AhR Activation Research Articles

Environmental behavior and safety of polyhalogenated carbazoles (PHCZs): A review.

Polyhalogenated carbazoles (PHCZs) are well-known as emergent environmental contaminants. Given their wide distribution in the environment and structural similarity with dioxins and dioxin-like chemicals (DLCs), the environmental behavior and ecological risks of these chemicals have become the major issue concerned by the governments and scientists. Since the initial report of PHCZ residues in the environment in the 1980s, over 20 PHCZ congeners with different residual levels had been identified in various environmental media all over the world. Nevertheless, researches concerning the toxicological effects of PHCZs are of an urgent need for the relatively lagging study of their ecological risks. Currently, only limited evidence has indicated that PHCZs would pose dioxin-like toxicity, including developmental toxicity, cardiotoxicity, etc; and their toxicological effects were partially consistent with AhR activation. And yet, much remains to be done to fill in the knowledge gaps of their toxicological effects. In this review, the research progresses in environmental behavior and toxicology study of PHCZs were remarked; and the lack of current research, as well as future research prospects, were discussed.

Coffee induces AHR- and Nrf2-mediated transcription in intestinal epithelial cells

Coffee induces a health-promoting adaptive response of cells in the body. Here, we investigated enterocyte responses to AHR agonists in coffee and measured their transport across a polarized intestinal epithelium. AHR-activating potencies of Turkish, filter, and instant coffee were determined using DR CALUX® bioassay, before and after intestinal metabolization by Caco-2 cells. Furthermore, effects of coffee on induction of AHR- and Nrf2-pathway genes in Caco-2 cells were evaluated by real-time qPCR. Coffee samples showed considerable AHR-activating potencies in DR CALUX® bioassay (up to 79% of positive control activity). After incubation with Caco-2 cells, AHR activity of different coffees was between 35 and 64% of their initial value, suggesting rapid uptake and metabolization by epithelial cells. Expression of AHR-regulated gene CYP1A1 increased up to 41-fold and most Nrf2-pathway genes were up-regulated by coffee. This in vitro study may support the notion that coffee bioactives contribute to antioxidant defense and detoxification processes in vivo.

A toxicity pathway-oriented approach to develop adverse outcome pathway: AHR activation as a case study.

With numerous new chemicals introduced into the environment everyday, identification of their potential hazards to the environment and human health is a considerable challenge. Developing adverse outcome pathway (AOP) framework is promising in helping to achieve this goal as it can bring InVitro testing into toxicity measurement and understanding. To explore the toxic mechanism underlying environmental chemicals via the AOP approach, an integration of adequate experimental data with systems biology understanding is preferred. Here, we describe a novel method to develop reliable and sensible AOPs that relies on chemical-gene interactions, toxicity pathways, molecular regulations, phenotypes, and outcomes information obtained from comparative toxicogenomics database (CTD) and Ingenuity Pathway Analysis (IPA). Using Benzo(a)pyrene (BaP), a highly studied chemical as a stressor, we identified the pivotal IPA toxicity pathways, the molecular initiating event (MIE), and candidate key events (KEs) to structure AOPs in the liver and lung, respectively. Further, we used the corresponding CTD information of multiple typical AHR-ligands, including 2,3,7,8-tetrachlorodibenzoparadioxin (TCDD), valproic acid, quercetin, and particulate matter, to validate our AOP networks. Our approach is likely to speed up AOP development as providing a time- and cost-efficient way to collect all fragmented bioinformation in published studies. It also facilitates a better understanding of the toxic mechanism of environmental chemicals, and potentially brings new insights into the screening of critical paths in the AOP network.

AhR and IDO1 in pathogenesis of Covid-19 and the "Systemic AhR Activation Syndrome:" a translational review and therapeutic perspectives.

