Possible Involvement of the AH Receptor in the Induction of Cytochrome P-450IA1 Under Conditions of Hydrodynamic Shear in Microcarrier-Attached Hepatoma Cell Lines

Mufti, Naheed A.; Bleckwenn, Nicole A.; Babish, John G.; Shuler, Michael L.

doi:10.1006/bbrc.1995.1316

Cited by 37 publications

(29 citation statements)

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“…1). These findings are consistent with the results of Mufti et al (19) using an AHR-deficient murine cell line in a microcarrier-attached suspension culture system. Moreover, we demonstrated that a promoter fragment containing four canonical DREs was sufficient to induce reporter activity after cellular exposure to shear stress (Fig.…”

Section: Discussionsupporting

confidence: 92%

“…The fluid shear stress produced by this flow system is comparable with the levels of shear in the human arterial system (18) and within the range found to induce CYP1 gene expression (12,13,17). We first verified that our shear system could replicate the CYP1 response to fluid flow by using endothelial and hepatoma cell lines derived from several mammalian species (SI Table 1).…”

Section: Discussionmentioning

confidence: 55%

“…In this preliminary experiment, we identified previously characterized shear-responsive genes as TCDD-inducible (B.J.M., unpublished observation). A further survey of the literature revealed that the classical AHRregulated genes CYP1A1 and CYP1B1 are also highly inducible by shear stress (11)(12)(13)17). The overlap between shear and TCDD-induced gene expression changes led us to hypothesize that the AHR could act as a sensor for a shear-induced signal and mediate a transcriptional response to altered fluid flow in the vasculature.…”

Section: Discussionmentioning

confidence: 99%

“…In these preliminary gene expression studies, we identified a set of TCDD-inducible genes, which had been characterized as shear stress-inducible (B.J.M., unpublished observation). A subsequent survey of the literature revealed that classical AHRregulated genes, including CYP1A1 and CYP1B1, have also been documented to be highly induced by fluid flow (10)(11)(12)(13). Because of the key role of shear stress in vascular biology, we further explored the link between fluid flow and AHR signaling.…”

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confidence: 99%

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The Aryl hydrocarbon receptor is activated by modified low-density lipoprotein

McMillan

Bradfield²

2007

Proc. Natl. Acad. Sci. U.S.A.

115

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Endogenous activation of the aryl hydrocarbon receptor (AHR) is required for normal vascular development. This biology led us to investigate the interplay between the AHR and vascular physiology by using an in vitro model of fluid shear stress. Using this system, we show that fluid flow induces a robust AHR-mediated increase in CYP1 expression. Furthermore, we demonstrate that incubation with sheared bovine or human sera is sufficient for AHR activation, indicating that direct cellular exposure to shear stress is not required for this response. Fractionation of sera by size and density revealed the AHR-activating factor to be low-density lipoprotein (LDL). Purified LDL (0.1 mg/ml) from sheared sera induces a 6-fold increase in AHR-mediated signaling as compared with LDL purified from static sera. Similar results were obtained by exposing a purified fraction of LDL to fluid flow, suggesting that shear stress is capable of directly modifying LDL structure and/or function. In addition, we show that LDL can be converted to an AHR-activating species by conventional methods of lipoprotein modification, such as NaOCl oxidation. Finally, we demonstrate that an increased level of AHR-activating LDL is present in the sera of AHR null mice as compared with heterozygous littermates, suggesting a role for the Ahr locus in the physiological response to modified LDL in vivo. Overall, these data demonstrate a previously undescribed relationship between LDL modification and AHR biology and provide a potential explanation for the vascular abnormalities observed in AHR null mice.AHR ͉ cyp1a1 ͉ cyp1b1 ͉ shear stress T he aryl hydrocarbon receptor (AHR) is a ligand activated basic-helix-loop-helix (bHLH) transcription factor that belongs to the PAS (Per-Arnt-Sim) family of nuclear sensors (1). The AHR binds to numerous environmental contaminants including tetrachlorodibenzo-p-dioxin (TCDD), benzo[a]pyrene, and coplanar polychlorinated biphenyls (2). Upon binding these xenobiotic ligands, the AHR translocates to the nucleus and dimerizes with a second bHLH-PAS protein known as the Ah receptor nuclear translocator (ARNT). The AHR-ARNT complex can then bind to dioxin response elements (DREs) found upstream of target genes, leading to their transcriptional up-regulation. The CYP1 family of cytochrome P450s, CYP1A1, CYP1A2 and CYP1B1, as well as the phase II enzymes GST-A1 and NQO1 are among the genes that are regulated by the AHR (2, 3). The enzymes encoded by these genes display metabolic activity toward many AHR ligands, prompting the idea that this pathway represents an adaptive response to xenobiotic exposure (2, 3).In addition to its role in the metabolic response to xenobiotics, the AHR also has an important role in vascular development. Mice that harbor a null allele at the Ahr locus display a number of vascular phenotypes. These include a patent ductus venosus (DV), persistent hyaloid arteries in the eye, decreased hepatic perfusion, a confused vascularization in the corneal limbus, and cardiac hypertrophy (4-6). Experiments usi...

