Aryl Hydrocarbon Receptor-mediated Activity Research Articles

From farm to fork… and beyond! UV enhances Aryl hydrocarbon receptor-mediated activity of cruciferous vegetables in human intestinal cells upon colonic fermentation.

While the "farm to fork" strategy ticks many boxes in the sustainability agenda, it does not go far enough in addressing how we can improve crop nutraceutical quality. Here, we explored whether supplementary ultraviolet (UV) radiation exposure during growth of broccoli and Chinese cabbage can induce bioactive tryptophan- and glucosinolate-specific metabolite accumulation thereby enhancing Aryl hydrocarbon receptor (AhR) activation in human intestinal cells. By combining metabolomics analysis of both plant extracts and in vitro human colonic fermentation extracts with AhR reporter cell assay, we reveal that human colonic fermentation of UVB-exposed Chinese cabbage led to enhanced AhR activation in human intestinal cells by 23% compared to plants grown without supplementary UV. Thus, by exploring aspects beyond "from farm to fork", our study highlights a new strategy to enhance nutraceutical quality of Brassicaceae, while also providing new insights into the effects of cruciferous vegetables on human intestinal health.

Benzo[b]naphtho[d]thiophenes and naphthylbenzo[b]thiophenes: Their aryl hydrocarbon receptor-mediated activities and environmental presence

Polycyclic aromatic sulfur heterocyclic compounds (PASHs) belong among ubiquitous environmental pollutants; however, their toxic effects remain poorly understood. Here, we studied the aryl hydrocarbon receptor (AhR)-mediated activity of dibenzothiophene, benzo[b]naphtho[d]thiophenes, and naphthylbenzo[b]thiophenes, as well as their presence in two types of environmental matrices: river sediments collected from both rural and urban areas, and in airborne particulate matter (PM2.5) sampled in cities with different levels and sources of pollution. Benzo[b]naphtho[2,1-d]thiophene, benzo[b]naphtho[2,3-d]thiophene, 2,2-naphthylbenzo[b]thiophene, and 2,1-naphthylbenzo[b]thiophene were newly identified as efficient AhR agonists in both rat and human AhR-based reporter gene assays, with 2,2-naphthylbenzo[b]thiophene being the most potent compound identified in both species. Benzo[b]naphtho[1,2-d]thiophene and 3,2-naphthylbenzo[b]thiophene elicited AhR-mediated activity only in the rat liver cell model, while dibenzothiophene and 3,1-naphthylbenzo[b]thiophene were inactive in either cell type. Independently of their ability to activate the AhR, benzo[b]naphtho[1,2-d]thiophene, 2,1-naphthylbenzo[b]thiophene, 3,1-naphthylbenzo[b]thiophene, and 3,2-naphthylbenzo[b]thiophene inhibited gap junctional intercellular communication in a model of rat liver epithelial cells. Benzo[b]naphtho[d]thiophenes were dominant PASHs present in both PM2.5 and sediment samples, with benzo[b]naphtho[2,1-d]thiophene being the most abundant one, followed by benzo[b]naphtho[2,3-d]thiophene. The levels of naphthylbenzo[b]thiophenes were mostly low or below detection limit. Benzo[b]naphtho[2,1-d]thiophene and benzo[b]naphtho[2,3-d]thiophene were identified as the most significant contributors to the AhR-mediated activity in the environmental samples evaluated in this study. Both induced nuclear translocation of the AhR, and they induced CYP1A1 expression in a time-dependent manner, suggesting that their AhR-mediated activity may depend on the rate of their intracellular metabolism. In conclusion, some PASHs could be significant contributors to the overall AhR-mediated toxicity of complex environmental samples suggesting that more attention should be paid to the potential health impacts of this group of environmental pollutants.

