory cytokines, and restored the Th17/Treg balance, as demonstrated by the serum levels of their related cytokines (IL-17 and IL-10) and the proportion of each cell type. Tetrandrine inhibited Th17 cell differentiation and induced Treg cell differentiation in vitro . Notably, aryl hydrocarbon receptor (AhR) was proven to play a crucial role in tetrandrine-mediated T cell differentiation. The correlation between AhR activation, regulation of Th17/Treg and amelioration of arthritis by tetrandrine was verified in the CIA mice. Moreover, tetrandrine might be a ligand of AhR because it facilitated the expression of the AhR target gene cytochrome P450 1A1 (CYP1A1) and the activation of its downstream signaling pathways. Taken together, tetrandrine exerts its anti-arthritis efficacy by restoring Th17/Treg balance via AhR.

  18. Effects of enzyme induction on the distribution of the food carcinogen 2-amino-3,8-dimethyl-imidazo[4,5-ss]-quinoxaline (MeIQx) in Ah-receptor- responsive- and Ah-receptor-non-responsive mice.

    PubMed

    Vikse, R; Ingebrigtsen, K; Klungsøyr, L; Alexander, J

    1995-07-01

    The distribution of the food carcinogen 2-amino-3,8-dimethyl-imidazo[4,5-ss]quinoxaline (MeIQx) was studied in Ah-responsive-(C57BL/6J) and Ah-non-responsive mice (DBA/2N). The time dependent organ distribution of radioactivity after 14C-MeIQx (10 mg/kg) administration in C57BL/6J showed that at day 4 most of the radioactivity had been excreted and that the remaining radioactivity was found in liver, kidneys, lungs and spleen. C57BL/6J bound more radioactivity in the kidneys than the DBA/2N strain whereas approximately the same amount was left in the liver and lungs in both strains 4 days after MeIQx exposure. Liver microsomes of the two strains had approximately the same ability to activate MeIQx in the Ames Salmonella assay. beta-Naphthoflavone treatment of the animals greatly increased microsomal activating capacity, but only in the C57BL/6J strain. Isosafrole treatment of the animals only slightly increased the activating capacity, but particularly with microsomes from the DBA/2N strain, displacement of the putative inhibitory isosafrole metabolite greatly increased their activating capacity. In the whole animals pretreatment with beta-naphthoflavone, which induces P450IA only in the C57BL/6J strain, did not significantly change the amount of retained radioactivity in any of the strains. Isosafrole induces only P450IA2, the major N2-hydroxylating enzyme of heterocyclic amines, in both strains. Such pretreatment reduced the amount retained in the kidney of both strains whereas it reduced the retained amount of radioactivity in the liver with about 60% only in the Ah-non-responsive strain (DBA/2N). The effect of isosafrole did not persist when MeIQx was given three days after the last injection.(ABSTRACT TRUNCATED AT 250 WORDS)

  19. Siaα2-3Galβ1- Receptor Genetic Variants Are Associated with Influenza A(H1N1)pdm09 Severity.

    PubMed

    Maestri, Alvino; Sortica, Vinicius Albuquerque; Tovo-Rodrigues, Luciana; Santos, Mirleide Cordeiro; Barbagelata, Luana; Moraes, Milene Raiol; Alencar de Mello, Wyller; Gusmão, Leonor; Sousa, Rita Catarina Medeiros; Emanuel Batista Dos Santos, Sidney

    2015-01-01

    Different host genetic variants may be related to the virulence and transmissibility of pandemic Influenza A(H1N1)pdm09, influencing events such as binding of the virus to the entry receptor on the cell of infected individuals and the host immune response. In the present study, two genetic variants of the ST3GAL1 gene, which encodes the Siaα2-3Galβ1- receptor to which influenza A(H1N1)pdm09 virus binds for entry into the host cell, were investigated in an admixed Brazilian population. First, the six exons encoding the ST3GAL1 gene were sequenced in 68 patients infected with strain A(H1N1)pdm09. In a second phase of the study, the rs113350588 and rs1048479 polymorphisms identified in this sample were genotyped in a sample of 356 subjects from the northern and northeastern regions of Brazil with a diagnosis of pandemic influenza. Functional analysis of the polymorphisms was performed in silico and the influence of these variants on the severity of infection was evaluated. The results suggest that rs113350588 and rs1048479 may alter the function of ST3GAL1 either directly through splicing regulation alteration and/or indirectly through LD with SNP with regulatory function. In the study the rs113350588 and rs1048479 polymorphisms were in linkage disequilibrium in the population studied (D' = 0.65). The GC haplotype was associated with an increased risk of death in subjects with influenza (OR = 4.632, 95% CI = 2.10;1.21). The AT haplotype was associated with an increased risk of severe disease and death (OR = 1.993, 95% CI = 1.09;3.61 and OR 4.476, 95% CI = 2.37;8.44, respectively). This study demonstrated for the first time the association of ST3GAL1 gene haplotypes on the risk of more severe disease and death in patients infected with Influenza A(H1N1)pdm09 virus. PMID:26436774

  20. Effects of perfluoroalkyl acids on the function of the thyroid hormone and the aryl hydrocarbon receptor.

    PubMed

    Long, Manhai; Ghisari, Mandana; Bonefeld-Jørgensen, Eva Cecilie

    2013-11-01

    Perfluoroalkyl acids (PFAAs) are perfluorinated compounds that widely exist in the environment and can elicit adverse effects including endocrine disruption in humans and animals. This study investigated the effect of seven PFAAs on the thyroid hormone (TH) system assessing the proliferation of the 3,3',5-triiodo-L-thryonine (T3)-dependent rat pituitary GH3 cells using the T-screen assay and the effect on the aryl hydrocarbon receptor (AhR) transactivation in the AhR-luciferase reporter gene bioassay. A dose-dependent impact on GH3 cells was observed in the range 1×10(-9)-1×10(-4) M: seven PFAAs (perfluorooctane sulfonate (PFOS), perfluorohexane sulfonate (PFHxS), perfluorooctanoic acid, perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), perfluoroundecanoic acid (PFUnA), and perfluorododecanoic acid (PFDoA)) inhibited the GH3 cell growth, and four PFAAs (PFOS, PFHxS, PFNA, and PFUnA) antagonized the T3-induced GH3 cell proliferation. At the highest test concentration, PFHxS showed a further increase of the T3-induced GH3 growth. Among the seven tested PFAAs, only PFDoA and PFDA elicited an activating effect on the AhR. In conclusion, PFAAs possess in vitro endocrine-disrupting potential by interfering with TH and AhR functions, which need to be taken into consideration when assessing the impact on human health. PMID:23539207

  1. AhR-dependent secretion of PDGF-BB by human classically activated macrophages exposed to DEP extracts stimulates lung fibroblast proliferation

    SciTech Connect

    Jaguin, Marie; Fardel, Olivier; Lecureur, Valérie

    2015-06-15

    Lung diseases are aggravated by exposure to diesel exhaust particles (DEPs) found in air pollution. Macrophages are thought to play a crucial role in lung immune response to these pollutants, even if the mechanisms involved remain incompletely characterized. In the present study, we demonstrated that classically and alternative human macrophages (MΦ) exhibited increased secretion of PDGF-B in response to DEP extract (DEPe). This occurred via aryl hydrocarbon receptor (AhR)-activation because DEPe-induced PDGF-B overexpression was abrogated after AhR expression knock-down by RNA interference, in both M1 and M2 polarizing MΦ. In addition, TCDD and benzo(a)pyrene, two potent AhR ligands, also significantly increased mRNA expression of PDGF-B in M1 MΦ, whereas some weak ligands of AhR did not. We next evaluated the impact of conditioned media (CM) from MΦ culture exposed to DEPe or of recombinant PDGF-B onto lung fibroblast proliferation. The tyrosine kinase inhibitor, AG-1295, prevents phosphorylations of PDGF-Rβ, AKT and ERK1/2 and the proliferation of MRC-5 fibroblasts induced by recombinant PDGF-B and by CM from M1 polarizing MΦ, strongly suggesting that the PDGF-BB secreted by DEPe-exposed MΦ is sufficient to activate the PDGF-Rβ pathway of human lung fibroblasts. In conclusion, we demonstrated that human MΦ, whatever their polarization status, secrete PDGF-B in response to DEPe and that PDGF-B is a target gene of AhR. Therefore, induction of PDGF-B by DEP may participate in the deleterious effects towards human health triggered by such environmental urban contaminants. - Highlights: • PDGF-B expression and secretion are increased by DEPe exposure in human M1 and M2 MΦ. • DEPe-induced PDGF-B expression is aryl-hydrocarbon-dependent. • DEPe-exposed M1 MΦ secrete sufficient PDGF-B to increase lung fibroblast proliferation.

  2. Dioxin-Dependent and Dioxin-Independent Gene Batteries: Comparison of Liver and Kidney in AHR-Null Mice

    PubMed Central

    Boutros, Paul C.; Bielefeld, Kirsten A.; Pohjanvirta, Raimo; Harper, Patricia A.

    2009-01-01

    The aryl hydrocarbon receptor (AHR) is a widely expressed ligand-dependent transcription factor that mediates cellular responses to dioxins and other planar aromatic hydrocarbons. Ahr-null mice are refractory to the toxic effects of dioxin exposure. Although some mechanistic aspects of AHR activity are well understood, the tissue specificity of AHR effects remains unclear, both during development and following administration of exogenous ligands. To address the latter issue, we defined and compared transcriptional responses to dioxin exposure in the liver and kidney of wild-type and Ahr-null adult C57BL/6J mice treated with either 2,3,7,8-tetrachlorodibenzo-p-dioxin or corn-oil vehicle. In both tissues, essentially all effects of dioxin on hepatic mRNA levels were mediated by the AHR. Although 297 genes were altered by dioxin exposure in the liver, only 17 were changed in the kidney, including a number of well-established AHR target genes. Ahr genotype had a large effect in both tissues, profoundly remodeling both the renal and hepatic transcriptomes. Surprisingly, a large number of genes were affected by Ahr genotype in both tissues, suggesting the presence of a basal AHR gene battery. Alterations of the renal transcriptome in Ahr-null animals were associated with perturbation of specific functional pathways and enrichment of specific DNA motifs. Our results demonstrate the importance of intertissue comparisons, highlight the basal role of the AHR in liver and kidney, and support a role in development or normal physiology. PMID:19759094

  3. Alternative in vitro approach for assessing AHR-mediated CYP1A induction by dioxins in wild cormorant (Phalacrocorax carbo) population.

    PubMed

    Thuruthippallil, Leena Mol; Kubota, Akira; Kim, Eun-Young; Iwata, Hisato

    2013-06-18

    Our line of papers revealed that the common (great) cormorant (Phalacrocorax carbo) possesses two isoforms of the aryl hydrocarbon receptor (ccAHR1 and ccAHR2). This paper addresses in vitro tests of the ccAHR signaling pathways to solve two questions: (1) whether there are functional differences in the two ccAHR isoforms, and (2) whether a molecular perturbation, cytochrome P450 1A (ccCYP1A) induction, in the population-level can be predicted from the in vitro tests. The transactivation potencies mediated by ccAHR1 and ccAHR2 were measured in COS-7 cells treated with 15 selected dioxins and related compounds (DRCs), where ccAHR1 or ccAHR2 expression plasmid and ccCYP1A5 promoter/enhancer-linked luciferase reporter plasmid were transfected. For congeners that exhibited dose-dependent luciferase activities, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) relative potencies (REPs) and induction equivalency factors (IEFs) were estimated. ccAHR1-IEF profile was similar to WHO avian TCDD toxic equivalency factor (TEF) profile except for dioxin-like polychlorinated biphenyls that showed lower IEFs in ccAHR1-driven reporter assay. ccAHR2-IEF profile was different from WHO TEFs and ccAHR1-IEFs. Notably, 2,3,4,7,8-PeCDF was more potent than TCDD for ccAHR2-mediated response. Using ccAHR1- and ccAHR2-IEFs and hepatic DRC concentrations in the Lake Biwa cormorant population, total TCDD induction equivalents (IEQs) were calculated for each ccAHR-mediated response. Nonlinear regression analyses provided significant sigmoidal relationships of ccAHR1- and ccAHR2-derived IEQs with hepatic ccCYP1A5 mRNA levels, supporting the results of in vitro ccAHR-mediated TCDD dose-response curves. Collectively, our in vitro AHR reporter assay potentially could be an alternative to molecular epidemiology of the species of concern regarding CYP1A induction by AHR ligands. PMID:23676118

  4. Evaluation of the Ecotoxicity of Sediments from Yangtze River Estuary and Contribution of Priority PAHs to Ah Receptor-Mediated Activities

    PubMed Central

    Liu, Li; Chen, Ling; Shao, Ying; Zhang, Lili; Floehr, Tilman; Xiao, Hongxia; Yan, Yan; Eichbaum, Kathrin; Hollert, Henner; Wu, Lingling

    2014-01-01

    In this study, in vitro bioassays were performed to assess the ecotoxicological potential of sediments from Yangtze River estuary. The cytotoxicity and aryl hydrocarbon receptor (AhR)-mediated toxicity of sediment extracts with rainbow trout (Oncorhynchus mykiss) liver cells were determined by neutral red retention and 7-ethoxyresorufin-O-deethylase assays. The cytotoxicity and AhR-mediated activity of sediments from the Yangtze River estuary ranged from low level to moderate level compared with the ecotoxicity of sediments from other river systems. However, Yangtze River releases approximately 14 times greater water discharge compared with Rhine, a major river in Europe. Thus, the absolute pollution mass transfer of Yangtze River may be detrimental to the environmental quality of estuary and East China Sea. Effect-directed analysis was applied to identify substances causing high dioxin-like activities. To identify unknown substances contributing to dioxin-like potencies of whole extracts, we fractionated crude extracts by open column chromatography. Non-polar paraffinic components (F1), weakly and moderately polar components (F2), and highly polar substances (F3) were separated from each crude extract of sediments. F2 showed the highest dioxin-like activities. Based on the results of mass balance calculation of chemical toxic equivalent concentrations (TEQs), our conclusion is that priority polycyclic aromatic hydrocarbons indicated a low portion of bio-TEQs ranging from 1% to 10% of crude extracts. Further studies should be conducted to identify unknown pollutants. PMID:25111307

  5. Genetic variability of aryl hydrocarbon receptor (AhR)-mediated regulation of the human UDP glucuronosyltransferase (UGT) 1A4 gene

    SciTech Connect

    Erichsen, Thomas J.; Ehmer, Ursula; Kalthoff, Sandra; Lankisch, Tim O.; Mueller, Tordis M.; Munzel, Peter A.; Manns, Michael P.; Strassburg, Christian P.

    2008-07-15

    UDP glucuronosyltransferases (UGTs) play an important role for drug detoxification and toxicity. UGT function is genetically modulated by single nucleotide polymorphisms (SNPs) which lead to the expression of functionally altered protein, or altered expression levels. UGT1A4 activity includes anticonvulsants, antidepressants and environmental mutagens. In this study the induction of the human UGT1A4 gene and a potential influence of genetic variation in its promoter region were analyzed. SNPs at bp - 219 and - 163 occurred in 9% among 109 blood donors reducing UGT1A4 transcription by 40%. UGT1A4 transcription was dioxin inducible. Reporter gene experiments identified 2 xenobiotic response elements (XRE), which were functionally confirmed by mutagenesis analyses, and binding was demonstrated by electromobility shift assays. Constitutive human UGT1A4 gene expression and induction was aryl hydrocarbon receptor (AhR)-dependent, and reduced in the presence of SNPs at bp - 219 and - 163. AhR-mediated regulation of the human UGT1A4 gene by two XRE and a modulation by naturally occurring genetic variability by SNPs is demonstrated, which indicates gene-environment interaction with potential relevance for drug metabolism.

  6. Evaluation of the ecotoxicity of sediments from Yangtze river estuary and contribution of priority PAHs to ah receptor--mediated activities.

    PubMed

    Liu, Li; Chen, Ling; Shao, Ying; Zhang, Lili; Floehr, Tilman; Xiao, Hongxia; Yan, Yan; Eichbaum, Kathrin; Hollert, Henner; Wu, Lingling

    2014-01-01

    In this study, in vitro bioassays were performed to assess the ecotoxicological potential of sediments from Yangtze River estuary. The cytotoxicity and aryl hydrocarbon receptor (AhR)-mediated toxicity of sediment extracts with rainbow trout (Oncorhynchus mykiss) liver cells were determined by neutral red retention and 7-ethoxyresorufin-O-deethylase assays. The cytotoxicity and AhR-mediated activity of sediments from the Yangtze River estuary ranged from low level to moderate level compared with the ecotoxicity of sediments from other river systems. However, Yangtze River releases approximately 14 times greater water discharge compared with Rhine, a major river in Europe. Thus, the absolute pollution mass transfer of Yangtze River may be detrimental to the environmental quality of estuary and East China Sea. Effect-directed analysis was applied to identify substances causing high dioxin-like activities. To identify unknown substances contributing to dioxin-like potencies of whole extracts, we fractionated crude extracts by open column chromatography. Non-polar paraffinic components (F1), weakly and moderately polar components (F2), and highly polar substances (F3) were separated from each crude extract of sediments. F2 showed the highest dioxin-like activities. Based on the results of mass balance calculation of chemical toxic equivalent concentrations (TEQs), our conclusion is that priority polycyclic aromatic hydrocarbons indicated a low portion of bio-TEQs ranging from 1% to 10% of crude extracts. Further studies should be conducted to identify unknown pollutants.

  7. Regulation of Ahr signaling by Nrf2 during development: Effects of Nrf2a deficiency on PCB126 embryotoxicity in zebrafish (Danio rerio)

    PubMed Central

    Rousseau, Michelle E.; Sant, Karilyn E.; Borden, Linnea R.; Franks, Diana G.; Hahn, Mark E.; Timme-Laragy, Alicia R.

    2015-01-01

    The embryotoxicity of co-planar PCBs is regulated by the aryl hydrocarbon receptor (Ahr), and has been reported to involve oxidative stress. Ahr participates in crosstalk with another transcription factor, Nfe2l2, or Nrf2. Nrf2 binds to antioxidant response elements to regulate the adaptive response to oxidative stress. To explore aspects of the crosstalk between Nrf2 and Ahr and its impact on development, we used zebrafish (Danio rerio) with a mutated DNA binding domain in Nrf2a (nrf2afh318/fh318), rendering these embryos more sensitive to oxidative stress. Embryos were exposed to 2 nM or 5 nM PCB126 at 24 hours post fertilization (prim-5 stage of pharyngula) and examined for gene expression and morphology at 4 days post fertilization (dpf; protruding –mouth stage). Nrf2a mutant eleutheroembryos were more sensitive to PCB126 toxicity at 4 dpf, and in the absence of treatment also displayed some subtle developmental differences from wildtype embryos, including delayed inflation of the swim bladder and smaller yolk sacs. We used qPCR to measure changes in expression of the nrf gene family, keap1a, keap1b, the ahr gene family, and known target genes. cyp1a induction by PCB126 was enhanced in the Nrf2a mutants (156-fold in wildtypes vs. 228-fold in mutants exposed to 5 nM). Decreased expression of heme oxygenase (decycling) 1 (hmox1) in the Nrf2a mutants was accompanied by increased nrf2b expression. Target genes of Nrf2a and AhR2, NAD(P)H:quinone oxidoreductase 1 (nqo1) and glutathione S-transferase, alpha-like (gsta1), showed a 2-5-fold increase in expression in the Nrf2a mutants as compared to wildtype. This study elucidates the interaction between two important transcription factor pathways in the developmental toxicity of co-planar PCBs. PMID:26325326

  8. Metformin suppresses CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating aryl hydrocarbon receptor expression

    SciTech Connect

    Do, Minh Truong; Kim, Hyung Gyun; Tran, Thi Thu Phuong; Khanal, Tilak; Choi, Jae Ho; Chung, Young Chul; Jeong, Tae Cheon; Jeong, Hye Gwang

    2014-10-01

    Induction of cytochrome P450 (CYP) 1A1 and CYP1B1 by environmental xenobiotic chemicals or endogenous ligands through the activation of the aryl hydrocarbon receptor (AhR) has been implicated in a variety of cellular processes related to cancer, such as transformation and tumorigenesis. Here, we investigated the effects of the anti-diabetes drug metformin on expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and inducible conditions. Our results indicated that metformin down-regulated the expression of CYP1A1 and CYP1B1 in breast cancer cells under constitutive and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced conditions. Down-regulation of AhR expression was required for metformin-mediated decreases in CYP1A1 and CYP1B1 expression, and the metformin-mediated CYP1A1 and CYP1B1 reduction is irrelevant to estrogen receptor α (ERα) signaling. Furthermore, we found that metformin markedly down-regulated Sp1 protein levels in breast cancer cells. The use of genetic and pharmacological tools revealed that metformin-mediated down-regulation of AhR expression was mediated through the reduction of Sp1 protein. Metformin inhibited endogenous AhR ligand-induced CYP1A1 and CYP1B1 expression by suppressing tryptophan-2,3-dioxygenase (TDO) expression in MCF-7 cells. Finally, metformin inhibits TDO expression through a down-regulation of Sp1 and glucocorticoid receptor (GR) protein levels. Our findings demonstrate that metformin reduces CYP1A1 and CYP1B1 expression in breast cancer cells by down-regulating AhR signaling. Metformin would be able to act as a potential chemopreventive agent against CYP1A1 and CYP1B1-mediated carcinogenesis and development of cancer. - Graphical abstract: Schematic of the CYP1A1 and CYP1B1 gene regulation by metformin. - Highlights: • Metformin inhibits CYP1A1 and CYP1B1 expression. • Metformin down-regulates the AhR signaling. • Metformin reduces Sp1 protein expression. • Metformin suppresses TDO expression.

  9. Severe liver cirrhosis markedly reduces AhR-mediated induction of cytochrome P450 in rats by decreasing the transcription of target genes.

    PubMed

    Floreani, Maura; De Martin, Sara; Gabbia, Daniela; Barbierato, Massimo; Nassi, Alberto; Mescoli, Claudia; Orlando, Rocco; Bova, Sergio; Angeli, Paolo; Gola, Elisabetta; Sticca, Antonietta; Palatini, Pietro

    2013-01-01

    Although the induction of cytochrome P450 (CYP) has long been investigated in patients with cirrhosis, the question whether liver dysfunction impairs the response to CYP inducers still remains unresolved. Moreover, the mechanism underlying the possible effect of cirrhosis on induction has not been investigated. Since ethical constraints do not permit methodologically rigorous studies in humans, this question was addressed by investigating the effect of the prototypical inducer benzo[a]pyrene (BP) on CYP1A1 and CYP1A2 in cirrhotic rats stratified according to the severity of liver dysfunction. We simultaneously assessed mRNA level, protein expression and enzymatic activity of the CYP1A enzymes, as well as mRNA and protein expressions of the aryl hydrocarbon receptor (AhR), which mediates the BP effect. Basal mRNA and protein expressions of CYP1A1 were virtually absent in both healthy and cirrhotic rats. On the contrary, CYP1A2 mRNA, protein and enzyme activity were constitutively present in healthy rats and decreased significantly as liver function worsened. BP treatment markedly increased the concentrations of mRNA and immunodetectable protein, and the enzymatic activities of both CYP1A enzymes to similar levels in healthy and non-ascitic cirrhotic rats. Induced mRNA levels, protein expressions and enzymatic activities of both CYPs were much lower in ascitic rats and were proportionally reduced. Both constitutive and induced protein expressions of AhR were significantly lower in ascitic than in healthy rats. These results indicate that the inducibility of CYP1A enzymes is well preserved in compensated cirrhosis, whereas it is markedly reduced when liver dysfunction becomes severe. Induction appears to be impaired at the transcriptional level, due to the reduced expression of AhR, which controls the transcription of CYP1A genes.

  10. Environmental pollutants parathion, paraquat and bisphenol A show distinct effects towards nuclear receptors-mediated induction of xenobiotics-metabolizing cytochromes P450 in human hepatocytes.

    PubMed

    Vrzal, Radim; Zenata, Ondrej; Doricakova, Aneta; Dvorak, Zdenek

    2015-10-01

    Environmental pollutants parathion, bisphenol A and paraquat were not systematically studied towards the effects on the expression of phase I xenobiotics-metabolizing cytochromes P450 (CYPs). We monitored their effects on the expression of selected CYPs in primary cultures of human hepatocytes. Moreover, we investigated their effects on the receptors regulating these CYPs, particularly arylhydrocarbon receptor (AhR), pregnane X receptor (PXR) and glucocorticoid receptor (GR) by gene reporter assays. We found that parathion and bisphenol A are the activators of AhR. Moreover, they are the inducers of CYP1A1 mRNA in hepatoma cells HepG2 as well as in human hepatocytes by AhR-dependent mechanism via formation of AhR-DNA-binding complex, as revealed by gel shift assay. All three compounds possessed anti-glucocorticoid action as revealed by GR-dependent gene reporter assay and a decline in tyrosine aminotransferase (TAT) gene expression in human hepatocytes. Moreover, parathion and bisphenol A are the activators of PXR and inducers of CYP3A4 mRNA and protein in the primary cultures of human hepatocytes. In conclusion, the studied compounds displayed distinct activities towards nuclear receptors involved in many biological processes and these findings may help us to better understand their adverse actions in pathological states followed after their exposure. PMID:26196221

  11. The Aryl Hydrocarbon Receptor in Immunity: Tools and Potential.

    PubMed

    Esser, Charlotte

    2016-01-01

    The signaling pathway of the evolutionary old transcription factor AhR is inducible by a number of small molecular weight chemicals, including toxicants such as polycyclic aromatic hydrocarbons, bacterial toxic pigments, and physiological compounds such as tryptophan derivatives or dietary indoles. AhR activation is of immunological importance, but at the same time mediates toxicity of environmental pollutants, such as immunosuppression by dioxins. Measuring AhR activity and identification of ligands is thus of great interest for a variety of research fields. In this chapter, I briefly introduce the AhR signaling pathway, its role in immunology, and the tools and assays needed to analyze AhR signaling. Both are also needed when therapeutic applications are envisioned.

  12. Examination of the in vitro (anti)estrogenic, (anti)androgenic and (anti)dioxin-like activities of tetralin, indane and isochroman derivatives using receptor-specific bioassays.

    PubMed

    Schreurs, Richard H M M; Sonneveld, Edwin; van der Saag, Paul T; van der Burg, Bart; Seinen, Willem

    2005-04-10

    Molecules derived from tetralin, indane and isochroman are often used in the synthesis of fragrance materials. The two polycyclic musk fragrances AHTN (6-acetyl-1,1,2,4,4,7-hexamethyltetralin), HHCB (1,2,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethylcyclopenta-gamma-2-benzopyran) and ADBI (4-acetyl-1,1-dimethyl-6-tert-butylindane) are derived from tetralin, isochroman and indane, respectively. In previous studies, AHTN and HHCB have been shown to antagonize estrogen receptors (ERs), both in vitro and in vivo. Here, we used two newly developed reporter gene assays, to examine the agonistic and antagonistic properties of several indane, tetralin and isochroman derivatives towards the human androgen receptor (AR) and aryl hydrocarbon receptor (AhR). Additionally, we also assessed (anti)estrogenicity of these compounds. A number of compounds showed weak estrogenic activity towards the human ER alpha. Several compounds showed (anti)estrogenic effects, starting at a concentration of 0.1 microM. Surprisingly, almost all compounds were found to be AR antagonists, starting at 0.1 microM. None of the compounds tested, showed either agonism or antagonism towards the AhR. Non-specific effects via crosstalk of the AhR and the ER or AR can therefore be ruled out. As far as we are aware, molecules derived from indane, tetralin and isochroman showing direct interaction with the ER and AR have not been reported previously.

  13. Induction of mouse UDP-glucuronosyltransferase mRNA expression in liver and intestine by activators of aryl-hydrocarbon receptor, constitutive androstane receptor, pregnane X receptor, peroxisome proliferator-activated receptor alpha, and nuclear factor erythroid 2-related factor 2.

    PubMed

    Buckley, David B; Klaassen, Curtis D

    2009-04-01

    UDP-glucuronosyltransferases (UGTs) catalyze the addition of UDP-glucuronic acid to endo- and xenobiotics, enhancing their water solubility and elimination. Many exogenous compounds, such as microsomal enzyme inducers (MEIs), alter gene expression through xenobiotic-responsive transcription factors, namely, the aryl hydrocarbon receptor (AhR), constitutive androstane receptor (CAR), pregnane X receptor (PXR), peroxisome proliferator-activated receptor alpha (PPARalpha), and nuclear factor erythroid 2-related factor 2 (Nrf2). These transcription factors regulate xenobiotic-inducible expression of hepatic and intestinal biotransformation enzymes and transporters. The purpose of this study was to determine hepatic and intestinal inducibility of mouse Ugt mRNA by MEIs. Male C57BL/6 mice were treated for four consecutive days with activators of AhR [2,3,7,8-tetrachlorodibenzodioxin (TCDD), polychlorinated biphenyl 126, and beta-naphthoflavone], CAR [1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP), phenobarbital, and diallyl sulfide], PXR [pregnenolone-16alpha-carbonitrile (PCN), spironolactone, and dexamethasone], PPARalpha (clofibrate, ciprofibrate, and diethylhexylphthalate), and Nrf2 (oltipraz, ethoxyquin, and butylated hydroxyanisole), respectively. Ugt1a1 mRNA expression in liver was induced by activators of all five transcription factor pathways, Ugt1a5 by Nrf2 activators, Ugt1a6 by all the pathways except CAR, and Ugt1a9 by all the pathways except Nrf2. Ugt2b35 mRNA in liver was induced by AhR activators and Ugt2b36 by CAR and PPARalpha activators. Throughout the small and large intestine, the AhR ligand TCDD increased Ugt1a6 and Ugt1a7 mRNA. In small intestine, the PXR activator PCN increased Ugt1a1, Ugt1a6, Ugt1a7, Ugt2b34, and Ugt2b35 mRNA in the duodenum. In conclusion, chemical activation of AhR, CAR, PXR, PPARalpha, and Nrf2 in mouse results in induction of distinct Ugt gene sets in liver and intestine, predominantly the Ugt1a isoforms.

  14. Vitamin E Status and Metabolism in Adult and Aged Aryl Hydrocarbon Receptor Null Mice

    PubMed Central

    Traber, Maret G.; Mustacich, Debbie J.; Sullivan, Laura C.; Leonard, Scott W.; Ahern-Rindell, Amelia; Kerkvliet, Nancy

    2009-01-01

    The aryl hydrocarbon receptor (AhR) is involved in regulation of mechanisms for detoxification of xenobiotics, as well as vitamin A metabolism. Vitamin E is a fat-soluble nutrient whose metabolism is initialized via the cytochrome P450 system. Thus, AhR absence could alter hepatic regulation of α-tocopherol metabolism. To test this hypothesis, we assessed vitamin E status in adult (2–5 m) and old (21–22 m), wildtype and AhR-null mice. Plasma α-tocopherol concentrations in AhR null mice (2.3 ± 1.2 μmol/L, n= 19) were lower than those of wildtype mice (3.2 ± 1.2, n=17, P=0.0131); those in old mice (3.2 ± 1.2, n= 20) were higher than those of adults (2.2 ± 1.0, n=16, p=0.0075). Hepatic α-tocopherol concentrations were not different between genotypes, but were nearly double in old (32 ± 8 nmol/g, n=20) as compared with adult mice (17 ± 2, n=16, p<0.0001). Hepatic Cyp3a concentrations in AhR-null mice were greater than those in wildtypes (p=0.0011). Genotype (p=0.0047), sex (p<0.0001) and age (p<0.0001) were significant modifiers of liver α-tocopherol metabolite (α-CEHC) concentrations. In general, Cyp3a concentrations correlated with hepatic α-tocopherol (r= 0.3957, p<0.05) and α-CEHC (r=0.4260, p<0.05) concentrations. Since there were no significant genotype differences in the hepatic α- or γ-tocopherol concentrations, AhR null mice did not have dramatically altered vitamin E metabolism. Since they did have higher hepatic α-CEHC concentrations, these data suggest metabolism was up-regulated in the AhR null mice in order to maintain the hepatic tocopherol concentrations similar to those of wildtypes. PMID:20153623

  15. Characterisation of chlorinated, brominated and mixed halogenated dioxins, furans and biphenyls as potent and as partial agonists of the Aryl hydrocarbon receptor.

    PubMed

    Wall, Richard J; Fernandes, Alwyn; Rose, Martin; Bell, David R; Mellor, Ian R

    2015-03-01

    The Aryl hydrocarbon receptor (AhR) binds a variety of chlorinated and brominated dioxins, furans and biphenyls. Mixed halogenated variants have been recently identified in food at significant levels but full characterisation requires potency data in order to gauge their impact on risk assessment. Rat H4IIE and human MCF-7 cells were treated with various mixed halogenated ligands. Antagonist properties were measured by treating cells with various concentrations of TCDD in the presence of EC25 of the putative antagonist. Measurement of CYP1A1 RNA was used to quantify the potency of agonism and antagonism. The PXDDs were found to be slightly less potent than the corresponding fully chlorinated congeners with the exception of 2-B,3,7,8-TriCDD which was 2-fold more potent than TCDD. PXDFs and non-ortho-PXBs were found to be more potent than their chlorinated congeners whilst several mono-ortho-substituted PXBs were shown to have partial agonistic properties. REPs were produced for a range of mixed halogenated AhR-activating ligands providing a more accurate estimation of potency for risk assessment. Several environmentally abundant biphenyls were shown to be antagonists and reduce the ability of TCDD to induce CYP1A1. The demonstration of antagonism for AhR ligands represents a challenge for existing REP risk assessment schemes for AhR ligands.

