Aryl Hydrocarbon Receptor Agonist Activity Research Articles

Development of FluoAHRL: A Novel Synthetic Fluorescent Compound That Activates AHR and Potentiates Anti-Inflammatory T Regulatory Cells.

Aryl Hydrocarbon Receptor (AHR) ligands, upon binding, induce distinct gene expression profiles orchestrated by the AHR, leading to a spectrum of pro- or anti-inflammatory effects. In this study, we designed, synthesized and evaluated three indole-containing potential AHR ligands (FluoAHRL: AGT-4, AGT-5 and AGT-6). All synthesized compounds were shown to emit fluorescence in the near-infrared. Their AHR agonist activity was first predicted using in silico docking studies, and then confirmed using AHR luciferase reporter cell lines. FluoAHRLs were tested in vitro using mouse peritoneal macrophages and T lymphocytes to assess their immunomodulatory properties. We then focused on AGT-5, as it illustrated the predominant anti-inflammatory effects. Notably, AGT-5 demonstrated the ability to foster anti-inflammatory regulatory T cells (Treg) while suppressing pro-inflammatory T helper (Th)17 cells in vitro. AGT-5 actively induced Treg differentiation from naïve CD4+ cells, and promoted Treg proliferation, cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) expression and interleukin-10 (IL-10) production. The increase in IL-10 correlated with an upregulation of Signal Transducer and Activator of Transcription 3 (STAT3) expression. Importantly, the Treg-inducing effect of AGT-5 was also observed in human tonsil cells in vitro. AGT-5 showed no toxicity when applied to zebrafish embryos and was therefore considered safe for animal studies. Following oral administration to C57BL/6 mice, AGT-5 significantly upregulated Treg while downregulating pro-inflammatory Th1 cells in the mesenteric lymph nodes. Due to its fluorescent properties, AGT-5 could be visualized both in vitro (during uptake by macrophages) and ex vivo (within the lamina propria of the small intestine). These findings make AGT-5 a promising candidate for further exploration in the treatment of inflammatory and autoimmune diseases.

AhR Agonistic Components in Urban Particulate Matter Regulate Astrocytic Activation and Function.

Exposure to atmospheric particulate matter (PM) has been found to accelerate the onset of neurological disorders via the induction of detrimental neuroinflammatory responses. To reveal how astrocytes respond to urban atmospheric PM stimulation, a commercially available standard reference material (SRM1648a) was tested in this study on the activation of rat cortical astrocytes. The results showed that SRM1648a stimulation induced both A1 and A2 phenotypes in astrocytes, as characterized by the exposure concentration-dependent increases in Fkbp5, Sphk1, S100a10, and Il6 mRNA levels. Studying the functional alterations of astrocytes indicated that the neurotrophic factors of Gdnf and Ngf were transcriptionally upregulated due to astrocytic A2-type activation. SRM1648a also promoted autonomous motility of astrocytes and elevated the expressions of chemokines. The aryl hydrocarbon receptor (AhR) agonistic components, such as polycyclic aromatic hydrocarbons (PAHs), were recognized to greatly contribute to SRM1648a-induced effects on astrocytes, which was confirmed by the attenuation of PM-disturbed astrocytic effects via AhR blockage. This study, for the first time, uncovered the direct regulation of urban atmospheric PM on astrocytic activation and function and traced the containing bioactive components (e.g., PAHs) with AhR agonistic activity. The findings provided new knowledge on understanding the ambiguous neurological disturbance from ambient fine PM pollution.

Association between altered tryptophan metabolism, plasma aryl hydrocarbon receptor agonists, and inflammatory Chagas disease.

