Aryl hydrocarbon receptor-dependence of dioxin’s effects on constitutive mouse hepatic cytochromes P450 and growth hormone signaling components

Abstract

The aryl hydrocarbon receptor (AHR) has physiological roles in the absence of exposure to exogenous ligands and mediates adaptive and toxic responses to the environmental pollutant, 2,3,7,8-tetracholorodibenzo-p-dioxin (TCDD). A readily metabolized AHR agonist, 3-methylcholanthrene, disrupts expression of mouse hepatic growth hormone (GH) signaling components and suppresses cytochrome P450 2D9 (Cyp2d9), a male-specific gene controlled by pulsatile GH via signal transducer and activator of transcription 5b (STAT5b). Using TCDD as an essentially non-metabolized AHR agonist and Ahr −/− mice as the preferred model to determine the AHR-dependence of biological responses, we now show that two mouse hepatic STAT5b target genes, Cyp2d9 and major urinary protein 2 (Mup2), are suppressed by TCDD in an AHRdependent manner. TCDD also decreased hepatic mRNA levels for GH receptor, Janus kinase 2, and STAT5a/b with AHR-dependence. Without inducing selected hepatic inflammatory markers, TCDD caused AHR-dependent induction of Cyp1a1 and NADPH-cytochrome P450 oxidoreductase (Por) and suppression of Cyp3a11. In vehicle-treated mice, basal mRNA levels for CYP2D9, CYP3A11, POR, serum amyloid protein P, and MUP2 were influenced by Ahr genetic status. We conclude that AHR activation per se leads to dysregulation of hepatic GH signaling components and suppression of some, but not all, STAT5b target genes. Keywordsaryl hydrocarbon receptor; 2,3,7,8-tetrachlorodibenzo-p-dioxin; cytochrome P450; growth hormone receptor; Janus kinase 2; signal transducer and activator of transcription 5b; cytokineinducible Src homology 2 domain-containing protein; major urinary protein 2; inflammatory markers; NADPH-cytochrome P450 oxidoreductase

Downregulation of Mouse Hepatic CYP3A Protein by 3-Methylcholanthrene Does Not Require Cytochrome P450-Dependent Metabolism

“…Although we cannot rule out a role for MC metabolism by non-P450 pathways and/or in cell types other than hepatocytes, our findings are consistent with a key role for the parent drug MC in the CYP3A suppression response. This idea is also supported by our recent report that TCDD, an essentially nonmetabolized HAH-type AHR agonist, suppresses hepatic CYP3A11 mRNA levels in WT but not Ahr-null mice (Lee and Riddick, 2012). Our current working hypothesis is that the suppression of mouse hepatic Cyp3a11 by both TCDD and MC involves AHR activation by the parent compounds.…”

“…Hepatic levels of mRNAs encoding CYP3A11 and flavin-containing monooxygenase-3 (FMO3), normalized to b-actin as the internal reference standard, were determined by the comparative threshold cycle relative quantitation method on the ABI Prism 7500 Sequence Detection System (Applied Biosystems, Foster City, CA). The CYP3A11 primers and optimized assay conditions were validated in our previous study (Lee and Riddick, 2012), whereas the FMO3 primer sequences were derived from published work (Zhang et al, 2009).…”

“…TCDD, an essentially nonmetabolized AHR agonist, causes AHR-dependent suppression of hepatic CYP3A11 (Lee and Riddick, 2012) and CYP3A13 (Ovando et al, 2006) mRNA levels. Proteomic analysis showed that MC, a readily metabolized AHR agonist, decreases mouse hepatic CYP3A11/16/41 protein levels (Jenkins et al, 2006) and we found that the decrease in mouse hepatic Cyp3a11 expression caused by MC was particularly pronounced at the protein level (Lee et al, 2006).…”

The Role of Cytochrome P450–Dependent Metabolism in the Regulation of Mouse Hepatic Growth Hormone Signaling Components and Target Genes by 3-Methylcholanthrene

“…Hepatic levels of mRNAs encoding POR, CYP2D9, serum amyloid protein P (SAP), suppressor of cytokine signaling 3 (SOCS3), and GHR, normalized to b-actin as the internal reference standard, were determined by the comparative threshold cycle relative quantitation method on the ABI Prism 7500 Sequence Detection System (Applied Biosystems, Foster City, CA) using the primers and optimized assay conditions validated in our previous study (Lee and Riddick, 2012).…”

“…We have recently taken two approaches to answer this question. First, we used 2,3,7,8-tetrachlorodibenzo-pdioxin as an essentially nonmetabolized AHR agonist and Ahr-null mice to demonstrate that AHR activation per se decreases hepatic mRNA levels for GHR, JAK2, and STAT5a/b, as well as two STAT5b target genes, Cyp2d9 and Mup2 (Lee and Riddick, 2012). Second, in the present study, we make use of mice that are essentially devoid of hepatic microsomal P450-dependent metabolism because of hepatocyte-specific conditional deletion of NADPH-cytochrome P450 oxidoreductase (CPR or POR), the obligate electron transfer partner for all microsomal P450s.…”