Feedback control of AHR signalling regulates intestinal immunity

The aryl hydrocarbon receptor (AHR) recognizes xenobiotics as well as natural compounds such as tryptophan metabolites, dietary components and microbiota-derived factors, and it is important for maintenance of homeostasis at mucosal surfaces. AHR activation induces cytochrome P4501 (CYP1) enzymes, which oxygenate AHR ligands, leading to their metabolic clearance and detoxification. Thus, CYP1 enzymes have an important feedback role that curtails the duration of AHR signalling, but it remains unclear whether they also regulate AHR ligand availability in vivo. Here we show that dysregulated expression of Cyp1a1 in mice depletes the reservoir of natural AHR ligands, generating a quasi AHR-deficient state. Constitutive expression of Cyp1a1 throughout the body or restricted specifically to intestinal epithelial cells resulted in loss of AHR-dependent type 3 innate lymphoid cells and T helper 17 cells and increased susceptibility to enteric infection. The deleterious effects of excessive AHR ligand degradation on intestinal immune functions could be counter-balanced by increasing the intake of AHR ligands in the diet. Thus, our data indicate that intestinal epithelial cells serve as gatekeepers for the supply of AHR ligands to the host and emphasize the importance of feedback control in modulating AHR pathway activation

Does the managers´form of employment matter? : A qualitative study of the form of employment´s significance to a managerial appointment

Does the managers’ form of employment matter? A qualitative study of the form of employment’s significance to a managerial appointment The aim of this study is to examine the experiences and attitudes of managers with regards to the significance of the specific terms of their employment. To achieve this, a qualitative approach was deemed to be suitable and a thematic analysis was conducted. Ten middle managers at a Swedish County Council with contrasting terms of employment (permanent positions and short-term managerial appointments) were interviewed over a two week period. To complement the thematic analysis, a theoretic framework and previous research were considered and used. The study found that there is a strong preference among the managers towards a permanent form of employment rather than short-term managerial appointments. The analysis, on the other hand, found that the terms of employment had a negligible impact on the managers’ actual work, whereas it impacted the managers’ perception of their own ‘managerial value’ and their perceived importance to the organisation. On the other hand, our study has shown that the form of employment can also affect the work and organisation. Finally, the study found that this particular field of research is yet to be fully explored and thus further research is needed to reach a greater understanding of the effects of the form of employment to the individual manager, the organisation and its workers

Exposure to retene, fluoranthene, and their binary mixture causes distinct transcriptomic and apical outcomes in rainbow trout (Oncorhynchus mykiss) yolk sac alevins

Polycyclic aromatic hydrocarbons (PAHs) are widely spread environmental contaminants which affect developing organisms. It is known that improper activation of the aryl hydrocarbon receptor (AhR) by some PAHs contributes to toxicity, while other PAHs can disrupt cellular membrane function. The exact downstream mechanisms of AhR activation remain unresolved, especially with regard to cardiotoxicity. By exposing newly hatched rainbow trout alevins (Oncorhynchus mykiss) semi-statically to retene (32 µg l−1; AhR agonist), fluoranthene (50 µg l−1; weak AhR agonist and CYP1a inhibitor) and their binary mixture for 1, 3, 7 and 14 days, we aimed to uncover novel mechanisms of cardiotoxicity using a targeted microarray approach. At the end of the exposure, standard length, yolk area, blue sac disease (BSD) index and PAH body burden were measured, while the hearts were prepared for microarray analysis. Each exposure produced a unique toxicity profile. We observed that retene and the mixture, but not fluoranthene, significantly reduced growth by Day 14 compared to the control, while exposure to the mixture increased the BSD-index significantly from Day 3 onward. Body burden profiles were PAH-specific and correlated well with the exposure-specific upregulations of genes encoding for phase I and II enzymes. Exposure to the mixture over-represented pathways related to growth, amino acid and xenobiotic metabolism and oxidative stress responses. Alevins exposed to the individual PAHs displayed over-represented pathways involved in receptor signaling: retene downregulated genes with a role in G-protein signaling, while fluoranthene upregulated those involved in GABA signaling. Furthermore, exposure to retene and fluoranthene altered the expression of genes encoding for proteins involved in calcium- and potassium ion channels, which suggests affected heart structure and function. This study provides deeper understanding of the complexity of PAH toxicity and the necessity of investigating PAHs as mixtures and not as individual components.peerReviewe