Chagas disease causes a cardiac illness characterized by immunoinflammatory reactions leading to myocardial fibrosis and remodeling. The development of Chronic Chagas Cardiomyopathy (CCC) in some patients while others remain asymptomatic is not fully understood, but dysregulated inflammatory responses are implicated. The Aryl hydrocarbon receptor (AhR) plays a crucial role in regulating inflammation. Certain tryptophan (Trp) metabolites have been identified as AhR ligands with regulatory functions. We investigated AhR expression, agonist response, ligand production, and AhR-dependent responses, such as IDO activation and regulatory T (Treg) cells induction, in two T. cruzi-infected mouse strains (B6 and Balb/c) showing different polymorphisms in AhR. Furthermore, we assessed the metabolic profile of Trp catabolites and AhR agonistic activity levels in plasma samples from patients with chronic Chagas disease (CCD) and healthy donors (HD) using a luciferase reporter assay and liquid chromatography-mass spectrophotometry (LC-MS) analysis. T. cruzi-infected B6 mice showed impaired AhR-dependent responses compared to Balb/c mice, including reduced IDO activity, kynurenine levels, Treg cell induction, CYP1A1 up-regulation, and AhR expression following agonist activation. Additionally, B6 mice exhibited no detectable AhR agonist activity in plasma and displayed lower CYP1A1 up-regulation and AhR expression upon agonist activation. Similarly, CCC patients had decreased AhR agonistic activity in plasma compared to HD patients and exhibited dysregulation in Trp metabolic pathways, resulting in altered plasma metabolite profiles. Notably, patients with severe CCC specifically showed increased N-acetylserotonin levels in their plasma. The methods and findings presented here contribute to a better understanding of CCC development mechanisms and may identify potential specific biomarkers for T. cruzi infection and the severity of associated heart disease. These insights could be valuable in designing new therapeutic strategies. Ultimately, this research aims to establish the AhR agonistic activity and Trp metabolic profile in plasma as an innovative, non-invasive predictor of prognosis for chronic Chagas disease.

Crosstalk between aryl hydrocarbon receptor and Wnt/β-catenin signaling pathway: Possible culprit of di (2-ethylhexyl) phthalate-mediated cardiotoxicity in zebrafish larvae

Typical plasticizer di (2-ethylhexyl) phthalate (DEHP) has been demonstrated to induce cardiotoxicity in zebrafish, but the potential molecular mechanisms involved have not been fully elucidated. Aryl hydrocarbon receptor (AhR), an essential protein for inducing developmental abnormalities, has been demonstrated to be activated by DEHP in other species, but whether the AhR signaling pathway also contributes to DEHP-mediated cardiac developmental toxicity in zebrafish remains unclear. Firstly, molecular docking simulations initially confirmed the possibility that DEHP has AhR agonistic activity. To further confirm this conjecture, this work analyzed the changes of cardiac-related indexes in zebrafish stressed by DEHP at individual, protein, and gene levels. The results showed that DEHP mediated cardiac phenotypic developmental defects, increased CYP1A1 activity, and oxidative stress as well as significant changes in the expression levels of key proteins and genes of AhR, Wnt/β-catenin, and Nrf2-Keap1 signaling pathways. Notably, the addition of AhR inhibitors effectively alleviated the above negative effects, indicating that the AhR signaling pathway and its crosstalk with the Wnt/β-catenin signaling pathway is an essential pathway for DEHP-mediated cardiac developmental toxicity. Overall, this work enriches the molecular mechanism of DEHP-mediated cardiac developmental defects in zebrafish and provides a reliable biomarker for future environmental risk assessment of DEHP.

Switching on/off aryl hydrocarbon receptor and pregnane X receptor activities by chemically modified tryptamines

Microbial indoles have been demonstrated as selective or dual agonists and ligands of the pregnane X receptor (PXR) and aryl hydrocarbon receptor (AhR). However, structural determinants of microbial indoles selectivity towards both receptors remain elusive. Here, we studied the effects of existing and newly synthesized derivatives of indole microbial metabolite tryptamine on the activity of AhR and PXR receptors. We show that the elongation of indolyl-3-alkaneamine chain, indole N-methylation and conversion of indolyl-3-alkaneamines to oleamides resulted in a major increase of PXR activity and in parallel loss of AhR activity. Using reporter gene assays, RT-PCR and TR-FRET techniques, we have characterized in detail the activation of PXR by novel indolyl-3-alkanyl-oleamides, 1-methyltryptamine and 1-methyltryptamine-acetamide. As a proof of concept, we demonstrated anti-inflammatory and epithelial barrier-protective activity of lead derivatives in intestinal Caco-2 cells, employing the measurement of expression of pro-inflammatory chemokines, tight junction genes, trans-epithelial electric resistance TEER, and dextran-FITC permeability assay. In conclusion, we show that a subtle chemical modifications of simple microbial indole metabolite tryptamine, leads to substantial changes in AhR and PXR agonist activities.