  16. Certain photooxidized derivatives of tryptophan bind with very high affinity to the Ah receptor and are likely to be endogenous signal substances

    SciTech Connect

    Rannug, A.; Rannug, U.; Rosenkranz, H.S.; Winqvist, L.; Westerholm, R.; Agurell, E.; Grafstroem, A.K.

    1987-11-15

    The purpose of the present study was to determine whether ultraviolet light (UV) irradiation of amino acids produces compounds with affinity for the Ah receptor. Aqueous solutions of L-tryptophan were exposed to radiation from an unfiltered high-pressure mercury lamp. The photoproducts formed were solvent-extracted or concentrated on Sep-Pak C18 cartridges. The concentrated extracts or eluants were treated for their ability to compete with /sup 3/H-labeled 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Binding was assayed in liver cytosolic preparations from Sprague-Dawley rats using a technique based on hydroxylapatite separation. Photoproducts with receptor affinity were formed in a time-dependent manner. Histidine and tryptamine also gave products upon UV irradiation that competed with TCDD. Commercial tryptophan, at least aged, contained trace amounts of impurities with receptor affinity. Analysis by TLC and high-pressure liquid chromatography of the photo-products of tryptophan showed a minimum of three different binding compounds. Two of the products were studied in greater detail. One of them, showing UV absorbance and yellow fluorescence, gave a molecular ion (M+) of 284 and the other gave M+ 312 but showed little UV absorption and fluorescence. The concentration, based on mass spectrometry quantifications, of the two compounds that displaced more than 50% of TCDD was found to be extremely low, giving Kd values of 0.44 nM (M+ 312) and 0.07 nM (M+ 284). The existence of high affinity receptors for oxidized amino acids is postulated and their possible role in the proliferative cellular responses to TCDD and tryptophan is discussed briefly.

  17. Evidence for New Light-Independent Pathways for Generation of the Endogenous Aryl Hydrocarbon Receptor Agonist FICZ.

    PubMed

    Smirnova, Anna; Wincent, Emma; Vikström Bergander, Linda; Alsberg, Tomas; Bergman, Jan; Rannug, Agneta; Rannug, Ulf

    2016-01-19

    Activation of the aryl hydrocarbon receptor (AhR), a conserved transcription factor best known as a target for highly toxic halogenated substances such as dioxin, under normal xenobiotic-free conditions is of considerable scientific interest. We have demonstrated previously that a photoproduct of tryptophan, 6-formylindolo[3,2-b]carbazole (FICZ), fulfills the criteria for an endogenous ligand for this receptor and proposed that this compound is the enigmatic mediator of the physiological functions of AhR. Here, we describe novel light-independent pathways by which FICZ can be formed. The oxidant H2O2 was shown to convert tryptophan to FICZ on its own in the absence of light. The enzymatic deamination of tryptamine yielded indole-3-acetaldehyde (I3A), which then rearranged to FICZ and its oxidation product, indolo[3,2-b]carbazole-6-carboxylic acid (CICZ). Indole-3-pyruvate (I3P) also produced I3A, FICZ, and CICZ. Malassezia yeast species, which constitute a part of the normal skin microbiota, produce a number of AhR activators from tryptophan. We identified both FICZ and CICZ among those products. Formation of FICZ from tryptophan or I3P produces a complex mixture of indole derivatives, some of which are CYP1A1 inhibitors. These can hinder the cellular clearance of FICZ and thereby increase its power as an AhR agonist. We present a general molecular mechanism involving dehydrogenations and oxidative coupling for the formation of FICZ in which I3A is the important precursor. In conclusion, our results suggest that FICZ is likely to be formed systemically. PMID:26686552

  18. AHR2-Mediated Transcriptomic Responses Underlying the Synergistic Cardiac Developmental Toxicity of PAHs

    PubMed Central

    Jayasundara, Nishad; Van Tiem Garner, Lindsey; Meyer, Joel N.; Erwin, Kyle N.; Di Giulio, Richard T.

    2015-01-01

    Polycyclic aromatic hydrocarbons (PAHs) induce developmental defects including cardiac deformities in fish. The aryl hydrocarbon receptor (AHR) mediates the toxicity of some PAHs. Exposure to a simple PAH mixture during embryo development consisting of an AHR agonist (benzo(a)pyrene-BaP) with fluoranthene (FL), an inhibitor of cytochrome p450 1(CYP1)—a gene induced by AHR activation—results in cardiac deformities. Exposure to BaP or FL alone at similar concentrations alters heart rates, but does not induce morphological deformities. Furthermore, AHR2 knockdown prevents the toxicity of BaP + FL mixture. Here, we used a zebrafish microarray analysis to identify heart-specific transcriptomic changes during early development that might underlie cardiotoxicity of BaP + FL. We used AHR2 morphant embryos to determine the role of this receptor in mediating toxicity. Control and knockdown embryos at 36 h post-fertilization were exposed to DMSO, 100 μg/l BaP, 500 μg/l FL, or 100 μg/l BaP + 500 μg/l FL, and heart tissues for RNA were extracted at 2, 6, 12, and 18 h-post-exposure (hpe), prior to the appearance of cardiac deformities. Data show AHR2-dependent BaP + FL effects on expression of genes involved in protein biosynthesis and neuronal development in addition to signaling molecules and their associated molecular pathways. Ca2+-cycling and muscle contraction genes were the most significantly differentially expressed category of transcripts when comparing BaP + FL-treated AHR2 morphant and control embryos. These differences were most prominent at 2 and 6 hpe. Therefore, we postulate that BaP + FL may affect cellular Ca2+ levels and subsequently cardiac muscle function, potentially underlying BaP + FL cardiotoxicity. PMID:25412620

  19. Aryl hydrocarbon receptor activation by TCDD reduces inflammation associated with Crohn's disease.

    PubMed

    Benson, Jenna M; Shepherd, David M

    2011-03-01

    Crohn's disease results from a combination of genetic and environmental factors that trigger an inappropriate immune response to commensal gut bacteria. The aryl hydrocarbon receptor (AhR) is well known for its involvement in the toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), an environmental contaminant that affects people primarily through the diet. Recently, TCDD was shown to suppress immune responses by generating regulatory T cells (Tregs). We hypothesized that AhR activation dampens inflammation associated with Crohn's disease. To test this hypothesis, we utilized the 2,4,6-trinitrobenzenesulfonic acid (TNBS) murine model of colitis. Mice were gavaged with TCDD prior to colitis induction with TNBS. Several parameters were examined including colonic inflammation via histological and flow cytometric analyses. TCDD-treated mice recovered body weight faster and experienced significantly less colonic damage. Reduced levels of interleukin (IL) 6, IL-12, interferon-gamma, and tumor necrosis factor-α demonstrated suppression of inflammation in the gut following TCDD exposure. Forkhead box P3 (Foxp3)(egfp) mice revealed that TCDD increased the Foxp3+ Treg population in gut immune tissue following TNBS exposure. Collectively, these results suggest that activation of the AhR by TCDD decreases colonic inflammation in a murine model of colitis in part by generating regulatory immune cells. Ultimately, this work may lead to the development of more effective therapeutics for the treatment of Crohn's disease.

  20. Differences in the action of lower and higher chlorinated polychlorinated naphthalene (PCN) congeners on estrogen dependent breast cancer cell line viability and apoptosis, and its correlation with Ahr and CYP1A1 expression.

    PubMed

    Gregoraszczuk, Ewa L; Barć, Justyna; Falandysz, Jerzy

    2016-07-29

    There are data showing that exposition to PCNs mixture increased incidence of gastrointestinal and respiratory neoplasms, but data regarding incidence of hormone-dependent cancer so far not shown. The objective was to determine if exposure to single lower and higher chlorinated PCN congeners is associated with altered proliferation and apoptosis of estrogen dependent breast cancer cells, and whether such effects are related to induction of AhR and CYP1A1 protein expression. MCF-7 cells were exposed to PCN 34, 39, 42, 46, 48, 52, 53, 54, 66, 67, 70, 71, 73 and 74 at concentrations of 100-10,000pg/ml. We evaluated the action of these PCN congeners on cell proliferation, DNA fragmentation and caspase-8,-9 activity. AhR and CYP1A1 protein expression and CYP1A1 activity was evaluated at a concentration of 1000pg/ml. An opposite action of tri- to tetraCNs than of penta-to heptaCNs on cell proliferation and apoptosis was evident. Tetra PCNs increased cell proliferation, but had no effect on DNA fragmentation nor caspase activity. Fast induction of CYP1A1 protein expression under the influence of lower chlorinated PCNs suggests faster metabolism and a possible stimulatory action of locally formed metabolites on cell proliferation. None of the higher chlorinated PCNs affected cell proliferation but all higher chlorinated PCNs increased caspase-8 activity, and hexa PCNs also increased caspase-9 activity. The rapid activation of the Ah receptor and CYP1A1 protein expression by higher chlorinated PCNs point to their toxicity; however, it is not sufficient for potential carcinogenicity. Action of lower chlorinated naphthalenes metabolites should be explored.

  1. In Vitro Transformation of Chlorinated Parabens by the Liver S9 Fraction: Kinetics, Metabolite Identification, and Aryl Hydrocarbon Receptor Agonist Activity.

    PubMed

    Terasaki, Masanori; Wada, Takeshi; Nagashima, Satoshi; Makino, Masakazu; Yasukawa, Hiro

    2016-01-01

    We investigated the kinetics of in vitro transformation of a dichlorinated propyl paraben (2-propyl 3,5-dichloro-4-hydroxybenzoate; Cl2PP) by the rat liver S9 fraction and assessed the aryl hydrocarbon receptor (AhR) agonist activity of the metabolite products identified in HPLC and GC/MS analysis and by metabolite syntheses. The results indicated that the chlorination of Cl2PP reduced its degradation rate by approximately 40-fold. Two hydroxylated metabolite products showed AhR agonist activity of up to 39% of that of the parent Cl2PP when assessed in a yeast (YCM3) reporter gene assay. The determination of the metabolic properties of paraben bioaccumulation presented here provides further information on the value of risk assessments of chlorinated parabens as a means to ensure human health and environmental safety. PMID:27250800

  2. Progesterone, as well as 17β-estradiol, is important for regulating AHR battery homoeostasis in the rat uterus.

    PubMed

    Rataj, Felicitas; Möller, Frank Josef; Jähne, Maria; Hönscheid, Pia; Zierau, Oliver; Vollmer, Günter; Kretzschmar, Georg

    2015-03-01

    Several studies indicate that the aryl hydrocarbon receptor (AHR), which plays an important role in mediating the toxicity of many industrial chemicals, plays an important role in the physiology of female reproductive tract organs. This makes it likely that the AHR and additional components of the AHR signalling pathway are under the control of female sex steroids. In a previous study, we could already demonstrate the regulation of many members of the AHR battery by 17β-estradiol (E2) in the uterus of rats. In this study, we addressed the potential role of progesterone (P4) in this context. In a comparative approach using ovariectomized rats which were treated for 3 days with either vehicle control, E2, progesterone (P4) or the combination of both hormones in addition to sham-operated animals, we could demonstrate that in addition to E2, P4 is also an important factor in regulating AHR signalling in the rat uterus. P4 has effects similar to E2 on uterine Ahr, Arnt and Arnt2 mRNA levels, resulting in a downregulation of these genes, while the E2-mediated downregulation of key AHR response genes Cyp1a1, Gsta2 and Ugt1 is completely antagonized by P4. As with E2, P4 leads to an increase in uterine AHR levels, especially in the endometrial epithelium despite the decrease in corresponding mRNA levels. This indicates a complex gene-specific regulatory network involving E2, P4 and possibly AHR itself to maintain all components of the AHR signalling cascade at the required levels during all stages of the oestrous cycle and pregnancy.

  3. Dioxin mediates downregulation of the reduced folate carrier transport activity via the arylhydrocarbon receptor signalling pathway

    SciTech Connect

    Halwachs, Sandra; Lakoma, Cathleen; Gebhardt, Rolf; Schaefer, Ingo; Seibel, Peter; Honscha, Walther

    2010-07-15

    Dioxins such as 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD) are common environmental contaminants known to regulate several genes via activation of the transcription factor aryl hydrocarbon receptor (AhR) associated with the development of numerous adverse biological effects. However, comparatively little is known about the molecular mechanisms by which dioxins display their toxic effects in vertebrates. The 5' untranslated region of the hepatocellular Reduced folate carrier (Rfc1; Slc19a1) exhibits AhR binding sites termed dioxin responsive elements (DRE) that have as yet only been found in the promoter region of prototypical TCDD target genes. Rfc1 mediated transport of reduced folates and antifolate drugs such as methotrexate (MTX) plays an essential role in physiological folate homeostasis and MTX cancer chemotherapy. In order to determine whether this carrier represents a target gene of dioxins we have investigated the influence of TCDD on functional Rfc1 activity in rat liver. Pre-treatment of rats with TCDD significantly diminished hepatocellular Rfc1 uptake activity in a time- and dose-dependent manner. In further mechanistic studies we demonstrated that this reduction was due to TCDD-dependent activation of the AhR signalling pathway. We additionally showed that binding of the activated receptor to DRE motifs in the Rfc1 promoter resulted in downregulation of Rfc1 gene expression and reduced carrier protein levels. As downregulation of pivotal Rfc1 activity results in functional folate deficiency associated with an elevated risk of cardiovascular diseases or carcinogenesis, our results indicate that deregulation of this essential transport pathway represents a novel regulatory mechanism how dioxins display their toxic effects through the Ah receptor.

  4. Amino acid substitution D222N from fatal influenza infection affects receptor-binding properties of the influenza A(H1N1)pdm09 virus.

    PubMed

    Matos-Patrón, Adriana; Byrd-Leotis, Lauren; Steinhauer, David A; Barclay, Wendy S; Ayora-Talavera, Guadalupe

    2015-10-01

    We have analyzed the receptor binding profile of A(H1N1)pdm09 recombinant influenza viruses containing the amino acid substitution D222N which has been associated with a fatal case of infection. This mutation was investigated in conjunction with a secondary mutation, S185N. Using human tracheobronchial epithelial cells (HTBE), we found that single mutation D222N affects the binding and replication of the virus during initial stages of infection, with limited but preferred tropism to non-ciliated cells expressing α2,6-SA. However, in conjunction with the S185N change, the (D222N, S185N) virus shows a remarkable increase in binding and replication efficiency, with tropism for both ciliated and non-ciliated cells. Glycan microarray analysis demonstrated correlation between the binding profile and the cell tropism observed in the HTBE cells. These findings suggest that viruses with D222N required compensatory mutations such as S185N to maintain viral fitness, and in combination, affect the pathogenicity of the virus and the clinical outcome.

  5. Aryl Hydrocarbon Receptor Activation by TCDD Modulates Expression of Extracellular Matrix Remodeling Genes during Experimental Liver Fibrosis

    PubMed Central

    Lamb, Cheri L.; Cholico, Giovan N.; Perkins, Daniel E.; Fewkes, Michael T.; Oxford, Julia Thom; Lujan, Trevor J.; Morrill, Erica E.

    2016-01-01

    The aryl hydrocarbon receptor (AhR) is a soluble, ligand-activated transcription factor that mediates the toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Increasing evidence implicates the AhR in regulating extracellular matrix (ECM) homeostasis. We recently reported that TCDD increased necroinflammation and myofibroblast activation during liver injury elicited by carbon tetrachloride (CCl4). However, TCDD did not increase collagen deposition or exacerbate fibrosis in CCl4-treated mice, which raises the possibility that TCDD may enhance ECM turnover. The goal of this study was to determine how TCDD impacts ECM remodeling gene expression in the liver. Male C57BL/6 mice were treated for 8 weeks with 0.5 mL/kg CCl4, and TCDD (20 μg/kg) was administered during the last two weeks. Results indicate that TCDD increased mRNA levels of procollagen types I, III, IV, and VI and the collagen processing molecules HSP47 and lysyl oxidase. TCDD also increased gelatinase activity and mRNA levels of matrix metalloproteinase- (MMP-) 3, MMP-8, MMP-9, and MMP-13. Furthermore, TCDD modulated expression of genes in the plasminogen activator/plasmin system, which regulates MMP activation, and it also increased TIMP1 gene expression. These findings support the notion that AhR activation by TCDD dysregulates ECM remodeling gene expression and may facilitate ECM metabolism despite increased liver injury.

  6. Aryl Hydrocarbon Receptor Activation by TCDD Modulates Expression of Extracellular Matrix Remodeling Genes during Experimental Liver Fibrosis

    PubMed Central

    Lamb, Cheri L.; Cholico, Giovan N.; Perkins, Daniel E.; Fewkes, Michael T.; Oxford, Julia Thom; Lujan, Trevor J.; Morrill, Erica E.

    2016-01-01

    The aryl hydrocarbon receptor (AhR) is a soluble, ligand-activated transcription factor that mediates the toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Increasing evidence implicates the AhR in regulating extracellular matrix (ECM) homeostasis. We recently reported that TCDD increased necroinflammation and myofibroblast activation during liver injury elicited by carbon tetrachloride (CCl4). However, TCDD did not increase collagen deposition or exacerbate fibrosis in CCl4-treated mice, which raises the possibility that TCDD may enhance ECM turnover. The goal of this study was to determine how TCDD impacts ECM remodeling gene expression in the liver. Male C57BL/6 mice were treated for 8 weeks with 0.5 mL/kg CCl4, and TCDD (20 μg/kg) was administered during the last two weeks. Results indicate that TCDD increased mRNA levels of procollagen types I, III, IV, and VI and the collagen processing molecules HSP47 and lysyl oxidase. TCDD also increased gelatinase activity and mRNA levels of matrix metalloproteinase- (MMP-) 3, MMP-8, MMP-9, and MMP-13. Furthermore, TCDD modulated expression of genes in the plasminogen activator/plasmin system, which regulates MMP activation, and it also increased TIMP1 gene expression. These findings support the notion that AhR activation by TCDD dysregulates ECM remodeling gene expression and may facilitate ECM metabolism despite increased liver injury. PMID:27672655

  7. Toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in developing red seabream (Pagrus major) embryo: an association of morphological deformities with AHR1, AHR2 and CYP1A expressions.

    PubMed

    Yamauchi, Masanobu; Kim, Eun-Young; Iwata, Hisato; Shima, Yasuhiro; Tanabe, Shinsuke

    2006-11-16

    The toxicity of dioxins such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is mainly mediated by the aryl hydrocarbon receptor (AHR), which regulates the multiple target genes including cytochrome P4501A (CYP1A). In general, bony fishes, which possess at least two distinct AHRs are one of the most sensitive vertebrates to TCDD in early life stage. However, the physiological and toxicological roles of piscine multiple AHRs are not fully understood, especially in marine fish. To understand which AHR is responsible for TCDD toxicity in a marine fish species, we characterized the early life stage toxicity related to the expression of AHRs and CYP1A in red seabream (Pagrus major). The embryos at 10h post-fertilization (hpf) were treated with 0-100 microg/L TCDD for 80 min waterborne exposure. TCDD dose-dependently elicited developmental toxicities including mortality, yolk sac edema, retarded body growth, spinal deformity, reduced heart rate, shortened snout, underdeveloped fin, heart, and lower jaw. Intriguingly, hemorrhage and pericardium edema, typical TCDD developmental defects noticed in other fish species, were not found in red seabream until test termination. The EC(egg)50s for yolk sac edema, underdeveloped fin, and spinal deformity were 170, 240, and 340 pg/g, respectively. The LC(egg)50 was 360 pg/g embryo, indicating that this species is one of the most sensitive fishes to TCDD toxicity. The expression levels of rsAHR1, rsAHR2 and CYP1A mRNAs were also determined in different developmental stages. The rsAHR2 mRNA expression dose-dependently increased following TCDD exposure, while rsAHR1 mRNA level was not altered. Level of rsAHR2 mRNA measured by two-step real-time PCR was 30 times higher than rsAHR1 in embryos treated with the highest dose. Temporal patterns of rsAHR2 and CYP1A mRNAs were similar in TCDD-treated embryos, representing a significant positive correlation between rsAHR2 and CYP1A mRNA levels, but not between rsAHR1 and CYP1A. In comparison of

  8. Benzo[a]pyrene induces intercellular adhesion molecule-1 through a caveolae and aryl hydrocarbon receptor mediated pathway

    SciTech Connect

    Oesterling, Elizabeth; Toborek, Michal; Hennig, Bernhard

    2008-10-15

    Toxicologic and epidemiologic studies have linked benzo[a]pyrene (B[a]P) exposure with cardiovascular diseases such as atherosclerosis. The mechanisms of action leading to these diseases have not been fully understood. One key step in the development of atherosclerosis is vascular endothelial dysfunction, which is characterized by increased adhesiveness. To determine if B[a]P could lead to increased endothelial adhesiveness, the effects of B[a]P on human endothelial cell intercellular adhesion molecule-1 (ICAM-1) expression was investigated. B[a]P was able to increase ICAM-1 protein only after pretreatment with the aryl hydrocarbon receptor (AhR) agonist {beta}-naphthoflavone ({beta}-NF). Knockdown of AhR by siRNA or treatment with AhR antagonist {alpha}-naphthoflavone ({alpha}-NF) eliminated the induction of ICAM-1 from B[a]P, confirming the necessity of AhR in this process. Likewise, B[a]P only increased monocyte adhesion to the vascular endothelium when cells were pretreated with {beta}-NF. Experiments were done to define a signaling mechanism. B[a]P increased phosphorylation of MEK and p38-MAPK, and inhibitors to these proteins blunted the ICAM-1 induction. B[a]P was also able to increase AP-1 DNA binding and phosphorylation of cJun. Phosphorylation of cJun was disrupted by MEK and p38-MAPK inhibitors linking the signaling cascade. Finally, the importance of membrane microdomains, caveolae, was demonstrated by knockdown of the structural protein caveolin-1. Disruption of caveolae eliminated the B[a]P-induced ICAM-1 expression. These data suggest a possible pro-inflammatory mechanism of action of B[a]P involving caveolae, leading to increased vascular endothelial adhesiveness, and this inflammation may be a critical step in the development of B[a]P-induced atherosclerosis.

  9. The circadian clock circuitry and the AHR signaling pathway in physiology and pathology.

    PubMed

    Anderson, George; Beischlag, Timothy V; Vinciguerra, Manlio; Mazzoccoli, Gianluigi

    2013-05-15

    Life forms populating the Earth must face environmental challenges to assure individual and species survival. The strategies predisposed to maintain organismal homeostasis and grant selective advantage rely on anticipatory phenomena facing periodic modifications, and compensatory phenomena facing unpredictable changes. Biological processes bringing about these responses are respectively driven by the circadian timing system, a complex of biological oscillators entrained to the environmental light/dark cycle, and by regulatory and metabolic networks that precisely direct the body's adjustments to variations of external conditions and internal milieu. A critical role in organismal homeostatic functions is played by the aryl hydrocarbon receptor (AHR) complex, which senses environmental and endogenous compounds, influences metabolic responses controlling phase I/II gene expression, and modulates vital phenomena such as development, inflammation and adaptive immunity. A physiological cross-talk between circadian and AHR signaling pathways has been evidenced. The alteration of AHR signaling pathway deriving from genetic damage with polymorphisms or mutations, or produced by exogenous or endogenous AHR activation, and chronodisruption caused by mismatch between the body's internal clock and geophysical time/social schedules, are capable of triggering pathological mechanisms involved in metabolic, immune-related and neoplastic diseases. On the other hand, the molecular components of the circadian clock circuitry and AHR signaling pathway may represent useful tools for preventive interventions and valuable targets of therapeutic approaches.

  10. In silico predictive studies of mAHR congener binding using homology modelling and molecular docking.

    PubMed

    Panda, Roshni; Cleave, A Suneetha Susan; Suresh, P K

    2014-09-01

    The aryl hydrocarbon receptor (AHR) is one of the principal xenobiotic, nuclear receptor that is responsible for the early events involved in the transcription of a complex set of genes comprising the CYP450 gene family. In the present computational study, homology modelling and molecular docking were carried out with the objective of predicting the relationship between the binding efficiency and the lipophilicity of different polychlorinated biphenyl (PCB) congeners and the AHR in silico. Homology model of the murine AHR was constructed by several automated servers and assessed by PROCHECK, ERRAT, VERIFY3D and WHAT IF. The resulting model of the AHR by MODWEB was used to carry out molecular docking of 36 PCB congeners using PatchDock server. The lipophilicity of the congeners was predicted using the XLOGP3 tool. The results suggest that the lipophilicity influences binding energy scores and is positively correlated with the same. Score and Log P were correlated with r = +0.506 at p = 0.01 level. In addition, the number of chlorine (Cl) atoms and Log P were highly correlated with r = +0.900 at p = 0.01 level. The number of Cl atoms and scores also showed a moderate positive correlation of r = +0.481 at p = 0.01 level. To the best of our knowledge, this is the first study employing PatchDock in the docking of AHR to the environmentally deleterious congeners and attempting to correlate structural features of the AHR with its biochemical properties with regards to PCBs. The result of this study are consistent with those of other computational studies reported in the previous literature that suggests that a combination of docking, scoring and ranking organic pollutants could be a possible predictive tool for investigating ligand-mediated toxicity, for their subsequent validation using wet lab-based studies.

  11. Dioxin induces Ahr-dependent robust DNA demethylation of the Cyp1a1 promoter via Tdg in the mouse liver

    PubMed Central

    Amenya, Hesbon Z.; Tohyama, Chiharu; Ohsako, Seiichiroh

    2016-01-01

    The aryl hydrocarbon receptor (Ahr) is a highly conserved nuclear receptor that plays an important role in the manifestation of toxicity induced by polycyclic aromatic hydrocarbons. As a xenobiotic sensor, Ahr is involved in chemical biotransformation through activation of drug metabolizing enzymes. The activated Ahr cooperates with coactivator complexes to induce epigenetic modifications at target genes. Thus, it is conceivable that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a potent Ahr ligand, may elicit robust epigenetic changes in vivo at the Ahr target gene cytochrome P450 1a1 (Cyp1a1). A single dose of TCDD administered to adult mice induced Ahr-dependent CpG hypomethylation, changes in histone modifications, and thymine DNA glycosylase (Tdg) recruitment at the Cyp1a1 promoter in the liver within 24 hrs. These epigenetic changes persisted until 40 days post-TCDD treatment and there was Cyp1a1 mRNA hyperinduction upon repeat administration of TCDD at this time-point. Our demethylation assay using siRNA knockdown and an in vitro methylated plasmid showed that Ahr, Tdg, and the ten-eleven translocation methyldioxygenases Tet2 and Tet3 are required for the TCDD-induced DNA demethylation. These results provide novel evidence of Ahr-driven active DNA demethylation and epigenetic memory. The epigenetic alterations influence response to subsequent chemical exposure and imply an adaptive mechanism to xenobiotic stress. PMID:27713569

  12. Dioxin induces Ahr-dependent robust DNA demethylation of the Cyp1a1 promoter via Tdg in the mouse liver

    NASA Astrophysics Data System (ADS)

    Amenya, Hesbon Z.; Tohyama, Chiharu; Ohsako, Seiichiroh

    2016-10-01

    The aryl hydrocarbon receptor (Ahr) is a highly conserved nuclear receptor that plays an important role in the manifestation of toxicity induced by polycyclic aromatic hydrocarbons. As a xenobiotic sensor, Ahr is involved in chemical biotransformation through activation of drug metabolizing enzymes. The activated Ahr cooperates with coactivator complexes to induce epigenetic modifications at target genes. Thus, it is conceivable that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a potent Ahr ligand, may elicit robust epigenetic changes in vivo at the Ahr target gene cytochrome P450 1a1 (Cyp1a1). A single dose of TCDD administered to adult mice induced Ahr-dependent CpG hypomethylation, changes in histone modifications, and thymine DNA glycosylase (Tdg) recruitment at the Cyp1a1 promoter in the liver within 24 hrs. These epigenetic changes persisted until 40 days post-TCDD treatment and there was Cyp1a1 mRNA hyperinduction upon repeat administration of TCDD at this time-point. Our demethylation assay using siRNA knockdown and an in vitro methylated plasmid showed that Ahr, Tdg, and the ten-eleven translocation methyldioxygenases Tet2 and Tet3 are required for the TCDD-induced DNA demethylation. These results provide novel evidence of Ahr-driven active DNA demethylation and epigenetic memory. The epigenetic alterations influence response to subsequent chemical exposure and imply an adaptive mechanism to xenobiotic stress.

  13. Aryl hydrocarbon receptor-independent toxicity of weathered crude oil during fish development.

    PubMed

    Incardona, John P; Carls, Mark G; Teraoka, Hiroki; Sloan, Catherine A; Collier, Tracy K; Scholz, Nathaniel L

    2005-12-01

    Polycyclic aromatic hydrocarbons (PAHs), derived largely from fossil fuels and their combustion, are pervasive contaminants in rivers, lakes, and nearshore marine habitats. Studies after the Exxon Valdez oil spill demonstrated that fish embryos exposed to low levels of PAHs in weathered crude oil develop a syndrome of edema and craniofacial and body axis defects. Although mechanisms leading to these defects are poorly understood, it is widely held that PAH toxicity is linked to aryl hydrocarbon receptor (AhR) binding and cytochrome P450 1A (CYP1A) induction. Using zebrafish embryos, we show that the weathered crude oil syndrome is distinct from the well-characterized AhR-dependent effects of dioxin toxicity. Blockade of AhR pathway components with antisense morpholino oligonucleotides demonstrated that the key developmental defects induced by weathered crude oil exposure are mediated by low-molecular-weight tricyclic PAHs through AhR-independent disruption of cardiovascular function and morphogenesis. These findings have multiple implications for the assessment of PAH impacts on coastal habitats. PMID:16330359

  14. Aryl hydrocarbon receptor-independent toxicity of weathered crude oil during fish development.

    PubMed

    Incardona, John P; Carls, Mark G; Teraoka, Hiroki; Sloan, Catherine A; Collier, Tracy K; Scholz, Nathaniel L

    2005-12-01

    Polycyclic aromatic hydrocarbons (PAHs), derived largely from fossil fuels and their combustion, are pervasive contaminants in rivers, lakes, and nearshore marine habitats. Studies after the Exxon Valdez oil spill demonstrated that fish embryos exposed to low levels of PAHs in weathered crude oil develop a syndrome of edema and craniofacial and body axis defects. Although mechanisms leading to these defects are poorly understood, it is widely held that PAH toxicity is linked to aryl hydrocarbon receptor (AhR) binding and cytochrome P450 1A (CYP1A) induction. Using zebrafish embryos, we show that the weathered crude oil syndrome is distinct from the well-characterized AhR-dependent effects of dioxin toxicity. Blockade of AhR pathway components with antisense morpholino oligonucleotides demonstrated that the key developmental defects induced by weathered crude oil exposure are mediated by low-molecular-weight tricyclic PAHs through AhR-independent disruption of cardiovascular function and morphogenesis. These findings have multiple implications for the assessment of PAH impacts on coastal habitats.

  15. The tryptophan-derived endogenous arylhydrocarbon receptor ligand 6-formylindolo[3,2-b]carbazole (FICZ) is a nanomolar UVA-photosensitizer in epidermal keratinocytes

    PubMed Central

    Williams, Joshua D.; Cabello, Christopher M.; Qiao, Shuxi; Wondrak, Georg T.

    2014-01-01

    Endogenous UVA-chromophores may act as sensitizers of oxidative stress underlying cutaneous photoaging and photocarcinogenesis, but the molecular identity of non-DNA key chromophores displaying UVA-driven photodyamic activity in human skin remains largely undefined. Here we report that 6-formylindolo[3,2-b]carbazole (FICZ), a tryptophan photoproduct and endogenous high affinity aryl hydrocarbon receptor (AhR) agonist, acts as a nanomolar photosensitizer potentiating UVA-induced oxidative stress irrespective of AhR ligand activity. In human HaCaT and primary epidermal keratinocytes, photodynamic induction of apoptosis was elicited by the combined action of solar simulated UVA and FICZ, whereas exposure to the isolated action of UVA or FICZ did not impair viability. In a human epidermal tissue reconstruct, FICZ/UVA-cotreatment caused pronounced phototoxicity inducing keratinocyte cell death, and FICZ photodynamic activity was also substantiated in a murine skin exposure model. Array analysis revealed pronounced potentiation of cellular heat shock, ER stress, and oxidative stress response gene expression observed only upon FICZ/UVA-cotreatment. FICZ photosensitization caused intracellular oxidative stress, and comet analysis revealed introduction of formamidopyrimidine-DNA glycosylase (FPG)-sensitive oxidative DNA lesions suppressible by antioxidant cotreatment. Taken together, our data demonstrate that the endogenous AhR ligand FICZ displays nanomolar photodynamic activity representing a molecular mechanism of UVA-induced photooxidative stress potentially operative in human skin. PMID:25431849

  16. In vitro assessment of retinoic acid and aryl hydrocarbon receptor activity of treated effluent from 39 wastewater-treatment plants in Victoria, Australia.