Covid-19 is the acute illness caused by SARS-CoV-2 with initial clinical symptoms such as cough, fever, malaise, headache, and anosmia. After entry into cells, corona viruses (CoV) activate aryl hydrocarbon receptors (AhRs) by an indoleamine 2,3-dioxygenase (IDO1)-independent mechanism, bypassing the IDO1-kynurenine-AhR pathway. The IDO1-kynurenine-AhR signaling pathway is used by multiple viral, microbial and parasitic pathogens to activate AhRs and to establish infections. AhRs enhance their own activity through an IDO1-AhR-IDO1 positive feedback loop prolonging activation induced by pathogens. Direct activation of AhRs by CoV induces immediate and simultaneous up-regulation of diverse AhR-dependent downstream effectors, and this, in turn, results in a “Systemic AhR Activation Syndrome” (SAAS) consisting of inflammation, thromboembolism, and fibrosis, culminating in multiple organ injuries, and death. Activation of AhRs by CoV may lead to diverse sets of phenotypic disease pictures depending on time after infection, overall state of health, hormonal balance, age, gender, comorbidities, but also diet and environmental factors modulating AhRs. We hypothesize that elimination of factors known to up-regulate AhRs, or implementation of measures known to down-regulate AhRs, should decrease severity of infection. Although therapies selectively down-regulating both AhR and IDO1 are currently lacking, medications in clinical use such as dexamethasone may down-regulate both AhR and IDO1 genes, as calcitriol/vitamin D3 may down-regulate the AhR gene, and tocopherol/vitamin E may down-regulate the IDO1 gene. Supplementation of calcitriol should therefore be subjected to epidemiological studies and tested in prospective trials for prevention of CoV infections, as should tocopherol, whereas dexamethasone could be tried in interventional trials. Because lack of physical exercise activates AhRs via the IDO1-kynurenine-AhR signaling pathway increasing risk of infection, physical exercise should be encouraged during quarantines and stay-at-home orders during pandemic outbreaks. Understanding which factors affect gene expression of both AhR and IDO1 may help in designing therapies to prevent and treat humans suffering from Covid-19.

Aryl Hydrocarbon Receptor Activity in Hepatocytes Sensitizes to Hyperacute Acetaminophen-Induced Hepatotoxicity in Mice.

Background & AimsAcetaminophen (APAP)-induced liver injury is one of the most common causes of acute liver failure, however, a clear definition of sensitizing risk factors is lacking. Here, we investigated the role of the ligand-activated transcription factor aryl hydrocarbon receptor (Ahr) in APAP-induced liver injury. We hypothesized that Ahr, which integrates environmental, dietary, microbial and metabolic signals into complex cellular transcriptional programs, might act as a rheostat for APAP-toxicity.MethodsWildtype or conditional Ahr knockout mice lacking Ahr in hepatocytes (AlbΔ/ΔAhr) or myeloid cells (LysMΔ/ΔAhr) were treated with the specific Ahr ligand 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) together with APAP.ResultsAhr activation by ITE, which by itself was non-toxic, exacerbated APAP-induced hepatotoxicity compared to vehicle-treated controls, causing 80% vs. 0% mortality after administration of a normally sublethal APAP overdose. Of note, Ahr activation induced hepatocyte death even at APAP doses within the therapeutic range. Aggravated liver injury was associated with significant neutrophil infiltration; however, lack of Ahr in myeloid cells did not protect LysMΔ/ΔAhr mice from exacerbated APAP hepatotoxicity. In contrast, AlbΔ/ΔAhr mice were largely protected from ITE-induced aggravated liver damage, indicating that Ahr activation in hepatocytes, but not in myeloid cells, was instrumental for disease exacerbation. Mechanistically, Ahr activation fueled hepatic accumulation of toxic APAP metabolites by up-regulating expression of the APAP-metabolizing enzyme Cyp1a2, a direct Ahr downstream target.ConclusionsAhr activation in hepatocytes potentiates APAP-induced hepatotoxicity. Thus, individual exposition to environmental Ahr ligands might explain individual sensitivity to hyperacute liver failure.