Aryl hydrocarbon receptor activation-mediated vascular toxicity of ambient fine particulate matter: contribution of polycyclic aromatic hydrocarbons and osteopontin as a biomarker

BackgroundExposure to ambient fine particulate matter (PM2.5) is associated with vascular diseases. Polycyclic aromatic hydrocarbons (PAHs) in PM2.5 are highly hazardous; however, the contribution of PM2.5-bound PAHs to PM2.5-associated vascular diseases remains unclear. The ToxCast high-throughput in vitro screening database indicates that some PM2.5-bound PAHs activate the aryl hydrocarbon receptor (AhR). The present study investigated whether the AhR pathway is involved in the mechanism of PM2.5-induced vascular toxicity, identified the PAH in PM2.5 that was the major contributor of AhR activation, and identified a biomarker for vascular toxicity of PM2.5-bound PAHs.ResultsTreatment of vascular smooth muscle cells (VMSCs) with an AhR antagonist inhibited the PM2.5-induced increase in the cell migration ability; NF-κB activity; and expression of cytochrome P450 1A1 (CYP1A1), 1B1 (CYP1B1), interleukin-6 (IL-6), and osteopontin (OPN). Most PM2.5-bound PAHs were extracted into the organic fraction, which drastically enhanced VSMC migration and increased mRNA levels of CYP1A1, CYP1B1, IL-6, and OPN. However, the inorganic fraction of PM2.5 moderately enhanced VSMC migration and only increased IL-6 mRNA levels. PM2.5 increased IL-6 secretion through NF-κB activation; however, PM2.5 and its organic extract increased OPN secretion in a CYP1B1-dependent manner. Inhibiting CYP1B1 activity and silencing OPN expression prevented the increase in VSMC migration ability caused by PM2.5 and its organic extract. The AhR activation potencies of seven PM2.5-bound PAHs, reported in the ToxCast database, were strongly correlated with their capabilities of enhancing the migration ability of VSMCs. Benzo(k)fluoranthene (BkF) contributed the most to the AhR agonistic activity of ambient PM2.5-bound PAHs. The association between PM2.5-induced vascular toxicity, AhR activity, and OPN secretion was further verified in mice; PM2.5-induced intimal hyperplasia in pulmonary small arteries and OPN secretion were alleviated in mice with low AhR affinity. Finally, urinary concentrations of 1-hydroxypyrene, a major PAH metabolite, were positively correlated with plasma OPN levels in healthy humans.ConclusionsThe present study offers in vitro, animal, and human evidences supporting the importance of AhR activation for PM2.5-induced vascular toxicities and that BkF was the major contributor of AhR activation. OPN is an AhR-dependent biomarker of PM2.5-induced vascular toxicity. The AhR activation potency may be applied in the risk assessment of vascular toxicity in PAH mixtures.

Effects of benzotriazole UV stabilizers, UV-PS and UV-P, on the differentiation of splenic regulatory T cells via aryl hydrocarbon receptor

Benzotriazole UV stabilizers (BUVSs) are widely used as additives in various materials, including plastics, to prevent damage from UV-irradiation. However, despite the extensive usage of BUVSs, information on their toxicological properties is limited. In this study, we investigated the effect of BUVSs on the immune regulatory system via the aryl hydrocarbon receptor (AhR). A cell-based transactivation assay using DR-EcoScreen cells revealed that, among 13 BUVSs tested, UV-P, UV-PS, UV-9, and UV-090 activated AhR in a dose-dependent manner. In particular, the AhR agonistic activity of UV-PS was about 10-fold more potent than those of UV-P, UV-090, and UV-9, and UV-PS acted as a full agonist against AhR. In order to investigate the immune regulatory effects of these BUVSs, we orally treated C57BL/6 mice with UV-PS or UV-P (10, 30, and 100 mg/kg) and studied the differentiation of regulatory T cells (Tregs) in spleen cells. Flow-cytometry analysis revealed that the administration of UV-PS (30 and 100 mg/kg) or UV-P (100 mg/kg) significantly increased the population of CD4+-/CD25+-/Foxp3+ Tregs in the spleen. In addition, we found that the in vitro exposure of mouse splenocytes to UV-PS (10 and 30 μM) or UV-P (30 μM) as well as to TCDD (0.1 nM) significantly induced Tregs. Notably, the induction of Tregs was eliminated by co-treatment with an AhR antagonist, CH-223191, in each case. Taken together, these findings suggest that some BUVSs might induce Tregs through direct AhR activation and act as immunosuppressive modulators.

