Aryl Hydrocarbon Receptor Antagonist Research Articles

Novel Insights into Psychosis and Antipsychotic Interventions: From Managing Symptoms to Improving Outcomes

For the past 70 years, the dopamine hypothesis has been the key working model in schizophrenia. This has contributed to the development of numerous inhibitors of dopaminergic signaling and antipsychotic drugs, which led to rapid symptom resolution but only marginal outcome improvement. Over the past decades, there has been limited research on the quantifiable pathological changes in schizophrenia, including premature cellular/neuronal senescence, brain volume loss, the attenuation of gamma oscillations in electroencephalograms, and the oxidation of lipids in the plasma and mitochondrial membranes. We surmise that the aberrant activation of the aryl hydrocarbon receptor by toxins derived from gut microbes or the environment drives premature cellular and neuronal senescence, a hallmark of schizophrenia. Early brain aging promotes secondary changes, including the impairment and loss of mitochondria, gray matter depletion, decreased gamma oscillations, and a compensatory metabolic shift to lactate and lactylation. The aim of this narrative review is twofold: (1) to summarize what is known about premature cellular/neuronal senescence in schizophrenia or schizophrenia-like disorders, and (2) to discuss novel strategies for improving long-term outcomes in severe mental illness with natural senotherapeutics, membrane lipid replacement, mitochondrial transplantation, microbial phenazines, novel antioxidant phenothiazines, inhibitors of glycogen synthase kinase-3 beta, and aryl hydrocarbon receptor antagonists.

Discovery of Quinazoline and Quinoline-Based Small Molecules as Utrophin Upregulators via AhR Antagonism for the Treatment of Duchenne Muscular Dystrophy.

Duchenne muscular dystrophy (DMD) is a fatal muscle-wasting disease caused by the absence of a dystrophin protein. Elevating utrophin, a dystrophin paralogue, offers an alternative therapeutic strategy for treating DMD, irrespective of the mutation type. Herein, we report the design and synthesis of novel quinazoline and quinoline-based small molecules as potent utrophin modulators screened via high throughput In-Cell ELISA in C2C12 cells. Remarkably, lead molecule SG-02, identified from a library of 70 molecules, upregulates utrophin 2.7-fold at 800 nM in a dose-dependent manner, marking the highest upregulation within the nanomolar range. SG-02's efficacy was further validated through DMD patient-derived cells, demonstrating a significant 2.3-fold utrophin expression. Mechanistically, SG-02 functions as an AhR antagonist, with excellent binding affinity (Kd = 41.68 nM). SG-02 also enhances myogenesis, as indicated by an increased MyHC expression. ADME evaluation supports SG-02's oral bioavailability. Overall, SG-02 holds promise for addressing the global DMD population.

Benzo[a]pyrene evokes epithelial-mesenchymal transition and pulmonary fibrosis through AhR-mediated Nrf2-p62 signaling

Benzo[a]pyrene (BaP) and its metabolic end product benzo(a)pyren-7,8-dihydrodiol-9,10-epoxide (BPDE), are known toxic environmental pollutants. This study aimed to analyze whether sub-chronic BPDE exposure initiated pulmonary fibrosis and the potential mechanisms. In this work, male C57BL6/J mice were exposed to BPDE by dynamic inhalation exposure for 8 weeks. Our results indicated that sub-chronic BPDE exposure evoked pulmonary fibrosis and epithelial-mesenchymal transition (EMT) in mice. Both in vivo and in vitro, BPDE exposure promoted nuclear translocation of Snail. Further experiments indicated that nuclear factor erythroid 2-related factor 2 (Nrf2) and p62 were upregulated in BPDE-exposed alveolar epithelial cells. Moreover, Nrf2 siRNA transfection evidently attenuated BPDE-induced p62 upregulation. Besides, p62 shRNA inhibited BPDE-incurred Snail nuclear translocation and EMT. Mechanically, BPDE facilitated physical interaction between p62 and Snail in the nucleus, then repressed Snail protein degradation by p62-dependent autophagy-lysosome pathway, and finally upregulated transcriptional activity of Snail. Additionally, aryl hydrocarbon receptor (AhR) was activated in BPDE-treated alveolar epithelial cells. Dual-luciferase assay indicated activating AhR could bind to Nrf2 gene promoter. Moreover, pretreatment with CH223191 or α-naphthoflavone (α-NF), AhR antagonists, inhibited BPDE-activated Nrf2-p62 signaling, and alleviated BPDE-induced EMT and pulmonary fibrosis in mice. Taken together, AhR-mediated Nrf2-p62 signaling contributes to BaP-induced EMT and pulmonary fibrosis.

