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Bisphenol A and its analogues activate human pregnane X receptor.

ABSTRACT: Bisphenol A (BPA) is a base chemical used extensively in many consumer products. BPA and its analogues are present in environmental and human samples. Many endocrine-disrupting chemicals, including BPA, have been shown to activate the pregnane X receptor (PXR), a nuclear receptor that functions as a master regulator of xenobiotic metabolism. However, the detailed mechanism by which these chemicals activate PXR remains unknown.We investigated the mechanism by which BPA interacts with and activates PXR and examined selected BPA analogues to determine whether they bind to and activate PXR.Cell-based reporter assays, in silico ligand-PXR docking studies, and site-directed mutagenesis were combined to study the interaction between BPA and PXR. We also investigated the influence of BPA and its analogues on the regulation of PXR target genes in human LS180 cells.We found that BPA and several of its analogues are potent agonists for human PXR (hPXR) but do not affect mouse PXR activity. We identified key residues within hPXR's ligand-binding pocket that constitute points of interaction with BPA. We also deduced the structural requirements of BPA analogues that activate hPXR. BPA and its analogues can also induce PXR target gene expression in human LS180 cells.The present study advances our understanding of the mechanism by which BPA interacts with and activates human PXR. Activation of PXR by BPA may explain some of the adverse effects of BPA in humans.

SUBMITTER: Sui Y

PROVIDER: S-EPMC3295358 | biostudies-literature | 2012 Mar

REPOSITORIES: biostudies-literature

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Bisphenol A and its analogues activate human pregnane X receptor.

Sui Yipeng Y Ai Ni N Park Se-Hyung SH Rios-Pilier Jennifer J Perkins Jordan T JT Welsh William J WJ Zhou Changcheng C

Environmental health perspectives 20120103 3

<h4>Background</h4>Bisphenol A (BPA) is a base chemical used extensively in many consumer products. BPA and its analogues are present in environmental and human samples. Many endocrine-disrupting chemicals, including BPA, have been shown to activate the pregnane X receptor (PXR), a nuclear receptor that functions as a master regulator of xenobiotic metabolism. However, the detailed mechanism by which these chemicals activate PXR remains unknown.<h4>Objective</h4>We investigated the mechanism by ...[more]

PMID: 22214767

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Project description:Aim: Evidence suggests that bisphenol A diglycidyl ether (BADGE), bisphenol A (BPA), and BPA analogs can interfere with human male fertility. However, the effect directly on human sperm function is not known. The CatSper Ca2+ channel in human sperm controls important sperm functions and is necessary for normal male fertility. Environmental chemicals have been shown to activate CatSper and thereby affect Ca2+ signaling in human sperm. BPA has previously been investigated for effects on Ca2+ signaling human sperm, whereas the effects of other BPA analogs are currently unknown. The aim of this study is thus to characterize the effect of BADGE, BPA, and the eight analogs BPG, BPAF, BPC, BPB, BPBP, BPE, BPF, BPS on Ca2+ signaling, and CatSper in human sperm. Methods: Direct effects of the bisphenols on Ca2+ signaling in human sperm cells were evaluated using a Ca2+ fluorimetric assay measuring changes in intracellular Ca2+. Effects via CatSper were assessed using the specific CatSper inhibitor RU1968. Effects on human sperm function was assessed using an image cytometry-based acrosome reaction assay and the modified Kremer's sperm-mucus penetration assay. Results: At 10 ?M the bisphenols BPG, BPAF, BPC, BADGE, BPB, and BPBP induced Ca2+ signals in human sperm cells, whereas BPE, BPF, BPS, and BPA had no effect. The efficacy of the chemicals at 10 ?M is BPG > BPAF > BPC > BADGE > BPB > BPBP. Dose-response relations of BPG, BPAF, BPC, BADGE, BPB, and BPBP yielded EC50-values in the nM-?M range. The induced Ca2+ signals were almost completely abolished using the CatSper inhibitor RU1968, indicating an effect of the bisphenols on CatSper. All bisphenols, except BPBP, were found to dose-dependently inhibit progesterone-induced Ca2+ signals, with BPG and BPAF displaying inhibition even in low ?M doses. BPG and BPAF were shown to affect human sperm function in a progesterone-like manner. Conclusion: Our results show that the bisphenols BPG, BPAF, BPC, BADGE, BPB, and BPBP can affect Ca2+ signaling in human sperm cells through activation of CatSper. This could potentially disrupt human sperm function by interfering with normal CatSper-signaling and thus be a contributing factor in human infertility, either alone or in mixtures with other chemicals.