AHR agonistic effects of 6-PN contribute to potential beneficial effects of Hops extract

Hops (Humulus lupulus) is used as an alternative to hormone replacement therapy due to the phytoestrogen, 8-prenylnaringenin (8-PN). To examine the potential risks/benefits of hops extract and its compounds (8-PN and 6-prenylnaringenin, 6-PN), we aimed to evaluate the estrogen receptor α (ERα) and aryl hydrocarbon receptor (AHR) signaling pathways in human endometrial cancer cells. Hops extract, 8-PN and 6-PN showed estrogenic activity. Hops extract and 6-PN activated both ERα and AHR pathways. 6-PN increased the expression of the tumor suppressor gene (AHRR), and that of genes involved in the estrogen metabolism (CYP1A1, CYP1B1). Although 6-PN might activate the detoxification and genotoxic pathways of estrogen metabolism, hops extract as a whole only modulated the genotoxic pathway by an up-regulation of CYP1B1 mRNA expression. These data demonstrate the relevant role of 6-PN contained in the hops extract as potential modulator of estrogen metabolism due to its ERα and AHR agonist activity.

Abstract 11010: Aryl Hydrocarbon Receptor Plays an Essential Role in the Pathogenesis of Pulmonary Arterial Hypertension

Introduction: Pulmonary arterial hypertension (PAH) is a devastating disease characterized by arteriopathy in the small to medium-sized distal pulmonary arteries, often accompanied by infiltration of inflammatory cells. Aryl hydrocarbon receptor (AHR), a nuclear receptor/transcription factor, detoxifies xenobiotics and regulates the differentiation and function of various immune cells. However, the role of AHR in the pathogenesis of PAH is largely unknown. Hypothesis: We hypothesized that AHR has a role in PAH pathogenesis both in human and animal models. Methods: Wild type (WT) and Ahr -knockout ( Ahr -/- ) rats were either treated with AHR agonists or VEGFR2 inhibitors or fed a diet containing a Chinese herbal drug Qing-Dai, which may cause drug-induced PAH, in combination with hypoxia, and then subjected to hemodynamic analysis. Histological characteristics were analyzed in the lungs of these rats and patients with idiopathic PAH. RNA-seq and qRT-PCR were performed on specimens from SU5416/Hypoxia (SuHx) rats, including lungs and the peripheral blood mononuclear cells (PBMCs), and PBMCs of PAH patients. AHR agonistic activity in serum was judged by luciferase reporter assay. Results: WT rats treated with the potent endogenous AHR agonist (6-formylindolo[3,2-b]carbazole) in combination with hypoxia, developed severe pulmonary hypertension (PH) with plexiform-like lesions, whereas WT rats treated with the potent VEGFR2 inhibitors (KI8751 and TAK-593) did not. Ahr -/- rats did not develop PH in the SuHx rats. A diet containing Qing-Dai in combination with hypoxia led to development of PH in Ahr +/+ rats, but not in Ahr -/- rats. Several inflammatory signaling pathways were up-regulated in endothelial cells and PBMCs, which led to infiltration of CD4 + IL-21 + T cells and MRC1 + macrophages into vascular lesions in an AHR-dependent manner. AHR agonistic activity in serum was significantly higher in PAH patients than in healthy volunteers and was associated with poor prognosis of PAH. The expression of several AHR-related genes were changed in PBMCs of PAH patients. Conclusions: AHR plays crucial roles in the development and progression of PAH, indicating that this receptor is a potential target for both diagnosis and treatment of PAH.

The Aryl Hydrocarbon Receptor-Dependent TGF-α/VEGF-B Ratio Correlates With Disease Subtype and Prognosis in Multiple Sclerosis.