Chemical composition and in vitro aryl hydrocarbon receptor-mediated activity of atmospheric particulate matter at an urban, agricultural and industrial site in North Africa (Bizerte, Tunisia)

As recognized risk factor to pose a health threat to humans and wildlife globally, atmospheric particulate matter (PM) were collected from a North African coastal city (Bizerte, Tunisia) for one year, and were characterized for their chemical compositions, including mercury (HgPM), as well as organic contaminants (polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs)), organic carbon (OC) and organic nitrogen (ON), determined in a previous study. Then, we applied an in vitro reporter gene assay (DR-CALUX) to detect and quantify the dioxin-like activity of PM-associated organic contaminants. Results showed that average HgPM concentration over the entire sampling period was found to be 13.4 ± 12 pg m−3. Seasonal variation in the HgPM concentration was observed with lower values in spring and summer and higher values in winter and autumn due to the variation of meteorological conditions together with the emission sources. Principal component analysis suggested that fossil fuel combustion and a nearby cement factory were the dominant anthropogenic HgPM sources. Aryl hydrocarbon receptor (AhR)-mediated activities were observed in all organic extracts of atmospheric PM from Bizerte city (388.3–1543.6 fg m−3), and shows significant positive correlations with all PM-associated organic contaminants. A significant proportion of dioxin-like activity of PM was related to PAHs. The dioxin-like activity followed the same trend as PM-associated organic contaminants, with higher dioxin-like activity in the cold season than in the warm season, indicating the advantage and utility of the use of bioassays in risk assessment of complex environmental samples.

Structure-Dependent Modulation of Aryl Hydrocarbon Receptor-Mediated Activities by Flavonoids.

Dietary flavonoids are used in treatment of multiple diseases, and their antiinflammatory effects in the intestine are due, in part, to interactions with gut microflora and possibly due to modulation of aryl hydrocarbon receptor (AhR) signaling. In this study, we investigated the structure-dependent AhR activity of 14 flavonoids in Caco2 colon cancer cells using induction of CYP1A1 and UGT1A1 gene expression as endpoints. A major structural determinant for AhR activation was the number of hydroxyl groups where pentahydroxyflavonoids (with the exception of morin) > hexahydroxyflavonoids > tetra-/trihydroxyflavonoids, and some of the latter compounds such as apigenin exhibited AhR antagonist activity for induction of CYP1A1. Simulations suggest that while quercetin and apigenin interact primarily with the same residues, the strength of interactions between specific AhR residues with CYP1A1 agonist, quercetin, in comparison with CYP1A1 antagonist, apigenin, is different; thus, such interactions are presumably indicative of potential switches for modulating CYP1A1 activity. The structure-dependent effects of the hydroxyl flavonoids on induction of UGT1A1 were similar to that observed for induction of CYP1A1 except that luteolin and apigenin induced UGT1A1 levels similar to that observed for TCDD, whereas both compounds were AhR antagonists for CYP1A1. Thus, the effects of the flavonoids in Caco2 cells on Ah-responsiveness and interactions with butyrate were both ligand structure- and response-dependent and these activities are consistent with hydroxyflavonoids being selective AhR modulators.

Assessment of the aryl hydrocarbon receptor-mediated activities of polycyclic aromatic hydrocarbons in a human cell-based reporter gene assay

Activation of the aryl hydrocarbon receptor (AhR)-mediated activity is one of key events in toxicity of polycyclic aromatic hydrocarbons (PAHs). Although various classes of AhR ligands may differentially activate human and rodent AhR, there is presently a lack of data on the human AhR-inducing relative potencies (REPs) of PAHs. Here, we focused on estimation of the AhR-mediated activities of a large set of environmental PAHs in human gene reporter AZ-AhR cell line, with an aim to develop the human AhR-based REP values with potential implications for risk assessment of PAHs. The previously identified weakly active PAHs mostly failed to activate the AhR in human cells. The order for REPs of individual PAHs in human cells largely corresponded with the available data from rodent-based experimental systems; nevertheless, we identified differences up to one order of magnitude in REP values of PAHs between human and rodent cells. Higher REP values were found in human cells for some important environmental contaminants or suspected carcinogens, such as indeno[1,2,3-cd]pyrene, benz[a]anthracene or benzo[b]fluoranthene, while lower REP values were determined for methyl-substituted PAHs. Our results also indicate that a different rate of metabolism for individual PAHs in human vs. rodent cells may affect estimation of REP values in human cell-based assay, and potentially alter toxicity of some compounds, such as benzofluoranthenes, in humans. We applied the AZ-AhR assay to evaluation of the AhR-mediated activity of complex mixtures of organic compounds associated with diesel exhaust particles, and we identified the polar compounds present in these mixtures as being particularly highly active in human cells, as compared with rodent cells. The present data suggest that differences may exist between the AhR-mediated potencies of PAHs in human and rodent cells, and that the AhR-mediated effects of polar PAH derivatives and metabolites in human cell models deserve further attention.