Project description:Tetrabromobisphenol A (TBBPA) and tetrachlorobisphenol A (TCBPA) - both halogenated bisphenol (BPA) analogues - are suspected ligands of peroxisome proliferator-activated receptor gamma (PPARγ). While previous studies have shown that TBBPA and TCBPA activate PPARγ within cell-free assays, the downstream effects of TBBPA- and TCBPA-induced PPARγ activation on cellular transcription and physiology have not been thoroughly investigated. Therefore, the objective of this study was to determine whether exposure to TBBPA or TCBPA (either alone or in combination) alters levels of neutral lipids and fatty acid synthase (FASN) - an enzyme that catalyzes synthesis of long-chain saturated fatty acids - within intact cells in a PPARγ-dependent manner. For this study, we relied on human hepatocellular carcinoma (HepG2) cells as a model since these liver cells express basal levels of PPARγ and have been used to study lipoprotein metabolism and regulation of drug metabolizing enzymes. Although exposure to TBBPA and TCBPA alone did not affect cell viability nor neutral lipid and FASN levels in a concentration-dependent manner, exposure to binary mixtures of TBBPA and TCBPA resulted in a concentration-dependent decrease in cell viability in the absence of concentration-dependent effects on neutral lipid and FASN levels. Interestingly, exposure to TBBPA or TCBPA alone or as a mixture enhanced the effects of a reference PPARγ agonist (ciglitazone) and antagonist (GW 9662) on cell viability (but not neutral lipid levels), suggesting that these two halogenated BPA analogues may interact synergistically with ciglitazone and GW 9662 to induce cytotoxicity. However, overexpression of PPARγ did not mitigate nor enhance the effects of TBBPA - a potent PPARγ ligand predicted by ToxCast's cell-free competitive binding assays - on cell viability, neutral lipid levels, nor the cellular transcriptome. Overall, our findings suggest that halogenated BPA analogues such as TCBPA and TBBPA induce toxicity within HepG2 cells in a PPARγ-independent manner.

Project description:The xenobiotic receptors pregnane X receptor (PXR) and constitutive androstane receptor (CAR) are activated by structurally diverse chemicals to regulate the expression of target genes, and they have overlapping regulation in terms of ligands and target genes. Receptor-selective agonists are, therefore, critical for studying the overlapping function of PXR and CAR. An early effort identified 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde-O-(3,4-dichlorobenzyl)oxime (CITCO) as a selective human CAR (hCAR) agonist, and this has since been widely used to distinguish the function of hCAR from that of human PXR (hPXR). The selectivity was demonstrated in a green monkey kidney cell line, CV-1, in which CITCO displayed >100-fold selectivity for hCAR over hPXR. However, whether the selectivity observed in CV-1 cells also represented CITCO activity in liver cell models was not hitherto investigated. In this study, we showed that CITCO: 1) binds directly to hPXR; 2) activates hPXR in HepG2 cells, with activation being blocked by an hPXR-specific antagonist, SPA70; 3) does not activate mouse PXR; 4) depends on tryptophan-299 to activate hPXR; 5) recruits steroid receptor coactivator 1 to hPXR; 6) activates hPXR in HepaRG cell lines even when hCAR is knocked out; and 7) activates hPXR in primary human hepatocytes. Together, these data indicate that CITCO binds directly to the hPXR ligand-binding domain to activate hPXR. As CITCO has been widely used, its confirmation as a dual agonist for hCAR and hPXR is important for appropriately interpreting existing data and designing future experiments to understand the regulation of hPXR and hCAR. SIGNIFICANCE STATEMENT: The results of this study demonstrate that 6-(4-chlorophenyl)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde-O-(3,4-dichlorobenzyl)oxime (CITCO) is a dual agonist for human constitutive androstane receptor (hCAR) and human pregnane X receptor (hPXR). As CITCO has been widely used to activate hCAR, and hPXR and hCAR have distinct and overlapping biological functions, these results highlight the value of receptor-selective agonists and the importance of appropriately interpreting data in the context of receptor selectivity of such agonists.

Dataset Information

Bisphenol A and its analogues activate human pregnane X receptor.

Publications

Bisphenol A and its analogues activate human pregnane X receptor.

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