    PubMed

    Allinson, M; Shiraishi, F; Salzman, S A; Allinson, G

    2011-11-01

    This project involved the collection of final effluent samples from 39 wastewater-treatment plants (WWTPs) in Victoria, Australia, in late summer (late February to early March 2007). The 39 WWTPs included 15 lagoon-based plants and 24 with activated sludge-based processes. Samples were collected and subjected to measurement of retinoic acid receptor (RAR) and aryl hydrocarbon receptor (AhR) activity of the dissolved phase using yeast-based recombinant receptor-reporter gene bioassays. More than 90% of the effluents examined in this study elicited RAR activity (<0.5-198 ng/l a-t-RA equivalents [EQ]). All of the effluents had AhR activity (16-279 ng/l βNF EQ). Notwithstanding the paucity of comparative data, on the whole, the levels of RAR and AhR activity observed in this pilot survey of Victorian WWTP effluents were greater than those recently reported internationally. One assumption commonly made is that WWTP discharges will be diluted significantly in the receiving environment, further decreasing the potential risk of the discharges. Making this assumption may not be appropriate for some of Victoria's more ephemeral waterways or where effluent is discharged to an enclosed water body, such as a lake or terminal wetland. However, even where WWTP discharges represent all of the environmental flow in the warmer months, the observed RAR and AhR activity (as all-trans-retinoic acid (RA) and 2,3,7,8-tetrachloro-dibenzo-p-dioxin [TCDD] EQ, respectively) was still significantly lower than the concentrations of RA, and 2,3,7,8-TCCD known to cause developmental malformations in fish larvae after short-term exposure to these chemicals. Of perhaps greater concern, WWTP effluent can contain significant suspended solids (essentially biosolids), which may be a considerable sink for some hormonally active, hydrophobic compounds, and which may in turn increase the long-term exposure risk for aquatic fauna. Further studies of the nuclear and AhR activity of WWTP effluent suspended

  17. Dopamine D2-Receptor Antagonists Down-Regulate CYP1A1/2 and CYP1B1 in the Rat Liver.

    PubMed

    Harkitis, P; Daskalopoulos, E P; Malliou, F; Lang, M A; Marselos, M; Fotopoulos, A; Albucharali, G; Konstandi, M

    2015-01-01

    Dopaminergic systems regulate the release of several hormones including growth hormone (GH), thyroid hormones, insulin, glucocorticoids and prolactin (PRL) that play significant roles in the regulation of various Cytochrome P450 (CYP) enzymes. The present study investigated the role of dopamine D2-receptor-linked pathways in the regulation of CYP1A1, CYP1A2 and CYP1B1 that belong to a battery of genes controlled by the Aryl Hydrocarbon Receptor (AhR) and play a crucial role in the metabolism and toxicity of numerous environmental toxicants. Inhibition of dopamine D2-receptors with sulpiride (SULP) significantly repressed the constitutive and benzo[a]pyrene (B[a]P)-induced CYP1A1, CYP1A2 and CYP1B expression in the rat liver. The expression of AhR, heat shock protein 90 (HSP90) and AhR nuclear translocator (ARNT) was suppressed by SULP in B[a]P-treated livers, whereas the AhRR expression was increased by the drug suggesting that the SULP-mediated repression of the CYP1 inducibility is due to inactivation of the AhR regulatory system. At signal transduction level, the D2-mediated down-regulation of constitutive CYP1A1/2 and CYP1B1 expression appears to be mediated by activation of the insulin/PI3K/AKT pathway. PRL-linked pathways exerting a negative control on various CYPs, and inactivation of the glucocorticoid-linked pathways that positively control the AhR-regulated CYP1 genes, may also participate in the SULP-mediated repression of both, the constitutive and induced CYP1 expression. The present findings indicate that drugs acting as D2-dopamine receptor antagonists can modify several hormone systems that regulate the expression of CYP1A1, CYP1A2 and CYP1B1, and may affect the toxicity and carcinogenicity outcome of numerous toxicants and pre-carcinogenic substances. Therefore, these drugs could be considered as a part of the strategy to reduce the risk of exposure to environmental pollutants and pre-carcinogens. PMID:26466350

  18. Dopamine D2-Receptor Antagonists Down-Regulate CYP1A1/2 and CYP1B1 in the Rat Liver.

    PubMed

    Harkitis, P; Daskalopoulos, E P; Malliou, F; Lang, M A; Marselos, M; Fotopoulos, A; Albucharali, G; Konstandi, M

    2015-01-01

    Dopaminergic systems regulate the release of several hormones including growth hormone (GH), thyroid hormones, insulin, glucocorticoids and prolactin (PRL) that play significant roles in the regulation of various Cytochrome P450 (CYP) enzymes. The present study investigated the role of dopamine D2-receptor-linked pathways in the regulation of CYP1A1, CYP1A2 and CYP1B1 that belong to a battery of genes controlled by the Aryl Hydrocarbon Receptor (AhR) and play a crucial role in the metabolism and toxicity of numerous environmental toxicants. Inhibition of dopamine D2-receptors with sulpiride (SULP) significantly repressed the constitutive and benzo[a]pyrene (B[a]P)-induced CYP1A1, CYP1A2 and CYP1B expression in the rat liver. The expression of AhR, heat shock protein 90 (HSP90) and AhR nuclear translocator (ARNT) was suppressed by SULP in B[a]P-treated livers, whereas the AhRR expression was increased by the drug suggesting that the SULP-mediated repression of the CYP1 inducibility is due to inactivation of the AhR regulatory system. At signal transduction level, the D2-mediated down-regulation of constitutive CYP1A1/2 and CYP1B1 expression appears to be mediated by activation of the insulin/PI3K/AKT pathway. PRL-linked pathways exerting a negative control on various CYPs, and inactivation of the glucocorticoid-linked pathways that positively control the AhR-regulated CYP1 genes, may also participate in the SULP-mediated repression of both, the constitutive and induced CYP1 expression. The present findings indicate that drugs acting as D2-dopamine receptor antagonists can modify several hormone systems that regulate the expression of CYP1A1, CYP1A2 and CYP1B1, and may affect the toxicity and carcinogenicity outcome of numerous toxicants and pre-carcinogenic substances. Therefore, these drugs could be considered as a part of the strategy to reduce the risk of exposure to environmental pollutants and pre-carcinogens.

  19. Dopamine D2-Receptor Antagonists Down-Regulate CYP1A1/2 and CYP1B1 in the Rat Liver

    PubMed Central

    Harkitis, P.; Lang, M. A.; Marselos, M.; Fotopoulos, A.; Albucharali, G.; Konstandi, M.

    2015-01-01

    Dopaminergic systems regulate the release of several hormones including growth hormone (GH), thyroid hormones, insulin, glucocorticoids and prolactin (PRL) that play significant roles in the regulation of various Cytochrome P450 (CYP) enzymes. The present study investigated the role of dopamine D2-receptor-linked pathways in the regulation of CYP1A1, CYP1A2 and CYP1B1 that belong to a battery of genes controlled by the Aryl Hydrocarbon Receptor (AhR) and play a crucial role in the metabolism and toxicity of numerous environmental toxicants. Inhibition of dopamine D2-receptors with sulpiride (SULP) significantly repressed the constitutive and benzo[a]pyrene (B[a]P)-induced CYP1A1, CYP1A2 and CYP1B expression in the rat liver. The expression of AhR, heat shock protein 90 (HSP90) and AhR nuclear translocator (ARNT) was suppressed by SULP in B[a]P-treated livers, whereas the AhRR expression was increased by the drug suggesting that the SULP-mediated repression of the CYP1 inducibility is due to inactivation of the AhR regulatory system. At signal transduction level, the D2-mediated down-regulation of constitutive CYP1A1/2 and CYP1B1 expression appears to be mediated by activation of the insulin/PI3K/AKT pathway. PRL-linked pathways exerting a negative control on various CYPs, and inactivation of the glucocorticoid-linked pathways that positively control the AhR-regulated CYP1 genes, may also participate in the SULP-mediated repression of both, the constitutive and induced CYP1 expression. The present findings indicate that drugs acting as D2-dopamine receptor antagonists can modify several hormone systems that regulate the expression of CYP1A1, CYP1A2 and CYP1B1, and may affect the toxicity and carcinogenicity outcome of numerous toxicants and pre-carcinogenic substances. Therefore, these drugs could be considered as a part of the strategy to reduce the risk of exposure to environmental pollutants and pre-carcinogens. PMID:26466350

  20. Inhibition of AHR transcription by NF1C is affected by a single-nucleotide polymorphism, and is involved in suppression of human uterine endometrial cancer.

    PubMed

    Li, D; Takao, T; Tsunematsu, R; Morokuma, S; Fukushima, K; Kobayashi, H; Saito, T; Furue, M; Wake, N; Asanoma, K

    2013-10-10

    Involvement of the aryl hydrocarbon receptor (AHR) in carcinogenesis has been suggested in many studies. Upregulation of AHR has been reported in some cancer species, and an association between single-nucleotide polymorphisms (SNPs) of AHR and cancer risk or cancer development has also been reported. This evidence suggests the involvement of some specific SNPs in AHR transcriptional regulation in the process of carcinogenesis or cancer development, but there have been no studies to elucidate the mechanism involved. In this study, we identified the transcription factor Nuclear Factor 1-C (NF1C) as a candidate to regulate AHR transcription in a polymorphism-dependent manner. SNP rs10249788 was included in a consensus binding site for NF1C. Our results suggested that NF1C preferred the C allele to the T allele at rs10249788 for binding. Forced expression of NF1C suppressed the activity of the AHR promoter with C at rs10249788 stronger than that with T. Moreover, expression analysis of human uterine endometrial cancer (HEC) specimens showed greater upregulation of AHR and downregulation of NF1C than those of normal endometrium specimens. Sequence analysis showed HEC patients at advanced stages tended to possess T/T alleles more frequently than healthy women. We also demonstrated that NF1C suppressed proliferation, motility and invasion of HEC cells. This function was at least partially mediated by AHR. This study is the first to report that a polymorphism on the AHR regulatory region affected transcriptional regulation of the AHR gene in vitro. Because NF1C is a tumor suppressor, our new insights into AHR deregulation and its polymorphisms could reveal novel mechanisms of genetic susceptibility to cancer.

  1. Exposure to Endocrine Disruptors and Nuclear Receptors Gene Expression in Infertile and Fertile Men from Italian Areas with Different Environmental Features

    PubMed Central

    La Rocca, Cinzia; Tait, Sabrina; Guerranti, Cristiana; Busani, Luca; Ciardo, Francesca; Bergamasco, Bruno; Perra, Guido; Mancini, Francesca Romana; Marci, Roberto; Bordi, Giulia; Caserta, Donatella; Focardi, Silvano; Moscarini, Massimo; Mantovani, Alberto

    2015-01-01

    Internal levels of selected endocrine disruptors (EDs) (i.e., perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), di-2-ethylhexyl-phthalate (DEHP), mono-(2-ethylhexyl)-phthalate (MEHP), and bisphenol A (BPA)) were analyzed in blood/serum of infertile and fertile men from metropolitan, urban and rural Italian areas. PFOS and PFOA levels were also evaluated in seminal plasma. In peripheral blood mononuclear cells (PBMCs) of same subjects, gene expression levels of a panel of nuclear receptors (NRs), namely estrogen receptor α (ERα) estrogen receptor β (ERβ), androgen receptor (AR), aryl hydrocarbon receptor (AhR), peroxisome proliferator-activated receptor γ (PPARγ) and pregnane X receptor (PXR) were also assessed. Infertile men from the metropolitan area had significantly higher levels of BPA and gene expression of all NRs, except PPARγ, compared to subjects from other areas. Subjects from urban areas had significantly higher levels of MEHP, whereas subjects from rural area had higher levels of PFOA in both blood and seminal plasma. Interestingly, ERα, ERβ, AR, PXR and AhR expression is directly correlated with BPA and inversely correlated with PFOA serum levels. Our study indicates the relevance of the living environment when investigating the exposure to specific EDs. Moreover, the NRs panel in PBMCs demonstrated to be a potential biomarker of effect to assess the EDs impact on reproductive health. PMID:26445054

  2. Exposure to Endocrine Disruptors and Nuclear Receptors Gene Expression in Infertile and Fertile Men from Italian Areas with Different Environmental Features.

    PubMed

    La Rocca, Cinzia; Tait, Sabrina; Guerranti, Cristiana; Busani, Luca; Ciardo, Francesca; Bergamasco, Bruno; Perra, Guido; Mancini, Francesca Romana; Marci, Roberto; Bordi, Giulia; Caserta, Donatella; Focardi, Silvano; Moscarini, Massimo; Mantovani, Alberto

    2015-10-01

    Internal levels of selected endocrine disruptors (EDs) (i.e., perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), di-2-ethylhexyl-phthalate (DEHP), mono-(2-ethylhexyl)-phthalate (MEHP), and bisphenol A (BPA)) were analyzed in blood/serum of infertile and fertile men from metropolitan, urban and rural Italian areas. PFOS and PFOA levels were also evaluated in seminal plasma. In peripheral blood mononuclear cells (PBMCs) of same subjects, gene expression levels of a panel of nuclear receptors (NRs), namely estrogen receptor α (ERα) estrogen receptor β (ERβ), androgen receptor (AR), aryl hydrocarbon receptor (AhR), peroxisome proliferator-activated receptor γ (PPARγ) and pregnane X receptor (PXR) were also assessed. Infertile men from the metropolitan area had significantly higher levels of BPA and gene expression of all NRs, except PPARγ, compared to subjects from other areas. Subjects from urban areas had significantly higher levels of MEHP, whereas subjects from rural area had higher levels of PFOA in both blood and seminal plasma. Interestingly, ERα, ERβ, AR, PXR and AhR expression is directly correlated with BPA and inversely correlated with PFOA serum levels. Our study indicates the relevance of the living environment when investigating the exposure to specific EDs. Moreover, the NRs panel in PBMCs demonstrated to be a potential biomarker of effect to assess the EDs impact on reproductive health. PMID:26445054

  3. The Q-rich/PST domain of the AHR regulates both ligand-induced nuclear transport and nucleocytoplasmic shuttling

    PubMed Central

    Tkachenko, Anna; Henkler, Frank; Brinkmann, Joep; Sowada, Juliane; Genkinger, Doris; Kern, Christian; Tralau, Tewes; Luch, Andreas

    2016-01-01

    The aryl hydrocarbon receptor (AHR) shuttles continuously between cytoplasm and nucleus, unless ligand-binding triggers association with the AHR nuclear translocator (ARNT) and subsequent binding to cognate DNA motifs. We have now identified Val 647 as mandatory residue for export from the nucleus and AHR-function. This residue prevents inactivation of the receptor as a consequence of nuclear sequestration via constitutive import. Concomitantly mutants lacking this residue are exclusively localised in the nucleus. Although ligands accelerate nuclear import transiently, stable nuclear transition depends on a motif adjacent to Val 647 that comprises residues 650–661. Together, this defined region within the Q-rich domain regulates intracellular trafficking of the AHR in context of both nucleocytoplasmic shuttling and receptor activation. Nuclear export therefore depends on the previously characterised N-terminal NES and the newly identified motif that includes V647. Nucleocytoplasmic distribution of full-length human AHR is further affected by a section of the PST domain that shows sequence similarities with nuclear export signals. In concert, these motifs maintain a predominant cytoplasmic compartmentalisation, receptive for ligand binding. PMID:27535013

  4. The Q-rich/PST domain of the AHR regulates both ligand-induced nuclear transport and nucleocytoplasmic shuttling.

    PubMed

    Tkachenko, Anna; Henkler, Frank; Brinkmann, Joep; Sowada, Juliane; Genkinger, Doris; Kern, Christian; Tralau, Tewes; Luch, Andreas

    2016-01-01

    The aryl hydrocarbon receptor (AHR) shuttles continuously between cytoplasm and nucleus, unless ligand-binding triggers association with the AHR nuclear translocator (ARNT) and subsequent binding to cognate DNA motifs. We have now identified Val 647 as mandatory residue for export from the nucleus and AHR-function. This residue prevents inactivation of the receptor as a consequence of nuclear sequestration via constitutive import. Concomitantly mutants lacking this residue are exclusively localised in the nucleus. Although ligands accelerate nuclear import transiently, stable nuclear transition depends on a motif adjacent to Val 647 that comprises residues 650-661. Together, this defined region within the Q-rich domain regulates intracellular trafficking of the AHR in context of both nucleocytoplasmic shuttling and receptor activation. Nuclear export therefore depends on the previously characterised N-terminal NES and the newly identified motif that includes V647. Nucleocytoplasmic distribution of full-length human AHR is further affected by a section of the PST domain that shows sequence similarities with nuclear export signals. In concert, these motifs maintain a predominant cytoplasmic compartmentalisation, receptive for ligand binding. PMID:27535013

  5. Contribution of Priority PAHs and POPs to Ah Receptor-Mediated Activities in Sediment Samples from the River Elbe Estuary, Germany

    PubMed Central

    Otte, Jens C.; Keiter, Steffen; Faßbender, Christopher; Higley, Eric B.; Rocha, Paula Suares; Brinkmann, Markus; Wahrendorf, Dierk-Steffen; Manz, Werner; Wetzel, Markus A.; Braunbeck, Thomas; Giesy, John P.; Hecker, Markus; Hollert, Henner

    2013-01-01

    The estuary of the River Elbe between Hamburg and the North Sea (Germany) is a sink for contaminated sediment and suspended particulate matter (SPM). One major concern is the effect of human activities on the hydrodynamics, particularly the intensive dredging activities in this area that may result in remobilization of sediment-bound pollutants. The aim of this study was to identify pollutants contributing to the toxicological risk associated with re-suspension of sediments in the Elbe Estuary by use of an effect-directed analysis that combines chemical and biological analyses in with specific fractionation techniques. Sediments were collected from sites along the Elbe Estuary and a site from a small harbor basin of the Elbe Estuary that is known to be polluted. The sixteen priority EPA-PAHs were quantified in organic extracts of sediments. In addition, dioxin equivalents of sediments were investigated by use of the 7-ethoxyresorufin O-deethylase assay with RTL-W1 cells and the Ah receptor-mediated luciferase transactivation assay with H4IIE-luc cells. Quantification of the 16 priority PAHs revealed that sediments were moderately contaminated at all of the sites in the Elbe River Estuary (<0.02–0.906 µg/g dw). Sediments contained relatively small concentrations of dioxin equivalents (Bio-TEQ) with concentrations ranging from 15.5 to 322 pg/g dw, which were significantly correlated with dioxin equivalents calculated based on toxicity reference values and concentrations of PAH. The concentration of Bio-TEQ at the reference site exceeded 200,000 pg/g dw. In a potency balance the 16 PAHs explained between 47 and 118% of the Bio-TEQ in the luciferase assay, which can be explained by the constant input of PAHs bound to SPM from the upper course of the Elbe River into its estuary. Successful identification of a significant portion of dioxin-like activity to priority PAHs in complex environmental samples such as sediments has rarely been reported. PMID:24146763

  6. Aryl hydrocarbon receptor-dependent up-regulation of the heterodimeric amino acid transporter LAT1 (SLC7A5)/CD98hc (SLC3A2) by diesel exhaust particle extract in human bronchial epithelial cells.

    PubMed

    Le Vee, Marc; Jouan, Elodie; Lecureur, Valérie; Fardel, Olivier

    2016-01-01

    The heterodimeric L-type amino acid transporter (LAT) 1/CD98hc is overexpressed in lung cancers with a poor prognosis factor. Factors that contribute to LAT1/CD98hc overexpression in lung cells remain however to be determined, but the implication of atmospheric pollution can be suspected. The present study was therefore designed to analyze the effects of diesel exhaust particle (DEP) extract (DEPe) on LAT1/CD98hc expression in bronchial epithelial BEAS-2B cells. Exposure to DEPe up-regulated LAT1 and CD98hc mRNA levels in a concentration-dependent manner, with DEPe EC50 values (around 0.2 μg/mL) relevant to environmental situations. DEPe concomitantly induced LAT1/CD98hc protein expression and LAT1-mediated leucine accumulation in BEAS-2B cells. Inhibition of the aryl hydrocarbon receptor (AhR) pathway through the use of a chemical AhR antagonist or the siRNA-mediated silencing of AhR expression was next found to prevent DEPe-mediated induction of LAT1/CD98hc, indicating that this regulation depends on AhR, known to be activated by major chemical DEP components like polycyclic aromatic hydrocarbons. DEPe exposure was finally shown to induce mRNA expression and activity of matrix metalloproteinase (MMP)-2 in BEAS-2B cells, in a CD98hc/focal adhesion kinase (FAK)/extracellular regulated kinase (ERK) manner, thus suggesting that DEPe-mediated induction of CD98hc triggers activation of the integrin/FAK/ERK signaling pathway known to be involved in MMP-2 regulation. Taken together, these data demonstrate that exposure to DEPe induces functional overexpression of the amino acid transporter LAT1/CD98hc in lung cells. Such a regulation may participate to pulmonary carcinogenic effects of DEPs, owing to the well-documented contribution of LAT1 and CD98hc to cancer development.

  7. Reciprocal inhibiting interactive mechanism between the estrogen receptor and aryl hydrocarbon receptor signaling pathways in goldfish (Carassius auratus) exposed to 17β-estradiol and benzo[a]pyrene.

    PubMed

    Yan, Zhenhua; Lu, Guanghua; He, Junjie

    2012-06-01

    In the aquatic environment, both the estrogen receptor (ER) and aryl hydrocarbon receptor (AhR) responses are established biomarkers for assessing exposure to pollutants. These receptor responses can also be affected by the presence of other classes of pollutants and may result in misinterpretation of existing pollution. In this study, we investigated the interaction between ER-vitellogenin (VTG) and AhR-cytochrome P450 1A (CYP1A) signaling pathways in goldfish (Carassius auratus) after 10 days exposure to pollutants. 17β-Estradiol (E(2)) and benzo[a]pyrene (BaP) were selected as the ER and AhR agonists, respectively. The messenger RNA (mRNA) expression of ER-VTG and AhR-CYP1A in liver was determined using quantitative real-time polymerase chain reaction (QRT-PCR). VTG, endogenous E(2) and 7-ethoxyresorufin-O-deethylase (EROD) were also studied. Exposure to E(2) and BaP alone significantly induced the gene expression of ERα-VTG and AhR2-CYP1A, respectively. Moreover, the obvious expression of related proteins was also observed. However, these inductions were significantly reduced after combined exposure to E(2) and lower concentrations of BaP (20 and 50 μg/L), indicative of a reciprocal inhibiting ER-AhR interaction. However, high concentrations (100 μg/L) of BaP did not affect the E(2)-induced gene expression. Changes in VTG protein were in accordance with the expression of VTG mRNA, and more VTG protein was observed in liver than in serum. The induced endogenous E(2) levels were suppressed by the presence of BaP. While the gene expression of CYP1A showed a concentration-dependent increase, EROD induction exhibited a bell-shaped concentration-response curve. Taken together, these results demonstrate a reciprocal inhibiting mode of ER-AhR interactions and may lead to a possible underestimation of actual exposure.

  8. Expression of profibrotic growth factors and their receptors by mouse lung macrophages and fibroblasts under conditions of acute viral inflammation in influenza A/H5N1 virus.

    PubMed

    Anikina, A G; Shkurupii, V A; Potapova, O V; Kovner, A V; Shestopalov, A M

    2014-04-01

    Morphological signs of early interstitial fibrosis, developing under conditions of acute viral inflammation (postinfection days 1-14), were observed in C57Bl/6 mice infected with influenza A/H5N1 A/goose/Krasnoozerskoye/627/05 virus. The development of fibrosis was confirmed by an increase in the number of lung cells expressing TNF-α. These changes were recorded in the presence of a many-fold increase in the counts of macrophages and fibroblasts expressing FGF, EGF, and their receptors.

  9. Endocrine disrupting chemicals targeting estrogen receptor signaling: Identification and mechanisms of action

    PubMed Central

    Shanle, Erin K.; Xu, Wei

    2011-01-01

    Many endocrine disrupting chemicals (EDCs) adversely impact estrogen signaling by interacting with two estrogen receptors (ERs): ERα and ERβ. Though the receptors have similar ligand binding and DNA binding domains, ERα and ERβ have some unique properties in terms of ligand selectivity and target gene regulation. EDCs that target ER signaling can modify genomic and non-genomic ER activity through direct interactions with ERs, indirectly through transcription factors like the aryl hydrocarbon receptor (AhR), or through modulation of metabolic enzymes that are critical for normal estrogen synthesis and metabolism. Many EDCs act through multiple mechanisms as exemplified by chemicals that bind both AhR and ER, such as 3-methylcholanthrene. Other EDCs that target ER signaling include phytoestrogens, bisphenolics, and organochlorine pesticides and many alter normal ER signaling through multiple mechanisms. EDCs can also display tissue-selective ER agonist and antagonist activities similar to selective estrogen receptor modulators (SERMs) designed for pharmaceutical use. Thus, biological effects of EDCs need to be carefully interpreted because EDCs can act through complex tissue-selective modulation of ERs and other signaling pathways in vivo. Current requirements by the U.S. Environmental Protection Agency require some in vitro and cell-based assays to identify EDCs that target ER signaling through direct and metabolic mechanisms. Additional assays may be useful screens for identifying EDCs that act through alternative mechanisms prior to further in vivo study. PMID:21053929

  10. Metabolic control of type 1 regulatory (Tr1) cell differentiation by AHR and HIF1-α

    PubMed Central

    Mascanfroni, Ivan D.; Takenaka, Maisa C.; Yeste, Ada; Patel, Bonny; Wu, Yan; Kenison, Jessica E.; Siddiqui, Shafiuddin; Basso, Alexandre S.; Otterbein, Leo E.; Pardoll, Drew M.; Pan, Fan; Priel, Avner; Clish, Clary B.; Robson, Simon C.; Quintana, Francisco J.

    2015-01-01

    Our understanding of the pathways that regulate lymphocyte metabolism, as well as the effects of metabolism and its products on the immune response, is still limited. We report that a metabolic program controlled by the transcription factors hypoxia inducible factor-1α (HIF1-α) and aryl hydrocarbon receptor (AHR) supports the differentiation of type 1 regulatory (Tr1) cells. HIF1-α controls the early metabolic reprograming of Tr1 cells. At later time points, AHR promotes HIF1-α degradation and takes control of Tr1 cell metabolism. Extracellular adenosine triphosphate (eATP) and hypoxia, linked to inflammation, trigger AHR inactivation by HIF1-α and inhibit Tr1 cell differentiation. Conversely, CD39 promotes Tr1 cell differentiation by depleting eATP. CD39 also contributes to Tr1 suppressive activity by generating adenosine in cooperation with CD73 expressed by responder T cells and antigen presenting cells. These results suggest that HIF1-α and AHR integrate immunological, metabolic and environmental signals to regulate the immune response. PMID:26005855

  11. Intersection of AHR and Wnt Signaling in Development, Health, and Disease

    PubMed Central

    Schneider, Andrew J.; Branam, Amanda M.; Peterson, Richard E.

    2014-01-01

    The AHR (aryl hydrocarbon receptor) and Wnt (wingless-related MMTV integration site) signaling pathways have been conserved throughout evolution. Appropriately regulated signaling through each pathway is necessary for normal development and health, while dysregulation can lead to developmental defects and disease. Though both pathways have been vigorously studied, there is relatively little research exploring the possibility of crosstalk between these pathways. In this review, we provide a brief background on (1) the roles of both AHR and Wnt signaling in development and disease, and (2) the molecular mechanisms that characterize activation of each pathway. We also discuss the need for careful and complete experimental evaluation of each pathway and describe existing research that explores the intersection of AHR and Wnt signaling. Lastly, to illustrate in detail the intersection of AHR and Wnt signaling, we summarize our recent findings which show that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced disruption of Wnt signaling impairs fetal prostate development. PMID:25286307

  12. The aryl hydrocarbon receptor-mediated and genotoxic effects of fractionated extract of standard reference diesel exhaust particle material in pulmonary, liver and prostate cells.

    PubMed

    Pálková, Lenka; Vondráček, Jan; Trilecová, Lenka; Ciganek, Miroslav; Pěnčíková, Kateřina; Neča, Jiří; Milcová, Alena; Topinka, Jan; Machala, Miroslav

    2015-04-01

    Diesel exhaust particles (DEP) and the associated complex mixtures of organic pollutants, such as polycyclic aromatic hydrocarbons (PAHs), or their derivatives, have been suggested to exert deleterious effects on human health. We used a set of defined cellular models representing liver, lung and prostate tissues, in order to compare non-genotoxic and genotoxic effects of crude and fractionated extract of a standard reference DEP material - SRM 1650b. We focused on the aryl hydrocarbon receptor (AhR)-mediated activity, modulation of cell proliferation, formation of DNA adducts, oxidative DNA damage, and induction of DNA damage responses, including evaluation of apoptosis, and phosphorylation of p53 tumor suppressor and checkpoint kinases (Chk). Both PAHs and the polar aromatic compounds contributed to the AhR-mediated activity of DEP-associated organic pollutants. The principal identified AhR agonists included benzo[k]fluoranthene, indeno[1,2,3-c,d]pyrene, chrysene and several non-priority PAHs, including benzochrysenes and methylated PAHs. In contrast to PAHs, polar compounds contributed more significantly to overall formation of DNA adducts associated with phosphorylation of p53, Chk1 or Chk2, and partly with apoptosis. Therefore, more attention should be paid to identification of DEP-associated polar organic compounds, contributing to the AhR activation and cytotoxic/genotoxic effects of complex airborne mixtures of organic contaminants produced by diesel engines.

  13. DNA Methylation of the Aryl Hydrocarbon Receptor Repressor Associations with Cigarette Smoking and Subclinical Atherosclerosis

    PubMed Central

    Reynolds, Lindsay M.; Wan, Ma; Ding, Jingzhong; Taylor, Jackson R.; Mstat, Kurt Lohman; Su, Dan; Bennett, Brian D.; Porter, Devin K.; Gimple, Ryan; Pittman, Gary S.; Wang, Xuting; Howard, Timothy D.; Siscovick, David; Psaty, Bruce M.; Shea, Steven; Burke, Gregory L.; Jacobs, David R.; Rich, Stephen S.; Hixson, James E.; Stein, James H.; Stunnenberg, Hendrik; Barr, R. Graham; Kaufman, Joel D.; Post, Wendy S.; Hoeschele, Ina; Herrington, David M.; Bell, Douglas A.; Liu, Yongmei

    2015-01-01

    Background Tobacco smoke contains numerous agonists of the aryl-hydrocarbon receptor (AhR) pathway, and activation of the AhR pathway was shown to promote atherosclerosis in mice. Intriguingly, cigarette smoking is most strongly and robustly associated with DNA modifications to an AhR pathway gene, the aryl-hydrocarbon receptor repressor (AHRR). We hypothesized that altered AHRR methylation in monocytes, a cell type sensitive to cigarette smoking and involved in atherogenesis, may be a part of the biological link between cigarette smoking and atherosclerosis. Methods and Results DNA methylation profiles of AHRR in monocytes (542 CpG sites ± 150kb of AHRR, using Illumina 450K array) were integrated with smoking habits and ultrasound-measured carotid plaque scores from 1,256 participants of the Multi-Ethnic Study of Atherosclerosis (MESA). Methylation of cg05575921 significantly associated (p = 6.1×10−134) with smoking status (current vs. never). Novel associations between cg05575921 methylation and carotid plaque scores (p = 3.1×10−10) were identified, which remained significant in current and former smokers even after adjusting for self-reported smoking habits, urinary cotinine, and well-known CVD risk factors. This association replicated in an independent cohort using hepatic DNA (n = 141). Functionally, cg05575921 was located in a predicted gene expression regulatory element (enhancer), and had methylation correlated with AHRR mRNA profiles (p = 1.4×10−17) obtained from RNA sequencing conducted on a subset (n = 373) of the samples. Conclusions These findings suggest AHRR methylation may be functionally related to AHRR expression in monocytes, and represents a potential biomarker of subclinical atherosclerosis in smokers. PMID:26307030

  14. AHR2 morpholino knockdown reduces the toxicity of total particulate matter to zebrafish embryos.

    PubMed

    Massarsky, Andrey; Bone, Audrey J; Dong, Wu; Hinton, David E; Prasad, G L; Di Giulio, Richard T

    2016-10-15

    The zebrafish embryo has been proposed as a 'bridge model' to study the effects of cigarette smoke on early development. Previous studies showed that exposure to total particulate matter (TPM) led to adverse effects in developing zebrafish, and suggested that the antioxidant and aryl hydrocarbon receptor (AHR) pathways play important roles. This study investigated the roles of these two pathways in mediating TPM toxicity. The study consisted of four experiments. In experiment I, zebrafish embryos were exposed from 6h post fertilization (hpf) until 96hpf to TPM0.5 and TPM1.0 (corresponding to 0.5 and 1.0μg/mL equi-nicotine units) in the presence or absence of an antioxidant (N-acetyl cysteine/NAC) or a pro-oxidant (buthionine sulfoximine/BSO). In experiment II, TPM exposures were performed in embryos that were microinjected with nuclear factor erythroid 2-related factor 2 (Nrf2), AHR2, cytochrome P450 1A (CYP1A), or CYP1B1 morpholinos, and deformities were assessed. In experiment III, embryos were exposed to TPM, and embryos/larvae were collected at 24, 48, 72, and 96hpf to assess several genes associated with the antioxidant and AHR pathways. Lastly, experiment IV assessed the activity and protein levels of CYP1A and CYP1B1 after exposure to TPM. We demonstrate that the incidence of TPM-induced deformities was generally not affected by NAC/BSO treatments or Nrf2 knockdown. In contrast, AHR2 knockdown reduced, while CYP1A or CYP1B1 knockdowns elevated the incidence of some deformities. Moreover, as shown by gene expression the AHR pathway, but not the antioxidant pathway, was induced in response to TPM exposure, providing further evidence for its importance in mediating TPM toxicity.