Bioavailability-based assessment of aryl hydrocarbon receptor-mediated activity in Lake Tai Basin from Eastern China

Coupling polydimethylsiloxane (PDMS)-based equilibrium passive sampling with chemical and bioassay analysis, we assessed aryl hydrocarbon receptor (AhR)-mediated activity and contributing chemicals in sediment from Lake Tai Basin, Eastern China. The bioanalytical equivalent concentrations (BEQs) of AhR-active chemicals for the exhaustive (total burden) and PDMS extracts (bioavailable fractions) ranged from <9.5–300ng TCDD-EQ/kgdry weight (dw) and <0.096–2.2ng TCDD-EQ/kgdw, respectively, which were of average levels compared to those reported elsewhere. The total concentrations of PAHs in sediment and PDMS were 17–4700μg/kgdw and 0.61–10μg/kgdw, respectively. The majority of the exhaustive extracts subject to acid treatment showed >70% decline in AhR-mediated activity, suggesting the minor contribution by persistent AhR ligands. Targeted analysis of polycyclic aromatic hydrocarbons (PAHs) showed, however, that these chemicals contributed <40% to the overall effect in both exhaustive and PDMS extracts, indicating the presence of other labile AhR ligands. The concentrations of PAHs and BEQs of the AhR-mediated activity attributed to these chemicals in the exhaustive extracts can be back calculated from those in the PDMS extracts via a general organic carbon-PDMS partition coefficient. Similar quantitative conversion between PDMS and aquatic organisms was also verified for aquatic organisms via the lipid-PDMS partition coefficient. Therefore, our study provided a first insight into the quantitative links between bulk chemical burdens in sediment, chemical bioavailability, bioaccumulation potential and resulting mixture effects, as an integral part of predictive environmental risk assessment of contaminated sediment.

Regulation of Pregnane X Receptor (PXR) Function andUGT1A1Gene Expression by Posttranslational Modification of PXR Protein

Human UDP-glucuronosyltransferase (UGT) 1A1 is a critical enzyme responsible for detoxification and metabolism of endogenous and exogenous lipophilic compounds such as bilirubin. The present study shows how cyclin-dependent kinase (CDK) inhibitor roscovitine stimulated the expression of UGT1A1 in HepG2 cells. Pregnane X receptor (PXR)-mediated transactivation of UGT1A1 reporter gene was more prominently enhanced by roscovitine, compared with the basal-, constitutive androstane receptor (CAR)-, and aryl hydrocarbon receptor-mediated activities. We determined the regulatory mechanism of UGT1A1 expression through PXR's stimulation by roscovitine. Although phosphomimetic mutations at Thr290 and Thr408 retained the PXR protein in cytoplasm and attenuated the induction of UGT1A1 expression by both roscovitine and rifampicin, a mutation at Ser350 specifically reduced the activity of PXR induced by roscovitine. Immunoprecipitation analysis revealed that the T290D but not T408D mutant protein remained in cytoplasm by forming a complex with heat shock protein 90 and cytoplasmic CAR retention protein, whereas treatment with proteasome inhibitor MG-132 accumulated the T408D mutant protein in cytoplasm. Transfection with anti-CDK2 small interfering RNA (siRNA) but not anti-CDK1 or CDK5 siRNA led to enhanced expression of UGT1A1. S350D yellow fluorescent protein-PXR fusion protein could translocate from cytoplasm to nucleus similar to the wild-type protein but was detected as an acetylated protein, whose binding with retinoid X receptor (RXR) and histone deacetylase was impaired. Cotransfection with coactivator steroid receptor coactivator (SRC) 2 but not SRC-1 partly recovered its PXR activity. These results indicate that roscovitine stimulated the expression of UGT1A1 by inhibiting CDK2, which phosphorylated PXR at Ser350 to suppress binding with RXR and coactivator and maintain the acetylation of PXR protein.