Systematic identification of a nuclear receptor-enriched predictive signature for erastin-induced ferroptosis

Erastin, a synthetic lethal compound against cancer expressing an oncogenic RAS, inhibits cystine/glutamate antiporters and causes ferroptosis. However, despite recent evidence for the mechanisms underlying ferroptosis, molecular biomarkers of erastin-dependent ferroptosis have not been identified. Here, we employed isogenic lung cancer cell models to show that a redox imbalance leads to glutathione depletion and ferroptosis. Subsequent transcriptome analysis of pan-cancer cell lines revealed that the activity of transcription factors, including NRF2 and AhR, serve as important markers of erastin resistance. Based on the integrated expression of genes in the nuclear receptor meta-pathway (NRM), we constructed an NRM model and validated its robustness using an independent pharmacogenomics dataset. The NRM model was further evaluated by sensitivity tests on nine cancer cell lines for which erastin sensitivities had not been determined. Our pharmacogenomics approach has the potential to pave the way for the efficient classification of patients for therapeutic intervention using erastin.

CYP2S1 is a synthetic lethal target in BRAFV600E-driven thyroid cancers

BRAFV600E is the most common genetic alteration and has become a major therapeutic target in thyroid cancers; however, intrinsic feedback mechanism limited clinical use of BRAFV600E specific inhibitors. Synthetic lethal is a kind of interaction between two genes, where only simultaneously perturbing both of the genes can lead to lethality. Here, we identified CYP2S1 as a synthetic lethal partner of BRAFV600E in thyroid cancers. First, we found that CYP2S1 was highly expressed in papillary thyroid cancers (PTCs) compared to normal thyroid tissues, particularly in conventional PTCs (CPTCs) and tall-cell PTCs (TCPTCs), and its expression was positively associated with BRAFV600E mutation. CYP2S1 knockdown selectively inhibited cell proliferation, migration, invasion and tumorigenic potential in nude mice, and promoted cell apoptosis in BRAFV600E mutated thyroid cancer cells, but not in BRAF wild-type ones. Mechanistically, BRAFV600E-mediated MAPK/ERK cascade upregulated CYP2S1 expression by an AHR-dependent pathway, while CYP2S1 in turn enhanced transcriptional activity of AHR through its metabolites. This AHR/CYP2S1 feedback loop strongly amplified oncogenic role of BRAFV600E in thyroid cancer cells, thereby causing synthetic lethal interaction between CYP2S1 and BRAFV600E. Finally, we demonstrated CYP2S1 as a potential therapeutic target in both BRAFV600E-drived xenograft and transgenic mouse models by targetedly delivering CYP2S1-specific siRNA. Altogether, our data demonstrate CYP2S1 as a synthetic lethal partner of BRAFV600E in thyroid cancers, and indicate that targeting CYP2S1 will provide a new therapeutic strategy for BRAFV600E mutated thyroid cancers.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) dysregulates hepatic one carbon metabolism during the progression of steatosis to steatohepatitis with fibrosis in mice

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a persistent environmental contaminant, induces steatosis that can progress to steatohepatitis with fibrosis, pathologies that parallel stages in the development of non-alcoholic fatty liver disease (NAFLD). Coincidently, one carbon metabolism (OCM) gene expression and metabolites are often altered during NAFLD progression. In this study, the time- and dose-dependent effects of TCDD were examined on hepatic OCM in mice. Despite AhR ChIP-seq enrichment at 2 h, OCM gene expression was not changed within 72 h following a bolus dose of TCDD. Dose-dependent repression of methionine adenosyltransferase 1A (Mat1a), adenosylhomocysteinase (Achy) and betaine-homocysteine S-methyltransferase (Bhmt) mRNA and protein levels following repeated treatments were greater at 28 days compared to 8 days. Accordingly, levels of methionine, betaine, and homocysteic acid were dose-dependently increased, while S-adenosylmethionine, S-adenosylhomocysteine, and cystathionine exhibited non-monotonic dose-dependent responses consistent with regulation by OCM intermediates and repression of glycine N-methyltransferase (Gnmt). However, the dose-dependent effects on SAM-dependent metabolism of polyamines and creatine could not be directly attributed to alterations in SAM levels. Collectively, these results demonstrate persistent AhR activation disrupts hepatic OCM metabolism at the transcript, protein and metabolite levels within context of TCDD-elicited progression of steatosis to steatohepatitis with fibrosis.

Epigenetic Regulations of AhR in the Aspect of Immunomodulation.