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Section: Discussionsupporting

confidence: 92%

Section: Discussionmentioning

confidence: 55%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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The Aryl hydrocarbon receptor is activated by modified low-density lipoprotein

McMillan

Bradfield²

2007

Proc. Natl. Acad. Sci. U.S.A.

115

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“…Furthermore, the present investigation offers a relatively straightforward explanation for the reported ability of numerous structurally diverse chemicals to activate the AHR; such reports have been used to argue that the AHR binds ligands promiscuously (37). Our results also may explain why the addition of fresh Trp-containing medium to cell cultures (42), oxidative stress [e.g., by hyperoxia (43)], and hydrodynamic shear that gives rise to oxidized LDL (44,45), as well as why the addition of various complex mixtures, such as extracts of paper and ink, can activate the AHR (46,47).…”

Section: Discussionmentioning

confidence: 57%

Inhibition of cytochrome P4501-dependent clearance of the endogenous agonist FICZ as a mechanism for activation of the aryl hydrocarbon receptor

Wincent

Bengtsson²,

Bardbori³

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

182

215

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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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Modular bioreactor for primary human hepatocyte culture: Medium flow stimulates expression and activity of detoxification genes

Vinci

Duret

Klieber

et al. 2011

Biotechnology Journal

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Down-regulation of detoxification genes, notably cytochrome P450 (CYPs), in primary hepatocyte cultures is a long-standing and major concern. We evaluated the influence of medium flow in this model. Hepatocytes isolated from 12 different liver donors were cultured either in a multichamber modular bioreactor (MCmB, flow rate 250–500 μL/min) or under standard/static conditions, and the expression of 32 genes, enzyme activities and biological parameters were measured 7–21 days later. mRNA expression of genes involved in xenobiotic/drug metabolism and transport, including CYP1A1, 1A2, 2B6, 2C9, 3A4 (and activities for some of them), UDP-glucuronosyltransferase (UGT) 1A1, UGT2B4, UGT2B7, glutathione S-transferase (GSTα), and multidrug resistance protein 1 (MDR1) and MRP2, were specifically up-regulated by medium flow as compared with static controls in all cultures tested. In 2-week-old cultures, expression of detoxification genes reached levels close to or higher than those measured in freshly isolated hepatocytes. In contrast, CYP2D6 and most of other tested genes were not affected by medium flow. We conclude that medium flow specifically interferes with, and up-regulates, the activity of xenosensors and/or the expression of detoxification genes in primary human hepatocytes. Down-regulation of detoxification genes in conventional (static) cultures is therefore partly a consequence of the absence of medium circulation.

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Possible Involvement of the AH Receptor in the Induction of Cytochrome P-450IA1 Under Conditions of Hydrodynamic Shear in Microcarrier-Attached Hepatoma Cell Lines

Cited by 37 publications

References 0 publications

The Aryl hydrocarbon receptor is activated by modified low-density lipoprotein

The Aryl hydrocarbon receptor is activated by modified low-density lipoprotein

Inhibition of cytochrome P4501-dependent clearance of the endogenous agonist FICZ as a mechanism for activation of the aryl hydrocarbon receptor

Modular bioreactor for primary human hepatocyte culture: Medium flow stimulates expression and activity of detoxification genes

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