| 1. |
- Mohammadi Bardbori, Afshin
(författare)
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Influence of oxidative stress on aryl hydrocarbon receptor signaling
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The aryl hydrocarbon receptor (AHR), a multifunctional protein and a key regulator of drug metabolizing enzymes, belongs to the basic-helix-loop-helix (bHLH)/PAS (Per-Arnt-Sim) super-family of transcription factors. The AHR responds to exogenous and endogenous chemicals by induction or repression of a large number of genes involved in many physiological processes and normal development. The diverse spectrum of AHR activators from well-known planar hydrophobic halogenated aromatic hydrocarbons (HAHs) to chemical compounds whose structure and physicochemical properties are very different from classical AHR ligands suggests that the AHR has a tremendously promiscuous ligand binding pocket. Due to the absence of a 3D structure of the ligand binding domain, promiscuity of the AHR has remained elusive. However, increasing experimental evidence indicate that the non-typical AHR ligands might activate the AHR signaling pathway indirectly by inhibiting the metabolic turnover of an endogenous ligand of the AHR. Therefore, the objective of this thesis was to characterize the inhibition of degradation of 6-formylindolo[3,2-b]carbazole (FICZ), the suggested natural high affinity AHR ligand, as a mechanism that could explain the earlier described agonistic properties of structurally very diverse AHR activators. The obtained results show that FICZ is a potent AHR agonist in vitro and in vivo which can distribute to the body through systemic circulation and induce cytochrome P450 1A1 (CYP1A1) the prototypical AHR target in various organs. The studies presented in this thesis demonstrate that if the metabolic clearance of FICZ is compromised, femtomolar concentrations of FICZ are sufficient to activate AHR signaling. The AHR signaling pathway seems to be sensitive to oxidative stress but the redox regulation of AHR has not been well characterized. Studies on dioxin and other reactive oxygen species (ROS) producing agents have demonstrated that the AHR is a mediator of oxidative stress. Indeed, AHR works in close concert with the master regulator of antioxidant responses, nuclear factor (erythroid-derived 2)-like 2 (Nrf2). Multiple sources of ROS appear to be involved in modulating AHR signaling and probably via three major systems, microsomes, mitochondria and NADPH oxidase enzymes (NOXs). Furthermore, it has been observed that many environmental pollutants, including metals and other NOX-activators increase the levels of the diffusible molecule hydrogen peroxide (H2O2) and change the cellular redox status and thereby interfere with cell growth kinetics and the endogenous functions of the AHR. To increase the understanding of downstream adaptive responses to oxidative stress, including up-regulation of antioxidant genes and modulation of AHR signaling was another objective of this work. The findings demonstrate that superoxide anion (O2−.) or H2O2 produced by NOXs can negatively and positively modulate the AHR signaling pathway. The importance of cellular redox levels which can influence endogenously activated AHR signaling broadens our earlier knowledge and explains why many oxidants behave both as AHR antagonists and agonists. In summary, this thesis extends the mechanistic understanding of the promiscuity of AHR and provides important information with regard to the redox regulation of AHR endogenous signaling.
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| 2. |
- Mohammadi-Bardbori, Afshin, et al.
(författare)
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NADPH Oxidase-Dependent Mechanism Explains How Arsenic and Other Oxidants Can Activate Aryl Hydrocarbon Receptor Signaling
- 2015
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 28:12, s. 2278-2286
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Tidskriftsartikel (refereegranskat)abstract
- The mechanisms explaining arsenic toxicity are not well understood, but physiological consequences of stimulated aryl hydrocarbon receptor (AHR) signaling both directly and through cross-talk with other pathways have been indicated. The aim of this study was to establish how arsenic interacts with AHR-mediated transcription. The human hepatoma cell line (HepG2-XRE-Luc) carrying a luciferase reporter under the control of two AHR response elements (AHREs) and immortalized human keratinocytes (HaCaT) were exposed to sodium arsenite (NaAsO2; As3+), alone or in combination with the endogenous high affinity AHR ligand 6-formylindolo[3,2-b]carbazole (FICZ). Luciferase activity, cytochrome P4501A1 (CYP1A1) activity, oxidative stress-related responses, metabolic clearance of FICZ, and NADPH:oxidase (NOX) activity as well as nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-dependent gene expression were measured. Arsenic inhibited,CYP1A1 enzyme activity and reduced the metabolic clearance of FICZ. Arsenic also led to activated CYP1A1 transcription but only in cells grown in medium containing trace amounts of the endogenous ligand FICZ, pointing to an indirect mechanism of activation. Initially, arsenic caused dose-dependent inhibition of FICZ-activated AHR signaling, disturbed intracellular GSH status, and increased expression of oxidative stress-related genes. Silencing of NOX4, addition of N-acetylcystein, or pretreatment with arsenic itself attenuated the initial dose-dependent inhibition of AHR signaling. Arsenic pretreatment led to elevated GSH levels and sensitized the cells to ligaild-dependent AHR signaling, while silencing of Nrf2 significantly reduced arsenic-mediated, activation of the AHR. In addition, influence of NOX on AHR activation was also observed in cells treated with the SH-reactive metals cadmium, mercury, and nickel. Together, the results suggest that SH-reactive agents via a new and possibly general NOX/H2O2-dependent mechanism can interfere with the endogenous regulation of the AHR.