  15. AHR2 morpholino knockdown reduces the toxicity of total particulate matter to zebrafish embryos.

    PubMed

    Massarsky, Andrey; Bone, Audrey J; Dong, Wu; Hinton, David E; Prasad, G L; Di Giulio, Richard T

    2016-10-15

    The zebrafish embryo has been proposed as a 'bridge model' to study the effects of cigarette smoke on early development. Previous studies showed that exposure to total particulate matter (TPM) led to adverse effects in developing zebrafish, and suggested that the antioxidant and aryl hydrocarbon receptor (AHR) pathways play important roles. This study investigated the roles of these two pathways in mediating TPM toxicity. The study consisted of four experiments. In experiment I, zebrafish embryos were exposed from 6h post fertilization (hpf) until 96hpf to TPM0.5 and TPM1.0 (corresponding to 0.5 and 1.0μg/mL equi-nicotine units) in the presence or absence of an antioxidant (N-acetyl cysteine/NAC) or a pro-oxidant (buthionine sulfoximine/BSO). In experiment II, TPM exposures were performed in embryos that were microinjected with nuclear factor erythroid 2-related factor 2 (Nrf2), AHR2, cytochrome P450 1A (CYP1A), or CYP1B1 morpholinos, and deformities were assessed. In experiment III, embryos were exposed to TPM, and embryos/larvae were collected at 24, 48, 72, and 96hpf to assess several genes associated with the antioxidant and AHR pathways. Lastly, experiment IV assessed the activity and protein levels of CYP1A and CYP1B1 after exposure to TPM. We demonstrate that the incidence of TPM-induced deformities was generally not affected by NAC/BSO treatments or Nrf2 knockdown. In contrast, AHR2 knockdown reduced, while CYP1A or CYP1B1 knockdowns elevated the incidence of some deformities. Moreover, as shown by gene expression the AHR pathway, but not the antioxidant pathway, was induced in response to TPM exposure, providing further evidence for its importance in mediating TPM toxicity. PMID:27576004

  16. Synergistic induction of AHR regulated genes in developmental toxicity from co-exposure to two model PAHs in zebrafish

    PubMed Central

    Timme-Laragy, Alicia. R.; Cockman, Crystal. J.; Matson, Cole. W.; Di Giulio, Richard. T.

    2007-01-01

    Polycyclic aromatic hydrocarbons (PAHs) are pollutants created by the incomplete combustion of carbon, and are increasing in the environment largely due to the burning of fossil fuels. PAHs occur as complex mixtures, and some combinations have been shown to cause synergistic developmental toxicity in fish embryos, characterized by pericardial edema and craniofacial malformations. Previous studies have indicated that in the zebrafish model, this toxicity is mediated by the aryl hydrocarbon receptor 2 (AHR2), and enhanced by inhibition of CYP1A activity. In this study, we further examined this interaction of the model PAH and AHR agonist β-naphthoflavone (BNF) with and without the AHR partial agonist/antagonist and CYP1A inhibitor α-naphthoflavone (ANF) to determine 1) whether ANF was acting as an AHR antagonist, 2) what alterations BNF and ANF both alone and in combination had on mRNA expression of the AHR regulated genes cytochrome P450 (cyp) 1a, 1b1, and 1c1, and the AHR repressor (ahrr2) prior to vs. during deformity onset, and 3) compare CYP1A enzyme activity with mRNA induction. Zebrafish embryos were exposed from 24–48 or 24–96 hpf to BNF, 1–100 μg/L, ANF, 1–150 μg/L, a BNF+ANF co-exposure (1 μg/L + 100 μg/L), or a DMSO solvent control. RNA was extracted and examined by quantitative real time PCR. Both BNF and ANF each individually resulted in a dose dependent increase CYP1A, CYP1B1, CYP1C1, and AHRR2 mRNA, confirming their activities as AHR agonists. In the BNF+ANF co-exposures prior to deformity onset, expression of these genes was synergistic, and expression levels of the AHR regulated genes resembled the higher doses of BNF alone. Gene induction during deformities was also significantly increased in the co-exposure, but to a lesser magnitude than prior to deformity onset. EROD measurements of CYP1A activity showed ANF inhibited activity induction by BNF in the co-exposure group; this finding is not predicted by mRNA expression, which is

  17. γ-Tocotrienol upregulates aryl hydrocarbon receptor expression and enhances the anticancer effect of baicalein.

    PubMed

    Yamashita, Shuya; Baba, Kiwako; Makio, Akiko; Kumazoe, Motofumi; Huang, Yuhui; Lin, I-Chian; Bae, Jaehoon; Murata, Motoki; Yamada, Shuhei; Tachibana, Hirofumi

    2016-05-13

    Previous studies have identified biomolecules that mediate the physiological actions of food factors, such as amino acids, vitamins, fatty acids, minerals, plant polyphenols, and lactobacilli, suggesting that our bodies are equipped with an innate system that senses which food factors are required to maintain our health. However, the effects of environmental factors on food factor sensing (FFS) remains largely unknown. Tocotorienols (T3s), which belongs to the vitamin E family, possess several physiological functions, including cholesterol lowering and neuroprotective effects. Here, we investigated the effects of naturally abundant γ-T3 on FFS-related gene expressions in melanoma using a DNA chip. Our results showed that γ-T3 increased the expression level of aryl hydrocarbon receptor (AhR), a sensing molecule to plant polyphenol baicalein. The co-treatment with γ-T3 and baicalein enhanced the anti-proliferative activity of baicalein, accompanied by the downstream events of AhR-activation induced by baicalein. These data suggest that γ-T3 upregulates AhR expression and enhances its sensitivity to baicalein. PMID:27055589

  18. Receptor mechanisms and dose-response models for the effects of dioxins.

    PubMed Central

    Lucier, G W; Portier, C J; Gallo, M A

    1993-01-01

    There is increasing evidence that receptor-mediated events impact one or more stages responsible for tumor development in experimental animals and humans. Although many chemicals and endogenous hormones require receptor interactions as a necessary event in their carcinogenic activity, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and its structural analogs are the most visible examples of receptor-mediated carcinogens. TCDD, or dioxin as it is frequently called, interacts with the Ah receptor (AhR), which functions in a manner analogous to receptors for steroids. TCDD produces a wide spectrum of biochemical and toxic responses in in vitro and in vivo systems, and the Ah receptor is generally considered necessary for most if not all of these responses. Risk assessments for dioxin made by the United States and other countries throughout the world have been based on its carcinogenecity in experimental animals. Recently, epidemiology studies have indicated that TCDD is a human carcinogen at high doses. Because TCDD appears to be acting like a potent and persistent hormone agonist, it appears reasonable to incorporate mechanistic information on receptor-mediated events in risk assessments for TCDD. This information may be obtained from steroid receptor action and from molecular data on the Ah receptor. In this paper, we evaluate the scientific foundation on which mechanistic models for estimating dioxin's risks should be based. These models need to recognize the mechanisms possible for the diversity of biological responses that are initiated by a single receptor interacting with a single ligand. The U.S. EPA is currently reevaluating dioxin's risks by examining the possibility of developing biologically based models.(ABSTRACT TRUNCATED AT 250 WORDS) Images p36-a Figure 1. p42-a p42-b p42-c PMID:8390353

  19. Teratogenic impact of dioxin-activated AHR in laboratory animals

    EPA Science Inventory

    AHR and ARNT are expressed in mouse and human palatal shelves and in the urinary tract of the mouse fetus. AHR expression, translocation to the nucleus, binding to DRE, and activation are required for mediation of TCDD-induction of CP and HN. Although the human palate requires a ...

  20. Genome Editing of the CYP1A1 Locus in iPSCs as a Platform to Map AHR Expression throughout Human Development

    PubMed Central

    Smith, Brenden W.; Stanford, Elizabeth A.; Sherr, David H.; Murphy, George J.

    2016-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand activated transcription factor that increases the expression of detoxifying enzymes upon ligand stimulation. Recent studies now suggest that novel endogenous roles of the AHR exist throughout development. In an effort to create an optimized model system for the study of AHR signaling in several cellular lineages, we have employed a CRISPR/CAS9 genome editing strategy in induced pluripotent stem cells (iPSCs) to incorporate a reporter cassette at the transcription start site of one of its canonical targets, cytochrome P450 1A1 (CYP1A1). This cell line faithfully reports on CYP1A1 expression, with luciferase levels as its functional readout, when treated with an endogenous AHR ligand (FICZ) at escalating doses. iPSC-derived fibroblast-like cells respond to acute exposure to environmental and endogenous AHR ligands, and iPSC-derived hepatocytes increase CYP1A1 in a similar manner to primary hepatocytes. This cell line is an important innovation that can be used to map AHR activity in discrete cellular subsets throughout developmental ontogeny. As further endogenous ligands are proposed, this line can be used to screen for safety and efficacy and can report on the ability of small molecules to regulate critical cellular processes by modulating the activity of the AHR. PMID:27148368

  1. Aryl hydrocarbon receptor antagonism and its role in rheumatoid arthritis

    PubMed Central

    Nguyen, Nam Trung; Nakahama, Taisuke; Nguyen, Chi Hung; Tran, Trang Thu; Le, Van Son; Chu, Hoang Ha; Kishimoto, Tadamitsu

    2015-01-01

    Although rheumatoid arthritis (RA) is the most common autoimmune disease, affecting approximately 1% of the population worldwide, its pathogenic mechanisms are poorly understood. Tobacco smoke, an environmental risk factor for RA, contains several ligands of aryl hydrocarbon receptor (Ahr), also known as dioxin receptor. Ahr plays critical roles in the immune system. We previously demonstrated that Ahr in helper T-cells contributes to development of collagen-induced arthritis, a mouse model of RA. Other studies have shown that cigarette smoke condensate and pure Ahr ligands exacerbate RA by altering bone metabolism and inducing proinflammatory responses in fibroblast-like synoviocytes. Consistent with these findings, several Ahr antagonists such as α-naphthoflavone, resveratrol, and GNF351 reverse the effect of Ahr ligands in RA pathogenesis. In this review, we summarize the current knowledge of Ahr function in the immune system and the potential clinical benefits of Ahr antagonism in treating RA. PMID:27186143

  2. A novel prostate cancer therapeutic strategy using icaritin-activated arylhydrocarbon-receptor to co-target androgen receptor and its splice variants

    PubMed Central

    Sun, Feng; Indran, Inthrani R.; Zhang, Zhi Wei; Tan, M.H.Eileen; Li, Yu; Lim, Z.L.Ryan; Hua, Rui; Yang, Chong; Soon, Fen-Fen; Li, Jun; Xu, H.Eric; Cheung, Edwin; Yong, Eu-Leong

    2015-01-01

    Persistent androgen receptor (AR) signaling is the key driving force behind progression and development of castration-resistant prostate cancer (CRPC). In many patients, AR COOH-terminal truncated splice variants (ARvs) play a critical role in contributing to the resistance against androgen depletion therapy. Unfortunately, clinically used antiandrogens like bicalutamide (BIC) and enzalutamide (MDV), which target the ligand binding domain, have failed to suppress these AR variants. Here, we report for the first time that a natural prenylflavonoid, icaritin (ICT), can co-target both persistent AR and ARvs. ICT was found to inhibit transcription of key AR-regulated genes, such as KLK3 [prostate-specific antigen (PSA)] and ARvs-regulated genes, such as UBE2C and induce apoptosis in AR-positive prostate cancer (PC) cells. Mechanistically, ICT promoted the degradation of both AR and ARvs by binding to arylhydrocarbon-receptor (AhR) to mediate ubiquitin-proteasomal degradation. Therefore, ICT impaired AR transactivation in PC cells. Knockdown of AhR gene restored AR stability and partially prevented ICT-induced growth suppression. In clinically relevant murine models orthotopically implanted with androgen-sensitive and CRPC cells, ICT was able to target AR and ARvs, to inhibit AR signaling and tumor growth with no apparent toxicity. Our results provide a mechanistic framework for the development of ICT, as a novel lead compound for AR-positive PC therapeutics, especially for those bearing AR splice variants. PMID:25908644

  3. An in vitro rainbow trout cell bioassay for aryl hydrocarbon receptor-mediated toxins

    SciTech Connect

    Richter, C.A.; Tieber, V.L.; Giesy, J.P.; Denison, M.S.

    1997-03-01

    Halogenated aromatic hydrocarbons (HAHs) and other chemicals that act as aryl hydrocarbon (Ah) receptor (AhR) agonists cause a variety of toxicity effects. In sac fry of many fish species, these effects include blue-sac disease and mortality. Because HAHs occur in complex mixtures, their toxicity in the environment is difficult to predict. A bioassay useful in predicting AhR-mediated toxicity to fish was developed using the RTH-149 rainbow trout hepatoma cell line. Stable transfection of this cell line with the pGudLuc 1.1 plasmid, which contains a firefly luciferase reporter gene under the transcriptional regulation of dioxin responsive enhancers, has produced a recombinant cell line designated Remodulated Lightning Trout (RLT 2.0). The RLT 2.0 bioassay method detection limit for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is 4 pM. The responses of the RLT 2.0 bioassay to TCDD and several HAH congeners closely matched the responses observed in vivo in fish. The RLT 2.0 bioassay can provide an integrative measure of the total AhR-mediated toxic activity of complex mixtures to fish. The assay will be useful in screening environmental extracts, guiding chemical analysis, and interpreting the AhR-mediated mechanism of toxicity.

  4. Nuclear pyruvate kinase M2 complex serves as a transcriptional coactivator of arylhydrocarbon receptor.

    PubMed

    Matsuda, Shun; Adachi, Jun; Ihara, Masaru; Tanuma, Nobuhiro; Shima, Hiroshi; Kakizuka, Akira; Ikura, Masae; Ikura, Tsuyoshi; Matsuda, Tomonari

    2016-01-29

    Pyruvate kinase M2 (PKM2) and pyruvate dehydrogenase complex (PDC) regulate production of acetyl-CoA, which functions as an acetyl donor in diverse enzymatic reactions, including histone acetylation. However, the mechanism by which the acetyl-CoA required for histone acetylation is ensured in a gene context-dependent manner is not clear. Here we show that PKM2, the E2 subunit of PDC and histone acetyltransferase p300 constitute a complex on chromatin with arylhydrocarbon receptor (AhR), a transcription factor associated with xenobiotic metabolism. All of these factors are recruited to the enhancer of AhR-target genes, in an AhR-dependent manner. PKM2 contributes to enhancement of transcription of cytochrome P450 1A1 (CYP1A1), an AhR-target gene, acetylation at lysine 9 of histone H3 at the CYP1A1 enhancer. Site-directed mutagenesis of PKM2 indicates that this enhancement of histone acetylation requires the pyruvate kinase activity of the enzyme. Furthermore, we reveal that PDC activity is present in nuclei. Based on these findings, we propose a local acetyl-CoA production system in which PKM2 and PDC locally supply acetyl-CoA to p300 from abundant PEP for histone acetylation at the gene enhancer, and our data suggest that PKM2 sensitizes AhR-mediated detoxification in actively proliferating cells such as cancer and fetal cells.

  5. Nuclear pyruvate kinase M2 complex serves as a transcriptional coactivator of arylhydrocarbon receptor

    PubMed Central

    Matsuda, Shun; Adachi, Jun; Ihara, Masaru; Tanuma, Nobuhiro; Shima, Hiroshi; Kakizuka, Akira; Ikura, Masae; Ikura, Tsuyoshi; Matsuda, Tomonari

    2016-01-01

    Pyruvate kinase M2 (PKM2) and pyruvate dehydrogenase complex (PDC) regulate production of acetyl-CoA, which functions as an acetyl donor in diverse enzymatic reactions, including histone acetylation. However, the mechanism by which the acetyl-CoA required for histone acetylation is ensured in a gene context-dependent manner is not clear. Here we show that PKM2, the E2 subunit of PDC and histone acetyltransferase p300 constitute a complex on chromatin with arylhydrocarbon receptor (AhR), a transcription factor associated with xenobiotic metabolism. All of these factors are recruited to the enhancer of AhR-target genes, in an AhR-dependent manner. PKM2 contributes to enhancement of transcription of cytochrome P450 1A1 (CYP1A1), an AhR-target gene, acetylation at lysine 9 of histone H3 at the CYP1A1 enhancer. Site-directed mutagenesis of PKM2 indicates that this enhancement of histone acetylation requires the pyruvate kinase activity of the enzyme. Furthermore, we reveal that PDC activity is present in nuclei. Based on these findings, we propose a local acetyl-CoA production system in which PKM2 and PDC locally supply acetyl-CoA to p300 from abundant PEP for histone acetylation at the gene enhancer, and our data suggest that PKM2 sensitizes AhR-mediated detoxification in actively proliferating cells such as cancer and fetal cells. PMID:26405201

  6. The Effects of Chronic Lifelong Activation of the AHR Pathway by Industrial Chemical Pollutants on Female Human Reproduction

    PubMed Central

    Vacca, Margherita; Nardelli, Claudia; Castegna, Alessandra; Arnesano, Fabio; Carella, Nicola; Depalo, Raffaella

    2016-01-01

    Environmental chemicals, such as heavy metals, affect female reproductive function. A biological sensor of the signals of many toxic chemical compounds seems to be the aryl hydrocarbon receptor (AHR). Previous studies demonstrated the environmental of heavy metals in Taranto city (Italy), an area that has been influenced by anthropogenic factors such as industrial activities and waste treatments since 1986. However, the impact of these elements on female fertility in this geographic area has never been analyzed. Thus, in the present study, we evaluated the AHR pathway, sex steroid receptor pattern and apoptotic process in granulosa cells (GCs) retrieved from 30 women, born and living in Taranto, and 30 women who are living in non-contaminated areas (control group), who were undergoing in vitro fertilization (IVF) protocol. In follicular fluids (FFs) of both groups the toxic and essential heavy metals, such as chromiun (Cr), Manganese (Mn), iron (Fe), cobalt (Co), nickel (Ni), copper (Cu), zinc (Zn), cadmium (Cd) and lead (Pb), were also analyzed. Higher levels of Cr, Fe, Zn and Pb were found in the FFs of the women from Taranto as compared to the control group, as were the levels of AHR and AHR-dependent cytochrome P450 1A1 and 1B1; while CYP19A1 expression was decreased. The anti-apoptotic process found in the GCs of women fromTaranto was associated with the highest levels of progesterone receptor membrane component 1 (PGRMC1), a novel progesterone receptor, the expression of which is subjected to AHR activated by its highest affinity ligands (e.g., dioxins) or indirectly by other environmental pollutants, such as heavy metals. In conclusion, decreased production of estradiol and decreased number of retrieved mature oocytes found in women from Taranto could be due to chronic exposure to heavy metals, in particular to Cr and Pb. PMID:27008165

  7. An aryl hydrocarbon receptor ligand acts on dendritic cells and T cells to suppress the Th17 response in allergic rhinitis patients.

    PubMed

    Wei, Ping; Hu, Guo-Hua; Kang, Hou-Yong; Yao, Hong-Bing; Kou, Wei; Liu, Hong; Zhang, Cheng; Hong, Su-Ling

    2014-05-01

    A predominant Th17 population is a marker of allergic rhinitis (AR). The aryl hydrocarbon receptor (AhR) exhibits strong immunomodulation potential via regulation of the differentiation of T lymphocytes and dendritic cells (DCs) after activation by its ligand, such as 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE). The aim of this study was to analyze the effect of AhR on Th17 differentiation by investigating the action of ITE on DCs and CD4(+) T cells from patients with AR. In all, 26 AR patients and 12 healthy controls were included in this study. The expression of interleukin (IL)-1β, IL-6, IL-10, and IL-17 in the culture supernatant and the presence of Th17 cells in CD4(+) T cells and DC-CD4(+) T-cell co-culture system were measured before and after treatment with ITE. We show that ITE significantly induced cell secretion of IL-10 and inhibited IL-1β and IL-6 production in DCs, and promoted IL-10 production and suppressed IL-17 expression in CD4(+) T cells in vitro. It also suppressed the expansion of Th17 cells in vitro. Our work demonstrates that ITE acts on DCs and CD4(+) T cells to inhibit the Th17 response that suppresses AR; the AhR-DC-Th17 axis may be an important pathway in the treatment of AR. ITE, a nontoxic AhR ligand, attenuated the Th17 response; thus, it appears to be a promising therapeutic candidate for suppressing the inflammatory responses associated with AR.

  8. Isomer-nonspecific action of dichlorodiphenyltrichloroethane on aryl hydrocarbon receptor and G-protein-coupled receptor 30 intracellular signaling in apoptotic neuronal cells.

    PubMed

    Kajta, M; Litwa, E; Rzemieniec, J; Wnuk, A; Lason, W; Zelek-Molik, A; Nalepa, I; Grzegorzewska-Hiczwa, M; Tokarski, K; Golas, A; Guzik, E; Grochowalski, A; Szychowski, K A; Wojtowicz, A K

    2014-07-01

    Extended residual persistence of the pesticide dichlorodiphenyltrichloroethane (DDT) raises concerns about its long-term neurotoxic effects. Little is known, however, about DDT toxicity during the early stages of neural development. This study demonstrated that DDT-induced apoptosis of mouse embryonic neuronal cells is a caspase-9-, caspase-3-, and GSK-3β-dependent process, which involves p,p'-DDT-specific impairment of classical ERs. It also provided evidence for DDT-isomer-nonspecific alterations of AhR- and GPR30-mediated intracellular signaling, including changes in the levels of the receptor and receptor-regulated mRNAs, and also changes in the protein levels of the receptors. DDT-induced stimulation of AhR-signaling and reduction of GPR30-signaling were verified using selective ligands and specific siRNAs. Co-localization of the receptors was demonstrated with confocal microscopy, and the presence of functional GPR30 was detected by electrophysiology. This study demonstrates that stimulation of AhR-signaling and impairment of GPR30-signaling play important roles in the propagation of DDT-induced apoptosis during the early stages of neural development.

  9. AH Her Observing Campaign

    NASA Astrophysics Data System (ADS)

    Waagen, Elizabeth O.

    2013-05-01

    Dr. Juan Echevarria (Universidad Nacional Autónoma de México) and colleagues request AAVSO assistance in a campaign on the Z Cam-type cataclysmic variable AH Her being carried out 2013 May 29 - June 18. They will be making photometric and spectroscopic observations of AH Her using the 2.1m and 0.84m telescopes at San Pedro Martir Observatory (SPM). Their goal is to carry out a radial velocity study of the system components using modern detectors; no study of AH Her has been made since the one by Horne, Wade, and Szkody in 1980-1981 (1986MNRAS.219..791H). Photometry and spectroscopy are requested. AH Her, for decades a reasonably "regular" Z Cam system, began exhibiting significantly anomalous behavior in ~2007. Since then it has experienced brief periods of fairly typical behavior interspersed with more anomalous intervals, including some unprecedented behavior. Most recently, it has returned to a more normal pattern of outbursts shape-wise but it is not back to its normal amplitude or frequency. AAVSO data will be essential for correlation in order to determine the precise time(s) of minimum occurring during the campaign. Finder charts with sequences may be created using the AAVSO Variable Star Plotter (http://www.aavso.org/vsp). Observations should be submitted to the AAVSO International Database. See full Alert Notice for more details.

  10. Binding of Estrogenic Compounds to Recombinant Estrogen Receptor-α: Application to Environmental Analysis

    PubMed Central

    Pillon, Arnaud; Boussioux, Anne-Marie; Escande, Aurélie; Aït-Aïssa, Sélim; Gomez, Elena; Fenet, Hélène; Ruff, Marc; Moras, Dino; Vignon, Françoise; Duchesne, Marie-Josèphe; Casellas, Claude; Nicolas, Jean-Claude; Balaguer, Patrick

    2005-01-01

    Estrogenic activity in environmental samples could be mediated through a wide variety of compounds and by various mechanisms. High-affinity compounds for estrogen receptors (ERs), such as natural or synthetic estrogens, as well as low-affinity compounds such as alkylphenols, phthalates, and polychlorinated biphenyls are present in water and sediment samples. Furthermore, compounds such as polycyclic aromatic hydrocarbons, which do not bind ERs, modulate estrogen activity by means of the aryl hydrocarbon receptor (AhR). In order to characterize compounds that mediate estrogenic activity in river water and sediment samples, we developed a tool based on the ER-αligand-binding domain, which permitted us to estimate contaminating estrogenic compound affinities. We designed a simple transactivation assay in which compounds of high affinity were captured by limited amounts of recombinant ER-αand whose capture led to a selective inhibition of transactivation. This approach allowed us to bring to light that water samples contain estrogenic compounds that display a high affinity for ERs but are present at low concentrations. In sediment samples, on the contrary, we showed that estrogenic compounds possess a low affinity and are present at high concentration. Finally, we used immobilized recombinant ER-αto separate ligands for ER and AhR that are present in river sediments. Immobilized ER-α, which does not retain dioxin-like compounds, enabled us to isolate and concentrate ER ligands to facilitate their further analysis. PMID:15743715

  11. The aryl hydrocarbon receptor and glucocorticoid receptor interact to activate human metallothionein 2A

    SciTech Connect

    Sato, Shoko; Shirakawa, Hitoshi; Tomita, Shuhei; Tohkin, Masahiro; Gonzalez, Frank J.; Komai, Michio

    2013-11-15

    Although the aryl hydrocarbon receptor (AHR) and glucocorticoid receptor (GR) play essential roles in mammalian development, stress responses, and other physiological events, crosstalk between these receptors has been the subject of much debate. Metallothioneins are classic glucocorticoid-inducible genes that were reported to increase upon treatment with AHR agonists in rodent tissues and cultured human cells. In this study, the mechanism of human metallothionein 2A (MT2A) gene transcription activation by AHR was investigated. Cotreatment with 3-methylcholanthrene and dexamethasone, agonists of AHR and GR respectively, synergistically increased MT2A mRNA levels in HepG2 cells. MT2A induction was suppressed by RNA interference against AHR or GR. Coimmunoprecipitation experiments revealed a physical interaction between AHR and GR proteins. Moreover, chromatin immunoprecipitation assays indicated that AHR was recruited to the glucocorticoid response element in the MT2A promoter. Thus, we provide a novel mechanism whereby AHR modulates expression of human MT2A via the glucocorticoid response element and protein–protein interactions with GR. - Highlights: • Aryl hydrocarbon receptor forms a complex with glucocorticoid receptor in cells. • Human metallothionein gene is regulated by the AHR and GR interaction. • AHR–GR complex binds to glucocorticoid response element in metallothionein gene. • We demonstrated a novel transcriptional mechanism via AHR and GR interaction.

  12. Simulator for SUPO, a Benchmark Aqueous Homogeneous Reactor (AHR)

    SciTech Connect

    Klein, Steven Karl; Determan, John C.

    2015-10-14

    A simulator has been developed for SUPO (Super Power) an aqueous homogeneous reactor (AHR) that operated at Los Alamos National Laboratory (LANL) from 1951 to 1974. During that period SUPO accumulated approximately 600,000 kWh of operation. It is considered the benchmark for steady-state operation of an AHR. The SUPO simulator was developed using the process that resulted in a simulator for an accelerator-driven subcritical system, which has been previously reported.

  13. Long interspersed element-1 is differentially regulated by food-borne carcinogens via the aryl hydrocarbon receptor.

    PubMed

    Okudaira, N; Okamura, T; Tamura, M; Iijma, K; Goto, M; Matsunaga, A; Ochiai, M; Nakagama, H; Kano, S; Fujii-Kuriyama, Y; Ishizaka, Y

    2013-10-10

    A single human cell contains more than 5.0 × 10(5) copies of long interspersed element-1 (L1), 80-100 of which are competent for retrotransposition (L1-RTP). Recent observations have revealed the presence of de novo L1 insertions in various tumors, but little is known about its mechanism. Here, we found that 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3,8-dimethyl-imidazo[4,5-f]quinoxaline (MeIQx), food-borne carcinogens that are present in broiled meats, induced L1-RTP. This induction was dependent on a cellular cascade comprising the aryl hydrocarbon receptor (AhR), a mitogen-activated protein kinase, and CCAAT/enhancer-binding protein β. Notably, these compounds exhibited differential induction of L1-RTP. MeIQx-induced L1-RTP was dependent on AhR nuclear translocator 1 (ARNT1), a counterpart of AhR required for gene expression in response to environmental pollutants. By contrast, PhIP-induced L1-RTP did not require ARNT1 but was dependent on estrogen receptor α (ERα) and AhR repressor. In vivo studies using transgenic mice harboring the human L1 gene indicated that PhIP-induced L1-RTP was reproducibly detected in the mammary gland, which is a target organ of PhIP-induced carcinoma. Moreover, picomolar levels of each compound induced L1-RTP, which is comparable to the PhIP concentration detected in human breast milk. Data suggest that somatic cells possess machineries that induce L1-RTP in response to the carcinogenic compounds. Together with data showing that micromolar levels of heterocyclic amines (HCAs) were non-genotoxic, our observations indicate that L1-RTP by environmental compounds is a novel type of genomic instability, further suggesting that analysis of L1-RTP by HCAs is a novel approach to clarification of modes of carcinogenesis.

  14. Baicalin Protects Mice from Aristolochic Acid I-Induced Kidney Injury by Induction of CYP1A through the Aromatic Hydrocarbon Receptor

    PubMed Central

    Wang, Ke; Feng, Chenchen; Li, Chenggang; Yao, Jun; Xie, Xiaofeng; Gong, Likun; Luan, Yang; Xing, Guozhen; Zhu, Xue; Qi, Xinming; Ren, Jin

    2015-01-01

    Exposure to aristolochic acid I (AAI) can lead to aristolochic acid nephropathy (AAN), Balkan endemic nephropathy (BEN) and urothelial cancer. The induction of hepatic CYP1A, especially CYP1A2, was considered to detoxify AAI so as to reduce its nephrotoxicity. We previously found that baicalin had the strong ability to induce CYP1A2 expression; therefore in this study, we examined the effects of baicalin on AAI toxicity, metabolism and disposition, as well as investigated the underlying mechanisms. Our toxicological studies showed that baicalin reduced the levels of blood urea nitrogen (BUN) and creatinine (CRE) in AAI-treated mice and attenuated renal injury induced by AAI. Pharmacokinetic analysis demonstrated that baicalin markedly decreased AUC of AAI in plasma and the content of AAI in liver and kidney. CYP1A induction assays showed that baicalin exposure significantly increased the hepatic expression of CYP1A1/2, which was completely abolished by inhibitors of the Aromatic hydrocarbon receptor (AhR), 3ʹ,4ʹ-dimethoxyflavone and resveratrol, in vitro and in vivo, respectively. Moreover, the luciferase assays revealed that baicalin significantly increased the luciferase activity of the reporter gene incorporated with the Xenobiotic response elements recognized by AhR. In summary, baicalin significantly reduced the disposition of AAI and ameliorated AAI-induced kidney toxicity through AhR-dependent CYP1A1/2 induction in the liver. PMID:26204831

  15. Pleiotropic effects of gold(I) mixed-ligand complexes of 9-deazahypoxanthine on transcriptional activity of receptors for steroid hormones, nuclear receptors and xenoreceptors in human hepatocytes and cell lines.

    PubMed

    Kubešová, Kateřina; Trávníček, Zdeněk; Dvořák, Zdeněk

    2016-10-01

    Development of metal-based compounds is an important research avenue in anti-cancer and anti-inflammatory drug discovery. Here we examined the effects of three gold (I) mixed-ligand complexes with the general formula [Au(Ln)(PPh3)] (1, 2, 3) involving triphenylphosphine (PPh3) and a deprotonated form of O-substituted derivatives of 9-deazahypoxanthine (Ln) on the transcriptional activity of aryl hydrocarbon receptor (AhR), androgen receptor (AR), glucocorticoid receptor (GR), thyroid receptor (TR), pregnane X receptor (PXR) and vitamin D receptor (VDR), employing gene reporter assays. In addition, we measured mRNA (RT-PCR) and protein (western blot) expression of target genes for those receptors, including drug-metabolizing P450s, in primary human hepatocytes and cancer cell lines LS180 and HepG2. The tested compounds displayed anti-glucocorticoid effects, as revealed by inhibition of dexamethasone-inducible transcriptional activity of GR and down-regulation of tyrosine aminotransferase. All the compounds slightly and dose-dependently activated PXR and AhR, and moderately induced CYP3A4 and CYP1A1/2 genes in human hepatocytes and LS180 cells. The complexes antagonized basal and ligand-activated AR and VDR, indicating inverse agonist behaviour. Both basal and thyroid hormone-inducible transcriptional activity of TR was dose-dependently increased by all tested compounds. In contrast, the expression of SPOT14 mRNA was decreased by tested compounds in human hepatocytes and HepG2 cells. In conclusion, if intended for human pharmacotherapy, the potential of the complexes 1-3 to influence studied receptors should be taken in account. PMID:27318977

  16. AHR-5850: a potent anti-inflammatory compound.

    PubMed

    Sancilio, L F; Reese, D L; Cheung, S; Alphin, R S

    1977-03-01

    AHR-5850 is a non-steroidal anti-inflammatory compound possessing antipyretic and analgesic properties. AHR-5850 was 16.4 and 22.8 times more potent than phenylbutazone in suppressing acute (Evans blue-carrageenan pleural effusion) and chronic (adjuvant-induced arthritis) inflammation, respectively. The analgesic activity of AHR 5850 was 43 times that of acetylsalicylic acid in the Randall-Selitto assay, and 156 and 56.3 times more potent than phenylbutazone in the acetylcholine-induced abdominal constriction in mice and in the bradykinin-induced nociceptive response in dogs, respectively. Single-dose studies showed that AHR-5850 produced less gastric irritation than acetylsalicylic acid when applied topically to the exposed gastric mucosa of cats or when administered orally to rats and dogs. Upon subchronic oral administration to rats, the therapeutic ratio of AHR-5850 was twice that of phenylbutazone. This was based on the ratio of its potency relative to phenylbutazone in producing intestinal lesions to its anti-inflammatory potency relative to phenylbutazone in the adjuvant-induced arthritis.