Carnosol, a constituent of Zyflamend, inhibits aryl hydrocarbon receptor-mediated activation of CYP1A1 and CYP1B1 transcription and mutagenesis.

The aryl hydrocarbon receptor (AhR), a ligand-activated member of the basic helix-loop-helix family of transcription factors, plays a significant role in polycyclic aromatic hydrocarbon (PAH)-induced carcinogenesis. In the upper aerodigestive tract of humans, tobacco smoke, a source of PAHs, activates the AhR leading to increased expression of CYP1A1 and CYP1B1, which encode proteins that convert PAHs to genotoxic metabolites. Inhibitors of Hsp90 ATPase cause a rapid decrease in levels of AhR, an Hsp90 client protein, and thereby block PAH-mediated induction of CYP1A1 and CYP1B1. The main objective of this study was to determine whether Zyflamend, a polyherbal preparation, suppressed PAH-mediated induction of CYP1A1 and CYP1B1 and inhibited DNA adduct formation and mutagenesis. We also investigated whether carnosol, one of multiple phenolic antioxidants in Zyflamend, had similar inhibitory effects. Treatment of cell lines derived from oral leukoplakia (MSK-Leuk1) and skin (HaCaT) with benzo[a]pyrene (B[a]P), a prototypic PAH, induced CYP1A1 and CYP1B1 transcription, resulting in enhanced levels of message and protein. Both Zyflamend and carnosol suppressed these effects of B[a]P. Notably, both Zyflamend and carnosol inhibited Hsp90 ATPase activity and caused a rapid reduction in AhR levels. The formation of B[a]P-induced DNA adducts and mutagenesis was also inhibited by Zyflamend and carnosol. Collectively, these results show that Zyflamend and carnosol inhibit Hsp90 ATPase leading to reduced levels of AhR, suppression of B[a]P-mediated induction of CYP1A1 and CYP1B1, and inhibition of mutagenesis. Carnosol-mediated inhibition of Hsp90 ATPase activity can help explain the chemopreventive activity of herbs such as Rosemary, which contain this phenolic antioxidant.

Effect-directed analysis of contaminated sediment from the wastewater canal in Pancevo industrial area, Serbia

Wastewater canal (WWC) in Pancevo industrial area in Serbia, whose main environmental receptor is the River Danube, is a well known hot-spot of contamination. WWC sediments have been assessed by UNEP based on chemical target analysis. However, integrative biological data on exposure to hazardous compounds are only provided by the present study which aims at evaluating whether the monitored compounds sufficiently reflect potential hazards and to suggest additional compounds to include in monitoring and hazard assessment by applying effect-directed analysis (EDA) based on arylhydrocarbon receptor-mediated activity and cytotoxicity. Multistep NP-HPLC fractionation provided 18 fractions co-eluting with polychlorinated biphenyls (PCBs), polychlorinated dibenzo- p-dioxins (PCDDs), polycyclic aromatic hydrocarbons (PAHs) and more polar compounds. PAHs fractions exhibited great potencies to induce ethoxyresorufin- o-deethylase (EROD) in H4IIE rat hepatoma cell line expressed as 2,3,7,8-tetrachlorodibenzo- p-dioxin equivalents (TCDD-EQ) (0.1–34.6 × 10 3 pg g −1 dry weight). Chemical analysis of the most active fractions revealed great concentrations of PAHs (up to 292 × 10 2 ng g −1 sediment equivalents (SEQ)), methylated PAHs (up to 900 × 10 2 ng g −1 SEQ), and other alkyl-substituted PAHs. Only minor portions of biologically derived TCDD-EQs could be attributed to monitored PAHs with known relative potencies (REPs). We hypothesize that a major part of the activity is due to non-monitored alkylated and heterocyclic PAHs. Results of the cell cytotoxicity/proliferation assay on H4IIE cell line suggest the presence of sediment pollutants with pronounced potency to disturb cell growth.