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| 3. |
- Mohammadi-Bardbori, Afshin, et al.
(författare)
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Quercetin, Resveratrol, and Curcumin Are Indirect Activators of the Aryl Hydrocarbon Receptor (AHR)
- 2012
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 25:9, s. 1878-1884
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Tidskriftsartikel (refereegranskat)abstract
- Several polyphenols have been shown to activate the aryl hydrocarbon receptor (AHR) in spite of the fact that they bind to the receptor with low affinity. The aim of this study was to investigate whether quercetin (QUE), resveratrol (RES), and curcumin (CUR) interfere with the metabolic degradation of the suggested endogenous AHR ligand 6-formylindolo[3,2-b]carbazole (FICZ) and thereby indirectly activate the AHR. Using recombinant human enzyme, we confirmed earlier reported inhibitory effects of the polyphenols on cytochrome P4501A1 (CYP1A1) activity, and inhibition of metabolic clearance of FICZ was documented in FICZ-treated immortalized human keratinocytes (HaCaT). CYP1A1 activity was induced in HaCaT cells by all three compounds, and when they were added together with FICZ, a prolonged activation was observed after a dose-dependent inhibition period. The same pattern of responses was seen at the transcriptional level as determined with a CYP1A1 reporter assay in human liver hepatoma (HepG2) cells. To test the ability of the polyphenols to activate the AHR in the absence of FICZ, the cells were treated in medium, which in contrast to commercial batches of medium did not contain background levels of FICZ. Importantly, AHR activation was only observed in the commercial medium. Taken together, these findings suggest that QUE, RES, and CUR induce CYP1A1 in an indirect manner by inhibiting the metabolic turnover of FICZ. Humans are exposed to these compounds through the diet and nutritional supplements, and we propose that altered systemic levels of FICZ caused by such compounds may have physiological consequences.
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| 4. |
- Wincent, Emma, et al.
(författare)
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Inhibition of cytochrome P4501-dependent clearance of the endogenous agonist FICZ as a mechanism for activation of the aryl hydrocarbon receptor
- 2012
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 109:12, s. 4479-4484
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Tidskriftsartikel (refereegranskat)abstract
- Altered systemic levels of 6-formylindolo[3,2-b]carbazole (FICZ), an enigmatic endogenous ligand for the aryl hydrocarbon receptor (AHR), may explain adverse physiological responses evoked by small natural and anthropogenic molecules as well as by oxidative stress and light. We demonstrate here that several different chemical compounds can inhibit the metabolism of FICZ, thereby disrupting the autoregulatory feedback control of cytochrome P4501 systems and other proteins whose expression is regulated by AHR. FICZ is both the most tightly bound endogenous agonist for the AHR and an ideal substrate for cytochrome CYP1A1/1A2 and 1B1, thereby also participating in an autoregulatory loop that keeps its own steady-state concentration low. At very low concentrations FICZ influences circadian rhythms, responses to UV light, homeostasis associated with pro-and anti-inflammatory processes, and genomic stability. Here, we demonstrate that, if its metabolic clearance is compromised, femtomolar background levels of this compound in cell-culture medium are sufficient to up-regulate CYP1A1 mRNA and enzyme activity. The oxidants UVB irradiation and hydrogen peroxide and the model AHR antagonist 3'-methoxy-4'-nitroflavone all inhibited induction of CYP1A1 enzyme activity by FICZ or 2,3,7,8-tetrachlorodibenzo-p-dioxin, thereby subsequently elevating intracellular levels of FICZ and activating AHR. Taken together, these findings support an indirect mechanism of AHR activation, indicating that AHR activation by molecules with low affinity actually may reflect inhibition of FICZ metabolism and raising questions about the reported promiscuity of the AHR. Accordingly, we propose that prolonged induction of AHR activity through inhibition of CYP1 disturbs feedback regulation of FICZ levels, with potential detrimental consequences.
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