Environmental factors contribute to autoimmune disease manifestation, and as regarded today, AhR has become an important factor in studies of immunomodulation. Besides immunological aspects, AhR also plays a role in pharmacological, toxicological and many other physiological processes such as adaptive metabolism. In recent years, epigenetic mechanisms have provided new insight into gene regulation and reveal a new contribution to autoimmune disease pathogenesis. DNA methylation, histone modifications, chromatin alterations, microRNA and consequently non-genetic changes in phenotypes connect with environmental factors. Increasing data reveals AhR cross-roads with the most significant in immunology pathways. Although study on epigenetic modulations in autoimmune diseases is still not well understood, therefore future research will help us understand their pathophysiology and help to find new therapeutic strategies. Present literature review sheds the light on the common ground between remodeling chromatin compounds and autoimmune antibodies used in diagnostics. In the proposed review we summarize recent findings that describe epigenetic factors which regulate AhR activity and impact diverse immunological responses and pathological changes.

The integrated use of in silico methods for the hepatotoxicity potential of Piper methysticum

Herbal products as supplements and therapeutic intervention have been used for centuries. However, their toxicities are not completely evaluated and the mechanisms are not clearly understood. Dried rhizome of the plant kava (Piper methysticum) is used for its anxiolytic, and sedative effects. The drug is also known for its hepatotoxicity potential. Major constituents of the plant were identified as kavalactones, alkaloids and chalcones in previous studies. Kava hepatotoxicity mechanism and the constituent that causes the toxicity have been debated for decades. In this paper, we illustrated the use of computational tools for the hepatotoxicity of kava constituents. The proposed mechanisms and major constituents that are most probably responsible for the toxicity have been scrutinized. According to the experimental and prediction results, the kava constituents play a substantial role in hepatotoxicity by some means or other via glutathione depletion, CYP inhibition, reactive metabolite formation, mitochondrial toxicity and cyclooxygenase activity. Some of the constituents, which have not been tested yet, were predicted to involve mitochondrial membrane potential, caspase-3 stimulation, and AhR activity. Since Nrf2 activation could be favorable for prevention of hepatotoxicity, we also suggest that these compounds should undergo testing given that they were predicted not to be activating Nrf2. Among the major constituents, alkaloids appear to be the least studied and the least toxic group in general. The outcomes of the study could help to appreciate the mechanisms and to prioritize the kava constituents for further testing.

Dietary Supplementation of Foxtail Millet Ameliorates Colitis-Associated Colorectal Cancer in Mice via Activation of Gut Receptors and Suppression of the STAT3 Pathway.

Coarse cereal intake has been reported to be associated with reduced risk of colorectal cancer. However, evidence from intervention studies is absent and the molecular basis of this phenomenon remains largely unexplored. This study sought to investigate the effects of foxtail millet and rice, two common staple grains in Asia, on the progression of colitis-associated colorectal cancer (CAC) and define the mechanism involved. In total, 40 BALB/c mice were randomized into four groups. The Normal and azoxymethane/dextran sodium sulfate (AOM/DSS) groups were supplied with an AIN-93G diet, while the millet- and rice-treated groups were supplied with a modified AIN-93G diet. Compared to the AOM/DSS-induced CAC mice supplemented with rice, an increased survival rate, suppressed tumor burden, and reduced disease activity index were observed in the millet-treated group. The levels of IL-6 and IL-17 were decreased in the millet-treated group compared to both the AOM/DSS and AOM/DSS + rice groups. Millet treatment inhibited the phosphorylation of STAT3 and the related signaling proteins involved in cell proliferation, survival and angiogenesis. These beneficial effects were mediated by the activation of gut receptors AHR and GPCRs via the microbial metabolites (indole derivates and short-chain fatty acids) of foxtail millet. Moreover, millet-treatment increased the abundance of Bifidobacterium and Bacteroidales_S24-7 compared to the rice-treated mice. This study could help researchers to develop better dietary patterns that work against inflammatory bowel disease (IBD) and for CAC patients.