6PPDQ induces cardiomyocyte senescence via AhR/ROS-mediated autophagic flux blockage

Recently, attention has been drawn to the adverse outcomes of N-(1,3-Dimethylbutyl)-N′-phenyl-p-phenylenediamine quinone (6PPDQ) on human health, but its cardiac toxicity has been relatively understudied. This work aims to investigate the effects of 6PPDQ on differentiated H9c2 cardiomyocytes. Our findings demonstrated that exposure to 6PPDQ altered cellular morphology and disrupted the expression of cardiac-specific markers. Significantly, 6PPDQ exposure led to cardiomyocyte senescence, characterized by elevated β-Galactosidase activity, upregulation of cell cycle inhibitor, induction of DNA double-strand breaks, and remodeling of Lamin B1. Furthermore, 6PPDQ hindered autophagy flux by promoting the formation of autophagosomes while inhibiting the degradation of autolysosomes. Remarkably, restoration of autophagic flux using rapamycin counteracted 6PPDQ-induced cardiomyocyte senescence. Additionally, our study revealed that 6PPDQ significantly increased the ROS production. However, ROS scavenger effectively reduced the blockage of autophagic flux and cardiomyocyte senescence caused by 6PPDQ. Furthermore, we discovered that 6PPDQ activated the Aryl hydrocarbon receptor (AhR) signaling pathway. AhR antagonist was found to reverse the blockage of autophagy and alleviate cardiac senescence, while also reducing ROS levels in 6PPDQ-treated group. In conclusion, our research unveils that exposure to 6PPDQ induces ROS overproduction through AhR activation, leading to disruption of autophagy flux and ultimately contributing to cardiomyocyte senescence.

Jasmone, an essential oil component, is an antagonist of aryl hydrocarbon receptor in liver and intestinal cells

Jasmone, an essential oil component, is an antagonist of aryl hydrocarbon receptor in liver and intestinal cells

The Role of Endocrine Disruption Chemical-Regulated Aryl Hydrocarbon Receptor Activity in the Pathogenesis of Pancreatic Diseases and Cancer.

The aryl hydrocarbon receptor (AHR) serves as a ligand-activated transcription factor crucial for regulating fundamental cellular and molecular processes, such as xenobiotic metabolism, immune responses, and cancer development. Notably, a spectrum of endocrine-disrupting chemicals (EDCs) act as agonists or antagonists of AHR, leading to the dysregulation of pivotal cellular and molecular processes and endocrine system disruption. Accumulating evidence suggests a correlation between EDC exposure and the onset of diverse pancreatic diseases, including diabetes, pancreatitis, and pancreatic cancer. Despite this association, the mechanistic role of AHR as a linchpin molecule in EDC exposure-related pathogenesis of pancreatic diseases and cancer remains unexplored. This review comprehensively examines the involvement of AHR in EDC exposure-mediated regulation of pancreatic pathogenesis, emphasizing AHR as a potential therapeutic target for the pathogenesis of pancreatic diseases and cancer.