ObjectiveTo evaluate the aryl hydrocarbon receptor (AHR)-dependent transforming growth factor alpha (TGF-α)/vascular endothelial growth factor B (VEGF-B) ratio, which regulates the effects of metabolic, dietary, and microbial factors on acute and chronic CNS inflammation, as a potential marker in multiple sclerosis (MS).MethodsTGF-α, VEGF-B, and AHR agonistic activity were determined in serum of 252 patients with relapsing-remitting (RR) MS, primary and secondary progressive MS, as well as during active disease (clinically isolated syndrome [CIS] and RRMS relapse).ResultsThe TGF-α/VEGF-B ratio and AHR agonistic activity were decreased in all MS subgroups with a stable disease course as compared to controls. During active CNS inflammation in CIS and RRMS relapse, the TGF-α/VEGF-B ratio and AHR agonistic activity were increased. Conversely, in patients with minimal clinical impairment despite long-standing disease, the TGF-α/VEGF-B ratio and AHR agonistic activity were unaltered. Finally, the TGF-α/VEGF-B ratio and AHR agonistic activity correlated with neurologic impairment and time to conversion from CIS to MS.ConclusionsThe AHR-dependent TGF-α/VEGF-B ratio is altered in a subtype, severity, and disease activity–specific manner and correlates with time to conversion from CIS to MS. It may thus represent a novel marker and serve as additive guideline for immunomodulatory strategies in MS.Classification of EvidenceThis study provides Class III evidence that serum levels of AHR, TGF-α, and VEGF-B distinguish subtypes of MS and predict the severity and disease activity of MS.

3,3'-Diindolylmethane Exhibits Significant Metabolism after Oral Dosing in Humans.

3,3'-Diindolylmethane (DIM), a major phytochemical derived from ingestion of cruciferous vegetables, is also a dietary supplement. In preclinical models, DIM is an effective cancer chemopreventive agent and has been studied in a number of clinical trials. Previous pharmacokinetic studies in preclinical and clinical models have not reported DIM metabolites in plasma or urine after oral dosing, and the pharmacological actions of DIM on target tissues is assumed to be solely via the parent compound. Seven subjects (6 males and 1 female) ranging from 26-65 years of age, on a cruciferous vegetable-restricted diet prior to and during the study, took 2 BioResponse DIM 150-mg capsules (45.3 mg DIM/capsule) every evening for one week with a final dose the morning of the first blood draw. A complete time course was performed with plasma and urine collected over 48 hours and analyzed by UPLC-MS/MS. In addition to parent DIM, two monohydroxylated metabolites and 1 dihydroxylated metabolite, along with their sulfate and glucuronide conjugates, were present in both plasma and urine. Results reported here are indicative of significant phase 1 and phase 2 metabolism and differ from previous pharmacokinetic studies in rodents and humans, which reported only parent DIM present after oral administration. 3-((1H-indole-3-yl)methyl)indolin-2-one, identified as one of the monohydroxylated products, exhibited greater potency and efficacy as an aryl hydrocarbon receptor agonist when tested in a xenobiotic response element-luciferase reporter assay using Hepa1 cells. In addition to competitive phytochemical-drug adverse reactions, additional metabolites may exhibit pharmacological activity highlighting the importance of further characterization of DIM metabolism in humans. SIGNIFICANCE STATEMENT: 3,3'-Diindolylmethane (DIM), derived from indole-3-carbinol in cruciferous vegetables, is an effective cancer chemopreventive agent in preclinical models and a popular dietary supplement currently in clinical trials. Pharmacokinetic studies to date have found little or no metabolites of DIM in plasma or urine. In marked contrast, we demonstrate rapid appearance of mono- and dihydroxylated metabolites in human plasma and urine as well as their sulfate and glucuronide conjugates. The 3-((1H-indole-3-yl)methyl)indolin-2-one metabolite exhibited significant aryl hydrocarbon receptor agonist activity, emphasizing the need for further characterization of the pharmacological properties of DIM metabolites.

Aryl Hydrocarbon Receptor Plasma Agonist Activity Correlates With Disease Activity in Progressive MS.