A set of six Gene expression biomarkers and their thresholds identify rat liver tumorigens in short-term assays

Traditional methods for cancer risk assessment are retrospective, resource-intensive, and not feasible for the vast majority of environmental chemicals. In earlier studies, we used a set of six biomarkers to accurately identify liver tumorigens in transcript profiles derived from chemically-treated rats using either a Toxicological Priority Index (ToxPi) approach or using derived biomarker thresholds for cancer. The biomarkers consisting of 7–113 genes are used to predict the most common liver cancer molecular initiating events: genotoxicity, cytotoxicity and activation of the xenobiotic receptors AhR, CAR, ER, and PPARα. In the present study, we apply and evaluate the performance of these methods for cancer prediction in an independent rat liver study of 44 chemicals (6 h-7d exposures) examined by Affymetrix arrays. In the first approach, ToxPi ranking of biomarker scores consistently gave the highest scores to tumorigenic chemical-dose pairs; balanced accuracies for identification of liver tumorigenic chemicals were up to 89 %. The second approach used tumorigenic thresholds derived in the present study or from our earlier study that were set at the maximum value for chemical-dose exposures without detectable liver tumor outcomes. Using these thresholds, balanced accuracies were up to 90 %. Both approaches identified all tumorigenic chemicals. Almost all of the tumorigenic chemicals activated more than one MIE. We also compared biomarker responses between two types of profiling platforms (Affymetrix full-genome array, TempO-Seq 1500+ array containing ∼2600 genes) and found that the lack of the full set of biomarker genes on the 1500+ array resulted in decreased ability to identify chemicals that activate the MIEs. Overall, these results demonstrate that predictive approaches based on the 6 biomarkers could be used in short-term assays to identify chemicals and their doses that induce liver tumors, the most common endpoint in rodent bioassays.

620 UVB radiation with high photon density induces dendritic cell maturation and contributes to cutaneous immune suppression via Treg cell expansion

Phototherapy is an important treatment modality used in dermatology practice, and UVB radiation is often used for treating inflammatory skin disorders due to its immune suppressive effects. Currently, several UVB emitting radiation with different irradiance (W/cm2) are available. Clinical reports have shown different response profiles from devices emitting similar wavelengths at different irradiance. We previously showed that at equivalent fluence (mJ/cm2), high irradiance (HI) UVB imparted more immune suppressive effect as compared to its low irradiance (LI) counterpart. The current study was launched to explore the mechanisms involved. Using animal model, we showed that at equivalent fluence, HIUVB induced significantly higher immunosuppressive effect as compared to its LIUVB counterpart. Moreover, induction of more T regulatory cells (Treg) was noted in HIUVB treated group as compared to its LIUVB treated counterpart. It was recognized that dendritic cell (DC) maturation after UVB induces Treg cell proliferation. Using cell model, we showed that at equivalent fluence, HIUVB induced significantly higher HLA-DR, C80, CD86 expression on DCs as compared to its LIUVB counterpart. Additionally, at equivalent fluence, conditioned media (CM) from keratinocytes (KC) treated with HIUVB induced significantly more DC maturation as compared to CM derived from KC treated with equivalent fluence of LIUVB. Previously, aryl hydrocarbon receptor (Ahr) activation was shown to play a part on DC induced immune suppression. HIUVB treated DC and KC showed higher activation of Ahr as compared to their respective counterparts treated with equivalent fluence UVB at LI. Taken together, our results suggest that at equivalent fluence, HIUVB is more capable of immune suppression through induction of Treg expansion as compared to ts LIUVB counterpart, and differential DC modulation due to difference in UVB irradiance contributed significantly to the observed phenomenon.

A mixture of Lactobacillus sp. modulates the expression of inflammatory molecules, signalling kinases and nuclear receptors in LPS-treated Caco-2 cell culture model