The aryl hydrocarbon receptor differentially modulates the expression profile of antibody isotypes in a human B-cell line.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a persistent environmental contaminant and high affinity ligand for the aryl hydrocarbon receptor (AhR). In animal models, AhR activation by TCDD generally inhibits antibody secretion. However, it is less clear if this translates to human antibody production. Using a human Burkitt lymphoma B-cell line (CL-01) that can be stimulated to secrete Ig and undergo class switch recombination to other Ig isotypes, the current study evaluated the effects of AhR activation or antagonism on the human Ig isotypic expression profile with CD40L+IL-4 stimulation. Our results suggest that AhR agonists (TCDD and indirubin) have little to no effect on IgM or IgA secretion, which were also not induced with stimulation. However, AhR activation significantly inhibited stimulation-induced IgG secretion, an effect reversed by the AhR antagonist CH223191. Evaluation of Ig heavy chain (IgH) constant region gene expression (ie Cμ, Cγ1-4, Cα1-2, and Cε that encode for IgM, IgG1-4, IgA1-2, and IgE, respectively) demonstrated differential effects. While Cμ and Cα2 transcripts were unaffected by stimulation or AhR agonists, AhR activation significantly inhibited stimulation-induced Cγ2-4 and Cε mRNA transcripts, which was reversed by AhR antagonism. Notably, AhR antagonism in the absence of exogenous AhR ligands significantly increased IgG and IgA secretion as well as the expression of Cγ2-4 and Cε. These results suggest that modulation of AhR activity differentially alters the IgH isotypic expression profile and antibody secretion that may be partly dependent on cellular stimulation. Since a variety of chemicals from anthropogenic, industrial, pharmaceutical, dietary, and bacterial sources bind the AhR, the ability of environmental exposures to alter AhR activity (ie activate or inhibit) may have a direct influence on immune function and antibody-relevant disease conditions.

Indoxyl sulfate contributes to colorectal cancer cell proliferation and increased EGFR expression by activating AhR and Akt.

Although patients with chronic kidney disease (CKD) have a higher risk of colorectal cancer (CRC) aggravation, the connection between these two diseases is not well understood. Recent studies have shown that both CKD and CRC aggravation are closely related to an increased abundance of indole-producing Fusobacterium nucleatum in the gut. The indole absorbed from the gut is eventually metabolized to indoxyl sulfate in the liver. Since indoxyl sulfate is involved not only in accelerating CKD progression but also in the initiation and development of its associated complications, the present study aimed to clarify whether indoxyl sulfate induces the proliferation of CRC cells. This study found that indoxyl sulfate induced the proliferation of CRC-derived HCT-116 cells by activating the aryl hydrocarbon receptor (AhR) and the proto-oncogene Akt. The AhR antagonist CH223191 and Akt inhibitor MK2206 suppressed indoxyl sulfate-induced proliferation of HCT-116 cells. We also found that indoxyl sulfate upregulated epidermal growth factor receptor (EGFR) expression, which is associated with poor prognosis of CRC, whereas CH223191 and MK2206 repressed EGFR expression. Furthermore, indoxyl sulfate increased the sensitivity of CRC cells to EGF by upregulating EGFR expression. These findings suggest that indoxyl sulfate may be an important link between CKD and CRC aggravation.

Blockade of aryl hydrocarbon receptor restricts omeprazole-induced chronic kidney disease.

Chronic kidney disease (CKD) is the 16th leading cause of mortality worldwide. Clinical studies have raised that long-term use of omeprazole (OME) is associated with the morbidity of CKD. OME is commonly used in clinical practice to treat peptic ulcers and gastroesophageal reflux disease. However, the mechanism underlying renal failure following OME treatment remains mostly unknown and the rodent model of OME-induced CKD is yet to be established. We described the process of renal injury after exposure to OME in mice; the early renal injury markers were increased in renal tubular epithelial cells (RTECs). And after long-term OME treatment, the OME-induced CKD mice model was established. Herein, aryl hydrocarbon receptor (AHR) translocation appeared after exposure to OME in HK-2 cells. Then for both in vivo and in vitro, we found that Ahr-knockout (KO) and AHR small interfering RNA (siRNA) substantially alleviated the OME-induced renal function impairment and tubular cell damage. Furthermore, our data demonstrate that antagonists of AHR and CYP1A1 could attenuate OME-induced tubular cell impairment in HK-2 cells. Taken together, these data indicate that OME induces CKD through the activation of the AHR-CYP axis in RTECs. Our findings suggest that blocking the AHR-CYP1A1 pathway acts as a potential strategy for the treatment of CKD caused by OME. KEY MESSAGES: We provide an omeprazole-induced chronic kidney disease (CKD) mice model. AHR activation and translocation process was involved in renal tubular damage and promoted the occurrence of CKD. The process of omeprazole nephrotoxicity can be ameliorated by blockade of the AHR-CYP1A1 axis.