ObjectiveThe relationship between serum aryl hydrocarbon receptor (AHR) agonistic activity levels with disease severity, its modulation over the course of relapsing-remitting MS (RRMS), and its regulation in progressive MS (PMS) are unknown. Here, we report the analysis of AHR agonistic activity levels in cross-sectional and longitudinal serum samples of patients with RRMS and PMS.MethodsIn a cross-sectional investigation, a total of 36 control patients diagnosed with noninflammatory diseases, 84 patients with RRMS, 35 patients with secondary progressive MS (SPMS), and 41 patients with primary progressive MS (PPMS) were included in this study. AHR activity was measured in a cell-based luciferase assay and correlated with age, sex, the presence of disease-modifying therapies, Expanded Disability Status Scale scores, and disease duration. In a second longitudinal investigation, we analyzed AHR activity in 13 patients diagnosed with RRMS over a period from 4 to 10 years and correlated AHR agonistic activity with white matter atrophy and lesion load volume changes.ResultsIn RRMS, AHR ligand levels were globally decreased and associated with disease duration and neurologic disability. In SPMS and PPMS, serum AHR agonistic activity was decreased and correlated with disease severity. Finally, in longitudinal serum samples of patients with RRMS, decreased AHR agonistic activity was linked to progressive CNS atrophy and increased lesion load.ConclusionsThese findings suggest that serum AHR agonist levels negatively correlate with disability in RRMS and PMS and decrease longitudinally in correlation with MRI markers of disease progression. Thus, serum AHR agonistic activity may serve as novel biomarker for disability progression in MS.

Improvement in the screening performance of potential aryl hydrocarbon receptor ligands by using supervised machine learning

The aryl hydrocarbon receptor (AhR), which is a ligand-dependent transcription factor, plays a crucial role in the regulation of xenobiotic metabolism. There are a large number of artificial or natural molecules in the environment that can activate AhR. In this study, we developed a virtual screening procedure to identify potential ligands of AhR. One structure-based method and two ligand-based methods were used for the virtual screening procedure. The results showed that the precision rate (0.96) and recall rate (0.64) of our procedure were significantly higher than those of a procedure used in a previous study, which suggests that supervised machine learning techniques can greatly improve the performance of virtual screening. Moreover, a pesticide dataset including 777 frequently used pesticides was screened. Seventy-seven pesticides were identified as potential AhR ligands by all three screening methods, among which 12 have never been previously reported as AhR agonists. Two non-agonist AhR ligands and 14 of the 77 pesticides were randomly selected for testing by in vitro and in vivo assays. All 14 pesticides showed different degrees of AhR agonistic activity, and none of the two non-agonist AhR ligand pesticides showed AhR agonistic activity, which suggests that our procedure had good robustness. Four of the pesticides were reported as AhR agonists for the first time, suggesting that these pesticides may need further toxicity assessment. In general, our procedure is a rapid, powerful and computationally inexpensive tool for predicting chemicals with AhR agonistic activity, which could be useful for environmental risk prediction and management.

Diosmin restores the skin barrier by targeting the aryl hydrocarbon receptor in atopic dermatitis

BackgroundAtopic dermatitis (AD) is an inflammatory chronic skin disease that is characterized by the dysfunction or lack of skin barrier proteins. Recent studies have proposed that the pharmacological upregulation of skin barrier proteins is an effective treatment for AD. Aryl hydrocarbon receptor (AhR) is a transcription factor that positively regulates the expression of skin barrier proteins upon its activation. PurposeThis study aimed to identify AhR agonists from phytochemicals and investigate its effect on skin barrier restoration as well as its mechanisms of action in AD. Study designA publicly available assay database and HaCaT cells stably transduced with a luciferase gene driven by an AhR-target gene promoter (CYP1A1) were used to screen for the activity of AhR agonists from phytochemicals. Normal human epidermal keratinocytes (NHEKs) and a human skin equivalent (HSE) model were used to investigate the effect of AhR agonists on skin restoration and its underlying mechanisms. MethodsA Gaussia luciferase assaywas performed to screen for AhR agonist activity. Western blotting, qRT-PCR analysis, immunofluorescence, drug affinity responsive target stability assay, and siRNA-mediated AhR knockdown were performed in NHEKs. Hematoxylin and eosin staining was performed to measure epidermal thickness in the HSE model. ResultsDiosmin, a potential AhR agonist derived from natural products, upregulated the expression of skin barrier proteins (filaggrin and loricrin) and their upstream regulator (OVOL1) in NHEKs. Diosmin treatment also increased epidermal thickness in the HSE model. In addition, incubating NHEKs with diosmin restored the expression of skin barrier proteins and mRNAs that were suppressed by Th2 cytokines and inhibited STAT3 phosphorylation that was induced by Th2 cytokines. Diosmin also upregulated the expression of NQO1, a negative regulator of STAT3. Immunofluorescence results showed that diosmin stimulated AhR nuclear translocation, and the drug affinity responsive target stability assay revealed that this phytochemical directly bound to AhR. Furthermore, AhR knockdown abolished diosmin-induced filaggrin and loricrin expression. ConclusionThese results suggest that diosmin is a potential treatment for AD that targets AhR.