  17. Biological effects of 6-formylindolo[3,2-b]carbazole (FICZ) in vivo are enhanced by loss of CYP1A function in an Ahr2-dependent manner.

    PubMed

    Wincent, Emma; Kubota, Akira; Timme-Laragy, Alicia; Jönsson, Maria E; Hahn, Mark E; Stegeman, John J

    2016-06-15

    6-Formylindolo[3,2-b]carbazole (FICZ) is a potent aryl hydrocarbon receptor (AHR) agonist that is efficiently metabolized by AHR-regulated cytochrome P4501 enzymes. FICZ is a proposed physiological AHR ligand that induces its own degradation as part of a regulatory negative feedback loop. In vitro studies in cells show that CYP1 inhibition in the presence of FICZ results in enhanced AHR activation, suggesting that FICZ accumulates in the cell when its metabolism is blocked. We used zebrafish (Danio rerio) embryos to investigate the in vivo effects of FICZ when CYP1A is knocked down or inhibited. Embryos were injected with morpholino antisense oligonucleotides targeting CYP1A (CYP1A-MO), Ahr2, or a combination of both. FICZ exposure of non-injected embryos or embryos injected with control morpholino had little effect. In CYP1A-MO-injected embryos, however, FICZ dramatically increased mortality, incidence and severity of pericardial edema and circulation failure, reduced hatching frequency, blocked swim bladder inflation, and strongly potentiated expression of Ahr2-regulated genes. These effects were substantially reduced in embryos with a combined knockdown of Ahr2 and CYP1A, indicating that the toxicity was mediated at least partly by Ahr2. Co-exposure to the CYP1 inhibitor alpha-naphthoflavone (αNF) and FICZ had similar effects as the combination of CYP1A-MO and FICZ. HPLC analysis of FICZ-exposed embryos showed increased levels of FICZ after concomitant CYP1A-MO injection or αNF co-exposure. Together, these results show that a functioning CYP1/AHR feedback loop is crucial for regulation of AHR signaling by a potential physiological ligand in vivo and further highlights the role of CYP1 enzymes in regulating biological effects of FICZ.

  18. Genotoxicity but not the AhR-mediated activity of PAHs is inhibited by other components of complex mixtures of ambient air pollutants.

    PubMed

    Líbalová, Helena; Krčková, Simona; Uhlířová, Kateřina; Milcová, Alena; Schmuczerová, Jana; Ciganek, Miroslav; Kléma, Jiri; Machala, Miroslav; Šrám, Radim J; Topinka, Jan

    2014-03-21

    In this study, we compared the genotoxicity and aryl hydrocarbon receptor (AhR)-dependent transcriptional changes of selected target genes in human lung epithelial A549 cells incubated for 24 h, either with extractable organic matter (EOMs) from airborne particles <2.5 μm (PM2.5) collected at four localities from heavily polluted areas of the Czech Republic or two representative toxic polycyclic aromatic hydrocarbons (PAHs) present in EOMs, benzo[a]pyrene (B[a]P) and benzo[k]fluoranthene (B[k]F). Genotoxic effects were determined using DNA adduct analysis or analysis of expression of selected AhR-related genes involved in bioactivation of PAHs (CYP1A1, CYP1B1) and transcriptional repression (TIPARP). Sampled localities differing in the extent and source of air pollution did not exhibit substantially different genotoxicity. DNA adduct levels induced by three subtoxic EOM concentrations were relatively low (1-5 adducts/10(8) nucleotides), compared to levels induced by similar concentrations of B[a]P, while B[k]F gave very low DNA adduct levels. Here, we compared genotoxicity and gene deregulation induced by complex mixtures containing PAHs with the effects of the comparable concentrations of individual PAHs. Our results suggested inhibition of formation of B[a]P-induced DNA adducts compared to individual B[a]P, probably attributable to competitive inhibition by other non-genotoxic EOM components. In contrast, induction of AhR target genes appeared not to be antagonized by the components of complex mixtures, as induction of CYP1A1, CYP1B1 and TIPARP transcripts reached maximum levels induced by PAHs.

  19. Structural identification of Diindole agonists of the aryl hydrocarbon receptor derived from degradation of indole-3-pyruvic acid.

    PubMed

    Chowdhury, Goutam; Dostalek, Miroslav; Hsu, Erin L; Nguyen, Linh P; Stec, Donald F; Bradfield, Christopher A; Guengerich, F Peter

    2009-12-01

    Aerobic incubation of the tryptophan transamination/oxidation product indole-3-pyruvic acid (I3P) at pH 7.4 and 37 degrees C yielded products with activity as Ah receptor (AHR) agonists. The extracts were fractionated using HPLC and screened for AHR agonist activity. Two compounds were identified as agonists: 1,3-di(1H-indol-3-yl)propan-2-one (1) and 1-(1H-indol-3-yl)-3-(3H-indol-3-ylidene) propan-2-one (2), with the potency of 2 being 100-fold > 1 [ Nguyen et al. ( 2009 ) Chem. Res. Toxicol. , DOI: 10.1021/tx900043s . ]. Both 1 and 2 showed UV spectra indicative of indole. The molecular formulas were established by high-resolution mass spectrometry (HRMS), and the structures were determined by a combination of NMR methods, including (1)H, natural abundance (13)C, and two-dimensional methods. An intermediate in the oxidation of I3P to 1 is 3-hydroxy-2,4-di(1H-indol-3-yl)butanal (HRMS established the presence of a compound with the formula C(20)H(19)N(2)O(2)). Compound 1 was converted to 2 in air or (faster) with mild oxidants, and 2 could be further oxidized to 1,3-di(3H-indol-3-ylidene)propan-2-one. Determination of the structures allowed estimation of the molar Ah receptor agonist activity of these natural products, similar in potency to known classical AHR inducers. PMID:19860413

  20. Expression and distribution patterns of Mas-related gene receptor subtypes A-H in the mouse intestine: inflammation-induced changes.

    PubMed

    Avula, Leela Rani; Buckinx, Roeland; Favoreel, Herman; Cox, Eric; Adriaensen, Dirk; Van Nassauw, Luc; Timmermans, Jean-Pierre

    2013-05-01

    Mas-related gene (Mrg) receptors constitute a subfamily of G protein-coupled receptors that are implicated in nociception, and are as such considered potential targets for pain therapies. Furthermore, some Mrgs have been suggested to play roles in the regulation of inflammatory responses to non-immunological activation of mast cells and in mast cell-neuron communication. Except for MrgD, E and F, whose changed expression has been revealed during inflammation in the mouse intestine in our earlier studies, information concerning the remaining cloned mouse Mrg subtypes in the gastrointestinal tract during (patho) physiological conditions is lacking. Therefore, the present study aimed at identifying the presence and putative function of these remaining cloned Mrg subtypes (n = 19) in the (inflamed) mouse intestine. Using reverse transcriptase-PCR, quantitative-PCR and multiple immunofluorescence staining with commercial and newly custom-developed antibodies, we compared the ileum and the related dorsal root ganglia (DRG) of non-inflamed mice with those of two models of intestinal inflammation, i.e., intestinal schistosomiasis and 2,4,6-trinitrobenzene sulfonic acid-induced ileitis. In the non-inflamed ileum and DRG, the majority of the Mrg subtypes examined were sparsely expressed, showing a neuron-specific expression pattern. However, significant changes in the expression patterns of multiple Mrg subtypes were observed in the inflamed ileum; for instance, MrgA4, MrgB2and MrgB8 were expressed in a clearly increased number of enteric sensory neurons and in nerve fibers in the lamina propria, while de novo expression of MrgB10 was observed in enteric sensory neurons and in newly recruited mucosal mast cells (MMCs). The MrgB10 expressing MMCs were found to be in close contact with nerve fibers in the lamina propria. This is the first report on the expression of all cloned Mrg receptor subtypes in the (inflamed) mouse intestine. The observed changes in the expression and

  1. USE OF THE TEQ MODEL FOR ASSESSING AHR MEDIATED TOXICITY RISKS TO POPULATIONS OF LAKE TROUT AND OTHER SPECIES IN LAKE ONTARIO

    EPA Science Inventory

    The toxicity equivalence (TEQ) model for assessing aryl hydrocarbon receptor (AHR) mediated toxicity risks associated with polyhalogenated aromatic chemicals structurally similar to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has been applied to human health risks for more than 15...

  2. Sequence and in vitro function of chicken, ring-necked pheasant, and Japanese quail AHR1 predict in vivo sensitivity to dioxins.

    PubMed

    Farmahin, Reza; Wu, Dongmei; Crump, Doug; Hervé, Jessica C; Jones, Stephanie P; Hahn, Mark E; Karchner, Sibel I; Giesy, John P; Bursian, Steven J; Zwiernik, Matthew J; Kennedy, Sean W

    2012-03-01

    There are large differences in sensitivity to the toxic and biochemical effects of dioxins and dioxin-like compounds (DLCs) among vertebrates. Previously, we demonstrated that the difference in sensitivity between domestic chicken (Gallus gallus domesticus) and common tern (Sterna hirundo) to aryl hydrocarbon receptor 1 (AHR1)-dependent changes in gene expression following exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is based upon the identities of the amino acids at two sites within the ligand binding domain of AHR1 (chicken--highly sensitive; Ile324_Ser380 vs common tern--250-fold less sensitive than chicken; Val325_Ala381). Here, we tested the hypotheses that (i) the sensitivity of other avian species to TCDD, 2,3,4,7,8-pentachlorodibenzofuran (PeCDF), and 2,3,7,8-tetrachlorodibenzofuran (TCDF) is also determined by the amino acids at sites that are equivalent to sites 324 and 380 in chicken, and (ii) Ile324_Ala380 and Val324_Ser380 genotypes confer intermediate sensitivity to DLCs in birds. We compared ligand-induced transactivation function of full-length AHR1s from chicken, common tern, ring-necked pheasant (Phasianus colchicus; Ile324_Ala380) and Japanese quail (Coturnix japonica; Val324_Ala380), and three Japanese quail AHR1 mutants. The results support our hypothesis that avian species can be grouped into three general classes of sensitivity to DLCs. Both AHR1 genotype and in vitro transactivation assays predict in vivo sensitivity. Contrary to the assumption that TCDD is the most potent DLC, PeCDF was more potent than TCDD at activating Japanese quail (13- to 26-fold) and common tern (23- to 30-fold) AHR1. Our results support and expand previous in vitro and in vivo work that demonstrated ligand-dependent species differences in AHR1 affinity. The findings and methods will be of use for DLC risk assessments.

  3. Silencing of miR-124 induces neuroblastoma SK-N-SH cell differentiation, cell cycle arrest and apoptosis through promoting AHR.

    PubMed

    Huang, Tsui-Chin; Chang, Hsin-Yi; Chen, Cheng-Yu; Wu, Pei-Yi; Lee, Hsinyu; Liao, Yung-Feng; Hsu, Wen-Ming; Huang, Hsuan-Cheng; Juan, Hsueh-Fen

    2011-11-16

    Neuroblastoma is the most common extracranial solid tumor in children. We investigate whether miR-124, the abundant neuronal miRNA, plays a pivotal role in neuroblastoma. Knockdown of miR-124 promotes neuroblastoma SK-N-SH cell differentiation, cell cycle arrest and apoptosis. Further miR-124 is predicted to target aryl hydrocarbon receptor (AHR) which may promote neuroblastoma cell differentiation. We validate that miR-124 may suppress the expression of AHR by targeting its 3'-UTR. These results suggest that miR-124 could serve as a potential therapeutic target of neuroblastoma.

  4. Regulation of pregnane-X-receptor, CYP3A and P-glycoprotein genes in the PCB-resistant killifish (Fundulus heteroclitus) population from New Bedford Harbor.

    PubMed

    Gräns, Johanna; Wassmur, Britt; Fernández-Santoscoy, María; Zanette, Juliano; Woodin, Bruce R; Karchner, Sibel I; Nacci, Diane E; Champlin, Denise; Jayaraman, Saro; Hahn, Mark E; Stegeman, John J; Celander, Malin C

    2015-02-01

    Killifish survive and reproduce in the New Bedford Harbor (NBH) in Massachusetts (MA), USA, a site severely contaminated with polychlorinated biphenyls (PCBs) for decades. Levels of 22 different PCB congeners were analyzed in liver from killifish collected in 2008. Concentrations of dioxin-like PCBs in liver of NBH killifish were ∼400 times higher, and the levels of non-dioxin-like PCBs ∼3000 times higher than in killifish from a reference site, Scorton Creek (SC), MA. The NBH killifish are known to be resistant to the toxicity of dioxin-like compounds and to have a reduced aryl hydrocarbon receptor (AhR) signaling response. Little is known about the responses of these fish to non-dioxin-like PCBs, which are at extraordinarily high levels in NBH fish. In mammals, some non-dioxin-like PCB congeners act through nuclear receptor 1I2, the pregnane-X-receptor (PXR). To explore this pathway in killifish, a PXR cDNA was sequenced and its molecular phylogenetic relationship to other vertebrate PXRs was determined. Killifish were also collected in 2009 from NBH and SC, and after four months in the laboratory they were injected with a single dose of either the dioxin-like PCB 126 (an AhR agonist) or the non-dioxin-like PCB 153 (a mammalian PXR agonist). Gills and liver were sampled three days after injection and transcript levels of genes encoding PXR, cytochrome P450 3A (CYP3A), P-glycoprotein (Pgp), AhR2 and cytochrome P450 1A (CYP1A) were measured by quantitative PCR. As expected, there was little effect of PCB exposure on mRNA expression of AhR2 or CYP1A in liver and gills of NBH fish. In NBH fish, but not in SC fish, there was increased mRNA expression of hepatic PXR, CYP3A and Pgp upon exposure to either of the two PCB congeners. However, basal PXR and Pgp mRNA levels in liver of NBH fish were significantly lower than in SC fish. A different pattern was seen in gills, where there were no differences in basal mRNA expression of these genes between the two

  5. Inhibition of aryl hydrocarbon receptor transactivation and DNA adduct formation by CYP1 isoform-selective metabolic deactivation of benzo[a]pyrene

    SciTech Connect

    Endo, Kaori; Uno, Shigeyuki; Seki, Taiichiro; Ariga, Toyohiko; Kusumi, Yoshiaki; Mitsumata, Masako; Yamada, Sachiko; Makishima, Makoto

    2008-07-15

    Benzo[a]pyrene (BaP), a polyaromatic hydrocarbon produced by the combustion of cigarettes and coke ovens, is a known procarcinogen. BaP activates the aryl hydrocarbon receptor (AhR) and induces the expression of a battery of genes, including CYP1A1, which metabolize BaP to toxic compounds. The possible role of CYP1 enzymes in mediating BaP detoxification or metabolic activation remains to be elucidated. In this study, we assessed the effects of CYP1 enzymes (CYP1A1, CYP1A2 and CYP1B1) on BaP-induced AhR transactivation and DNA adduct formation in HEK293 cells and HepG2 cells. Transfection of CYP1A1 and CYP1B1, but not CYP1A2, suppressed BaP-induced activation of AhR. Expression of CYP1A1 and CYP1A2, but not CYP1B1, inhibited DNA adduct formation in BaP-treated HepG2 cells. These results indicate that CYP1A1 and CYP1B1 play a role in deactivation of BaP on AhR and that CYP1A1 and CYP1A2 are involved in BaP detoxification by suppressing DNA adduct formation. BaP treatment did not induce DNA adduct formation in HEK293 cells, even after transfection of CYP1 enzymes, suggesting that expression of CYP1 enzymes is not sufficient for DNA adduct formation. Lower expression of epoxide hydrolase and higher expression of glutathione S-transferase P1 (GSTP1) and GSTM1/M2 were observed in HEK293 cells compared with HepG2 cells. Dynamic expression of CYP1A1, CYP1A2 and CYP1B1 along with expression of other enzymes such as epoxide hydrolase and phase II enzymes may determine the detoxification or metabolic activation of BaP.

  6. PAH-CALUX, an optimized bioassay for AhR-mediated hazard identification of polycyclic aromatic hydrocarbons (PAHs) as individual compounds and in complex mixtures.

    PubMed

    Pieterse, B; Felzel, E; Winter, R; van der Burg, B; Brouwer, A

    2013-10-15

    Polycyclic aromatic hydrocarbons (PAHs) represent a class of ubiquitously occurring environmental compounds that are implicated in a wide range of toxicological effects. Routine measurement of PAH contamination generally involves chemical analytical analysis of a selected group of representatives, for example, EPA-16, which may result in underestimation of the PAH-related toxicity of a sample. Many high molecular weight PAHs are known ligands of the aryl hydrocarbon receptor (AhR), a nuclear receptor that mediates toxic effects related to these compounds. Making use of this property we developed a PAH CALUX assay, a mammalian, H4IIe- cell-based reporter assay for the hazard identification of total PAH mixtures. The PAH CALUX reporter cell line allows for specific, rapid (4 h exposure time) and reliable quantification of AhR-induced luciferase induction relative to benzo[a]pyrene (BaP), which is used as a positive reference PAH congener. Full dose response relationships with inductions over 100-fold were reached within only 2 h of exposure to BaP. The PAH CALUX is highly sensitive, that is, using a 4 h exposure time, a limit of detection (LOD) of 5.2 × 10(-11) M BaP was achieved, and highly accurate, that is, a repeatability of 5.9% and a reproducibility of 6.6% were established. Screening of a selection of PAHs that were prioritized by the European Union and/or the U.S. Environmental Protection Agency showed that the PAH CALUX bioassay has a high predictability, particularly for carcinogenic PAHs. Experiments with synthetic mixtures and reference materials containing complex PAH mixtures show the suitability of the assay for these types of applications. Moreover, the presented results suggest that application of the PAH CALUX will result in a lower risk of underestimation of the toxicity of a sample than chemical analytical approaches that focus on a limited set of prioritized compounds.

  7. The aryl hydrocarbon receptor: a perspective on potential roles in the immune system

    PubMed Central

    Stevens, Emily A; Mezrich, Joshua D; Bradfield, Christopher A

    2009-01-01

    The aryl hydrocarbon receptor (AHR) is a protein best known for its role in mediating toxicity. Over 30 years of research has uncovered additional roles for the AHR in xenobiotic metabolism and normal vascular development. Activation of the AHR has long been known to cause immunotoxicity, including thymic involution. Recent data suggesting a role for the AHR in regulatory T-cell (Treg) and T-helper 17 (Th17) cell development have only added to the excitement about this biology. In this review, we will attempt to illustrate what is currently known about AHR biology in the hope that data from fields as diverse as evolutionary biology and pharmacology will help elucidate the mechanism by which AHR modifies immune responses. We also will discuss the complexities of AHR pharmacology and genetics that may influence future studies of AHR in the immune system. PMID:19538249

  8. The aryl hydrocarbon receptor agonist benzo(a)pyrene reactivates LINE-1 in HepG2 cells through canonical TGF-β1 signaling: implications in hepatocellular carcinogenesis.

    PubMed

    Reyes-Reyes, Elsa M; Ramos, Irma N; Tavera-Garcia, Marco A; Ramos, Kenneth S

    2016-01-01

    Long interspersed nuclear element-1 (L1) is a genetic element that mobilizes throughout the mammalian genome via retrotransposition and damages host DNA via mutational insertions, chromosomal rearrangements, and reprogramming of gene expression. The cellular mechanisms responsible for aberrant L1 expression during cancer pathogenesis are unclear. Previously, we have shown that L1 reactivation in several human cell lines is dependent upon the activation of aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor member of the PAS superfamily of proteins. We also showed that ectopic expression of L1 reprograms the HepG2 genome leading to epithelial-to-mesenchymal transition (EMT). Here we present evidence that reactivation of L1 and modulation of EMT in HepG2 cells by the AhR ligand benzo(a)pyrene (BaP) is effected through the canonical TGF-β1 signaling pathway. BaP increased TGF-β1 mRNA, SMAD2 phosphorylation and decreased expression of E-Cadherin. The functional relevance of these interactions and the involvement of TGFBR1/ALK5 and SMAD2/3 were confirmed by siRNA interference. Furthermore, expression of L1-encoded ORF1p was positively correlated with the activation of TGF-β1 signaling in human hepatocarcinoma samples at various stages of malignant progression. These results indicate that ligand-mediated AhR activation regulates L1 via canonical TGF-β1 signaling and raise important questions about the molecular etiology of human hepatocarcinomas.

  9. The aryl hydrocarbon receptor agonist benzo(a)pyrene reactivates LINE-1 in HepG2 cells through canonical TGF-β1 signaling: implications in hepatocellular carcinogenesis.

    PubMed

    Reyes-Reyes, Elsa M; Ramos, Irma N; Tavera-Garcia, Marco A; Ramos, Kenneth S

    2016-01-01

    Long interspersed nuclear element-1 (L1) is a genetic element that mobilizes throughout the mammalian genome via retrotransposition and damages host DNA via mutational insertions, chromosomal rearrangements, and reprogramming of gene expression. The cellular mechanisms responsible for aberrant L1 expression during cancer pathogenesis are unclear. Previously, we have shown that L1 reactivation in several human cell lines is dependent upon the activation of aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor member of the PAS superfamily of proteins. We also showed that ectopic expression of L1 reprograms the HepG2 genome leading to epithelial-to-mesenchymal transition (EMT). Here we present evidence that reactivation of L1 and modulation of EMT in HepG2 cells by the AhR ligand benzo(a)pyrene (BaP) is effected through the canonical TGF-β1 signaling pathway. BaP increased TGF-β1 mRNA, SMAD2 phosphorylation and decreased expression of E-Cadherin. The functional relevance of these interactions and the involvement of TGFBR1/ALK5 and SMAD2/3 were confirmed by siRNA interference. Furthermore, expression of L1-encoded ORF1p was positively correlated with the activation of TGF-β1 signaling in human hepatocarcinoma samples at various stages of malignant progression. These results indicate that ligand-mediated AhR activation regulates L1 via canonical TGF-β1 signaling and raise important questions about the molecular etiology of human hepatocarcinomas. PMID:27293999

  10. The aryl hydrocarbon receptor agonist benzo(a)pyrene reactivates LINE-1 in HepG2 cells through canonical TGF-β1 signaling: implications in hepatocellular carcinogenesis

    PubMed Central

    Reyes-Reyes, Elsa M; Ramos, Irma N; Tavera-Garcia, Marco A; Ramos, Kenneth S

    2016-01-01

    Long interspersed nuclear element-1 (L1) is a genetic element that mobilizes throughout the mammalian genome via retrotransposition and damages host DNA via mutational insertions, chromosomal rearrangements, and reprogramming of gene expression. The cellular mechanisms responsible for aberrant L1 expression during cancer pathogenesis are unclear. Previously, we have shown that L1 reactivation in several human cell lines is dependent upon the activation of aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor member of the PAS superfamily of proteins. We also showed that ectopic expression of L1 reprograms the HepG2 genome leading to epithelial-to-mesenchymal transition (EMT). Here we present evidence that reactivation of L1 and modulation of EMT in HepG2 cells by the AhR ligand benzo(a)pyrene (BaP) is effected through the canonical TGF-β1 signaling pathway. BaP increased TGF-β1 mRNA, SMAD2 phosphorylation and decreased expression of E-Cadherin. The functional relevance of these interactions and the involvement of TGFBR1/ALK5 and SMAD2/3 were confirmed by siRNA interference. Furthermore, expression of L1-encoded ORF1p was positively correlated with the activation of TGF-β1 signaling in human hepatocarcinoma samples at various stages of malignant progression. These results indicate that ligand-mediated AhR activation regulates L1 via canonical TGF-β1 signaling and raise important questions about the molecular etiology of human hepatocarcinomas. PMID:27293999

  11. TCDD dysregulation of 13 AHR-target genes in rat liver

    SciTech Connect

    Watson, John D.; Prokopec, Stephenie D.; Smith, Ashley B.; Okey, Allan B.; Pohjanvirta, Raimo; Boutros, Paul C.

    2014-02-01

    Despite several decades of research, the complete mechanism by which 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and other xenobiotic agonists of the aryl hydrocarbon receptor (AHR) cause toxicity remains unclear. While it has been shown that the AHR is required for all major manifestations of toxicity, the specific downstream changes involved in the development of toxic phenotypes remain unknown. Here we examine a panel of 13 genes that are AHR-regulated in many species and tissues. We profiled their hepatic mRNA abundances in two rat strains with very different sensitivities to TCDD: the TCDD-sensitive Long–Evans (Turku/AB; L–E) and the TCDD-resistant Han/Wistar (Kuopio; H/W). We evaluated doses ranging from 0 to 3000 μg/kg at 19 h after TCDD exposure and time points ranging from 1.5 to 384 h after exposure to 100 μg/kg TCDD. Twelve of 13 genes responded to TCDD in at least one strain, and seven of these showed statistically significant inter-strain differences in the time course analysis (Aldh3a1, Cyp1a2, Cyp1b1, Cyp2a1, Fmo1, Nfe2l2 and Nqo1). Cyp2s1 did not respond to TCDD in either rat strain. Five genes exhibited biphasic responses to TCDD insult (Ahrr, Aldh3a1, Cyp1b1, Nfe2l2 and Nqo1), suggesting a secondary event, such as association with additional transcriptional modulators. Of the 12 genes that responded to TCDD during the dose–response analysis, none had an ED{sub 50} equivalent to that of Cyp1a1, the most sensitive gene in this study, while nine genes responded to doses at least 10–100 fold higher, in at least one strain (Ahrr (L–E), Aldh3a1 (both), Cyp1a2 (both), Cyp1b1 (both), Cyp2a1 (L–E), Inmt (both), Nfe2l2 (L–E), Nqo1 (L–E) and Tiparp (both)). These data shed new light on the association of the AHR target genes with TCDD toxicity, and in particular the seven genes exhibiting strain-specific differences represent strong candidate mediators of Type-II toxicities. - Highlights: • NanoString measured hepatic mRNA molecules

  12. Differential modulation of expression of nuclear receptor mediated genes by tris(2-butoxyethyl) phosphate (TBOEP) on early life stages of zebrafish (Danio rerio).

    PubMed

    Ma, Zhiyuan; Yu, Yijun; Tang, Song; Liu, Hongling; Su, Guanyong; Xie, Yuwei; Giesy, John P; Hecker, Markus; Yu, Hongxia

    2015-12-01

    As one substitute for phased-out brominated flame retardants (BFRs), tris(2-butoxyethyl) phosphate (TBOEP) is frequently detected in aquatic organisms. However, knowledge about endocrine disrupting mechanisms associated with nuclear receptors caused by TBOEP remained restricted to results from in vitro studies with mammalian cells. In the study, results of which are presented here, embryos/larvae of zebrafish (Danio rerio) were exposed to 0.02, 0.1 or 0.5μM TBOEP to investigate expression of genes under control of several nuclear hormone receptors (estrogen receptors (ERs), androgen receptor (AR), thyroid hormone receptor alpha (TRα), mineralocorticoid receptor (MR), glucocorticoid receptor (GR), aryl hydrocarbon (AhR), peroxisome proliferator-activated receptor alpha (PPARα), and pregnane×receptor (P×R)) pathways at 120hpf. Exposure to 0.5μM TBOEP significantly (p<0.05, one-way analysis of variance) up-regulated expression of estrogen receptors (ERs, er1, er2a, and er2b) genes and ER-associated genes (vtg4, vtg5, pgr, ncor, and ncoa3), indicating TBOEP modulates the ER pathway. In contrast, expression of most genes (mr, 11βhsd, ube2i,and adrb2b) associated with the mineralocorticoid receptor (MR) pathway were significantly down-regulated. Furthermore, in vitro mammalian cell-based (MDA-kb2 and H4IIE-luc) receptor transactivation assays, were also conducted to investigate possible agonistic or antagonistic effects on AR- and AhR-mediated pathways. In mammalian cells, none of these pathways were affected by TBOEP at the concentrations studied. Receptor-mediated responses (in vivo) and mammalian cell lines receptor binding assay (in vitro) combined with published information suggest that TBOEP can modulate receptor-mediated, endocrine process (in vivo/in vitro), particularly ER and MR. PMID:26562049

  13. A Mutant Ahr Allele Protects the Embryonic Kidney from Hydrocarbon-Induced Deficits in Fetal Programming

    PubMed Central

    Nanez, Adrian

    2011-01-01

    Background: The use of experimental model systems has expedited the elucidation of pathogenetic mechanisms of renal developmental disease in humans and the identification of genes that orchestrate developmental programming during nephrogenesis. Objectives: We conducted studies to evaluate the role of AHR polymorphisms in the disruption of renal developmental programming by benzo(a)pyrene (BaP). Methods: We used metanephric cultures of C57BL/6J (C57) mice expressing the Ahrb-1 allele and B6.D2N-Ahrd/J (D2N) mice expressing a mutant allele deficient in ligand binding (Ahrd) to investigate molecular mechanisms of renal development. Deficits in fetal programming were evaluated in the offspring of pregnant mice treated with BaP during nephrogenesis. Results: Hydrocarbon challenge of metanephri from C57 mice altered Wilms’ tumor suppressor gene (Wt1) mRNA splice variant ratios and reduced mRNAs of the Wt1 transcriptional targets syndecan-1 (Sdc1) paired box gene 2 (Pax2), epidermal growth factor receptor (Egfr), and retinoic acid receptor, alpha (Rarα). These changes correlated with down-regulation of effectors of differentiation [secreted frizzled-related sequence protein 1 (Sfrp1), insulin-like growth factor 1 receptor (Igf1r), wingless-related MMTV-integration site 4 (Wnt4), Lim homeobox protein 1 (Lhx1), E-cadherin]. In contrast, metanephri from D2N mice were spared hydrocarbon-induced changes in Wt1 splice variant ratios and deficits of differentiation. We observed similar patterns of dysmorphogenesis and progressive loss of renal function at postnatal weeks 7 and 52 in the offspring of pregnant C57 but not D2N mice gavaged with 0.1 or 0.5 mg/kg BaP on gestation days 10–13. Conclusions: These findings support a functional link between AHR and WT1 in the regulation of renal morphogenesis and raise important questions about the contribution of human AHR polymorphisms to the fetal origins of adult-onset kidney disease. PMID:21803694

  14. In silico analysis of the interaction of avian aryl hydrocarbon receptors and dioxins to decipher isoform-, ligand-, and species-specific activations.

    PubMed

    Hirano, Masashi; Hwang, Ji-Hee; Park, Hae-Jeong; Bak, Su-Min; Iwata, Hisato; Kim, Eun-Young

    2015-03-17

    The aryl hydrocarbon receptor (AHR) mediates toxic responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and other dioxin-like compounds (DLCs). Avian species possess multiple AHR isoforms (AHR1, AHR1β, and AHR2) that exhibit species- and isoform-specific responses to ligands. To account for the ligand preference in terms of the structural features of avian AHRs, we generated in silico homology models of the ligand-binding domain of avian AHRs based on holo human HIF-2α (PDB entry 3H7W ). Molecular docking simulations of TCDD and other DLCs with avian AHR1s and AHR2s using ASEDock indicated that the interaction energy increased with the number of substituted chlorine atoms in congeners, supporting AHR transactivation potencies and World Health Organization TCDD toxic equivalency factors of congeners. The potential interaction energies of an endogenous AHR ligand, 6-formylindolo [3,2-b] carbazole (FICZ) to avian AHRs were lower than those of TCDD, which was supported by a greater potency of FICZ for in vitro AHR-mediated transactivation than TCDD. The molecular dynamics simulation revealed that mean square displacements in Ile324 and Ser380 of TCDD-bound AHR1 of the chicken, the most sensitive species to TCDD, were smaller than those in other avian AHR1s, suggesting that the dynamic stability of these amino acid residues contribute to TCDD preference. For avian AHR2, the corresponding residues (Val/Ser or Val/Ala type) were not responsible for differential TCDD sensitivity. Application of the three-dimensional reference interaction site model showed that the stabilization of TCDD binding to avian AHRs may be due to the solvation effect depending on the characteristics of two amino acids corresponding to Ile324 and Ser380 in chicken AHR1. This study demonstrates that in silico simulations of AHRs and ligands could be used to predict isoform-, ligand-, and species-specific interactions.