Abstract The treatment of intestinal inflammation pathologies (also known as Inflammatory Bowel Diseases, IBD) has included a large variety of strategies, from pharmaceutical to traditional medicine and dietary therapies. In the last years, numerous efforts were undertaken to demonstrate the health promoting activities of probiotics in intestinal inflammation and more other pathologic conditions. The aim of our study was to evaluate the effects of a probiotic mixture of Lactobacillus sp. on the inflammatory mediators and signalling pathways as well as nuclear receptors in colonic Caco-2 cells. Human adenocarcinoma Caco-2 cells were challenged in vitro with lipopolysaccharide (LPS) for 4 hours for the induction of inflammation. The LPS-treated cells were cultured for additional 24 hours in the presence of Lactobacillus (Lb) mixture (3 x108 CFU/mL total Lb). Genomic and proteomic array approaches were used to analyse the profile expression of 18 key genes and their proteins involved in intestinal inflammatory response (chemokines, adhesion molecules, growth factors and matrix metalloproteinases inhibitors) as well as signalling markers (Akt, GSK) and nuclear receptors (NF-kB/RELA, Nrf2, AhR). Our study demonstrated that the probiotic Lactobacillus mixture could decrease LPS-induced inflammatory mediator expressions (chemokines, growth factors and matrix metalloproteinases inhibitor) at gene and protein level. This down-regulation exerted by Lb. mix in LPS-treated Caco-2 cells seemed to be regulated through inhibition of both the PI3K/AKT and NF-κB signalling pathways. Additionally, AhR activation induced by LPS was reduced by probiotic mixture under the level of LPS-treated cells. These beneficial effects of Lactobacillus mixture support their use as inflammatory modulators in intestinal disorders.

SAT-133 Breast Tumor Kinase (Brk/PTK6) Mediates Triple Negative Breast Cancer Cell Migration and Taxol Resistance via SH2 Domain-Dependent Activation of RhoA and AhR

Triple negative breast cancer (TNBC) patients have higher recurrence rates and a worse prognosis relative to patients diagnosed with other breast cancer subtypes. Protein tyrosine kinase 6 (PTK6; also called Brk), a soluble tyrosine kinase, is overexpressed in 86% of breast cancer patients, however its precise function in the context of TNBC is poorly defined. PTK6 expression is elevated in TNBC models in response to both cellular and endocrine stress, coordinated transcriptionally by the Hypoxia-Inducible Factors (HIFs) and glucocorticoid receptor (GR). We showed previously that PTK6 expression, but not its intrinsic kinase activity, is required for breast cancer cell motility. To further delineate the mechanisms of PTK6 signaling, we created kinase-intact domain structure mutants of PTK6 via in frame deletions of the N-terminal SH3 or SH2 domains. MDA-MB-231 cells expressing a PTK6 variant lacking the SH2 domain (SH2-del PTK6) were less responsive to growth factor-stimulated cell motility relative to wild type or kinase dead (KM) controls. To identify signal transduction pathways activated in TNBC cells harboring PTK6 domain mutants, we used a reverse phase protein array (RPPA), which revealed that the SH2 domain of PTK6 mediates TNBC cell motility via activation of the RhoA and/or AhR signaling pathways. Moreover, in TNBC cells, including a taxane-refractory TNBC model, addition of AhR or Rho inhibitors to paclitaxel (Taxol) enhanced cytotoxicity. Together, these studies reveal that the SH2-domain of PTK6 is an effector of advanced cancer phenotypes in GR+ TNBC cells and identify RhoA and AhR as novel therapeutic targets in PTK6+ tumors.

Abstract PR05: Blockade of AHR activation by IDO/TDO-derived kynurenine restricts cancer immune suppression