Alhagi honey polysaccharide ameliorates ulcerative colitis by modulating gut microbiota–tryptophan metabolism via AhR activation

AbstractAlhagi honey (AH) is produced in arid and hot areas of Central Asia, and its polysaccharides (AP) are widely known for their activity in the treatment of intestinal diseases such as diarrhea. However, the therapeutic potential and mechanism of AP in ulcerative colitis (UC) remain unclear. Here, AH polysaccharide‐2 (AP2), a polysaccharide with the highest content in AP, was isolated and evaluated for its effects on dextran sulfate sodium (DSS)‐induced UC in mice. AP2 was found to alleviate UC symptoms and regulate gut microbiota dysbiosis by decreasing Helicobacter levels and increasing Lactobacillus levels. Analysis of PICRUSt2 predicted that AP2 may regulate carbohydrate and amino acid metabolism, and metabolomic analysis confirmed that AP2 promotes the metabolism of tryptophan to produce kynurenic acid (kyna). Moreover, kyna acted as an aryl hydrocarbon receptor (AhR) ligand, which activated AhR to increase the expression of the tight junction proteins claudin‐1 and occludin. Interestingly, AP2 showed similar effects in protecting the intestinal barrier and alleviating colitis as the AhR agonist 6‐formylindolo[3,2‐b]carbazole, and the AhR antagonist CH223191 partially blocked the therapeutic effect of AP2 in UC mice, indicating that the anti‐UC effect of AP2 was AhR dependent. These findings demonstrate that AP2 alleviates UC by regulating the gut microbiota and promoting tryptophan metabolism to generate kyna‐activated AhR. The insights gained from this study could help in the future development of AP2 as a drug candidate or functional food for the treatment of UC.

Chronic aryl hydrocarbon receptor activity impairs muscle mitochondrial function with tobacco smoking.

Accumulating evidence has demonstrated that chronic tobacco smoking directly contributes to skeletal muscle dysfunction independent of its pathological impact to the cardiorespiratory systems. The mechanisms underlying tobacco smoke toxicity in skeletal muscle are not fully resolved. In this study, the role of the aryl hydrocarbon receptor (AHR), a transcription factor known to be activated with tobacco smoke, was investigated. AHR related gene (mRNA) expression was quantified in skeletal muscle from adult controls and patients with chronic obstructive pulmonary disease (COPD), as well as mice with and without cigarette smoke exposure. Utilizing both skeletal muscle-specific AHR knockout mice exposed to chronic repeated (5days per week for 16weeks) cigarette smoke and skeletal muscle-specific expression of a constitutively active mutant AHR in healthy mice, a battery of assessments interrogating muscle size, contractile function, mitochondrial energetics, and RNA sequencing were employed. Skeletal muscle from COPD patients (N=79, age=67.0±8.4years) had higher levels of AHR (P=0.0451) and CYP1B1 (P<0.0001) compared to healthy adult controls (N=16, age=66.5±6.5years). Mice exposed to cigarette smoke displayed higher expression of Ahr (P=0.008), Cyp1b1 (P<0.0001), and Cyp1a1 (P<0.0001) in skeletal muscle compared to air controls. Cigarette smoke exposure was found to impair skeletal muscle mitochondrial oxidative phosphorylation by ~50% in littermate controls (Treatment effect, P<0.001), which was attenuated by deletion of the AHR in muscle in male (P=0.001), but not female, mice (P=0.37), indicating there are sex-dependent pathological effects of smoking-induced AHR activation in skeletal muscle. Viral mediated expression of a constitutively active mutant AHR in the muscle of healthy mice recapitulated the effects of cigarette smoking by decreasing muscle mitochondrial oxidative phosphorylation by ~40% (P=0.003). These findings provide evidence linking chronic AHR activation secondary to cigarette smoke exposure to skeletal muscle bioenergetic deficits in male, but not female, mice. AHR activation is a likely contributor to the decline in muscle oxidative capacity observed in smokers and AHR antagonism may provide a therapeutic avenue aimed to improve muscle function in COPD.