  15. The Search for Endogenous Activators of the Aryl Hydrocarbon Receptor

    PubMed Central

    Nguyen, Linh P.; Bradfield, Christopher A.

    2008-01-01

    In its simplest aspect, this review is an attempt to describe the major ligand classes of the aryl hydrocarbon receptor (AHR). A grander objective is to provide models that may help define the physiological activator or “endogenous ligand” of the AHR. We begin by presenting evidence that supports a developmental function for the AHR. This is followed by proposing mechanisms by which an endogenous ligand and consequent AHR activation might be important during normal physiology and development. With this background, we then present a survey of the known xenobiotic, endogenous, dietary and “un-conventional” activators of the AHR. When possible, this includes information about their induction potency, receptor binding affinity and potential for exposure. Because of the essential function of the AHR in embryonic development, we discuss the candidacy of each of these compounds as physiologically important activators. PMID:18076143

  16. Novel cDNA sequences of aryl hydrocarbon receptors and gene expression in turtles (Chrysemys picta and Pseudemys scripta) exposed to different environments

    PubMed Central

    Marquez, Emily C.; Traylor-Knowles, Nikki; Novillo-Villajos, Apolonia; Callard, Ian P.

    2011-01-01

    Reproductive changes have been observed in painted turtles from a site with known contamination located on Cape Cod, MA, USA. We hypothesize that these changes are caused by exposure to endocrine-disrupting compounds and that genes involved in reproduction are affected. The aryl hydrocarbon receptor (AHR) is an orphan receptor that is activated by environmental contaminants. AHR mRNA was measured in turtles exposed to soil collected from a contaminated site. Adult turtles were trapped from the study site (Moody Pond, MP) or a reference site and exposed to laboratory environments containing soil from either site. The red-eared slider was used to assess neonatal exposure to soil and water from the sites. The environmental exposures occurred over a 13-month period. Juveniles showed an age-dependent increase in brain AHR1. Juvenile turtles exposed to the MP environment had elevated gonadal AHR1. Adult turtles exposed to the MP environment showed significantly decreased brain AHR2. The painted turtle AHR is the first complete reptile AHR cDNA sequence. Phylogenetic analysis of the painted turtle AHR showed that it clusters with other AHR2s. Partial AHR1 and partial AHR2 cDNA sequences were cloned from the red-eared slider. MEME analysis identified 18 motifs in the turtle AHRs, showing high conservation between motifs that overlapped functional regions in both AHR isoforms. PMID:21763458

  17. Atropine-resistant effects of the muscarinic agonists McN-A-343 and AHR 602 on cardiac performance and the release of noradrenaline from sympathetic nerves of the perfused rabbit heart

    PubMed Central

    Fozard, J.R.; Muscholl, E.

    1974-01-01

    1 The effects of 4-(m-chlorophenylcarbamoyloxy)-2-butynyltrimethylammonium chloride (McN-A-343) and N-benzyl-3-pyrrolidyl acetate methobromide (AHR 602) on cardiac performance and noradrenaline release from terminal sympathetic fibres were measured in isolated perfused hearts of rabbits. 2 In the presence of sufficient atropine to block muscarinic receptors, high concentrations of McN-A-343 and AHR 602 caused no cardiac stimulation and there was no increase in the resting output of noradrenaline into the perfusates. 3 McN-A-343 and AHR 602 increased both the mechanical responses and the transmitter overflow evoked by electrical stimulation of the sympathetic nerves (SNS) but inhibited both parameters during perfusion with 1,1-dimethyl-4-phenylpiperazinium (DMPP). The effects were atropine-resistant and qualitatively similar to those seen with cocaine. Hexamethonium inhibited DMPP, but affected neither SNS per se nor the facilitatory effects of McN-A-343 and AHR 602 on SNS. 4 McN-A-343, cocaine and desipramine (but not AHR 602 or hexamethonium) blocked the net cardiac noradrenaline uptake and increased the positive chronotropic effect of noradrenaline. 5 Prior perfusion with concentrations of cocaine and desipramine sufficient to block uptake reduced or abolished the facilitatory effects of both McN-A-343 and AHR 602 on SNS. 6 Cocaine, McN-A-343 and AHR 602 displayed local anaesthetic properties on the guinea-pig wheal and frog nerve plexus tests, and their relative potencies in this respect were similar to those for inhibition of DMPP-evoked transmitter overflow. Hexamethonium did not produce local anaesthesia. 7 The results indicate that the facilitated release of noradrenaline after SNS and the inhibition of release after DMPP produced by McN-A-343 and AHR 602 are the result of their combined local anaesthetic action and inhibition of amine uptake. PMID:4447857

  18. In Utero and Lactational Exposure to PCBs in Mice: Adult Offspring Show Altered Learning and Memory Depending on Cyp1a2 and Ahr Genotypes

    PubMed Central

    Curran, Christine P.; Genter, Mary Beth; Patel, Krishna V.; Schaefer, Tori L.; Skelton, Matthew R.; Williams, Michael T.; Vorhees, Charles V.

    2011-01-01

    Background: Both coplanar and noncoplanar polychlorinated biphenyls (PCBs) exhibit neurotoxic effects in animal studies, but individual congeners do not always produce the same effects as PCB mixtures. Humans genetically have > 60-fold differences in hepatic cytochrome P450 1A2 (CYP1A2)-uninduced basal levels and > 12-fold variability in aryl hydrocarbon receptor (AHR)affinity; because CYP1A2 is known to sequester coplanar PCBs and because AHR ligands include coplanar PCBs, both genotypes can affect PCB response. Objectives: We aimed to develop a mouse paradigm with extremes in Cyp1a2 and Ahr genotypes to explore genetic susceptibility to PCB-induced developmental neurotoxicity using an environmentally relevant mixture of PCBs. Methods: We developed a mixture of eight PCBs to simulate human exposures based on their reported concentrations in human tissue, breast milk, and food supply. We previously characterized specific differences in PCB congener pharmacokinetics and toxicity, comparing high-affinity–AHR Cyp1a2 wild-type [Ahrb1_Cyp1a2(+/+)], poor-affinity–AHR Cyp1a2 wild-type [Ahrd_Cyp1a2(+/+)], and high-affinity–AHR Cyp1a2 knockout [Ahrb1_Cyp1a2(–/–)] mouse lines [Curran CP, Vorhees CV, Williams MT, Genter MB, Miller ML, Nebert DW. 2011. In utero and lactational exposure to a complex mixture of polychlorinated biphenyls: toxicity in pups dependent on the Cyp1a2 and Ahr genotypes. Toxicol Sci 119:189–208]. Dams received a mixture of three coplanar and five noncoplanar PCBs on gestational day 10.5 and postnatal day (PND) 5. In the present study we conducted behavioral phenotyping of exposed offspring at PND60, examining multiple measures of learning, memory, and other behaviors. Results: We observed the most significant deficits in response to PCB treatment in Ahrb1_Cyp1a2(–/–) mice, including impaired novel object recognition and increased failure rate in the Morris water maze. However, all PCB-treated genotypes showed significant differences on

  19. Hepatic Aryl Hydrocarbon Receptor Attenuates Fibroblast Growth Factor 21 Expression.

    PubMed

    Girer, Nathaniel G; Murray, Iain A; Omiecinski, Curtis J; Perdew, Gary H

    2016-07-15

    The Aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor involved in many physiological processes. Several studies indicate that AHR is also involved in energy homeostasis. Fibroblast growth factor 21 (FGF21) is an important regulator of the fasting and feeding responses. When administered to various genetic and diet-induced mouse models of obesity, FGF21 can attenuate obesity-associated morbidities. Here, we explore the role of AHR in hepatic Fgf21 expression through the use of a conditional, hepatocyte-targeted AHR knock-out mouse model (Cre(Alb)Ahr(Fx/Fx)). Compared with the congenic parental strain (Ahr(Fx/Fx)), non-fasted Cre(Alb)Ahr(Fx/Fx) mice exhibit a 4-fold increase in hepatic Fgf21 expression, as well as elevated expression of the FGF21-target gene Igfbp1 Furthermore, in vivo agonist activation of AHR reduces hepatic Fgf21 expression during a fast. The Fgf21 promoter contains several putative dioxin response elements (DREs). Using EMSA, we demonstrate that the AHR-ARNT heterodimer binds to a specific DRE that overlaps binding sequences for peroxisome proliferator-activated receptor α (PPARα), carbohydrate response element-binding protein (ChREBP), and cAMP response element-binding protein, hepatocyte specific (CREBH). In addition, we reveal that agonist-activated AHR impairs PPARα-, ChREBP-, and CREBH-mediated promoter activity in Hepa-1 cells. Accordingly, agonist treatment in Hepa-1 cells ablates potent ER stress-driven Fgf21 expression, and pre-treatment with AHR antagonist blocks this effect. Finally, we show that pre-treatment of primary human hepatocytes with AHR agonist diminishes PPARα-, glucose-, and ER stress-driven induction of FGF21 expression, indicating the effect is not mouse-specific. Together, our data show that AHR contributes to hepatic energy homeostasis, partly through the regulation of FGF21 expression and signaling. PMID:27226639

  20. A novel 4 S [3H]beta-naphthoflavone-binding protein in liver cytosol of female Sprague-Dawley rats treated with aryl hydrocarbon receptor agonists.

    PubMed Central

    Brauze, D; Malejka-Giganti, D

    2000-01-01

    beta-Naphthoflavone (beta-NF) is a widely used inducer of phase-I and phase-II enzymes controlled by aryl hydrocarbon receptor (AhR). Studies of competitive binding with (3)H-labelled 2,3,7, 8-tetrachlorodibenzo-p-dioxin (TCDD), 3-methylcholanthrene (3-MC) and benzo[a]pyrene (B[a]P) have shown that beta-NF is a high-affinity ligand for AhR and also for polycyclic aromatic hydrocarbon (PAH)-binding protein, both soluble proteins of rat liver in 8 S and 4 S fractions, respectively, of sucrose gradients. This study examined binding of [(3)H]beta-NF to liver cytosolic proteins of female Sprague-Dawley rats. Treatment of rats with beta-NF, 3-MC, TCDD or alpha-naphthoflavone (alpha-NF) increased the specific [(3)H]beta-NF binding to liver cytosol up to 125-fold that of vehicle (corn oil)-treated rats (<100 fmol/mg of protein). Sucrose gradients revealed a large 4 S and a small 8 S peak of radioactivity from [(3)H]beta-NF binding to cytosols of beta-NF-, 3-MC-, TCDD- or alpha-NF-treated rats. Whereas co-incubation with the unlabelled beta-NF eliminated both peaks, co-incubation with 2,3, 7,8-tetrachlorodibenzofuran (TCDF) eliminated only the 8 S peak. The sucrose density gradient from [(3)H]TCDD binding to cytosol of beta-NF- or TCDD-treated rats yielded a small 4 S and a larger 8 S peak; only the latter was abolished by co-incubation with TCDF. Thus, the patterns of sedimentation, distribution and elimination of radioactivity from the 8 S fraction of the liver cytosols from beta-NF-, 3-MC-, TCDD- or alpha-NF-treated rats were characteristic for the AhR, whereas those from the 4 S fraction appeared specific for [(3)H]beta-NF binding. The data indicate that potent AhR agonists, TCDD, 3-MC and beta-NF, and to a lesser extent alpha-NF, a weak AhR agonist, induce a 4 S [(3)H]beta-NF-binding protein in liver cytosol of female rats. alpha-NF, beta-NF and 3-MC were effective competitors (80-85% inhibition) of the [(3)H]beta-NF-specific binding to the beta-NF-, 3 MC- or TCDD

  1. Knockout of the aryl hydrocarbon receptor results in distinct hepatic and renal phenotypes in rats and mice

    SciTech Connect

    Harrill, Joshua A.; Hukkanen, Renee R.; Lawson, Marie; Martin, Greg; Gilger, Brian; Soldatow, Valerie; LeCluyse, Edward L.; Budinsky, Robert A.; Rowlands, J. Craig; Thomas, Russell S.

    2013-10-15

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor which plays a role in the development of multiple tissues and is activated by a large number of ligands, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In order to examine the roles of the AHR in both normal biological development and response to environmental chemicals, an AHR knockout (AHR-KO) rat model was created and compared with an existing AHR-KO mouse. AHR-KO rats harboring either 2-bp or 29-bp deletion mutation in exon 2 of the AHR were created on the Sprague–Dawley genetic background using zinc-finger nuclease (ZFN) technology. Rats harboring either mutation type lacked expression of AHR protein in the liver. AHR-KO rats were also insensitive to thymic involution, increased hepatic weight and the induction of AHR-responsive genes (Cyp1a1, Cyp1a2, Cyp1b1, Ahrr) following acute exposure to 25 μg/kg TCDD. AHR-KO rats had lower basal expression of transcripts for these genes and also accumulated ∼ 30–45-fold less TCDD in the liver at 7 days post-exposure. In untreated animals, AHR-KO mice, but not AHR-KO rats, had alterations in serum analytes indicative of compromised hepatic function, patent ductus venosus of the liver and persistent hyaloid arteries in the eye. AHR-KO rats, but not AHR-KO mice, displayed pathological alterations to the urinary tract: bilateral renal dilation (hydronephrosis), secondary medullary tubular and uroepithelial degenerative changes and bilateral ureter dilation (hydroureter). The present data indicate that the AHR may play significantly different roles in tissue development and homeostasis and toxicity across rodent species. - Highlights: • An AHR knockout rat was generated on a Sprague–Dawley outbred background. • AHR-KO rats lack expression of AHR protein. • AHR-KO rats are insensitive to TCDD-mediated effects. • Data suggests difference in the role of AHR in tissue development of rats and mice. • Abnormalities in vascular

  2. Adaptation of the human aryl hydrocarbon receptor to sense microbiota-derived indoles

    PubMed Central

    Hubbard, Troy D.; Murray, Iain A.; Bisson, William H.; Lahoti, Tejas S.; Gowda, Krishne; Amin, Shantu G.; Patterson, Andrew D.; Perdew, Gary H.

    2015-01-01

    Ligand activation of the aryl hydrocarbon (AHR) has profound effects upon the immunological status of the gastrointestinal tract, establishing and maintaining signaling networks, which facilitate host-microbe homeostasis at the mucosal interface. However, the identity of the ligand(s) responsible for such AHR-mediated activation within the gut remains to be firmly established. Here, we combine in vitro ligand binding, quantitative gene expression, protein-DNA interaction and ligand structure activity analyses together with in silico modeling of the AHR ligand binding domain to identify indole, a microbial tryptophan metabolite, as a human-AHR selective agonist. Human AHR, acting as a host indole receptor may exhibit a unique bimolecular (2:1) binding stoichiometry not observed with typical AHR ligands. Such bimolecular indole-mediated activation of the human AHR within the gastrointestinal tract may provide a foundation for inter-kingdom signaling between the enteric microflora and the immune system to promote commensalism within the gut. PMID:26235394

  3. Knockout of the aryl hydrocarbon receptor results in distinct hepatic and renal phenotypes in rats and mice.

    PubMed

    Harrill, Joshua A; Hukkanen, Renee R; Lawson, Marie; Martin, Greg; Gilger, Brian; Soldatow, Valerie; Lecluyse, Edward L; Budinsky, Robert A; Rowlands, J Craig; Thomas, Russell S

    2013-10-15

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor which plays a role in the development of multiple tissues and is activated by a large number of ligands, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In order to examine the roles of the AHR in both normal biological development and response to environmental chemicals, an AHR knockout (AHR-KO) rat model was created and compared with an existing AHR-KO mouse. AHR-KO rats harboring either 2-bp or 29-bp deletion mutation in exon 2 of the AHR were created on the Sprague-Dawley genetic background using zinc-finger nuclease (ZFN) technology. Rats harboring either mutation type lacked expression of AHR protein in the liver. AHR-KO rats were also insensitive to thymic involution, increased hepatic weight and the induction of AHR-responsive genes (Cyp1a1, Cyp1a2, Cyp1b1, Ahrr) following acute exposure to 25 μg/kg TCDD. AHR-KO rats had lower basal expression of transcripts for these genes and also accumulated ~30-45-fold less TCDD in the liver at 7 days post-exposure. In untreated animals, AHR-KO mice, but not AHR-KO rats, had alterations in serum analytes indicative of compromised hepatic function, patent ductus venosus of the liver and persistent hyaloid arteries in the eye. AHR-KO rats, but not AHR-KO mice, displayed pathological alterations to the urinary tract: bilateral renal dilation (hydronephrosis), secondary medullary tubular and uroepithelial degenerative changes and bilateral ureter dilation (hydroureter). The present data indicate that the AHR may play significantly different roles in tissue development and homeostasis and toxicity across rodent species. PMID:23859880

  4. Xeno-sensing activity of the aryl hydrocarbon receptor in human pluripotent stem cell-derived hepatocyte-like cells

    PubMed Central

    Kim, Hye-Min; Kim, Ji-Woo; Choi, Youngjun; Chun, Hang-Suk; Im, Ilkyun; Han, Yong-Mahn; Song, Chang-Woo; Yoon, Seokjoo; Park, Han-Jin

    2016-01-01

    Although hepatocyte-like cells derived from human pluripotent stem cells (hPSC-HLCs) are considered a promising model for predicting hepatotoxicity, their application has been restricted because of the low activity of drug metabolizing enzymes (DMEs). Here we found that the low expression of xenobiotic receptors (constitutive androstane receptor, CAR; and pregnane X receptor, PXR) contributes to the low activity of DMEs in hPSC-HLCs. Most CAR- and PXR-regulated DMEs and transporters were transcriptionally down-regulated in hPSC-HLC. Transcriptional expression of CAR and PXR was highly repressed in hPSC-HLCs, whereas mRNA levels of aryl hydrocarbon receptor (AHR) were comparable to those of adult liver. Furthermore, ligand-induced transcriptional activation was observed only at AHR in hPSC-HLCs. Bisulfite sequencing analysis demonstrated that promoter hypermethylation of CAR and PXR was associated with diminished transcriptional activity in hPSC-HLCs. Treatment with AHR-selective ligands increased the transcription of AHR-dependent target genes by direct AHR-DNA binding at the xenobiotic response element. In addition, an antagonist of AHR significantly inhibited AHR-dependent target gene expression. Thus, AHR may function intrinsically as a xenosensor as well as a ligand-dependent transcription factor in hPSC-HLCs. Our results indicate that hPSC-HLCs can be used to screen toxic substances related to AHR signaling and to identify potential AHR-targeted therapeutics. PMID:26899675

  5. Simulation studies for wells AH-4bis/AH-17 and AH-18, Ahuachapan Geothermal Field

    SciTech Connect

    Monterrosa, Manuel Ernesto

    1996-01-24

    Well AH-4bis, at the Ahuachapan Geothermal Field is planned to be drilled on the same pad as the former AH-4. A simulation study was carried out for two casing dameters 13 5/8 and 9 5/8” in order to estimate its production and to know its economic feasibility. The simulation results indcate a high probability of production in the range of 7 Mwe, equivalent to 120 kg/s total mass flow rate, 1250 kJ/kg at 6 bar-a for the new well AH-4bis. Well AH- 17 is good producer, during 1991 after ten years of production, the well was shut-in due to silica scaling problems. A wellbore simulation was carried out in order to predict the new production conditions after the work-over, mainly to estimate the water flow rate in order to reduce the silica scaling. The results indicate a very low water flow rate. The match between the simulated and measured production curves after the work-over was successful. The well AH-18 is located at the southern part of the actual bore field. CEL is planning to expand the borefield at this area and it is neccessary to estimate the possible production condtions at that zone. The results indicate a high probabilty of production at that area. The power potential is estimated at 3.5 Mwe per well at WHP 6 bar-a and the wells will not require induction.

  6. In Utero and Lactational Exposure to a Complex Mixture of Polychlorinated Biphenyls: Toxicity in Pups Dependent on the Cyp1a2 and Ahr Genotypes

    PubMed Central

    Curran, Christine P.; Vorhees, Charles V.; Williams, Michael T.; Genter, Mary Beth; Miller, Marian L.; Nebert, Daniel W.

    2011-01-01

    Polychlorinated biphenyls (PCBs) are persistent toxic pollutants occurring as complex mixtures in the environment. Humans are known genetically to have > 60-fold differences in hepatic cytochrome P450 1A2 (CYP1A2) levels and > 12-fold differences in aryl hydrocarbon receptor (AHR) affinity, both of which could affect PCB pharmacokinetics. Thus, we compared Ahrb1_Cyp1a2(+/+) high-affinity AHR wild-type, Ahrd_Cyp1a2(+/+) poor affinity AHR wild-type, Ahrb1_Cyp1a2(−/−) knockout, and Ahrd_Cyp1a2(−/−) knockout mouse lines. We chose a mixture of three coplanar and five noncoplanar PCBs to reproduce that seen in human tissues, breast milk, and the food supply. The mixture was given by gavage to the mother on gestational day 10.5 (GD10.5) and postnatal day 5 (PND5); tissues were collected from pups and mothers at GD11.5, GD18.5, PND6, PND13, and PND28. Ahrb1_Cyp1a2(−/−) pups showed lower weight at birth and slower rate of growth postnatally. Absence of CYP1A2 resulted in significant splenic atrophy at PND13 and PND28. Presence of high-affinity AHR enhanced thymic atrophy and liver hypertrophy in the pups. Concentrations of each congener were analyzed at all time points: maximal noncoplanar congener levels in maternal tissues were observed from GD18 until PND6, whereas the highest levels in pups were found between PND6 and PND28. Coplanar PCB concentrations were generally higher in Ahrd-containing pup tissues; these findings are consistent with earlier studies demonstrating the crucial importance of AHR-mediated inducible CYP1 in the gastrointestinal tract as a means of detoxication of oral planar polycyclic aromatic hydrocarbons. PMID:20961953

  7. AHR2 knockdown prevents PAH-mediated cardiac toxicity and XRE- and ARE-associated gene induction in zebrafish (Danio rerio)

    SciTech Connect

    Van Tiem, Lindsey A.; Di Giulio, Richard T.

    2011-08-01

    Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental contaminants often present in aquatic systems as complex mixtures. Embryonic fish are sensitive to the developmental toxicity of some PAHs, but the exact mechanisms involved in this toxicity are still unknown. This study explored the role of the aryl hydrocarbon receptor (AHR) in the oxidative stress response of zebrafish to the embryotoxicity of select PAHs. Embryos were exposed to two PAHs, benzo[k]fluoranthene (BkF; a strong AHR agonist) and fluoranthene (FL; a cytochrome P4501A (CYP1A) inhibitor), alone and in combination. CYP1A, CYP1B1, CYP1C1, and redox-responsive genes glutathione s-transferase pi 2 (GSTp2), glutathione peroxidase 1 (GPx1), the glutamate-cysteine ligase catalytic subunit (GCLc), MnSOD and CuZnSOD mRNA expression was examined. CYP1 activity was measured via an in vivo ethoxyresorufin-O-deethlyase (EROD) activity assay, and the area of the pericardium was measured as an index of cardiotoxicity. BkF or FL alone caused no deformities whereas BkF + FL resulted in extreme pericardial effusion. BkF induced CYP activity above controls and co-exposure with FL inhibited this activity. BkF induced expression of all three CYPs, GSTp2, and GCLc. BkF + FL caused greater than additive induction of the three CYPs, GSTp2, GPx1, and GCLc but had no effect on MnSOD or CuZnSOD. AHR2 knockdown protected against the cardiac deformities caused by BkF + FL and significantly inhibited the induction of the CYPs, GSTp2, GPx1, and GCLc after BkF + FL compared to non-injected controls. These results further show the protective role of AHR2 knockdown against cardiotoxic PAHs and the role of AHR2 as a mediator of redox-responsive gene induction. - Research Highlights: > Co-exposure of the PAHs BkF and FL causes cardiotoxicity in zebrafish. > BkF and FL co-exposure upregulates certain XRE- and ARE-associated genes. > AHR2 knockdown prevents the deformities caused by BkF and FL co-exposure. > AHR2

  8. Ahr2-dependence of PCB126 effects on the swim bladder in relation to expression of CYP1 and cox-2 genes in developing zebrafish

    SciTech Connect

    Jönsson, Maria E.; Kubota, Akira; Timme-Laragy, Alicia R.; Woodin, Bruce; Stegeman, John J.

    2012-12-01

    The teleost swim bladder is assumed a homolog of the tetrapod lung. Both swim bladder and lung are developmental targets of persistent aryl hydrocarbon receptor (AHR) agonists; in zebrafish (Danio rerio) the swim bladder fails to inflate with exposure to 3,3′,4,4′,5-pentachlorobiphenyl (PCB126). The mechanism for this effect is unknown, but studies have suggested roles of cytochrome P450 1 (CYP1) and cyclooxygenase 2 (Cox-2) in some Ahr-mediated developmental effects in zebrafish. We determined relationships between swim bladder inflation and CYP1 and Cox-2 mRNA expression in PCB126-exposed zebrafish embryos. We also examined effects on β-catenin dependent transcription, histological effects, and Ahr2 dependence of the effect of PCB126 on swim bladder using morpholinos targeting ahr2. One-day-old embryos were exposed to waterborne PCB126 or carrier (DMSO) for 24 h and then held in clean water until day 4, a normal time for swim bladder inflation. The effects of PCB126 were concentration-dependent with EC{sub 50} values of 1.4 to 2.0 nM for induction of the CYP1s, 3.7 and 5.1 nM (or higher) for cox-2a and cox-2b induction, and 2.5 nM for inhibition of swim bladder inflation. Histological defects included a compaction of the developing bladder. Ahr2-morpholino treatment rescued the effect of PCB126 (5 nM) on swim bladder inflation and blocked induction of CYP1A, cox-2a, and cox-2b. With 2 nM PCB126 approximately 30% of eleutheroembryos failed to inflate the swim bladder, but there was no difference in CYP1 or cox-2 mRNA expression between those embryos and embryos showing inflated swim bladder. Our results indicate that PCB126 blocks swim bladder inflation via an Ahr2-mediated mechanism. This mechanism seems independent of CYP1 or cox-2 mRNA induction but may involve abnormal development of swim bladder cells. -- Highlights: ► PCB126 caused cellular changes in the developing swim bladder. ► Swim bladder inflation was not related to expression of CYP1 or cox

  9. The role of endogenous aryl hydrocarbon receptor signaling in cardiovascular physiology

    PubMed Central

    Zhang, Nan

    2011-01-01

    The aryl hydrocarbon receptor (AHR) is an orphan nuclear receptor with a primary function of mediating xenobiotic metabolism through transcriptional activation of Phase I and Phase II drug-metabolizing enzymes. Although no high-affinity physiological activators of AHR have been discovered, the endogenous signaling of the AHR pathway is believed to play an important role in the development and function of the cardiovascular system, based on the observations on ahr gene-deficient mice. The AHR knockout mice develop cardiac hypertrophy, abnormal vascular structure in multiple organs and altered blood pressure depending on their host environment. In this review, the endogenous role of AHR in cardiovascular physiology, including heart function, vascular development and blood pressure regulation has been summarized and discussed. PMID:21814412

  10. The role of endogenous aryl hydrocarbon receptor signaling in cardiovascular physiology.

    PubMed

    Zhang, Nan

    2011-04-01

    The aryl hydrocarbon receptor (AHR) is an orphan nuclear receptor with a primary function of mediating xenobiotic metabolism through transcriptional activation of Phase I and Phase II drug-metabolizing enzymes. Although no high-affinity physiological activators of AHR have been discovered, the endogenous signaling of the AHR pathway is believed to play an important role in the development and function of the cardiovascular system, based on the observations on ahr gene-deficient mice. The AHR knockout mice develop cardiac hypertrophy, abnormal vascular structure in multiple organs and altered blood pressure depending on their host environment. In this review, the endogenous role of AHR in cardiovascular physiology, including heart function, vascular development and blood pressure regulation has been summarized and discussed.

  11. 75 FR 49550 - Fifth Meeting: RTCA Special Committee 219: Attitude and Heading Reference System (AHRS)

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-08-13

    ... Federal Aviation Administration Fifth Meeting: RTCA Special Committee 219: Attitude and Heading Reference...: Notice of RTCA Special Committee 219: Attitude and Heading Reference System (AHRS). SUMMARY: The FAA is... Heading Reference System (AHRS). DATES: The meeting will be held September 14-16, 2010 from 9 a.m. to 5...