Endogenous AhR agonist FICZ accumulates in rainbow trout (Oncorhynchus mykiss) alevins exposed to a mixture of two PAHs, retene and fluoranthene

Multiple studies have reported synergized toxicity of PAH mixtures in developing fish larvae relative to the additive effect of the components. From a toxicological perspective, multiple mechanisms are known to contribute to synergism, such as altered toxicodynamics and kinetics, as well as increased oxidative stress. An understudied contributor to synergism is the accumulation of endogenous metabolites, for example: the aryl hydrocarbon receptor 2 (AhR2) agonist and tryptophan metabolite 6-Formylindolo(3,2-b)carbazole (FICZ). Fish larvae exposed to FICZ, alongside knock-down of cytochrome p450 (cyp1a), has been reported to induced symptoms of toxicity similar to those observed following exposure to PAHs or the dioxin 2,3,7,8-tetrachlorodibenzo-p-dioxin. Here, we explored if FICZ accumulates in newly hatched rainbow trout alevins (Oncorhynchus mykiss) exposed to two PAHs with different properties: retene (potent AhR2 agonist) and fluoranthene (weak AhR2 agonist and Cyp1a inhibitor), either alone or as a binary mixture for 3 and 7 days. We found that exposure to the mixture resulted in accumulation of endogenous FICZ, synergized the blue sac disease index (BSD), and altered the body burden profiles of the PAHs, when compared to the alevins exposed to the individual components. It is thus very plausible that accumulation of endogenously derived FICZ contributed to the synergized BSD index and toxicity in exposed alevins. Accumulation of endogenously derived FICZ is a novel finding that extends our general understanding on PAHs toxicity in developing fish larvae, while at the same time highlighting why environmental risk assessment of PAHs should not be based solely results from the assessment of individual compounds.peerReviewe

PBTK modeled perfluoroalkyl acid kinetics in zebrafish eleutheroembryos suggests impacts on bioconcentrations by chorion porosity dynamics

International audienceThe zebrafish eleutheroembryo (zfe) is widely used as a model to characterize the toxicity of chemicals. However, analytical methods are still missing to measure organ concentrations. Therefore, physiologically-based toxicokinetic (PBTK) modeling may overcome current limitations to help understand the relationship between toxic effects and internal exposure in various organs.A previous PBTK model has been updated to include the chorionic transport barrier and its permeabilization, hatching dynamics within a zfe population over development, and active mediated transport mechanisms. The zfe PBTK model has been calibrated using measured time-dependent internal concentrations of PFBA, PFHxS, PFOA, and PFOS in a zfe population and evaluated using external datasets from the literature.Calibration was successful with 96% of the predictions falling within a 2-fold range of the observed concentrations. The external dataset was correctly estimated with about 50% of the predictions falling within a factor of 3 of the observed data and 10% of the predictions are out of the 10-fold error. The calibrated model suggested that active mediated transport differs between PFAS with a sulfonic and carboxylic acid functional end groups.This PBTK model predicts well the fate of PFAS with various physicochemical properties in zfe. Therefore, this model may improve the use of zfe as an alternative model in toxicokinetic-toxicodynamic studies and help to refine and reduce zfe-based experiments, while giving insights into the internal kinetics of chemicals

Perfluorooctanesulfonic acid modulates barrier function and systemic T cell homeostasis during intestinal inflammation.

The intestinal epithelium is continuously exposed to deleterious environmental factors which might cause aberrant immune responses leading to inflammatory disorders. However, what environmental factors might contribute to disease are yet poorly understood. Here, to overcome the lack of in vivo models suitable for screening of environmental factors we used zebrafish reporters of intestinal inflammation. Using zebrafish, we interrogated the immunomodulatory effects of polyfluoroalkyl substances (PFAS), which have been positively associated with ulcerative colitis incidence. Exposure with perfluorooctanesulfonic acid (PFOS) during TNBS-induced inflammation enhances the expression of proinflammatory cytokines as well as neutrophil recruitment to the intestine of zebrafish larvae, which was validated in TNBS-induced colitis mice models. Moreover, PFOS exposure in mice undergoing colitis resulted in neutrophil-dependent increased intestinal permeability and enhanced PFOS translocation into circulation. Finally, this was associated with a neutrophil dependent expansion of systemic CD4+ T cells. Thus, our results indicate that PFOS worsens inflammation-induced intestinal damage with disruption of T cell homeostasis beyond the gut and provides a novel in vivo toolbox to screen for pollutants affecting intestinal homeostasis