Wogonin improves colitis by activating the AhR pathway to regulate the plasticity of ILC3/ILC1

BackgroundIntestinal barrier dysfunction caused by the disrupted balance of group 3 innate lymphoid cells (ILC3)/group 1 innate lymphoid cells (ILC1) is a significant feature in the pathogenesis of inflammatory bowel disease (IBD). Activation of aryl hydrocarbon receptor (AhR) signaling contributes to the maintenance of ILC3/ILC1 balance. Wogonin, a natural flavonoid from Scutellaria baicalensis Georgi, can repair intestinal mucosal damage of IBD. However, it remains unclear if wogonin can exert a therapeutic effect by activating the AhR pathway to regulate the plasticity of ILC3/ILC1. PurposeIn this study, we investigated the immunomodulatory effects of wogonin on IBD and its potential mechanisms in vitro and in vivo. Study design and MethodsChronic colitis was induced by four cycles of 2 % DSS treatment in mice. 20 mg kg-1/day wogonin was administrated by oral gavage and mice were treated intraperitoneally with 10 mg kg-1/2 days CH223191 to block the AhR pathway. Colon tissues were processed for histopathological examination and evaluation of the epithelial barrier function by immunohistochemistry. The activation of the AhR pathway and the plasticity of ILC3/ILC1 were determined by western blot and flow cytometry. Then, we also detected the intestinal microflora and their metabolites by 16 s sequencing and non-targeted Metabolomics analysis. Furthermore, an in vitro culture system consisting of MNK3 cells and NCM460 cells, and a CETSA assay were performed to confirm the molecular mechanism. ResultsWogonin ameliorated histological severity of the colon, decreased the secretion of inflammatory factors, and increased tight junction proteins in colitis mice. These effects are associated with the tendency of conversion from ILC3 to ILC1 prevented by wogonin, which was offset by AhR antagonist CH223191. In addition, wogonin exerted the curative effect by altering gut microbiota to produce metabolites such as Kynurenic acid, and 1H-Indole-3-carboxaldehyde as AhR endogenous ligands. In vitro data further verified that wogonin as an exogenous ligand directly binds to the structural domain of AhR by CETSA. Also, the supernatant of MNK-3 cells stimulated with wogonin enhanced expression of Occludin and Claudin1 in NCM460 cells induced by LPS. ConclusionCumulatively, our study illustrated that wogonin improved the outcomes of DSS-induced chronic colitis via regulating the plasticity of ILC3/ILC1. Its specific mechanism is to binding to AhR directly, and to activate the AhR pathway indirectly by altering the tryptophan metabolisms of gut microbiota.

PHGDH knockdown increases sensitivity to SR1, an aryl hydrocarbon receptor antagonist, in colorectal cancer by activating the autophagy pathway.

Colorectal cancer (CRC) has emerged as the third most prevalent and second deadliest cancer worldwide. Metabolic reprogramming is a key hallmark of cancer cells. Phosphoglycerate dehydrogenase (PHGDH) is over-expressed in multiple cancers, including CRC. Although the role of PHGDH in metabolism has been extensively investigated, its effects on CRC development remains to be elucidated. In the present study, it was demonstrated that PHGDH expression was significantly up-regulated in colorectal cancer. PHGDH expression was positively correlated with that of the aryl hydrocarbon receptor (AhR) and its target genes, CYP1A1 and CYP1B1, in CRC cells. Knockdown of PHGDH reduced AhR levels and activity, as well as the ratio of reduced to oxidized glutathione. The selective AhR antagonist stemregenin 1 induced cell death through reactive oxygen species-dependent autophagy in CRC cells. PHGDH knockdown induced CRC cell sensitivity to stemregenin 1 via the autophagy pathway. Our findings suggest that PHGDH modulates AhR signaling and the redox-dependent autophagy pathway in CRC, and that the combination of inhibition of both PHGDH and AhR may be a novel therapeutic strategy for CRC.