  12. Accurate Orientation Estimation Using AHRS under Conditions of Magnetic Distortion

    PubMed Central

    Yadav, Nagesh; Bleakley, Chris

    2014-01-01

    Low cost, compact attitude heading reference systems (AHRS) are now being used to track human body movements in indoor environments by estimation of the 3D orientation of body segments. In many of these systems, heading estimation is achieved by monitoring the strength of the Earth's magnetic field. However, the Earth's magnetic field can be locally distorted due to the proximity of ferrous and/or magnetic objects. Herein, we propose a novel method for accurate 3D orientation estimation using an AHRS, comprised of an accelerometer, gyroscope and magnetometer, under conditions of magnetic field distortion. The system performs online detection and compensation for magnetic disturbances, due to, for example, the presence of ferrous objects. The magnetic distortions are detected by exploiting variations in magnetic dip angle, relative to the gravity vector, and in magnetic strength. We investigate and show the advantages of using both magnetic strength and magnetic dip angle for detecting the presence of magnetic distortions. The correction method is based on a particle filter, which performs the correction using an adaptive cost function and by adapting the variance during particle resampling, so as to place more emphasis on the results of dead reckoning of the gyroscope measurements and less on the magnetometer readings. The proposed method was tested in an indoor environment in the presence of various magnetic distortions and under various accelerations (up to 3 g). In the experiments, the proposed algorithm achieves <2° static peak-to-peak error and <5° dynamic peak-to-peak error, significantly outperforming previous methods. PMID:25347584

  13. Loss of HIF-1α in macrophages attenuates AhR/ARNT-mediated tumorigenesis in a PAH-driven tumor model.

    PubMed

    Henke, Nina; Ferreirós, Nerea; Geisslinger, Gerd; Ding, Martina G; Essler, Silke; Fuhrmann, Dominik C; Geis, Theresa; Namgaladze, Dmitry; Dehne, Nathalie; Brüne, Bernhard

    2016-05-01

    Activation of hypoxia-inducible factor (HIF) and macrophage infiltration of solid tumors independently promote tumor progression. As little is known how myeloid HIF affects tumor development, we injected the polycyclic aromatic hydrocarbon (PAH) and procarcinogen 3-methylcholanthrene (MCA; 100 μg/100 μl) subcutaneously into myeloid-specific Hif-1α and Hif-2α knockout mice (C57BL/6J) to induce fibrosarcomas (n = 16). Deletion of Hif-1α but not Hif-2α in macrophages diminished tumor outgrowth in the MCA-model. While analysis of the tumor initiation phase showed comparable inflammation after MCA-injection, metabolism of MCA was impaired in the absence of Hif-1α. An ex vivo macrophage/fibroblast coculture recapitulated reduced DNA damage after MCA-stimulation in fibroblasts of cocultures with Hif-1α LysM-/- macrophages compared to wild type macrophages. A loss of myeloid Hif-1α decreased RNA levels of arylhydrocarbon receptor (AhR)/arylhydrocarbon receptor nuclear translocator (ARNT) targets such as Cyp1a1 because of reduced Arnt but unchanged Ahr expression. Cocultures using Hif-1α LysM-/- macrophages stimulated with the carcinogen 7,12-dimethylbenz[a]anthracene (DMBA; 2 μg/ml) also attenuated a DNA damage response in fibroblasts, while the DNA damage-inducing metabolite DMBA-trans-3,4-dihydrodiol remained effective in the absence of Hif-1α. In chemical-induced carcinogenesis, HIF-1α in macrophages maintains ARNT expression to facilitate PAH-biotransformation. This implies a metabolic activation of PAHs in stromal cells, i.e. myeloid-derived cells, to be crucial for tumor initiation. PMID:27015123

  14. Loss of HIF-1β in macrophages attenuates AhR/ARNT-mediated tumorigenesis in a PAH-driven tumor model

    PubMed Central

    Henke, Nina; Ferreirós, Nerea; Geisslinger, Gerd; Ding, Martina G.; Essler, Silke; Fuhrmann, Dominik C.; Geis, Theresa; Namgaladze, Dmitry; Dehne, Nathalie; Brüne, Bernhard

    2016-01-01

    Activation of hypoxia-inducible factor (HIF) and macrophage infiltration of solid tumors independently promote tumor progression. As little is known how myeloid HIF affects tumor development, we injected the polycyclic aromatic hydrocarbon (PAH) and procarcinogen 3-methylcholanthrene (MCA; 100 μg/100 μl) subcutaneously into myeloid-specific Hif-1α and Hif-2α knockout mice (C57BL/6J) to induce fibrosarcomas (n = 16). Deletion of Hif-1α but not Hif-2α in macrophages diminished tumor outgrowth in the MCA-model. While analysis of the tumor initiation phase showed comparable inflammation after MCA-injection, metabolism of MCA was impaired in the absence of Hif-1α. An ex vivo macrophage/fibroblast coculture recapitulated reduced DNA damage after MCA-stimulation in fibroblasts of cocultures with Hif-1αLysM−/− macrophages compared to wild type macrophages. A loss of myeloid Hif-1α decreased RNA levels of arylhydrocarbon receptor (AhR)/arylhydrocarbon receptor nuclear translocator (ARNT) targets such as Cyp1a1 because of reduced Arnt but unchanged Ahr expression. Cocultures using Hif-1αLysM−/− macrophages stimulated with the carcinogen 7,12-dimethylbenz[a]anthracene (DMBA; 2 μg/ml) also attenuated a DNA damage response in fibroblasts, while the DNA damage-inducing metabolite DMBA-trans-3,4-dihydrodiol remained effective in the absence of Hif-1α. In chemical-induced carcinogenesis, HIF-1α in macrophages maintains ARNT expression to facilitate PAH-biotransformation. This implies a metabolic activation of PAHs in stromal cells, i.e. myeloid-derived cells, to be crucial for tumor initiation. PMID:27015123

  15. An Aryl Hydrocarbon Receptor-Mediated Amplification Loop That Enforces Cell Migration in ER−/PR−/Her2− Human Breast Cancer Cells

    PubMed Central

    Novikov, Olga; Wang, Zhongyan; Stanford, Elizabeth A.; Parks, Ashley J.; Ramirez-Cardenas, Alejandra; Landesman, Esther; Laklouk, Israa; Sarita-Reyes, Carmen; Gusenleitner, Daniel; Li, Amy; Monti, Stefano; Manteiga, Sara; Lee, Kyongbum

    2016-01-01

    The endogenous ligand-activated aryl hydrocarbon receptor (AHR) plays an important role in numerous biologic processes. As the known number of AHR-mediated processes grows, so too does the importance of determining what endogenous AHR ligands are produced, how their production is regulated, and what biologic consequences ensue. Consequently, our studies were designed primarily to determine whether ER−/PR−/Her2− breast cancer cells have the potential to produce endogenous AHR ligands and, if so, how production of these ligands is controlled. We postulated that: 1) malignant cells produce tryptophan-derived AHR ligand(s) through the kynurenine pathway; 2) these metabolites have the potential to drive AHR-dependent breast cancer migration; 3) the AHR controls expression of a rate-limiting kynurenine pathway enzyme(s) in a closed amplification loop; and 4) environmental AHR ligands mimic the effects of endogenous ligands. Data presented in this work indicate that primary human breast cancers, and their metastases, express high levels of AHR and tryptophan-2,3-dioxygenase (TDO); representative ER−/PR−/Her2− cell lines express TDO and produce sufficient intracellular kynurenine and xanthurenic acid concentrations to chronically activate the AHR. TDO overexpression, or excess kynurenine or xanthurenic acid, accelerates migration in an AHR-dependent fashion. Environmental AHR ligands 2,3,7,8-tetrachlorodibenzo[p]dioxin and benzo[a]pyrene mimic this effect. AHR knockdown or inhibition significantly reduces TDO2 expression. These studies identify, for the first time, a positive amplification loop in which AHR-dependent TDO2 expression contributes to endogenous AHR ligand production. The net biologic effect of AHR activation by endogenous ligands, which can be mimicked by environmental ligands, is an increase in tumor cell migration, a measure of tumor aggressiveness. PMID:27573671

  16. Identification of aryl hydrocarbon receptor binding targets in mouse hepatic tissue treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin

    SciTech Connect

    Lo, Raymond; Celius, Trine; Forgacs, Agnes L.; Dere, Edward; MacPherson, Laura; Harper, Patricia; Zacharewski, Timothy; Matthews, Jason

    2011-11-15

    Genome-wide, promoter-focused ChIP-chip analysis of hepatic aryl hydrocarbon receptor (AHR) binding sites was conducted in 8-week old female C57BL/6 treated with 30 {mu}g/kg/body weight 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for 2 h and 24 h. These studies identified 1642 and 508 AHR-bound regions at 2 h and 24 h, respectively. A total of 430 AHR-bound regions were common between the two time points, corresponding to 403 unique genes. Comparison with previous AHR ChIP-chip studies in mouse hepatoma cells revealed that only 62 of the putative target genes overlapped with the 2 h AHR-bound regions in vivo. Transcription factor binding site analysis revealed an over-representation of aryl hydrocarbon response elements (AHREs) in AHR-bound regions with 53% (2 h) and 68% (24 h) of them containing at least one AHRE. In addition to AHREs, E2f-Myc activator motifs previously implicated in AHR function, as well as a number of other motifs, including Sp1, nuclear receptor subfamily 2 factor, and early growth response factor motifs were also identified. Expression microarray studies identified 133 unique genes differentially regulated after 4 h treatment with TCDD. Of which, 39 were identified as AHR-bound genes at 2 h. Ingenuity Pathway Analysis on the 39 AHR-bound TCDD responsive genes identified potential perturbation in biological processes such as lipid metabolism, drug metabolism, and endocrine system development as a result of TCDD-mediated AHR activation. Our findings identify direct AHR target genes in vivo, highlight in vitro and in vivo differences in AHR signaling and show that AHR recruitment does not necessarily result in changes in target gene expression. -- Highlights: Black-Right-Pointing-Pointer ChIP-chip analysis of hepatic AHR binding after 2 h and 24 h of TCDD. Black-Right-Pointing-Pointer We identified 1642 and 508 AHR-bound regions at 2 h and 24 h. Black-Right-Pointing-Pointer 430 regions were common to both time points and highly enriched with

  17. Expression and function of ryanodine receptor related pathways in PCB tolerant Atlantic killifish (Fundulus heteroclitus) from New Bedford Harbor, MA, USA

    PubMed Central

    Fritsch, Erika B.; Stegeman, John J.; Goldstone, Jared V.; Nacci, Diane E.; Champlin, Denise; Jayaraman, Saro; Connon, Richard E.; Pessah, Isaac N.

    2015-01-01

    Atlantic killifish (Fundulus heteroclitus) thrive in New Bedford Harbor (NBH), MA, highly contaminated with polychlorinated biphenyls (PCBs). Resident killifish have evolved tolerance to dioxin-like (DL) PCBs, whose toxic effects through the aryl hydrocarbon receptor (AhR) are well studied. In NBH, non-dioxin like PCBs (NDL PCBs), which lack activity toward the AhR, vastly exceed levels of DL congeners yet how killifish counter NDL toxic effects has not been explored. In mammals and fish, NDL PCBs are potent activators of ryanodine receptors (RyR), Ca2+ release channels necessary for a vast array of physiological processes. In the current study we compared the expression and function of RyR related pathways in NBH killifish with killifish from the reference site at Scorton Creek (SC, MA). Relative to the SC fish, adults from NBH displayed increased levels of skeletal muscle RyR1 protein, and increased levels of FK506-binding protein 12 kDa (FKBP12), an accessory protein essential for NDL PCB-triggered changes in RyR channel function. In accordance with increased RyR1 levels, NBH killifish displayed increased maximal ligand binding, increased maximal response to Ca2+ activation and increased maximal response to activation by the NDL PCB congener PCB 95. Compared to SC, NBH embryos and larvae had increased levels of mtor and ryr2 transcripts at multiple stages of development, and generations, while levels of serca2 were decreased at 9 days post-fertilization in the F1 and F2 generations. These findings suggest that there are compensatory and heritable changes in RyR mediated Ca2+ signaling proteins or potential signaling partners in NBH killifish. PMID:25546006

  18. Novel role of hnRNP-A2/B1 in modulating aryl hydrocarbon receptor ligand sensitivity.

    PubMed

    Cho, See-Wun; Suzuki, Ken-ichi; Miura, Yoshiaki; Miyazaki, Tatsuhiko; Nose, Masato; Iwata, Hisato; Kim, Eun-Young

    2015-11-01

    The aryl hydrocarbon receptor (AHR) is responsible for susceptibility to its ligand-dependent responses. However, the effect of non-AHR factors is less clear. To explore the non-AHR factors, we used two mouse strains with different AHR genetic variants, namely C3H/lpr and MRL/lpr strains with Ala and Val as the 375th amino acid residue, respectively. To assess the contribution of AHR alone, COS-7 cells transiently expressing AHR from each strain were treated with 6-formylindolo[3,2-b]carbazole (FICZ) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and xenobiotic-responsive element (XRE)-driven reporter gene activities were measured. FICZ-EC50 values for the C3H/lpr and MRL/lpr AHR-mediated transactivation were 0.023 and 0.046 nM, respectively, indicating a similar susceptibility in both AHR genotypes. In contrast, C3H/lpr AHR was fourfold more sensitive to TCDD than MRL/lpr AHR. By a pull-down assay using a XRE-containing PCR product as bait and the hepatic nuclear extracts of both FICZ-treated mouse strains, we identified two interacting proteins as heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP-A2) and its splicing variant (hnRNP-A2b). Immunoprecipitation assays demonstrated the AHR interaction with hnRNP-A2/B1. When hnRNP-A2 was co-expressed with the MRL/lpr or C3H/lpr AHR in COS-7, FICZ treatment decreased EC50 to about threefold in both AHR genotypes, compared with EC50 in AHR alone. Similarly, hnRNP-A2b co-expression also lowered the FICZ-EC50 values. In TCDD-treated COS-7, responses depended on the AHR genotype; while no change in TCDD-EC50 was observed for C3H/lpr AHR when hnRNP-A2 was co-expressed, the value was reduced to nearly tenfold for MRL/lpr AHR. Co-transfection with hnRNP-A2b attenuated the AHR sensitivity to TCDD. In conclusion, the hnRNP-A2/B1 interacting with AHR may be a modulator of the AHR ligand sensitivity.

  19. Ahr2-dependence of PCB126 effects on the swim bladder in relation to expression of CYP1 and cox-2 genes in developing zebrafish.

    PubMed

    Jönsson, Maria E; Kubota, Akira; Timme-Laragy, Alicia R; Woodin, Bruce; Stegeman, John J

    2012-12-01

    The teleost swim bladder is assumed a homolog of the tetrapod lung. Both swim bladder and lung are developmental targets of persistent aryl hydrocarbon receptor (AHR(2)) agonists; in zebrafish (Danio rerio) the swim bladder fails to inflate with exposure to 3,3',4,4',5-pentachlorobiphenyl (PCB126). The mechanism for this effect is unknown, but studies have suggested roles of cytochrome P450 1 (CYP1) and cyclooxygenase 2 (Cox-2) in some Ahr-mediated developmental effects in zebrafish. We determined relationships between swim bladder inflation and CYP1 and Cox-2 mRNA expression in PCB126-exposed zebrafish embryos. We also examined effects on β-catenin dependent transcription, histological effects, and Ahr2 dependence of the effect of PCB126 on swim bladder using morpholinos targeting ahr2. One-day-old embryos were exposed to waterborne PCB126 or carrier (DMSO) for 24h and then held in clean water until day 4, a normal time for swim bladder inflation. The effects of PCB126 were concentration-dependent with EC(50) values of 1.4 to 2.0 nM for induction of the CYP1s, 3.7 and 5.1 nM (or higher) for cox-2a and cox-2b induction, and 2.5 nM for inhibition of swim bladder inflation. Histological defects included a compaction of the developing bladder. Ahr2-morpholino treatment rescued the effect of PCB126 (5 nM) on swim bladder inflation and blocked induction of CYP1A, cox-2a, and cox-2b. With 2nM PCB126 approximately 30% of eleutheroembryos(3) failed to inflate the swim bladder, but there was no difference in CYP1 or cox-2 mRNA expression between those embryos and embryos showing inflated swim bladder. Our results indicate that PCB126 blocks swim bladder inflation via an Ahr2-mediated mechanism. This mechanism seems independent of CYP1 or cox-2 mRNA induction but may involve abnormal development of swim bladder cells.

  20. Pluripotency factors and Polycomb Group proteins repress aryl hydrocarbon receptor expression in murine embryonic stem cells.

    PubMed

    Ko, Chia-I; Wang, Qin; Fan, Yunxia; Xia, Ying; Puga, Alvaro

    2014-01-01

    The aryl hydrocarbon receptor (AHR) is a transcription factor and environmental sensor that regulates expression of genes involved in drug-metabolism and cell cycle regulation. Chromatin immunoprecipitation analyses, Ahr ablation in mice and studies with orthologous genes in invertebrates suggest that AHR may also play a significant role in embryonic development. To address this hypothesis, we studied the regulation of Ahr expression in mouse embryonic stem cells and their differentiated progeny. In ES cells, interactions between OCT3/4, NANOG, SOX2 and Polycomb Group proteins at the Ahr promoter repress AHR expression, which can also be repressed by ectopic expression of reprogramming factors in hepatoma cells. In ES cells, unproductive RNA polymerase II binds at the Ahr transcription start site and drives the synthesis of short abortive transcripts. Activation of Ahr expression during differentiation follows from reversal of repressive marks in Ahr promoter chromatin, release of pluripotency factors and PcG proteins, binding of Sp factors, establishment of histone marks of open chromatin, and engagement of active RNAPII to drive full-length RNA transcript elongation. Our results suggest that reversible Ahr repression in ES cells holds the gene poised for expression and allows for a quick switch to activation during embryonic development.

  1. Using Nuclear Receptor Activity to Stratify Hepatocarcinogens

    PubMed Central

    Shah, Imran; Houck, Keith; Judson, Richard S.; Kavlock, Robert J.; Martin, Matthew T.; Reif, David M.; Wambaugh, John; Dix, David J.

    2011-01-01

    Background Nuclear receptors (NR) are a superfamily of ligand-activated transcription factors that control a range of cellular processes. Persistent stimulation of some NR is a non-genotoxic mechanism of rodent liver cancer with unclear relevance to humans. Here we report on a systematic analysis of new in vitro human NR activity data on 309 environmental chemicals in relationship to their liver cancer-related chronic outcomes in rodents. Results The effects of 309 environmental chemicals on human constitutive androstane receptors (CAR/NR1I3), pregnane X receptor (PXR/NR1I2), aryl hydrocarbon receptor (AhR), peroxisome proliferator-activated receptors (PPAR/NR1C), liver X receptors (LXR/NR1H), retinoic X receptors (RXR/NR2B) and steroid receptors (SR/NR3) were determined using in vitro data. Hepatic histopathology, observed in rodents after two years of chronic treatment for 171 of the 309 chemicals, was summarized by a cancer lesion progression grade. Chemicals that caused proliferative liver lesions in both rat and mouse were generally more active for the human receptors, relative to the compounds that only affected one rodent species, and these changes were significant for PPAR (p0.001), PXR (p0.01) and CAR (p0.05). Though most chemicals exhibited receptor promiscuity, multivariate analysis clustered them into relatively few NR activity combinations. The human NR activity pattern of chemicals weakly associated with the severity of rodent liver cancer lesion progression (p0.05). Conclusions The rodent carcinogens had higher in vitro potency for human NR relative to non-carcinogens. Structurally diverse chemicals with similar NR promiscuity patterns weakly associated with the severity of rodent liver cancer progression. While these results do not prove the role of NR activation in human liver cancer, they do have implications for nuclear receptor chemical biology and provide insights into putative toxicity pathways. More importantly, these findings suggest the

  2. Genome-wide RNAi high-throughput screen identifies proteins necessary for the AHR-dependent induction of CYP1A1 by 2,3,7,8-tetrachlorodibenzo-p-dioxin.

    PubMed

    Solaimani, Parrisa; Damoiseaux, Robert; Hankinson, Oliver

    2013-11-01

    The aryl hydrocarbon receptor (AHR) has a plethora of physiological roles, and upon dysregulation, carcinogenesis can occur. One target gene of AHR encodes the xenobiotic and drug-metabolizing enzyme CYP1A1, which is inducible by the environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) via the AHR. An siRNA library targeted against over 5600 gene candidates in the druggable genome was used to transfect mouse Hepa-1 cells, which were then treated with TCDD, and subsequently assayed for CYP1A1-dependent ethoxyresorufin-o-deethylase (EROD) activity. Following redundant siRNA activity (RSA) statistical analysis, we identified 93 hits that reduced EROD activity with a p value ≤ .005 and substantiated 39 of these as positive hits in a secondary screening using endoribonuclease-prepared siRNAs (esiRNAs). Twelve of the corresponding gene products were subsequently confirmed to be necessary for the induction of CYP1A1 messenger RNA by TCDD. None of the candidates were deficient in aryl hydrocarbon nuclear translocator expression. However 6 gene products including UBE2i, RAB40C, CRYGD, DCTN4, RBM5, and RAD50 are required for the expression of AHR as well as for induction of CYP1A1. We also found 2 gene products, ARMC8 and TCF20, to be required for the induction of CYP1A1, but our data are ambiguous as to whether they are required for the expression of AHR. In contrast, SIN3A, PDC, TMEM5, and CD9 are not required for AHR expression but are required for the induction of CYP1A1, implicating a direct role in Cyp1a1 transcription. Our methods, although applied to Cyp1a1, could be modified for identifying proteins that regulate other inducible genes. PMID:23997114

  3. Genome-Wide RNAi High-Throughput Screen Identifies Proteins Necessary for the AHR-Dependent Induction of CYP1A1 by 2,3,7,8-Tetrachlorodibenzo-p-dioxin

    PubMed Central

    Hankinson, Oliver

    2013-01-01

    The aryl hydrocarbon receptor (AHR) has a plethora of physiological roles, and upon dysregulation, carcinogenesis can occur. One target gene of AHR encodes the xenobiotic and drug-metabolizing enzyme CYP1A1, which is inducible by the environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) via the AHR. An siRNA library targeted against over 5600 gene candidates in the druggable genome was used to transfect mouse Hepa-1 cells, which were then treated with TCDD, and subsequently assayed for CYP1A1-dependent ethoxyresorufin-o-deethylase (EROD) activity. Following redundant siRNA activity (RSA) statistical analysis, we identified 93 hits that reduced EROD activity with a p value ≤ .005 and substantiated 39 of these as positive hits in a secondary screening using endoribonuclease-prepared siRNAs (esiRNAs). Twelve of the corresponding gene products were subsequently confirmed to be necessary for the induction of CYP1A1 messenger RNA by TCDD. None of the candidates were deficient in aryl hydrocarbon nuclear translocator expression. However 6 gene products including UBE2i, RAB40C, CRYGD, DCTN4, RBM5, and RAD50 are required for the expression of AHR as well as for induction of CYP1A1. We also found 2 gene products, ARMC8 and TCF20, to be required for the induction of CYP1A1, but our data are ambiguous as to whether they are required for the expression of AHR. In contrast, SIN3A, PDC, TMEM5, and CD9 are not required for AHR expression but are required for the induction of CYP1A1, implicating a direct role in Cyp1a1 transcription. Our methods, although applied to Cyp1a1, could be modified for identifying proteins that regulate other inducible genes. PMID:23997114

  4. RESULTS FROM THE AHS PESTICIDE EXPOSURE STUDY

    EPA Science Inventory

    The Agricultural Health Study/Pesticide Exposure Study (AHS/PES) measured exposures resulting from agricultural use of 2,4-D and chlorpyrifos for a subset of applicators in the AHS cohort. Through on-farm measurements and observations, data collected in the exposure study will...

  5. Impaired glucose and lipid metabolism in ageing aryl hydrocarbon receptor deficient mice.

    PubMed

    Biljes, Daniel; Hammerschmidt-Kamper, Christiane; Kadow, Stephanie; Diel, Patrick; Weigt, Carmen; Burkart, Volker; Esser, Charlotte

    2015-01-01

    Disturbed homeostasis of glucose and lipid metabolism are dominant features of the so-called metabolic syndrome (MetS) and can increase the risk for the development of type 2 diabetes (T2D), a severe metabolic disease. T2D prevalence increases with age. The aryl hydrocarbon receptor (AHR) is a sensor of small molecules including dietary components. AHR has been identified as potential regulator of glucose homeostasis and lipid metabolism. Epidemiologically, exposure to xenobiotic AHR ligands such as polycyclic aromatic hydrocarbons is linked to T2D. We assess here the potential role of the AHR in disturbances of glucose and lipid metabolism in young (age 2-5 months) and old (age > 1,5 years) AHR-deficient (AHR KO) mice. Fasted young wildtype (WT) and AHR-KO mice displayed similar blood glucose kinetics after challenge with intra-peritoneal glucose injection. However, old AHR-KO mice showed lower tolerance than WT to i.p. administered glucose, i.e. glucose levels rose higher and returned more slowly to normal levels. Old mice had overall higher insulin levels than young mice, and old AHR-KO had a somewhat disturbed insulin kinetic in the serum after glucose challenge. Surprisingly, young AHR-KO mice had significantly lower triglycerides, cholesterol, high density lipoprotein values than WT, i.e., a dyslipidemic profile. With ageing, AHR-KO and WT mice did not differ in these lipid levels, except for slightly reduced levels of triglycerides and cholesterol. In conclusion, our findings in AHR KO mice suggest that AHR expression is relevant for the maintenance of glucose and lipid homeostasis in old mice.

  6. Impaired glucose and lipid metabolism in ageing aryl hydrocarbon receptor deficient mice

    PubMed Central

    Biljes, Daniel; Hammerschmidt-Kamper, Christiane; Kadow, Stephanie; Diel, Patrick; Weigt, Carmen; Burkart, Volker; Esser, Charlotte

    2015-01-01

    Disturbed homeostasis of glucose and lipid metabolism are dominant features of the so-called metabolic syndrome (MetS) and can increase the risk for the development of type 2 diabetes (T2D), a severe metabolic disease. T2D prevalence increases with age. The aryl hydrocarbon receptor (AHR) is a sensor of small molecules including dietary components. AHR has been identified as potential regulator of glucose homeostasis and lipid metabolism. Epidemiologically, exposure to xenobiotic AHR ligands such as polycyclic aromatic hydrocarbons is linked to T2D. We assess here the potential role of the AHR in disturbances of glucose and lipid metabolism in young (age 2-5 months) and old (age > 1,5 years) AHR-deficient (AHR KO) mice. Fasted young wildtype (WT) and AHR-KO mice displayed similar blood glucose kinetics after challenge with intra-peritoneal glucose injection. However, old AHR-KO mice showed lower tolerance than WT to i.p. administered glucose, i.e. glucose levels rose higher and returned more slowly to normal levels. Old mice had overall higher insulin levels than young mice, and old AHR-KO had a somewhat disturbed insulin kinetic in the serum after glucose challenge. Surprisingly, young AHR-KO mice had significantly lower triglycerides, cholesterol, high density lipoprotein values than WT, i.e., a dyslipidemic profile. With ageing, AHR-KO and WT mice did not differ in these lipid levels, except for slightly reduced levels of triglycerides and cholesterol. In conclusion, our findings in AHR KO mice suggest that AHR expression is relevant for the maintenance of glucose and lipid homeostasis in old mice. PMID:26664351

  7. AHR Over-Expression in Papillary Thyroid Carcinoma: Clinical and Molecular Assessments in a Series of Italian Acromegalic Patients with a Long-Term Follow-Up

    PubMed Central

    Mian, Caterina; Ceccato, Filippo; Barollo, Susi; Watutantrige-Fernando, Sara; Albiger, Nora; Regazzo, Daniela; de Lazzari, Paola; Pennelli, Gianmaria; Rotondi, Sandra; Nacamulli, Davide; Pelizzo, Maria Rosa; Jaffrain-Rea, Marie-Lise; Grimaldi, Franco; Occhi, Gianluca; Scaroni, Carla

    2014-01-01

    Aim Acromegaly reportedly carries an increased risk of malignant and benign thyroid tumors, with a prevalence of thyroid cancer of around 3–7%. Germline mutations in the aryl-hydrocarbon receptor (AHR) interacting protein (AIP) have been identified in familial forms of acromegaly. The molecular and endocrine relationships between follicular thyroid growth and GH-secreting pituitary adenoma have yet to be fully established. Our aim was to study the prevalence of differentiated thyroid cancer (DTC) in acromegaly, focusing on the role of genetic events responsible for the onset of thyroid cancer. Methods Germline mutations in the AIP gene were assessed in all patients; BRAF and H-N-K RAS status was analyzed by direct sequencing in thyroid specimens, while immunohistochemistry was used to analyze the protein expression of AIP and AHR. A set of PTCs unrelated to acromegaly was also studied. Results 12 DTCs (10 papillary and 2 follicular carcinomas) were identified in a cohort of 113 acromegalic patients. No differences in GH/IGF-1 levels or disease activity emerged between patients with and without DTC, but the former were older and more often female. BRAF V600E was found in 70% of the papillary thyroid cancers; there were no RAS mutations. AIP protein expression was similar in neoplastic and normal cells, while AHR protein was expressed more in PTCs carrying BRAF mutations than in normal tissue, irrespective of acromegaly status. Conclusions The prevalence of DTC in acromegaly is around 11% and endocrinologists should bear this in mind, especially when examining elderly female patients with uninodular goiter. The DTC risk does not seem to correlate with GH/IGF-1 levels, while it may be associated with BRAF mutations and AHR over-expression. Genetic or epigenetic events probably play a part in promoting thyroid carcinoma. PMID:25019383

  8. 2-(4-Amino-3-methylphenyl)-5-fluorobenzothiazole is a ligand and shows species-specific partial agonism of the aryl hydrocarbon receptor

    SciTech Connect

    Bazzi, Rana; Bradshaw, Tracey D.; Rowlands, J. Craig; Stevens, Malcolm F.G.; Bell, David R.

    2009-05-15

    2-(4-Amino-3-methylphenyl)-5-fluorobenzothiazole (5F 203) and related compounds are a series of anti-cancer candidate pharmaceuticals, that have been shown to activate the AhR. We show that these compounds are high-affinity ligands for the rat AhR, but a quantitative assay for their ability to induce CYP1A1 RNA in H4IIEC3 cells, a measure of activation of the AhR, showed a poor relationship between affinity for the AhR and ability to induce CYP1A1 RNA. 5F 203, an agonist with low potency, was able to antagonise the induction of CYP1A1 RNA by TCDD, while IH 445, a potent agonist, did not antagonise the induction of CYP1A1 RNA by TCDD, and Schild analysis confirmed 5F 203 to be a potent antagonist of the induction of CYP1A1 RNA by TCDD in H4IIEC3 cells. In contrast, several benzothiazoles show potent induction of CYP1A1 RNA in human MCF-7 cells, and 5F 203 is unable to detectably antagonise the induction of CYP1A1 RNA in MCF-7 cells, showing a species difference in antagonism. Evaluation of the anti-proliferative activity of benzothiazoles showed that the ability to agonise the AhR correlated with growth inhibition both in H4IIEC3 cells for a variety of benzothiazoles, and between H4IIEC3 and MCF-7 cells for 5F 203, suggesting an important role of agonism of the AhR in the anti-proliferative activity of benzothiazoles.

  9. Insulin like growth factor 2 regulation of aryl hydrocarbon receptor in MCF-7 breast cancer cells

    SciTech Connect

    Tomblin, Justin K.; Salisbury, Travis B.

    2014-01-17

    Highlights: •IGF-2 stimulates concurrent increases in AHR and CCND1 expression. •IGF-2 promotes the binding of AHR to the endogenous cyclin D1 promoter. •AHR knockdown inhibits IGF-2 stimulated increases in CCND1 mRNA and protein. •AHR knockdown inhibits IGF-2 stimulated increases in MCF-7 proliferation. -- Abstract: Insulin like growth factor (IGF)-1 and IGF-2 stimulate normal growth, development and breast cancer cell proliferation. Cyclin D1 (CCND1) promotes cell cycle by inhibiting retinoblastoma protein (RB1). The aryl hydrocarbon receptor (AHR) is a major xenobiotic receptor that also regulates cell cycle. The purpose of this study was to investigate whether IGF-2 promotes MCF-7 breast cancer proliferation by inducing AHR. Western blot and quantitative real time PCR (Q-PCR) analysis revealed that IGF-2 induced an approximately 2-fold increase (P < .001) in the expression of AHR and CCND1. Chromatin immunoprecipitation (ChIP), followed by Q-PCR indicated that IGF-2 promoted (P < .001) a 7-fold increase in AHR binding on the CCND1 promoter. AHR knockdown significantly (P < .001) inhibited IGF-2 stimulated increases in CCND1 mRNA and protein. AHR knockdown cells were less (P < .001) responsive to the proliferative effects of IGF-2 than control cells. Collectively, our findings have revealed a new regulatory mechanism by which IGF-2 induction of AHR promotes the expression of CCND1 and the proliferation of MCF-7 cells. This previously uncharacterized pathway could be important for the proliferation of IGF responsive cancer cells that also express AHR.

  10. AhV_aPA-induced vasoconstriction involves the IP₃Rs-mediated Ca²⁺ releasing.

    PubMed

    Zeng, Fuxing; Zou, Zhisong; Niu, Liwen; Li, Xu; Teng, Maikun

    2013-08-01

    AhV_aPA, the acidic PLA₂ purified from Agkistrodon halys pallas venom, was previously reported to possess a strong enzymatic activity and can remarkably induce a further contractile response on the 60 mM K⁺-induced contraction with an EC₅₀ in 369 nM on mouse thoracic aorta rings. In the present study, we found that the p-bromo-phenacyl-bromide (pBPB), which can completely inhibit the enzymatic activity of AhV_aPA, did not significantly reduce the contractile response on vessel rings induced by AhV_aPA, indicating that the vasoconstrictor effects of AhV_aPA are independent of the enzymatic activity. The inhibitor experiments showed that the contractile response induced by AhV_aPA is mainly attributed to the Ca²⁺ releasing from Ca²⁺ store, especially sarcoplasmic reticulum (SR). Detailed studies showed that the Ca²⁺ release from SR is related to the activation of inositol trisphosphate receptors (IP₃Rs) rather than ryanodine receptors (RyRs). Furthermore, the vasoconstrictor effect could be strongly reduced by pre-incubation with heparin, indicating that the basic amino acid residues on the surface of AhV_aPA may be involved in the interaction between AhV_aPA and the molecular receptors. These findings offer new insights into the functions of snake PLA₂ and provide a novel pathogenesis of A. halys pallas venom.

  11. An Aryl Hydrocarbon Receptor from the Salamander Ambystoma mexicanum Exhibits Low Sensitivity to 2,3,7,8-Tetrachlorodibenzo-p-dioxin.

    PubMed

    Shoots, Jenny; Fraccalvieri, Domenico; Franks, Diana G; Denison, Michael S; Hahn, Mark E; Bonati, Laura; Powell, Wade H

    2015-06-01

    Structural features of the aryl hydrocarbon receptor (AHR) can underlie species- and population-specific differences in its affinity for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). These differences often explain variations in TCDD toxicity. Frogs are relatively insensitive to dioxin, and Xenopus AHRs bind TCDD with low affinity. Weak TCDD binding results from the combination of three residues in the ligand-binding domain: A354 and A370, and N325. Here we sought to determine whether this mechanism of weak TCDD binding is shared by other amphibian AHRs. We isolated an AHR cDNA from the Mexican axolotl (Ambystoma mexicanum). The encoded polypeptide contains identical residues at positions that confer low TCDD affinity to X. laevis AHRs (A364, A380, and N335), and homology modeling predicts they protrude into the binding cavity. Axolotl AHR bound one-tenth the TCDD of mouse AHR in velocity sedimentation analysis, and in transactivation assays, the EC50 for TCDD was 23 nM, similar to X. laevis AHR1β (27 nM) and greater than AHR containing the mouse ligand-binding domain (0.08 nM). Sequence, modeled structure, and function indicate that axolotl AHR binds TCDD weakly, predicting that A. mexicanum lacks sensitivity toTCDD toxicity. We hypothesize that this characteristic of axolotl and Xenopus AHRs arose in a common ancestor of the Caudata and Anura.

  12. An Aryl Hydrocarbon Receptor from the Salamander Ambystoma mexicanum Exhibits Low Sensitivity to 2,3,7,8-Tetrachlorodibenzo-p-dioxin.

    PubMed

    Shoots, Jenny; Fraccalvieri, Domenico; Franks, Diana G; Denison, Michael S; Hahn, Mark E; Bonati, Laura; Powell, Wade H

    2015-06-01

    Structural features of the aryl hydrocarbon receptor (AHR) can underlie species- and population-specific differences in its affinity for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). These differences often explain variations in TCDD toxicity. Frogs are relatively insensitive to dioxin, and Xenopus AHRs bind TCDD with low affinity. Weak TCDD binding results from the combination of three residues in the ligand-binding domain: A354 and A370, and N325. Here we sought to determine whether this mechanism of weak TCDD binding is shared by other amphibian AHRs. We isolated an AHR cDNA from the Mexican axolotl (Ambystoma mexicanum). The encoded polypeptide contains identical residues at positions that confer low TCDD affinity to X. laevis AHRs (A364, A380, and N335), and homology modeling predicts they protrude into the binding cavity. Axolotl AHR bound one-tenth the TCDD of mouse AHR in velocity sedimentation analysis, and in transactivation assays, the EC50 for TCDD was 23 nM, similar to X. laevis AHR1β (27 nM) and greater than AHR containing the mouse ligand-binding domain (0.08 nM). Sequence, modeled structure, and function indicate that axolotl AHR binds TCDD weakly, predicting that A. mexicanum lacks sensitivity toTCDD toxicity. We hypothesize that this characteristic of axolotl and Xenopus AHRs arose in a common ancestor of the Caudata and Anura. PMID:25941739

  13. Two years after the Hebei Spirit oil spill: residual crude-derived hydrocarbons and potential AhR-mediated activities in coastal sediments.