Aryl Hydrocarbon Receptor (AHR) Ligands as Selective AHR Modulators (SAhRMs).

The aryl hydrocarbon receptor (AhR) was first identified as the intracellular protein that bound and mediated the toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin) and dioxin-like compounds (DLCs). Subsequent studies show that the AhR plays an important role in maintaining cellular homeostasis and in pathophysiology, and there is increasing evidence that the AhR is an important drug target. The AhR binds structurally diverse compounds, including pharmaceuticals, phytochemicals and endogenous biochemicals, some of which may serve as endogenous ligands. Classification of DLCs and non-DLCs based on their persistence (metabolism), toxicities, binding to wild-type/mutant AhR and structural similarities have been reported. This review provides data suggesting that ligands for the AhR are selective AhR modulators (SAhRMs) that exhibit tissue/cell-specific AhR agonist and antagonist activities, and that their functional diversity is similar to selective receptor modulators that target steroid hormone and other nuclear receptors.

Exposure to the AhR agonist cyprodinil impacts the cardiac development and function of zebrafish larvae.

Cyprodinil is a broad-spectrum pyrimidine amine fungicide that has been reportedly used worldwide. However, toxicity studies of cyprodinil on aquatic organisms, specifically zebrafish (Danio rerio), are lacking. In our present study, we predicted cyprodinil binding to the aryl hydrocarbon receptor (AhR) by using molecular docking simulation. Then, we used recombinant HepG2 cells and Tg(cyp1a1-12DRE:egfp) transgenic zebrafish to further assess the AhR agonistic activity of cyprodinil. Besides, the significant upregulation of cyp1a1 further verified that statement. Moreover, we found that zebrafish exposure to cyprodinil induced developmental toxicity in the larvae, particularly during cardiac development. The expression levels of cardiac development-related genes, namely tbx5, nkx2.5, gata4, and tnnt2, were markedly altered, which might cause the adverse effects of cyprodinil on cardiac function and development. In summary, we found that cyprodinil, as an AhR agonist, induced development toxicity in zebrafish larvae, especially on cardiac. Data here can assess the potential effects on organisms in the aquatic environment and promote the regulation and safe use of cyprodinil.

Discovery of 2-aryl and 2-pyridinylbenzothiazoles endowed with antimicrobial and aryl hydrocarbon receptor agonistic activities

Benzothiazole is a privileged scaffold in medicinal chemistry present in diverse bioactive compounds with multiple pharmacological applications such as analgesic, anticonvulsant, antidiabetic, anti-inflammatory, anticancer and radioactive amyloidal imagining agents. We reported in this work the study of sixteen functionalized 2-aryl and 2-pyridinylbenzothiazoles as antimicrobial agents and as aryl hydrocarbon receptor (AhR) modulators. The antimicrobial activity against Gram-positive (S. aureus and M. luteus) and Gram-negative (P. aeruginosa, S. enterica and E. coli) pathogens yielded MIC ranging from 3.13 to 50 μg/mL and against the yeast C. albicans, the benzothiazoles displayed MIC from 12.5 to 100 μg/mL. All compounds showed promising antibiofilm activity against S. aureus and P. aeruginosa. The arylbenzothiazole 12 displayed the greatest biofilm eradication in S. aureus (74%) subsequently verified by fluorescence microscopy. The ability of benzothiazoles to modulate AhR expression was evaluated in a cell-based reporter gene assay. Six benzothiazoles (7, 8-10, 12, 13) induced a significant AhR-mediated transcription and interestingly compound 12 was also the strongest AhR-agonist identified. Structure-activity relationships are suggested herein for the AhR-agonism and antibiofilm activities. Furthermore, in silico predictions revealed a good ADMET profile and druglikeness for the arylbenzothiazole 12 as well as binding similarities to AhR compared with the endogenous agonist FICZ.