TCDD and CH223191 Alter T Cell Balance but Fail to Induce Anti-Inflammatory Response in Adult Lupus Mice.

Aryl hydrocarbon receptor (AhR) responds to endogenous and exogenous ligands as a cytosolic receptor, transcription factor, and E3 ubiquitin ligase. Several studies support an anti-inflammatory effect of AhR activation. However, exposure to the AhR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) during early stages of development results in an autoimmune phenotype and exacerbates lupus. The effects of TCDD on lupus in adults with pre-existing autoimmunity have not been described. We present novel evidence that AhR stimulation by TCDD alters T cell responses but fails to impact lupus-like disease using an adult mouse model. Interestingly, AhR antagonist CH223191 also changed T cell balance in our model. We next developed a conceptual framework for identifying cellular and molecular factors that contribute to physiological outcomes in lupus and created models that describe cytokine dynamics that were fed into a system of differential equations to predict the kinetics of T follicular helper (Tfh) and regulatory T (Treg) cell populations. The model predicted that Tfh cells expanded to larger values following TCDD exposure compared with vehicle and CH223191. Following the initial elevation, both Tfh and Treg cell populations continuously decayed over time. A function based on the ratio of predicted Treg/Tfh cells showed that Treg cells exceed Tfh cells in all groups, with TCDD and CH223191 showing lower Treg/Tfh cell ratios than the vehicle and that the ratio is relatively constant over time. We conclude that AhR ligands did not induce an anti-inflammatory response to attenuate autoimmunity in adult lupus mice. This study challenges the dogma that TCDD supports an immunosuppressive phenotype.

The Role of the Aryl Hydrocarbon Receptor in Vascular Factors Related to Preeclampsia in a Smoking Mouse Model.

Smoking cigarettes is known to lower the risk of preeclampsia. The objective of this study is to evaluate the effect of smoking on the expression of soluble FMS-like tyrosine kinase-1 (sFlt-1), vascular endothelial growth factor (VEGF), and endoglin (sEng)-1 and the role of the aryl hydrocarbon receptor (AhR) in pregnant mice. We developed a smoking mouse model using a gas-filling system. One or two cigarettes per day were exposed to each of the five pregnant mice for five days a week throughout pregnancy. AhR agonist and antagonist were injected. Serum levels and expression in the placenta of sFlt-1, VEGF, and sEng-1 were analyzed and compared among the cigarette smoke and no-exposure groups after delivery. Compared to the no-smoke exposure group, the serum level of sFlt-1 was significantly decreased in the two-cigarette-exposed group (p < 0.001). When the AhR antagonist was added to the two-cigarette-exposed group, sFlt-1 levels were significantly increased compared to the two-cigarette group (p = 0.002). The levels of sFlt-1 in the AhR antagonist group did not change regardless of two-cigarette exposure (p = 0.064). With the AhR agonist, sFlt-1 decreased significantly compared to the control (p = 0.001) and AhR antagonist group (p = 0.002). The sFlt-1 level was significantly decreased after the injection of the AhR agonist compared to the control group (p = 0.001). Serum levels of VEGF were significantly decreased in the one-cigarette-exposed group compared to the control group; however, there was no difference between the control and the two-cigarette-exposed groups. The placental expression of sFlt-1, VEGF, and sEng were inconsistent. This study offers insights into the potential role of AhR on antiangiogenic sFlt-1 associated with preeclampsia. It may support the invention of a new treatment strategy for preeclampsia using AhR activation.