    PubMed

    Hong, Seongjin; Khim, Jong Seong; Ryu, Jongseong; Park, Jinsoon; Song, Sung Joon; Kwon, Bong-Oh; Choi, Kyungho; Ji, Kyunghee; Seo, Jihyun; Lee, Sangwoo; Park, Jeongim; Lee, Woojin; Choi, Yeyong; Lee, Kyu Tae; Kim, Chan-Kook; Shim, Won Joon; Naile, Jonathan E; Giesy, John P

    2012-02-01

    The Hebei Spirit oil spill occurred in December 2007 approximately 10 km off the coast of Taean, South Korea, on the Yellow Sea. However, the exposure and potential effects remain largely unknown. A total of 50 surface and subsurface sediment samples were collected from 22 sampling locations at the spill site in order to determine the concentration, distribution, composition of residual crudes, and to evaluate the potential ecological risk after two years of oil exposure. Samples were extracted and analyzed for 16 polycyclic aromatic hydrocarbons (PAHs), 20 alkyl-PAHs, 15 aliphatic hydrocarbons, and total petroleum hydrocarbons using GC-MSD. AhR-mediated activity associated with organic sediment extracts was screened using the H4IIE-luc cell bioassay. The response of the benthic invertebrate community was assessed by mapping the macrobenthic fauna. Elevated concentrations of residual crudes from the oil spill were primarily found in muddy bottoms, particularly in subsurface layers. In general, the bioassay results were consistent with the chemistry data in a dose-dependent manner, although the mass-balance was incomplete. More weathered samples containing greater fractions of alkylated PAHs exhibited greater AhR activity, due to the occurrence of recalcitrant AhR agonists present in residual oils. The macrobenthic population distribution exhibits signs of species-specific tolerances and/or recolonization of certain species such as Batillaria during weathering periods. Although the Hebei Spirit oil spill was a severe oil exposure, it appears the site is recovering two years later.

  14. Characterization testing of a 40 Ahr bipolar nickel hydrogen battery

    NASA Technical Reports Server (NTRS)

    Brewer, Jeffrey C.; Manzo, Michelle A.; Gahn, Randall F.

    1989-01-01

    In a continuing effort to develop NiH2 bipolar technology to a point where it can be used efficiently in space flight, testing of a second 40 Ahr, 10-cell bipolar battery has begun. This battery has undergone extensive characterization testing to determine the effects of such operating parameters as charge and discharge rates, temperature, and pressure. The fundamental design of this actively cooled bipolar battery is the same as the first battery. Most of the individual components, however, are from different manufacturers. Different testing procedures as well as certain unique battery characteristics make it difficult to directly compare the two sets of results. In general, the performance of this battery throughout characterization produced expected results. The main differences seen between the first and second batteries occurred during the high-rate discharge portion of the test matrix. The first battery also had poor high-rate discharge results, although better than those of the second battery. Minor changes were made to the battery frame design used for the first battery in an attempt to allow better gas access to the reaction sites for the second build and hopefully improve performance. The changes, however, did not improve the performance of the second battery and could have possibly contributed to the poorer performance that was observed. There are other component differences that could have contributed to the poorer performance of the second battery. The H2 electrode in the second battery was constructed with a Goretex backing which could have limited the high-rate current flow. The gas screen in the second battery had a larger mesh which again could have limited the high-rate current flow. Small scale 2 x 2 batteries are being tested to evaluate the effects of the component variations.

  15. In vitro and in silico derived relative effect potencies of ah-receptor-mediated effects by PCDD/Fs and PCBs in rat, mouse, and guinea pig CALUX cell lines.

    PubMed

    Ghorbanzadeh, Mehdi; van Ede, Karin I; Larsson, Malin; van Duursen, Majorie B M; Poellinger, Lorenz; Lücke-Johansson, Sandra; Machala, Miroslav; Pěnčíková, Kateřina; Vondráček, Jan; van den Berg, Martin; Denison, Michael S; Ringsted, Tine; Andersson, Patrik L

    2014-07-21

    For a better understanding of species-specific relative effect potencies (REPs), responses of dioxin-like compounds (DLCs) were assessed. REPs were calculated using chemical-activated luciferase gene expression assays (CALUX) derived from guinea pig, rat, and mouse cell lines. Almost all 20 congeners tested in the rodent cell lines were partial agonists and less efficacious than 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). For this reason, REPs were calculated for each congener using concentrations at which 20% of the maximal TCDD response was reached (REP20TCDD). REP20TCDD values obtained for PCDD/Fs were comparable with their toxic equivalency factors assigned by the World Health Organization (WHO-TEF), while those for PCBs were in general lower than the WHO-TEF values. Moreover, the guinea pig cell line was the most sensitive as indicated by the 20% effect concentrations of TCDD of 1.5, 5.6, and 11.0 pM for guinea pig, rat, and mouse cells, respectively. A similar response pattern was observed using multivariate statistical analysis between the three CALUX assays and the WHO-TEFs. The mouse assay showed minor deviation due to higher relative induction potential for 2,3,7,8-tetrachlorodibenzofuran and 2,3,4,6,7,8-hexachlorodibenzofuran and lower for 1,2,3,4,6,7,8-heptachlorodibenzofuran and 3,3',4,4',5-pentachlorobiphenyl (PCB126). 2,3,7,8-Tetrachlorodibenzofuran was more than two times more potent in the mouse assay as compared with that of rat and guinea pig cells, while measured REP20TCDD for PCB126 was lower in mouse cells (0.05) as compared with that of the guinea pig (0.2) and rat (0.07). In order to provide REP20TCDD values for all WHO-TEF assigned compounds, quantitative structure-activity relationship (QSAR) models were developed. The QSAR models showed that specific electronic properties and molecular surface characteristics play important roles in the AhR-mediated response. In silico derived REP20TCDD values were generally consistent with the WHO

  16. An altered hydrotropic response (ahr1) mutant of Arabidopsis recovers root hydrotropism with cytokinin

    PubMed Central

    Saucedo, Manuel; Ponce, Georgina; Campos, María Eugenia; Eapen, Delfeena; García, Edith; Luján, Rosario; Sánchez, Yoloxóchitl; Cassab, Gladys I.

    2012-01-01

    Roots are highly plastic and can acclimate to heterogeneous and stressful conditions. However, there is little knowledge of the effect of moisture gradients on the mechanisms controlling root growth orientation and branching, and how this mechanism may help plants to avoid drought responses. The aim of this study was to isolate mutants of Arabidopsis thaliana with altered hydrotropic responses. Here, altered hydrotropic response 1 (ahr1), a semi-dominant allele segregating as a single gene mutation, was characterized. ahr1 directed the growth of its primary root towards the source of higher water availability and developed an extensive root system over time. This phenotype was intensified in the presence of abscisic acid and was not observed if ahr1 seedlings were grown in a water stress medium without a water potential gradient. In normal growth conditions, primary root growth and root branching of ahr1 were indistinguishable from those of the wild type (wt). The altered hydrotropic growth of ahr1 roots was confirmed when the water-rich source was placed at an angle of 45° from the gravity vector. In this system, roots of ahr1 seedlings grew downward and did not display hydrotropism; however, in the presence of cytokinins, they exhibited hydrotropism like those of the wt, indicating that cytokinins play a critical role in root hydrotropism. The ahr1 mutant represents a valuable genetic resource for the study of the effects of cytokinins in the differential growth of hydrotropism and control of lateral root formation during the hydrotropic response. PMID:22442413

  17. Effect-directed analysis and mixture effects of AhR-active PAHs in crude oil and coastal sediments contaminated by the Hebei Spirit oil spill.

    PubMed

    Hong, Seongjin; Lee, Sangwoo; Choi, Kyungho; Kim, Gi Beum; Ha, Sung Yong; Kwon, Bong-Oh; Ryu, Jongseong; Yim, Un Hyuk; Shim, Won Joon; Jung, Jinho; Giesy, John P; Khim, Jong Seong

    2015-04-01

    The major AhR-active PAHs were identified in crude oil and oil-contaminated sediments by use of effect-directed analysis. As part of the study, an enhanced potency balance analysis was conducted by establishing the novel relative potency values of (alkyl)-PAHs from the H4IIE-luc bioassay. Silica gel column fractionation of crude oil resulted in greater AhR-mediated potencies in fractions of aromatics (F2) and resins (F3), and such trend was also observed for field collected sediment samples. AhR-mediated potencies of six F2 sub-fractions from HPLC indicated that the majority of F2 responses were attributable to 3-4 ring aromatics. Target PAHs including C4-phenanthrene, C1-chrysene, and C3-chrysene in sediments explained ∼ 18% of the bioassay-derived TCDD-EQs, however, the unknown AhR agonists and potential mixture effects remain in question. Overall, the AhR-potency and antagonistic potential of residual oil in sediment tended to decrease over time, thus monitoring of weathering process would be key for the post management of oil-contaminated sites. PMID:25645060

  18. Effect-directed analysis and mixture effects of AhR-active PAHs in crude oil and coastal sediments contaminated by the Hebei Spirit oil spill.

    PubMed

    Hong, Seongjin; Lee, Sangwoo; Choi, Kyungho; Kim, Gi Beum; Ha, Sung Yong; Kwon, Bong-Oh; Ryu, Jongseong; Yim, Un Hyuk; Shim, Won Joon; Jung, Jinho; Giesy, John P; Khim, Jong Seong

    2015-04-01

    The major AhR-active PAHs were identified in crude oil and oil-contaminated sediments by use of effect-directed analysis. As part of the study, an enhanced potency balance analysis was conducted by establishing the novel relative potency values of (alkyl)-PAHs from the H4IIE-luc bioassay. Silica gel column fractionation of crude oil resulted in greater AhR-mediated potencies in fractions of aromatics (F2) and resins (F3), and such trend was also observed for field collected sediment samples. AhR-mediated potencies of six F2 sub-fractions from HPLC indicated that the majority of F2 responses were attributable to 3-4 ring aromatics. Target PAHs including C4-phenanthrene, C1-chrysene, and C3-chrysene in sediments explained ∼ 18% of the bioassay-derived TCDD-EQs, however, the unknown AhR agonists and potential mixture effects remain in question. Overall, the AhR-potency and antagonistic potential of residual oil in sediment tended to decrease over time, thus monitoring of weathering process would be key for the post management of oil-contaminated sites.

  19. Inherent and benzo[a]pyrene-induced differential aryl hydrocarbon receptor signaling greatly affects life span, atherosclerosis, cardiac gene expression, and body and heart growth in mice.

    PubMed

    Kerley-Hamilton, Joanna S; Trask, Heidi W; Ridley, Christian J A; Dufour, Eric; Lesseur, Corina; Ringelberg, Carol S; Moodie, Karen L; Shipman, Samantha L; Korc, Murray; Gui, Jiang; Shworak, Nicholas W; Tomlinson, Craig R

    2012-04-01

    Little is known of the environmental factors that initiate and promote disease. The aryl hydrocarbon receptor (AHR) is a key regulator of xenobiotic metabolism and plays a major role in gene/environment interactions. The AHR has also been demonstrated to carry out critical functions in development and disease. A qualitative investigation into the contribution by the AHR when stimulated to different levels of activity was undertaken to determine whether AHR-regulated gene/environment interactions are an underlying cause of cardiovascular disease. We used two congenic mouse models differing at the Ahr gene, which encodes AHRs with a 10-fold difference in signaling potencies. Benzo[a]pyrene (BaP), a pervasive environmental toxicant, atherogen, and potent agonist for the AHR, was used as the environmental agent for AHR activation. We tested the hypothesis that activation of the AHR of different signaling potencies by BaP would have differential effects on the physiology and pathology of the mouse cardiovascular system. We found that differential AHR signaling from an exposure to BaP caused lethality in mice with the low-affinity AHR, altered the growth rates of the body and several organs, induced atherosclerosis to a greater extent in mice with the high-affinity AHR, and had a huge impact on gene expression of the aorta. Our studies also demonstrated an endogenous role for AHR signaling in regulating heart size. We report a gene/environment interaction linking differential AHR signaling in the mouse to altered aorta gene expression profiles, changes in body and organ growth rates, and atherosclerosis.

  20. Generation of Tg(cyp1a:gfp) Transgenic Zebrafish for Development of a Convenient and Sensitive In Vivo Assay for Aryl Hydrocarbon Receptor Activity.

    PubMed

    Xu, Hongyan; Li, Caixia; Li, Yan; Ng, Grace Hwee Boon; Liu, Chunsheng; Zhang, Xiaoyan; Gong, Zhiyuan

    2015-12-01

    Both dioxins/dioxin-like compounds and polycyclic aromatic hydrocarbons (PAHs) are persistent organic pollutants and cause multiple adverse health effects on human and wildlife. Cyp1a is the most commonly used biomarker induced by these pollutants through activation of the aryl hydrocarbon receptor (AhR) pathway. Here we generated Tg(cyp1a:gfp) transgenic zebrafish for establishing a convenient in vivo assay for analysing these xenobiotic compounds. The Tg(cyp1a:gfp) larvae at 4 day post-fertilization were tested with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), and GFP induction was observed mainly in the kidney, liver and gut. Similar GFP expression was also induced strongly by two dioxin-like chemicals, co-planar polychlorinated biphenyl (PCB126) and polychlorinated dibenzo-p-furan (PeCDF) and relatively weakly by two PAHs, 3-methylcholanthrene (3-MC) and benzo[a]pyrene (BAP). The lowest observed effective concentration (LOEC) of TCDD was estimated to be ∼1 pM and the EC50 (effective concentration to induce GFP in 50 % of Tg(cyp1a:gfp) larvae) was ∼10 pM. PCB126 and PeCDF had ∼10× lower potencies in GFP induction than TCDD, while the potencies for 3-MC and BAP were at least 1000× lower. The sensitivity of Tg(cyp1a:gfp) larvae to respond TCDD was also favourable compared to that of ethoxyresorufin-O-deethylase (EROD) assay in both zebrafish larvae and adult livers. As GFP-based assay in transgenic zebrafish can be easily accommodated in multi-well dishes, the Tg(cyp1a:gfp) zebrafish should provide not only a valuable biomonitoring tool for aquatic contaminants but also a potential high-throughput chemical screening platform for identification of new AhR agonists.

  1. Role of zebrafish cytochrome P450 CYP1C genes in the reduced mesencephalic vein blood flow caused by activation of AHR2

    SciTech Connect

    Kubota, Akira; Stegeman, John J.; Woodin, Bruce R.; Iwanaga, Toshihiko; Harano, Ryo; Peterson, Richard E.; Hiraga, Takeo; Teraoka, Hiroki

    2011-06-15

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) causes various signs of toxicity in early life stages of vertebrates through activation of the aryl hydrocarbon receptor (AHR). We previously reported a sensitive and useful endpoint of TCDD developmental toxicity in zebrafish, namely a decrease in blood flow in the dorsal midbrain, but downstream genes involved in the effect are not known. The present study addressed the role of zebrafish cytochrome P450 1C (CYP1C) genes in association with a decrease in mesencephalic vein (MsV) blood flow. The CYP1C subfamily was recently discovered in fish and includes the paralogues CYP1C1 and CYP1C2, both of which are induced via AHR2 in zebrafish embryos. We used morpholino antisense oligonucleotides (MO or morpholino) to block initiation of translation of the target genes. TCDD-induced mRNA expression of CYP1Cs and a decrease in MsV blood flow were both blocked by gene knockdown of AHR2. Gene knockdown of CYP1C1 by two different morpholinos and CYP1C2 by two different morpholinos, but not by their 5 nucleotide-mismatch controls, was effective in blocking reduced MsV blood flow caused by TCDD. The same CYP1C-MOs prevented reduction of blood flow in the MsV caused by {beta}-naphthoflavone (BNF), representing another class of AHR agonists. Whole-mount in situ hybridization revealed that mRNA expression of CYP1C1 and CYP1C2 was induced by TCDD most strongly in branchiogenic primordia and pectoral fin buds. In situ hybridization using head transverse sections showed that TCDD increased the expression of both CYP1Cs in endothelial cells of blood vessels, including the MsV. These results indicate a potential role of CYP1C1 and CYP1C2 in the local circulation failure induced by AHR2 activation in the dorsal midbrain of the zebrafish embryo. - Research Highlights: > We examine the roles of zebrafish CYP1C1 and CYP1C2 in TCDD developmental toxicity. > TCDD induces mRNA expression of both CYP1Cs in the mesencephalic vein. > Knockdown of each

  2. Phenotype refinement strengthens the association of AHR and CYP1A1 genotype with caffeine consumption.

    PubMed

    McMahon, George; Taylor, Amy E; Davey Smith, George; Munafò, Marcus R

    2014-01-01

    Two genetic loci, one in the cytochrome P450 1A1 (CYP1A1) and 1A2 (CYP1A2) gene region (rs2472297) and one near the aryl-hydrocarbon receptor (AHR) gene (rs6968865), have been associated with habitual caffeine consumption. We sought to establish whether a more refined and comprehensive assessment of caffeine consumption would provide stronger evidence of association, and whether a combined allelic score comprising these two variants would further strengthen the association. We used data from between 4,460 and 7,520 women in the Avon Longitudinal Study of Parents and Children, a longitudinal birth cohort based in the United Kingdom. Self-report data on coffee, tea and cola consumption (including consumption of decaffeinated drinks) were available at multiple time points. Both genotypes were individually associated with total caffeine consumption, and with coffee and tea consumption. There was no association with cola consumption, possibly due to low levels of consumption in this sample. There was also no association with measures of decaffeinated drink consumption, indicating that the observed association is most likely mediated via caffeine. The association was strengthened when a combined allelic score was used, accounting for up to 1.28% of phenotypic variance. This was not associated with potential confounders of observational association. A combined allelic score accounts for sufficient phenotypic variance in caffeine consumption that this may be useful in Mendelian randomization studies. Future studies may therefore be able to use this combined allelic score to explore causal effects of habitual caffeine consumption on health outcomes. PMID:25075865

  3. Phenotype Refinement Strengthens the Association of AHR and CYP1A1 Genotype with Caffeine Consumption

    PubMed Central

    McMahon, George; Taylor, Amy E.; Davey Smith, George; Munafò, Marcus R.

    2014-01-01

    Two genetic loci, one in the cytochrome P450 1A1 (CYP1A1) and 1A2 (CYP1A2) gene region (rs2472297) and one near the aryl-hydrocarbon receptor (AHR) gene (rs6968865), have been associated with habitual caffeine consumption. We sought to establish whether a more refined and comprehensive assessment of caffeine consumption would provide stronger evidence of association, and whether a combined allelic score comprising these two variants would further strengthen the association. We used data from between 4,460 and 7,520 women in the Avon Longitudinal Study of Parents and Children, a longitudinal birth cohort based in the United Kingdom. Self-report data on coffee, tea and cola consumption (including consumption of decaffeinated drinks) were available at multiple time points. Both genotypes were individually associated with total caffeine consumption, and with coffee and tea consumption. There was no association with cola consumption, possibly due to low levels of consumption in this sample. There was also no association with measures of decaffeinated drink consumption, indicating that the observed association is most likely mediated via caffeine. The association was strengthened when a combined allelic score was used, accounting for up to 1.28% of phenotypic variance. This was not associated with potential confounders of observational association. A combined allelic score accounts for sufficient phenotypic variance in caffeine consumption that this may be useful in Mendelian randomization studies. Future studies may therefore be able to use this combined allelic score to explore causal effects of habitual caffeine consumption on health outcomes. PMID:25075865

  4. The aryl hydrocarbon receptor interacts with ATP5{alpha}1, a subunit of the ATP synthase complex, and modulates mitochondrial function

    SciTech Connect

    Tappenden, Dorothy M.; Lynn, Scott G.; Crawford, Robert B.; Lee, KangAe; Vengellur, Ajith; Kaminski, Norbert E.; Thomas, Russell S.; LaPres, John J.

    2011-08-01

    Dioxins, including 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD), produce a wide range of toxic effects in mammals. Most, if not all, of these toxic effects are regulated by the aryl hydrocarbon receptor (AHR). The AHR is a ligand activated transcription factor that has been shown to interact with numerous proteins capable of influencing the receptor's function. The ability of secondary proteins to alter AHR-mediated transcriptional events, a necessary step for toxicity, led us to determine whether additional interacting proteins could be identified. To this end, we have employed tandem affinity purification (TAP) of the AHR in Hepa1c1c7 cells. TAP of the AHR, followed by mass spectrometry (MS) identified ATP5{alpha}1, a subunit of the ATP synthase complex, as a strong AHR interactor in the absence of ligand. The interaction was lost upon exposure to TCDD. The association was confirmed by co-immunoprecipitation in multiple cell lines. In addition, cell fractionation experiments showed that a fraction of the AHR is found in the mitochondria. To ascribe a potential functional role to the AHR:ATP5{alpha}1 interaction, TCDD was shown to induce a hyperpolarization of the mitochondrial membrane in an AHR-dependent and transcription-independent manner. These results suggest that a fraction of the total cellular AHR pool is localized to the mitochondria and contributes to the organelle's homeostasis. - Highlights: > The AHR interacts with the mitochondrial protein, ATP5{alpha}1. > Cell fractionation experiments show that the AHR can be found in the mitochondria. > TCDD-exposure induces a hyperpolarization of the mitochondrial inner membrane. > The hyperpolarization is AHR-dependent. > The hyperpolarization occurs without altering ATP levels within the cell.

  5. The aryl hydrocarbon receptor-dependent disruption of contact inhibition in rat liver WB-F344 epithelial cells is linked with induction of survivin, but not with inhibition of apoptosis.

    PubMed

    Svobodová, Jana; Kabátková, Markéta; Šmerdová, Lenka; Brenerová, Petra; Dvořák, Zdeněk; Machala, Miroslav; Vondráček, Jan

    2015-07-01

    Inhibition of apoptosis by the ligands of the aryl hydrocarbon receptor (AhR) has been proposed to play a role in their tumor promoting effects on liver parenchymal cells. However, little is presently known about the impact of toxic AhR ligands, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on apoptosis in other liver cell types, such as in liver epithelial/progenitor cells. In the present study, we focused on the effects of TCDD on apoptosis regulation in a model of liver progenitor cells, rat WB-F344 cell line, during the TCDD-elicited release from contact inhibition. The stimulation of cell proliferation in this cell line was associated with deregulated expression of a number of genes known to be under transcriptional control of the Hippo signaling pathway, a principal regulatory pathway involved in contact inhibition of cell proliferation. Interestingly, we found that mRNA and protein levels of survivin, a known Hippo target, which plays a role both in cell division and inhibition of apoptosis, were significantly up-regulated in rat liver epithelial cell model, as well as in undifferentiated human liver HepaRG cells. Using the short interfering RNA-mediated knockdown, we confirmed that survivin plays a central role in cell division of WB-F344 cells. When evaluating the effects of TCDD on apoptosis induction by camptothecin, a genotoxic topoisomerase I inhibitor, we observed that the pre-treatment of WB-F344 cells with TCDD increased number of cells with apoptotic nuclear morphology, and it potentiated cleavage of both caspase-3 and poly(ADP-ribose) polymerase I. This indicated that despite the observed up-regulation of survivin, apoptosis induced by the genotoxin was potentiated in the model of rat liver progenitor cells. The present results indicate that, unlike in hepatocytes, AhR agonists may not prevent induction of apoptosis elicited by DNA-damaging agents in a model of rat liver progenitor cells.

  6. Action spectrum of melanoblast maturation and involvement of the aryl hydrocarbon receptor.

    PubMed

    Nakamura, Motoki; Nishida, Emi; Morita, Akimichi

    2016-08-01

    The aryl hydrocarbon receptor (AHR) mediates melanocyte activation and skin tanning. We hypothesized that the AHR also mediates melanoblast-to-melanocyte maturation. In a cloned cell line, NCCmelb4, derived from mouse neural crest cells, we investigated AHR expression in melanoblasts stimulated by UV irradiation and AHR agonists. We irradiated the cells with UV, ranging from 280 to 380 nm in 10-nm increments, using a multiwavelength irradiation spectral apparatus. Tyrosinase expression significantly increased with bimodal peaks at 310 and 360 nm. Although melanoblast activation peaked 48 hours after irradiation, the most suitable irradiation interval was 24 hours. AHR expression significantly increased at 360 nm, but not at 310 nm. The AHR agonist, VAF347, and water-soluble tobacco smoke extract induced melanoblast maturation and AHR activation. The culture supernatant derived from the NS47 fibroblast cell line also induced melanoblast maturation and AHR activation. These findings suggest that UV and environmental stimulation of melanoblast-to-melanocyte maturation are enhanced via the AHR pathway. PMID:27539901

  7. Regulation of zebrafish CYP3A65 transcription by AHR2

    SciTech Connect

    Chang, Chin-Teng; Chung, Hsin-Yu; Su, Hsiao-Ting; Tseng, Hua-Pin; Tzou, Wen-Shyong; Hu, Chin-Hwa

    2013-07-15

    CYP3A proteins are the most abundant CYPs in the liver and intestines, and they play a pivotal role in drug metabolism. In mammals, CYP3A genes are induced by various xenobiotics through processes mediated by PXR. We previously identified zebrafish CYP3A65 as a CYP3A ortholog that is constitutively expressed in gastrointestinal tissues, and is upregulated by treatment with dexamethasone, rifampicin or tetrachlorodibenzo-p-dioxin (TCDD). However, the underlying mechanism of TCDD-mediated CYP3A65 transcription is unclear. Here we generated two transgenic zebrafish, Tg(CYP3A65S:EGFP) and Tg(CYP3A65L:EGFP), which contain 2.1 and 5.4 kb 5′ flanking sequences, respectively, of the CYP3A65 gene upstream of EGFP. Both transgenic lines express EGFP in larval gastrointestinal tissues in a pattern similar to that of the endogenous CYP3A65 gene. Moreover, EGFP expression can be significantly induced by TCDD exposure during the larval stage. In addition, EGFP expression can be stimulated by kynurenine, a putative AHR ligand produced during tryptophan metabolism. AHRE elements in the upstream regulatory region of the CYP3A65 gene are indispensible for basal and TCDD-induced transcription. Furthermore, the AHR2 DNA and ligand-binding domains are required to mediate effective CYP3A65 transcription. AHRE sequences are present in the promoters of many teleost CYP3 genes, but not of mammalian CYP3 genes, suggesting that AHR/AHR2-mediated transcription is likely a common regulatory mechanism for teleost CYP3 genes. It may also reflect the different environments that terrestrial and aquatic organisms encounter. - Highlights: • Tg(CYP3A65:EGFP) and CYP3A65 exhibits identical expression pattern. • CYP3A65 can be significantly induced by TCDD or kynurenine. • The AHRE elements are required to mediate CYP3A65 transcription. • The AHR2 DNA and ligand-binding domains are required for CYP3A65 transcription. • AHRE elements are present in many teleost CYP3 genes, but not in

  8. Effects of tris(1,3-dichloro-2-propyl) phosphate and triphenyl phosphate on receptor-associated mRNA expression in zebrafish embryos/larvae.

    PubMed

    Liu, Chunsheng; Wang, Qiangwei; Liang, Kang; Liu, Jingfu; Zhou, Bingsheng; Zhang, Xiaowei; Liu, Hongling; Giesy, John P; Yu, Hongxia

    2013-03-15

    Tris(1,3-dichloro-2-propyl) phosphate (TDCPP) and triphenyl phosphate (TPP) are frequently detected in biota, including fish. However, knowledge of the toxicological and molecular effects of these currently used flame retardants is limited. In the present study, an in vivo screening approach was developed to evaluate effects of TDCPP and TPP on developmental endpoints and receptor-associated expression of mRNA in zebrafish embryos/larvae. Exposure to TDCPP or TPP resulted in significantly smaller rates of hatching and survival, in dose- and time-dependent manners. The median lethal concentration (LC(50)) was 7.0 mg/L for TDCPP and 29.6 mg/L for TPP at 120 hour post-fertilization (hpf). Real-time PCR revealed alterations in expression of mRNAs involved in aryl hydrocarbon receptors (AhRs)-, peroxisome proliferator-activated receptor alpha (PPARα)-, estrogenic receptors (ERs)-, thyroid hormone receptor alpha (TRα)-, glucocorticoid receptor (GR)-, and mineralocorticoid receptor (MR)-centered gene networks. Exposure to positive control chemicals significantly altered abundances of mRNA in corresponding receptor-centered gene networks, a result that suggests that it is feasible to use zebrafish embryos/larvae to evaluate effects of chemicals on mRNA expression in these gene networks. Exposure to TDCPP altered transcriptional profiles in all six receptor-centered gene networks, thus exerting multiple toxic effects. TPP was easily metabolized and its potency to change expression of mRNA involved in receptor-centered gene networks was weaker than that of TDCPP. The PPARα- and TRα-centered gene networks might be the primary pathways affected by TPP. Taken together, these results demonstrated that TDCPP and TPP could alter mRNA expression of genes involved in the six receptor-centered gene networks in zebrafish embryos/larvae, and TDCPP seemed to have higher potency in changing the mRNA expression of these genes. PMID:23306105

  9. Toxicity of teriflunomide in aryl hydrocarbon receptor deficient mice.

    PubMed

    Redaelli, Chiara; Gaffarogullari, Ece Cazibe; Brune, Maik; Pilz, Caroline; Becker, Simon; Sonner, Jana; Jäschke, Andres; Gröne, Hermann-Josef; Wick, Wolfgang; Platten, Michael; Lanz, Tobias Volker

    2015-12-01

    The intracellular transcription factor aryl hydrocarbon receptor (AHR) is bound and activated by xenobiotics, thereby promoting their catabolism by inducing expression of cytochrome P450 oxidase (CYP) genes through binding xenobiotic response elements (XRE) in their promoter region. In addition, it is involved in several cellular pathways like cell proliferation, differentiation, regeneration, tumor invasiveness and immune responses. Several pharmaceutical compounds like benzimidazoles activate the AHR and induce their own metabolic degradation. Using newly generated XRE-reporter mice, which allow in vivo bioluminescence imaging of AHR activation, we show here that the AHR is activated in vivo by teriflunomide (TER), which has recently been approved for the treatment of multiple sclerosis. While we did not find any evidence that the AHR mediates the immunomodulatory effects of TER, AHR activation led to metabolism and detoxification of teriflunomide, most likely via CYP. Mice deficient for the AHR show higher blood levels of teriflunomide, suffer from enhanced thrombo- and leukopenia and elevated liver enzymes as well as from severe gastrointestinal ulcers and bleeding which are lethal after 8-11 days of treatment. Leukopenia, acute liver damage and diarrhea have also been described as common side effects in human trials with TER. These data suggest that the AHR is relevant for detoxification not only of environmental toxins but also of drugs in clinical use, with potential implications for the application of AHR-modifying therapies in conjunction to TER in humans. The XRE-reporter mouse is a useful novel tool for monitoring AHR activation using in vivo imaging. PMID:26341389

  10. The role of the aryl hydrocarbon receptor in the female reproductive system.

    PubMed

    Hernández-Ochoa, Isabel; Karman, Bethany N; Flaws, Jodi A

    2009-02-15

    In recent years, many studies have emphasized how changes in aryl hydrocarbon receptor (AHR)-mediated gene expression result in biological effects, raising interest in this receptor as a regulator of normal biological function. This review focuses on what is known about the role of the AHR in the female reproductive system, which includes the ovaries, Fallopian tubes or oviduct, uterus and vagina. This review also focuses on the role of the AHR in reproductive outcomes such as cyclicity, senescence, and fertility. Specifically, studies using potent AHR ligands, as well as transgenic mice lacking the AHR-signaling pathway are discussed from a viewpoint of understanding the endogenous role of this ligand-activated transcription factor in the female reproductive lifespan. Based on findings highlighted in this paper, it is proposed that the AHR has a role in physiological functions including ovarian function, establishment of an optimum environment for fertilization, nourishing the embryo and maintaining pregnancy, as well as in regulating reproductive lifespan and fertility. The mechanisms by which the AHR regulates female reproduction are poorly understood, but it is anticipated that new models and the ability to generate specific gene deletions will provide powerful experimental tools for better understanding how alterations in AHR pathways result in functional changes in the female reproductive system.

  11. The Role of the Aryl Hydrocarbon Receptor in the Female Reproductive System

    PubMed Central

    Hernández-Ochoa, Isabel; Karman, Bethany N.; Flaws, Jodi A.

    2009-01-01

    In recent years, many studies have emphasized how changes in aryl hydrocarbon receptor (AHR)-mediated gene expression result in biological effects, raising interest in this receptor as a regulator of normal biological function. This review focuses on what is known about the role of the AHR in the female reproductive system, which includes the ovaries, fallopian tubes or oviduct, uterus and vagina. This review also focuses on the role of the AHR in reproductive outcomes such as cyclicity, senescence, and fertility. Specifically, studies using potent AHR ligands, as well as transgenic mice lacking the AHR signaling pathway are discussed from a viewpoint of understanding the endogenous role of this ligand-activated transcription factor in the female reproductive lifespan. Based on findings highlighted in this paper, it is proposed that the AHR has a role in physiological functions including ovarian function, establishment of an optimum environment for fertilization, nourishing the embryo and maintaining pregnancy, as well as in regulating reproductive lifespan and fertility. The mechanisms by which the AHR regulates female reproduction are poorly understood, but it is anticipated that new models and the ability to generate specific gene deletions will provide powerful experimental tools for better understanding how alterations in AHR pathways result in functional changes in the female reproductive system. PMID:18977336