Abstract Immune checkpoint blockade (ICB) has been shown to convey significant clinical activity across a spectrum of malignancies, yet there is now recognition that multiple mechanisms of resistance can impair response. The catabolism of tryptophan into metabolites known as kynurenines (Kyn) by enzymes such as indoleamine 2,3-dioxygenase (IDO) or tryptophan 2,3-dioxygenase (TDO) plays a major suppressive role. Recently it was shown that Kyn acts as an endogenous agonist of the Aryl hydrocarbon receptor (AhR). In order to gain a better understanding of this pathway, we sought to characterize the mechanisms of immunosuppression associated with AhR and evaluate its potential as therapeutic target. Gene-expression analysis of IDO-overexpressing melanomas (B16-IDO vs. B16-WT) demonstrated reduced expression levels of Type 1 inflammatory genes, including IFNy, TNF, GzmB, and CD40. In addition, B16-IDO presents higher infiltration of tumor-associated macrophages TAMs, which upregulate the AHR as well as classic AhR-regulated genes (Cyp1a1 and Cyp1b1) and are differentially skewed towards an immunosuppressive M2 phenotype. Tumor-antigen specific CD8+T cells show reduced expression of activation markers (GzmB and CD44) and proliferation rate when primed by Kyn-treated antigen-presenting cells. In addition, TAMs from B16-IDO tumors suppressed activation of CD8+T cells in vitro and their depletion delayed tumor growth. When B16-IDO cells are implanted in mice depleted of Foxp3 expressing cells, TAMs do not accumulate. Treatment of B16-IDO tumors with an AhR-specific antagonist (CH-223191) upregulates MHC II in APCs, activation markers in CD8s, and reduced frequency of T-regs in B16-IDO tumors. AhR inhibition slows tumor growth and prolongs survival of tumors with active IDO/TDO/Kyn pathway (B16-IDO and B16-TDO), and this is enhanced when PD-1 blockade is used in combination. In summary, our findings demonstrate that targeting the Kyn pathway through AhR inhibition could overcome key suppressive mechanisms and sensitize tumors to ICB. This abstract is also being presented as Poster A57. Citation Format: Luis F. Campesato, Sadna Budhu, Mathieu Gigoux, Jeremy Tchaicha, Stephane Pourpe, Cailian Liu, Dmitriy Zamarin, Mark G. Manfredi, Karen McGovern, Jedd D. Wolchok, Taha Merghoub. Blockade of AHR activation by IDO/TDO-derived kynurenine restricts cancer immune suppression [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr PR05.

Effects of Hippeastrum reticulatum on memory, spatial learning and object recognition in a scopolamine-induced animal model of Alzheimer’s disease

Context The methanol extracts from Hippeastrum reticulatum (L'Hér.) Herb. (Amaryllidaceae) (HR) display acetylcholinesterase inhibitory (AChEI) activity. Objective AChEI of alkaloids isolated from HR bulbs and the ameliorating effects of the alkaloid fraction (AHR) on memory and cognitive dysfunction in scopolamine-treated mice were investigated. Materials and methods Alkaloids were isolated by column chromatography and identified by spectroscopy. AChEI was evaluated using the modified Ellman’s method. Sixty Swiss male mice were randomly divided into six groups, received samples for 15 days. Normal group received saline, scopolamine-treated group scopolamine (1.5 mg/kg, intraperitoneal injection). Test groups received AHR (5, 10 and 15 mg/kg, per os) and positive control group donepezil (5 mg/kg, per os), administered 1 h before the test, scopolamine was injected 30 min prior to testing. The cognitive-enhancing activity of AHR against scopolamine-induced memory impairments was investigated using Y-maze, the novel object recognition test (NORT) and the Morris water maze (MWM) test. Results Seven alkaloids were isolated for the first time from the genus Hippeastrum: trans-dihydronarciclasine (1), N-chloromethylnarcissidinium (2), narciprimin (3), narciclasine-4-O-β-d-xylopyranoside (4), N-methyltyramine (5), 3β,11α-dihydroxy-1,2-dehydrocrinane (6) and brunsvigine (7); three are new compounds (2, 5, 6). Among them, 2–3 and 5–6 showed AChEI in vitro with IC50 values of 29.1, 46.4, 70.1 and 104.5 µg/mL, respectively. The anti-AChEI of 2, 5 and 6 are reported for the first time. In in vivo test, AHR (15 mg/kg) significantly increased in spontaneous alternation performance in the Y-maze test (p < 0.01), it significantly increased the time spent exploring the novel object (p < 0.05) comparison with scopolamine-treated group. The administration of AHR at doses 10 and 15 mg/kg significantly decreased escapes latency and swimming distance to the platform on day 6 compared to these in day 1 (p < 0.01 and p < 0.05, respectively). Conclusions AHR could be a potential candidate of future trials for treatment of memory and cognitive dysfunction in Alzheimer’s disease.

