Benzo[a]pyrene (BaP) is a well-studied pro-carcinogen that is metabolically activated by cytochrome P450 enzymes. Cytochrome P4501A1 (CYP1A1) has been considered to play a central role in the activation step, which is essential for the formation of DNA adducts. This enzyme is strongly induced by many different chemical agents, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), which binds to the aryl hydrocarbon receptor (AhR). Therefore, AhR activators are suspected to have the potential to aggravate the toxicity of BaP through the induction of CYP1A1. Besides, CYP1A1 inhibitors, including its substrates, are estimated to have preventive effects against BaP toxicity. However, strangely, increased hepatic BaP–DNA adduct levels have been reported in Cyp1a1 knockout mice. Moreover, numerous reports describe that concomitant treatment of AhR activators reduced BaP–DNA adduct formation. In an experiment using several human cell lines, TCDD had diverse modulatory effects on BaP–DNA adducts, both enhancing and inhibiting their formation. In this review, we focus on the factors that could influence the BaP–DNA adduct formation. To interpret these complicated outcomes, we propose a hypothesis that CYP1A1 is a key enzyme for both generation and reduction of (±)-anti-benzo[a]pyrene-7,8-diol-9,10-epoxide (BPDE), the major carcinogenic intermediate of BaP. Conversely, CYP1B1 is thought to contribute only to the metabolic activation of BaP related to carcinogenesis.

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The aryl hydrocarbon receptor (AhR) plays a suppressive role in cecal carcinogenesis by CUL4B/AhR-mediated ubiquitylation and degradation of β-catenin, which is activated by xenobiotics and natural ligands. AhR-deficient (AhR(-)(/-)) mice develop cecal tumors with severe inflammation. To elucidate whether the tumors develop autonomously in AhR(-/-) mice due to impaired β-catenin degradation or in association with accelerated inflammation, we performed two kinds of experiments using germ-free (GF) AhR(-/-) mice and compound mutant mice lacking genes for AhR and apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), which plays an essential role in caspase-1 activation in inflammasomes. Both GF AhR(-/-) and AhR(-/-)•ASC(-/-) mice showed considerably reduced tumor development compared with that in AhR(-/-) mice albeit in a 'cancer-prone' state with aberrant β-catenin accumulation. Blocking of the interleukin (IL)-1β signaling pathway by treatment with a caspase-1 inhibitor, YVAD, reduced cecal tumorigenesis in AhR(-/-) mice. Signal transducers and activators of transcription 3 (STAT3) activation was detected in the cecal epithelium of the AhR(-/-) mice due to enhanced IL-6 production. An inhibitor of the STAT3 signaling pathway, AG490 suppressed the tumor formation. ASC-mediated inflammation was also found to play a critical role in tumor development in Apc(Min/+) mice, a mouse model of familial adenomatous polyposis. Collectively, these results revealed an important role of the bacteria-triggered or ASC-mediated inflammation signaling pathway in the intestinal tumorigenesis of mice and suggest a possible chemical therapeutic intervention, including AhR ligands and inhibitors of the inflammation pathway.

This assay system will be a valuable tool to detect agonists as well as antagonists of corticosteroid receptors, in the fields of drug discovery and the assessment of environmental pollutants.

Endocrine disruptors are exogenous substances that act like hormones in the endocrine system and disrupt the physiologic function of endogenous hormones. In the present study, we established reporter yeast strains (Saccharomyces cerevisiae) expressing human estrogen receptors, ERalpha or ERbeta. These strains contain a reporter plasmid carrying an estrogen responsive element (ERE) upstream of the beta-galactosidase gene, and a plasmid expressing a steroid receptor coactivator, SRC-1e. Using these reporter strains, we demonstrated dose-dependent estrogenic activities of different categories of ligands, a natural hormone, 17beta-estradiol (E2); a synthetic drug, diethylstilbestrol (DES); phytoestrogens, genistein, daizein and emodin; and an environmental endocrine disrupter, bisphenol A. EC(50) values of E2 for ERalpha and ERbeta are 5.31 x 10(-10) and 5.85 x 10(-10) M, respectively. We also demonstrated that these yeasts were applicable for measuring estrogenic activities of environmental water samples. Most downstream sites of a river showed similar activity in both ERalpha and ERbeta assays. These yeast strains are useful and convenient for detecting and comparing the estrogenic ligand activities of environmental samples in response to ERalpha and ERbeta.

Abstract. Telomerase activation, known to be stimulated by estrogen, is essential for cellular immortalization and transformation, both of which play a role in tumorigenesis. Dioxin and dioxin-like compounds have been shown to induce endometriosis and promote estrogen-dependent tumors. In this study, we show that either 2,3,7,8-tetrachlorodibenzo-pdioxin (TCDD) or a combination of TCDD and 17-ß estradiol (E2) increase telomerase activity and the expression of the human telomerase catalytic subunit (hTERT) in human choriocarcinoma (BeWo) cells. Compared with estrogen or TCDD alone, the combination treatment did not show an additive effect. Likewise, treatment with either E2 or TCDD increased DNA synthesis and the cell population in S-phase, as detected by FACS analysis. However, following treatment with the E2 and TCDD combination, the proportion of cells in S-phase was actually lower than in cells treated with TCDD alone. These results suggest that TCDD alone mimics estrogenic action in telomerase activation and cell proliferation but, in the presence of estrogen, TCDD-induced actions were partially counteracted. E2 and TCDD also induced c-Myc, which is a transcriptional activator of hTERT in BeWo, but neither of these agents induced telomerase activity in HO15.19 c-myc-null cells. In contrast, only TCDD upregulated telomerase in TGR-1 cells, which are c-Myc expressing but lacking ER expression. The findings suggest that TCDD induces telomerase activity mediated through AhR signaling and/or ER-independent c-Myc signaling. The present study provides insight into the mechanism of promoter activity of TCDD in estrogen-related tumors.

The present assay yeasts may be valuable tools for subtype-specific assessments of unidentified environmental ligand chemicals and receptor-specific pharmaceuticals.

Background: Sex steroid hormone receptors are classified into three classes of receptors: estrogen receptors (ER) α and β, androgen receptor (AR), and progesterone receptor (PR). They belong to the nuclear receptor superfamily and activate their downstream genes in a ligand-dependent manner. Since sex steroid hormones are involved in a wide variety of physiological processes and cancer development, synthetic chemical substances that exhibit sex steroid hormone activities have been applied as pharmaceuticals and consumed in large amounts worldwide. They are potentially hazardous contaminants as endocrine disruptors in the environment because they may induce inappropriate gene expression mediated by sex steroid hormone receptors in vivo. Results: To develop simple reporter gene assays with enhanced sensitivity for the detection of sex steroid hormones, we newly established mutant yeast strains lacking the CWP and PDR genes encoding cell wall mannoproteins and plasma membrane drug efflux pumps, respectively, and expressing human ERα, ERβ, AR, and PR. Reporter gene assays with mutant yeast strains responded to endogenous and synthetic ligands more strongly than those with wild-type strains. Sex steroid hormone activities in some pharmaceutical oral tablets and human urine were also detectable in these yeast assays. Conclusions: Yeast reporter gene assay systems for all six steroid hormone receptors, including previously established glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) assay yeasts, are now available. Environmental endocrine disrupters with steroid hormone activity will be qualitatively detectable by simple and easy procedures. The yeast-based reporter gene assay will be valuable as a primary screening tool to detect and evaluate steroid hormone activities in various test samples. Our assay system will strongly support the detection of agonists, antagonists, and inverse agonists of steroid hormone receptors in the field of novel drug discovery and assessments of environmental pollutants.