The suggested physiologic aryl hydrocarbon receptor activator and cytochrome P4501 substrate 6-formylindolo[3,2-b]carbazole is present in humans

Dioxins and other polycyclic aromatic compounds formed during the combustion of waste and fossil fuels represent a risk to human health, as well as to the well being of our environment. Compounds of this nature exert carcinogenic and endocrine-disrupting effects in experimental animals by binding to the orphan aryl hydrocarbon receptor (AhR). Understanding the mechanism of action of these pollutants, as well as the physiological role(s) of the AhR, requires identification of the endogenous ligand(s) of this receptor. We reported earlier that activation of AhR by ultraviolet radiation is mediated by the chromophoric amino acid tryptophan (Trp), and we suggested that a new class of compounds derived from Trp, in particular 6-formylindolo[3,2-b]carbazole (FICZ), acts as natural high affinity ligands for this receptor. Here we describe seven new FICZ-derived indolo[3,2-b]carbazole-6-carboxylic acid metabolites and two sulfoconjugates, and we demonstrate the following. (i) FICZ is formed efficiently by photolysis of Trp upon exposure to visible light. (ii) FICZ is an exceptionally good substrate for cytochromes P450 (CYP) 1A1, 1A2, and 1B1, and its hydroxylated metabolites are remarkably good substrates for the sulfotransferases (SULTs) 1A1, 1A2, 1B1, and 1E1. Finally, (iii) sulfoconjugates of phenolic metabolites of FICZ are present in human urine. Our findings indicate that formylindolo[3,2-b]carbazols are the most potent naturally occurring activators of the AhR signaling pathway and may be the key substrates of the CYP1 and SULT1 families of enzymes. These conclusions contradict the widespread view that xenobiotic compounds are the major AhR ligands and CYP1 substrates

Biological effects of 6-formylindolo[3,2-b]carbazole (FICZ) in vivo are enhanced by loss of CYP1A function in an Ahr2-dependent manner

6-Formylindolo[3,2-b]carbazole (FICZ) is a potent aryl hydrocarbon receptor (AHR) agonist that is efficiently metabolized by AHR-regulated cytochrome P4501 enzymes. FICZ is a proposed physiological AHR ligand that induces its own degradation as part of a regulatory negative feedback loop. In vitro studies in cells show that CYP1 inhibition in the presence of FICZ results in enhanced AHR activation, suggesting that FICZ accumulates in the cell when its metabolism is blocked. We used zebrafish (Danio rerio) embryos to investigate the in vivo effects of FICZ when CYP1A is knocked down or inhibited. Embryos were injected with morpholino antisense oligonucleotides targeting CYP1A (CYP1A-MO), Ahr2, or a combination of both. FICZ exposure of non-injected embryos or embryos injected with control morpholino had little effect. In CYP1A-MO-injected embryos, however, FICZ dramatically increased mortality, incidence and severity of pericardial edema and circulation failure, reduced hatching frequency, blocked swim bladder inflation, and strongly potentiated expression of Ahr2-regulated genes. These effects were substantially reduced in embryos with a combined knockdown of Ahr2 and CYP1A, indicating that the toxicity was mediated at least partly by Ahr2. Co-exposure to the CYP1 inhibitor alpha-naphthoflavone (αNF) and FICZ had similar effects as the combination of CYP1A-MO and FICZ. HPLC analysis of FICZ-exposed embryos showed increased levels of FICZ after concomitant CYP1A-MO injection or αNF co-exposure. Together, these results show that a functioning CYP1/AHR feedback loop is crucial for regulation of AHR signaling by a potential physiological ligand in vivo and further highlights the role of CYP1 enzymes in regulating biological effects of FICZ