Arsenic (III) or/and copper (II) exposure induce immunotoxicity through trigger oxidative stress, inflammation and immune imbalance in the bursa of chicken

The environmental hazards of arsenic (As) and copper (Cu) contamination have swept through quite a few districts worldwide. Whereas, molecular mechanisms involved in As- and Cu-induced immunotoxicity in Gallus gallus bursa of Fabricius (BF) are complex and elusive. Male Hy-line chickens were exposed to arsenic trioxide (As2O3; 30mg/kg) and copper sulfate (CuSO4; 300mg/kg) alone or in combination, respectively, to examine the potential ecotoxicity of them. The ions homeostasis and BF index of chicken had distinct changes after As or/and Cu exposure. Moreover, As or/and Cu treatment significantly increased the MDA content and NOS activity, and simultaneously resulted in reductions in CAT and AHR activities. Subsequently, it was further exhibited up-regulations of nuclear factor-κB (NF-κB), inflammatory mediators and pro-inflammation cytokines accompanied by depletion of anti-inflammatory cytokines and severe pathological conditions. Moreover, decreased ratio of IFN-γ/IL-4 and increased level of IL-17 illustrated an imbalance of the immune response. Meanwhile, incremental mRNA transcription and protein levels of heat shock proteins (HSPs) alleviated toxicity caused by As or/and Cu. Importantly, exposure to both contaminants significantly soared the BF injury in comparison with exposure to As or Cu alone. All these results illustrated that exposure to As2O3 or/and CuSO4 elicited BF tissue damage and ions changes, and its severity was associated with prolonged persistence of oxidative damage, accompanied by a dysregulated immune response which played a vital role in inflammatory injury. Additionally, combined management of As2O3 and CuSO4 could exacerbate BF injury.

Pilot study of global endocrine disrupting activity in Iowa public drinking water utilities using cell-based assays

Some anthropogenic substances in drinking water are known or suspected endocrine disrupting compounds (EDCs), but EDCs are not routinely measured. We conducted a pilot study of 10 public drinking water utilities in Iowa, where common contaminants (e.g., pesticides) are suspected EDCs. Raw (untreated) and finished (treated) drinking water samples were collected in spring and fall and concentrated using solid phase extraction. We assessed multiple endocrine disrupting activities using novel mammalian cell-based assays that express nuclear steroid receptors (aryl hydrocarbon [AhR], androgenic [AR], thyroid [TR], estrogenic [ER] and glucocorticoid [GR]). We quantified each receptor's activation relative to negative controls and compared activity by season and utility/sample characteristics. Among 62 samples, 69% had AhR, 52% AR, 3% TR, 2% ER, and 0% GR activity. AhR and AR activities were detected more frequently in spring (p =0 .002 and < 0.001, respectively). AR activity was more common in samples of raw water (p =0 .02) and from surface water utilities (p =0 .05), especially in fall (p =0 .03). Multivariable analyses suggested spring season, surface water, and nitrate and disinfection byproduct concentrations as determinants of bioactivity. Our results demonstrate that AR and AhR activities are commonly found in Iowa drinking water, and that their detection varies by season and utility/sample characteristics. Screening EDCs with cell-based bioassays holds promise for characterizing population exposure to diverse EDCs mixtures.

Kynurenine signaling through the aryl hydrocarbon receptor: Implications for aging and healthspan.

The tryptophan metabolite kynurenine increases with aging and inflammation, and appears to contribute directly to the development and progression of several age-related conditions. Kynurenine is now known to signal through the aryl hydrocarbon receptor (Ahr) to modulate levels of reactive oxygen species (ROS). The Ahr promoter region contains several sites for NF-kB binding, indicating that inflammation is a key factor modulating Ahr expression. Furthermore, kynurenine activation of Ahr is observed to stimulate expression of the enzyme IDO1, which generates kynurenine by degrading tryptophan, representing a positive feedback loop that may link inflammation with ROS production. On the other hand, the antioxidant system-inducing transcription factor Nrf2 can be stimulated by Ahr, and Nrf2 can itself activate Ahr expression. The balance between pro- and antioxidant functions of Ahr mediated by kynurenine may therefore regulate healthy versus unhealthy aging in different tissues and organ systems. Potential therapeutic approaches to target this pathway include exercise to alter kynurenine production or molecules such as metformin or resveratrol that may suppress Ahr activity.