Halogenation of common phenolic household and personal care product ingredients enhances their AhR-modulating capacity

The Aryl Hydrocarbon Receptor (AhR), a ligand-activated transcription factor, orchestrates responses to numerous structurally diverse endogenous and exogenous ligands. In addition to binding various xenobiotics, AhR also recognizes endocrine disruptors, particularly those featuring chlorinated or brominated aromatic structures. There is limited data available on the impact of common household and personal care product ingredients let alone their halogenated transformation products. Herein we bridge this knowledge gap by preparing a library of chlorinated and brominated parabens, bisphenols, UV filters, and nonylphenols. An evaluation of total of 125 compounds for agonistic and antagonistic activity on AhR unveiled a low micromolar agonist, Cl2BPAF with an EC50 of 13 μM. Moreover, our study identified several AhR antagonists, with BrBzP emerging as the most potent with an IC50 of 8.9 μM. To further investigate the functional implications of these compounds, we subjected the most potent agonist and antagonist to a functional assay involving cytokine secretion from peripheral blood mononuclear cells and compared their activity with the commercially available AhR agonist and antagonist. Cl2BPAF exhibited an overall immunosuppressive effect by reducing the secretion of proinflammatory cytokines, including IL-6, IFN-γ, and TNF-α, while BrBzP displayed opposite effects, leading to an increase of those cytokines. Notably, the immunomodulatory effects of Cl2BPAF surpassed those of ITE, a bona fide AhR agonist, while the impact of BrBzP exceeded that of CH223191, a bona fide AhR antagonist. In summary, our study underscores the potential influence of halogenated transformation products on the AhR pathway and, consequently, their role in shaping the immune responses.

The role of the aryl hydrocarbon receptor in mediating the effects of mono(2-ethylhexyl) phthalate in mouse ovarian antral follicles†.

Di(2-ethylhexyl) phthalate (DEHP) is a pervasive environmental toxicant used in the manufacturing of numerous consumer products, medical supplies, and building materials. DEHP is metabolized to mono(2-ethylhexyl) phthalate (MEHP). MEHP is an endocrine disruptor that adversely affects folliculogenesis and steroidogenesis in the ovary, but its mechanism of action is not fully understood. Thus, we tested the hypothesis that the aryl hydrocarbon receptor (AHR) plays a functional role in MEHP-mediated disruption of folliculogenesis and steroidogenesis. CD-1 mouse antral follicles were isolated and cultured with MEHP (0-400μM) in the presence or absence of the AHR antagonist CH223191 (1μM). MEHP treatment reduced follicle growth over a 96-h period, and this effect was partially rescued by co-culture with CH223191. MEHP exposure alone increased expression of known AHR targets, cytochrome P450 (CYP) enzymes Cyp1a1 and Cyp1b1, and this induction was blocked by CH223191. MEHP reduced media concentrations of estrone and estradiol compared to control. This effect was mitigated by co-culture with CH223191. Moreover, MEHP reduced the expression of the estrogen-sensitive genes progesterone receptor (Pgr) and luteinizing hormone/choriogonadotropin receptor (Lhcgr) and co-treatment with CH223191 blocked this effect. Collectively, these data indicate that MEHP activates the AHR to impair follicle growth and reduce estrogen production and signaling in ovarian antral follicles.

Therapeutic effects of in vivo administration of an inhibitor of tryptophan 2,3-dioxygenase (680c91) for the treatment of fibroids: a preclinical study

ObjectiveFibroids are characterized by marked overexpression of Tryptophan 2,3 dioxygenase (TDO2). The objective of this study was to determine the effectiveness of in vivo administration of an inhibitor of TDO2 (680C91) on fibroid size and gene expression. DesignAnimal and ex vivo human study. SubjectsSCID mice bearing human fibroid xenografts treated with vehicle and TDO2 inhibitor. InterventionDaily intraperitoneal administration of 680C91 or vehicle for two months and in vitro studies with fibroid explants. Main Outcome MeasuresTumor weight and gene expression profile of xenografts and in vitro mechanistic experiments using fibroid explants. ResultsCompound 680C91 was well-tolerated with no effects on blood chemistry and body weight. Treatment of mice with 680C91 resulted in 30% reduction in the weight of fibroid xenografts after two months of treatment and as expected lower levels of kynurenine, the byproduct of tryptophan degradation and an endogenous ligand of AhR (aryl hydrocarbon receptor) in the xenografts. The expression of CYP1B1 (cytochrome P450 family 1 subfamily B member 1), TGF-β3 (transforming growth factor beta 3), FN1 (fibronectin), CDK2 (cyclin-dependent kinase 2), E2F1 (E2F transcription factor 1), IL-8 (interleukin 8) and SPARC (secreted protein acidic and cysteine rich) mRNA, were lower in the xenografts of mice treated with 680C91 as compared with vehicle controls. Similarly, the protein abundance of collagen, FN1, CYP1B1 and SPARC were lower in the xenografts of 680C91 treated mice as compared with vehicle controls. Immunohistochemical analysis of xenografts indicated decreased expression of collagen, Ki67 and E2F1 but no significant changes in cleaved caspase 3 expression in mice treated with 680C91. The levels of kynurenine in the xenografts showed a direct correlation with the tumor weight and FN1 levels. In vitro studies with fibroid explants showed a significant induction of CYP1B1, TGF-β3, FN1, CDK2, E2F1, IL8 and SPARC mRNA by tryptophan which could be blocked by co-treatment with 680C91 and the AhR antagonist CH-223191. ConclusionThe results indicate that correction of aberrant tryptophan catabolism in fibroids could be an effective treatment through its effect to reduce cell proliferation and ECM accumulation.