Aryl hydrocarbon receptor-mediated activity of atmospheric particulate matter from an urban and a rural site in Switzerland

Atmospheric particulate matter (PM) is an air-suspended mixture of solid and liquid particles that vary in size, shape, and chemical composition. Long-term exposure to elevated concentrations of fine atmospheric particles is considered to pose a health threat to humans and animals. In this context, it has been hypothesized that toxic chemicals such as polycyclic aromatic hydrocarbons (PAHs) play an important role. Some PAHs are known to be carcinogenic and it has been shown that carcinogenic effects of PAHs are mediated by the aryl hydrocarbon receptor (AhR). In this study, PM1 was collected at a rural and an urban traffic site during an intense winter smog period, in which concentration of PM1 often exceeded 50 μg m −3. We applied an in vitro reporter gene assay (DR-CALUX) to detect and quantify PM1-associated chemicals that induce AhR-mediated gene expression. This activity was expressed as CALUX equivalents of 2,3,7,8-tetrachlorodibenzodioxin (PM-TCDD-CEQs). In addition, concentrations of PAHs in the PM1 extracts were determined using gas chromatography/high-resolution mass spectrometry. Concentrations of PM-TCDD-CEQs ranged from 10 to 85 pg m −3 and from 19 to 87 pg m −3 at the urban and rural site, respectively. By the use of known relative potency factors, the measured concentration of a PAH was converted into a PAH-TCDD-CEQ concentration. ΣPAH-TCDD-CEQ and PM-TCDD-CEQ were highly correlated at both sites ( r 2 = 0.90 and 0.69). The calculated ΣPAH-TCDD-CEQs explain between 2% and 20% of the measured PM-TCDD-CEQs. Benzo[ k]fluoranthene was the most important PAH causing approximately 60% of the total ΣPAH-TCDD-CEQ activity. In contrast to NO, CO, PM10, and PM1, the concentration of PM-TCDD-CEQs showed no significant difference between the two sites. No indications were found that road traffic emissions caused elevated concentrations of PM-TCDD-CEQs at the urban traffic site.

Targeting of Aryl Hydrocarbon Receptor-Mediated Activation of Cyclooxygenase-2 Expression by the Indole-3-Carbinol Metabolite 3,3′-Diindolylmethane in Breast Cancer Cells

Ligands of the aryl hydrocarbon receptor (AhR) include the environmental xenobiotic 2,3,7,8 tetrachlorodibenzo(p)dioxin (TCDD), polycyclic aryl hydrocarbons, and the dietary compounds 3, 3′-diindolylmethane (DIM), a condensation product of indol-3-carbinol found in Brassica vegetables, and the phytoalexin resveratrol (RES). The AhR and its cofactors regulate the expression of target genes at pentameric (GCGTG) xenobiotic responsive elements (XRE). Because the activation of cyclooxygenase-2 (COX-2) expression by AhR ligands may contribute to inflammation and tumorigenesis, we investigated the epigenetic regulation of the COX-2 gene by TCDD and the reversal effects of DIM in MCF-7 breast cancer cells. Results of DNA binding and chromatin immunoprecipitation (ChIP) studies documented that the treatment with TCDD induced the association of the AhR to XRE harbored in the COX-2 promoter and control CYP1A1 promoter oligonucleotides. The TCDD-induced binding of the AhR was reduced by small-interfering RNA for the AhR or the cotreatment with synthetic (3-methoxy-4-naphthoflavone) or dietary AhR antagonists (DIM, RES). In time course ChIP studies, TCDD induced the rapid (15 min) occupancy by the AhR, the histone acetyl transferase p300, and acetylated histone H4 (AcH4) at the COX-2 promoter. Conversely, the cotreatment of MCF-7 cells with DIM (10 μmol/L) abrogated the TCDD-induced recruitment of the AhR and AcH4 to the COX-2 promoter and the induction of COX-2 mRNA and protein levels. Taken together, these data suggest that naturally occurring modulators of the AhR such as DIM may be effective agents for dietary strategies against epigenetic activation of COX-2 expression by AhR agonists.

Heat shock protein 90 inhibitors suppress aryl hydrocarbon receptor-mediated activation of CYP1A1 and CYP1B1 transcription and DNA adduct formation.