Predictive modeling of aryl hydrocarbon receptor (AhR) agonism

The aryl hydrocarbon receptor (AhR) plays a key role in the regulation of gene expression in metabolic machinery and detoxification systems. In the recent years, this receptor has attracted interest as a therapeutic target for immunological, oncogenic and inflammatory conditions. In the present report, in silico and in vitro approaches were combined to study the activation of the AhR. To this end, a large database of chemical compounds with known AhR agonistic activity was employed to build 5 classifiers based on the Adaboost (AdB), Gradient Boosting (GB), Random Forest (RF), Multilayer Perceptron (MLP) and Support Vector Machine (SVM) algorithms, respectively. The built classifiers were examined, following a 10-fold external validation procedure, demonstrating adequate robustness and predictivity. These models were integrated into a majority vote based ensemble, subsequently used to screen an in-house library of compounds from which 40 compounds were selected for prospective in vitro experimental validation. The general correspondence between the ensemble predictions and the in vitro results suggests that the constructed ensemble may be useful in predicting the AhR agonistic activity, both in a toxicological and pharmacological context. A preliminary structure-activity analysis of the evaluated compounds revealed that all structures bearing a benzothiazole moiety induced AhR expression while diverse activity profiles were exhibited by phenolic derivatives.

Hypoxia modifies the response to flutamide and linuron in male three-spined stickleback (Gasterosteus aculeatus)

Hypoxia is a major stressor in aquatic environments and it is frequently linked with excess nutrients resulting from sewage effluent discharges and agricultural runoff, which often also contain complex mixtures of chemicals. Despite this, interactions between hypoxia and chemical toxicity are poorly understood. We exposed male three-spined stickleback during the onset of sexual maturation to a model anti-androgen (flutamide; 250 μg/L) and a pesticide with anti-androgenic activity (linuron; 250 μg/L), under either 97% or 56% air saturation (AS). We assessed the effects of each chemical, alone and in combination with reduced oxygen concentration, by measuring the transcription of spiggin in the kidney, as a marker of androgen signalling, and 11 genes in the liver involved in some of the molecular pathways hypothesised to be affected by the exposures. Spiggin transcription was strongly inhibited by flutamide under both AS conditions. In contrast, for linuron, a strong inhibition of spiggin was observed under 97% AS, but this effect was supressed under reduced air saturation, likely due to interactions between the hypoxia inducible factor and the aryl hydrocarbon receptor (AhR) pathways. In the liver, hypoxia inducible factor 1α was induced following exposure to both flutamide and linuron, however this was independent of the level of air saturation. This work illustrates the potential for interactions between hypoxia and pollutants with endocrine or AhR agonist activity to occur, with implications for risk assessment and management.

Exposure to Heptachlorodibenzo-p-dioxin (HpCDD) Regulates microRNA Expression in Human Lung Fibroblasts.

Benzo(ghi)perylene (BghiP) and 1,2,3,4,6,7,8-Heptachlorodibenzo-p-dioxin (HpCDD) were elevated in serum from personnel deployed to sites with open burn pits. Here, we investigated the ability of BghiP and HpCDD to regulate microRNA (miRNA) expression through the aryl hydrocarbon receptor (AHR). Human lung fibroblasts (HLFs) were exposed to BghiP and HpCDD. AHR activity was measured by reporter assay and gene expression. Deployment related miRNA were measured by quantitative polymerase chain reaction. AHR expression was depleted using siRNA. BghiP displayed weak AHR agonist activity. HpCDD induced AHR activity in a dose-dependent manner. Let-7d-5p, miR-103-3p, miR-107, and miR-144-3p levels were significantly altered by HpCDD. AHR knockdown attenuated these effects. These studies reveal that miRNAs previously identified in sera from personnel deployed to sites with open burn pits are altered by HpCDD exposure in HLFs.