The aryl hydrocarbon receptor controls mesenchymal stromal cell-mediated immunomodulation via ubiquitination of eukaryotic elongation factor-2 kinase

Mesenchymal stem cells (MSCs) have great therapeutic advantages due to their immunosuppressive properties. The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor whose signaling plays an important role in the immune system. AHR may be involved in the regulation of MSC-associated immunomodulatory functions. However, the mechanisms by which AHR controls the immunosuppressive functions of MSCs are not well understood. Here, we report that Ahr-deficient MSCs show decreased therapeutic efficacy against graft-versus-host disease (GVHD) compared to wild-type (WT)-MSCs. This was probably due to decreased iNOS protein expression, which is a key regulatory enzyme in MSC immunomodulation. The expression of eukaryotic elongation factor 2 kinase (eEF2K), which inhibits the elongation stage of protein synthesis, is significantly increased in the Ahr-deficient MSCs. Inhibition of eEF2K restored iNOS protein expression. AHR is known to act as an E3 ligase together with CUL4B. We observed constitutive binding of AHR to eEF2K. Consequently, ubiquitination and degradation of eEF2K were inhibited in Ahr-deficient MSCs and by the AHR antagonist CH223191 in WT-MSCs. In summary, AHR regulates the immunomodulatory functions of MSCs through ubiquitination of eEF2K, thereby controlling iNOS protein synthesis and its product, nitric oxide levels.

Inhibitory Effects of Urolithins, Bioactive Gut Metabolites from Natural Polyphenols, against Glioblastoma Progression

We previously reported that proinflammatory cytokines, particularly tumor necrosis factor (TNF)-α, promoted tumor migration, invasion, and proliferation, thus worsening the prognosis of glioblastoma (GBM). Urolithins, the potent metabolites produced by the gut from pomegranate polyphenols, have anticancer properties. To develop an effective therapy for GBM, this study aimed to study the effects of urolithins against GBM. Urolithin A and B significantly reduced GBM migration, reduced epithelial–mesenchymal transition, and inhibited tumor growth. Moreover, urolithin A and B inhibited TNF-α-induced vascular cell adhesion molecule (VCAM)-1 and programmed death ligand 1 (PD-L1) expression, thereby reducing human monocyte (HM) binding to GBM cells. Aryl hydrocarbon receptor (AhR) level had higher expression in patients with glioma than in healthy individuals. Urolithins are considered pharmacological antagonists of AhR. We demonstrated that the inhibition of AhR reduced TNF-α-stimulated VCAM-1 and PD-L1 expression. Furthermore, human macrophage condition medium enhanced expression of PD-L1 in human GBM cells. Administration of the AhR antagonist attenuated the enhancement of PD-L1, indicating the AhR modulation in GBM progression. The modulatory effects of urolithins in GBM involve inhibiting the Akt and epidermal growth factor receptor pathways. The present study suggests that urolithins can inhibit GBM progression and provide valuable information for anti-GBM strategy.