The aryl hydrocarbon receptor (AhR), a client protein of heat shock protein 90 (HSP90), plays a significant role in polycyclic aromatic hydrocarbon (PAH)-induced carcinogenesis. Tobacco smoke, a source of PAHs, activates the AhR, leading to enhanced transcription of CYP1A1 and CYP1B1, which encode proteins that convert PAHs to genotoxic metabolites. The main objectives of this study were to determine whether HSP90 inhibitors suppress PAH-mediated induction of CYP1A1 and CYP1B1 or block benzo(a)pyrene [B(a)P]-induced formation of DNA adducts. Treatment of cell lines derived from oral leukoplakia (MSK-Leuk1) or esophageal squamous cell carcinoma (KYSE450) with a saline extract of tobacco smoke, B(a)P, or dioxin induced CYP1A1 and CYP1B1 transcription, resulting in enhanced levels of message and protein. Inhibitors of HSP90 [17-allylamino-17-demethoxygeldanamycin (17-AAG); celastrol] suppressed these inductive effects of PAHs. Treatment with 17-AAG and celastrol also caused a rapid and marked decrease in amounts of AhR protein without modulating levels of HSP90. The formation of B(a)P-induced DNA adducts in MSK-Leuk1 cells was inhibited by 17-AAG, celastrol, and alpha-naphthoflavone, a known AhR antagonist. The reduction in B(a)P-induced DNA adducts was due, at least in part, to reduced metabolic activation of B(a)P. Collectively, these results suggest that 17-AAG and celastrol, inhibitors of HSP90, suppress the activation of AhR-dependent gene expression, leading, in turn, to reduced formation of B(a)P-induced DNA adducts. Inhibitors of HSP90 may have a role in chemoprevention in addition to cancer therapy.

Secondary Effects of Catalytic Diesel Particulate Filters: Reduced Aryl Hydrocarbon Receptor-Mediated Activity of the Exhaust

Diesel exhaust contains numerous toxic substances that show different modes of action such as triggering aryl hydrocarbon receptor (AhR)-mediated pathways. We investigated AhR-mediated activity of exhaust generated by a heavy-duty diesel engine operated with or without iron- or copper/iron-catalyzed diesel particulate filters (DPFs). AhR agonists were quantified using the DR-CALUX reporter gene assay (exposure of cells for 24 h). We found 54-60 ng 2,3,7,8-tetrachlorodibenzo-p-dioxin CALUX equivalents (TCDD-CEQs) per m3 of exhaust in unfiltered samples and 6-16 ng TCDD-CEQ m3 in DPF-treated samples. DPF applications decreased TCDD-CEQ concentrations by almost 90%. Concentrations of known AhR agonists were determined with GC/HRMS and converted to TCDD-CEQ concentrations using compound-specific relative potency values. The analyzed nine polycyclic aromatic hydrocarbons (PAHs) and the 172,3,7,8-chlorinated dibenzodioxins/furans (23,7,8-PCDD/Fs) contributed only marginally (0.6-1.6%) to the total agonist concentration. However, both DPFs also decreased concentrations of individual PAHs by 7(0-80%. Variation of the assay exposure time (8, 24, 48,72, and 96 h) revealed that AhR-mediated activity decreased over time and reached a plateau after 72 h, which was most likely due to biotransformation of AhR agonists by the exposed H4IIE cells. At the plateau, we measured 1-2 ng TCDD-CEQ m(-3) in both an unfiltered and a filtered exhaust sample. Our findings show that DPFs are a promising technology to detoxify diesel exhaust regarding compounds with AhR-mediated activity.

Transformation of Pyrene in Aqueous Chlorination in the Presence and Absence of Bromide Ion: Kinetics, Products, and Their Aryl Hydrocarbon Receptor-Mediated Activities

To assess the endocrine-disrupting activity stemming from the presence of pyrene in drinking water, the kinetics of chlorination of pyrene was investigated at room temperature, the products of its aqueous chlorination with and without bromide ion were identified, and their aryl hydrocarbon receptor (AhR)-mediated activities were determined. It was found that the presence of bromide ion greatly promoted the reaction rate of chlorination of pyrene accompanied with the formation of brominated products. While the main product was 1-Cl-pyrene without the addition of bromide ion, di-Br-pyrene and 1-Br-pyrene became the main products in the presence of bromide ion. GC-MS and NMR analysis identified three structures of dibromopyrene in chlorination with the addition of bromide ion as 1,3-di-Br-pyrene, 1,6-di-Br-pyrene, and 1,8-di-Br-pyrene, and their molar ratio was determined to be approximately 0.3:1:1. Finally, 1-Br-pyrene, 1,3-di-Br-pyrene, a mixture of 1,6-di-Br-pyrene and 1,8-di-Br-pyrene (di-Br-pyrene), 1-Cl-pyrene, and a mixture of 1,6-di-Cl-pyrene and 1,8-di-Cl-pyrene (di-Cl-pyrene) were fractionated by HPLC, and their AhR-mediated activities were assessed by a yeast assay. It was found that the effective molar concentrations (or mass concentration) showing half-maximal transcriptional response, EC50, for pyrene, 1-Br-pyrene, 1-Cl-pyrene, di-Cl-pyrene, and di-Br-pyrene were 5632 (1.14), 3089 (0.86), 1942 (0.46), 597.2 (0.21), and 147.3 (0.04) nM (mg/L), respectively.

Aryl hydrocarbon receptor-mediated activity of particulate organic matter from the Paso del Norte airshed along the U.S.-Mexico border.

In this study, we determined the biologic activity of dichloromethane-extracted particulate matter < 10 micro m in aerodynamic diameter (PM10) obtained from filters at three sites in the Paso del Norte airshed, which includes El Paso, Texas, USA; Juarez, Chihuahua, Mexico, and Sunland Park, New Mexico, USA. The extracts were rich in polycyclic aromatic hydrocarbons (PAHs) and had significant biologic activity, measured using two in vitro assay systems: ethoxyresorufin-(O-deethylase (EROD) induction and the aryl hydrocarbon-receptor luciferase reporter system. In most cases, both EROD (5.25 pmol/min/mg protein) and luciferase activities (994 relative light units/mg) were highest in extracts from the Advance site located in an industrial neighborhood in Juarez. These values represented 58% and 55%, respectively, of induction associated with 1 micro M ss-naphthoflavone exposures. In contrast, little activity was observed at the Northeast Clinic site in El Paso, the reference site. In most cases, luciferase and EROD activity from extracts collected from the Tillman Health Center site, situated in downtown El Paso, fell between those observed at the other two sites. Overall, a statistically significant correlation existed between PM10 and EROD and luciferase activities. Chemical analysis of extracts collected from the Advance site demonstrated that concentrations of most PAHs were higher than those reported in most other metropolitan areas in the United States. Calculations made with these data suggest a cancer risk of 5-12 cases per 100,000 people. This risk estimate, as well as comparisons with the work of other investigators, raises concern regarding the potential for adverse health effects to the residents of this airshed. Further work is needed to understand the sources, exposure, and effects of PM10 and particulate organic material in the Paso del Norte airshed.

Screening of the inhibitory effect of vegetable constituents on the aryl hydrocarbon receptor-mediated activity induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin.

The aryl hydrocarbon receptor (AhR) is a ligand-activated nuclear transcription factor that mediates responses to environmental contaminants such as dioxins, which have many adverse health effects. We performed a preliminary screening of the inhibitory effects of vegetable constituents on 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced activation of AhR using the AhR-based bioassay for dioxins, the Ah-Immunoassay. Ninety vegetable constituents including flavonoids, tannins, saponins, terpenes, etc., were assayed in vitro. Among them, flavones, flavonols, anthraquinones, piperine, coumestrol, brevifolincarboxylic acid, and resveratrol showed marked inhibitory effects on AhR-based bioassay activation by TCDD, and their effects were dose dependent. Curcumin, carnosol, and capsaicin also inhibited the activation of AhR in this assay, although to a lesser degree. These results suggest that several vegetable constituents might play a role in protection against dioxin toxicity.

ESTROGEN RECEPTOR- AND ARYL HYDROCARBON RECEPTOR-MEDIATED ACTIVITIES OF A COAL-TAR CREOSOTE

ESTROGEN RECEPTOR- AND ARYL HYDROCARBON RECEPTOR-MEDIATED ACTIVITIES OF A COAL-TAR CREOSOTE