Activation Of Nuclear Receptors Research Articles

Toxicity Assessment of Groundwater Contaminated by Petroleum Hydrocarbons at a Well-Characterized, Aged, Crude Oil Release Site.

Management of petroleum-impacted waters by monitored natural attenuation requires an understanding of the toxicology of both the original compounds released and the transformation products formed during natural breakdown. Here, we report data from a groundwater plume consisting of a mixture of crude oil compounds and transformation products in an effort to bridge the gap between groundwater quality information and potential biological effects of human exposures. Groundwater samples were characterized for redox processes, concentrations of nonvolatile dissolved organic carbon (NVDOC) and total petroleum hydrocarbons in the diesel range, as well as for activation of human nuclear receptors (hNR) and toxicologically relevant transcriptional pathways. Results show upregulation of several biological pathways, including peroxisome proliferator-activated receptor gamma and alpha, estrogen receptor alpha, and pregnane X receptor (PXR) with higher levels of hNR activity observed in more contaminated samples. Our study of affected groundwater contaminated by a crude-oil release 39 years ago shows these types of waters may have the potential to cause adverse impacts on development, endocrine, and liver functioning in exposed populations. Additionally, positive trends in activation of some of the molecular targets (e.g., PXR) with increasing NVDOC concentrations (including polar transformation products) demonstrate the importance of improving our understanding of the toxicity associated with the unknown transformation products present in hydrocarbon-impacted waters. Our results begin to provide insight into the potential toxicity of petroleum-impacted waters, which is particularly timely given the ubiquitous nature of waters impacted by petroleum contamination not only recently but also in the past and the need to protect drinking-water quality.

SIRT1 and Estrogen Signaling Cooperation for Breast Cancer Onset and Progression.

Breast cancer remains a significant female mortality cause. It constitutes a multifactorial disease for which research on environmental factors offers little help in predicting onset or progression. The pursuit for its foundations by analyzing hormonal changes as a motive for disease development, indicates that increased exposure to estrogens associates with increased risk. A prevalent number of breast cancer cases show dependence on the increased activity of the classic nuclear estrogen receptor (ER) for cell proliferation and survival. SIRT1 is a Type III histone deacetylase which is receiving increasing attention due to its ability to perform activities over relevant non-histone proteins and transcription factors. Interestingly, concomitant SIRT1 overexpression is commonly found in ER-positive breast cancer cases. Both proteins had been shown to directly interact, in a process related to altered intracellular signaling and aberrant transcription, then promoting tumor progression. Moreover, SIRT1 activities had been also linked to estrogenic effects through interaction with the G-protein coupled membrane bound estrogen receptor (GPER). This work aims to summarize present knowledge on the interplay between SIRT1 and ER/GPER for breast cancer onset and progression. Lastly, evidences on the ability of SIRT1 to interact with TGFß signaling, a concurrent pathway significantly involved in breast cancer progression, are reported. The potential of this research field for the development of innovative strategies in the assessment of orphan breast cancer subtypes, such as triple negative breast cancer (TNBC), is discussed.

Recent advances in understanding and managing cholesterol gallstones.

The high prevalence of cholesterol gallstones, the availability of new information about pathogenesis, and the relevant health costs due to the management of cholelithiasis in both children and adults contribute to a growing interest in this disease. From an epidemiologic point of view, the risk of gallstones has been associated with higher risk of incident ischemic heart disease, total mortality, and disease-specific mortality (including cancer) independently from the presence of traditional risk factors such as body weight, lifestyle, diabetes, and dyslipidemia. This evidence points to the existence of complex pathogenic pathways linking the occurrence of gallstones to altered systemic homeostasis involving multiple organs and dynamics. In fact, the formation of gallstones is secondary to local factors strictly dependent on the gallbladder (that is, impaired smooth muscle function, wall inflammation, and intraluminal mucin accumulation) and bile (that is, supersaturation in cholesterol and precipitation of solid crystals) but also to “extra-gallbladder” features such as gene polymorphism, epigenetic factors, expression and activity of nuclear receptors, hormonal factors (in particular, insulin resistance), multi-level alterations in cholesterol metabolism, altered intestinal motility, and variations in gut microbiota. Of note, the majority of these factors are potentially manageable. Thus, cholelithiasis appears as the expression of systemic unbalances that, besides the classic therapeutic approaches to patients with clinical evidence of symptomatic disease or complications (surgery and, in a small subgroup of subjects, oral litholysis with bile acids), could be managed with tools oriented to primary prevention (changes in diet and lifestyle and pharmacologic prevention in subgroups at high risk), and there could be relevant implications in reducing both prevalence and health costs.

Nuclear Receptors as Therapeutic Targets for Neurodegenerative Diseases: Lost in Translation.

Neurodegenerative diseases are characterized by a progressive loss of neurons that leads to a broad range of disabilities, including severe cognitive decline and motor impairment, for which there are no effective therapies. Several lines of evidence support a putative therapeutic role of nuclear receptors (NRs) in these types of disorders. NRs are ligand-activated transcription factors that regulate the expression of a wide range of genes linked to metabolism and inflammation. Although the activation of NRs in animal models of neurodegenerative disease exhibits promising results, the translation of this strategy to clinical practice has been unsuccessful. In this review we discuss the role of NRs in neurodegenerative diseases in light of preclinical and clinical studies, as well as new findings derived from the analysis of transcriptomic databases from humans and animal models. We discuss the failure in the translation of NR-based therapeutic approaches and consider alternative and novel research avenues in the development of effective therapies for neurodegenerative diseases.

In vivo positive mutagenicity of 1,4-dioxane and quantitative analysis of its mutagenicity and carcinogenicity in rats.

1,4-Dioxane is a widely used synthetic industrial chemical and its contamination of drinking water and food is a potential health concern. It induces liver tumors when administered in the drinking water to rats and mice. However, the mode of action (MOA) of the hepatocarcinogenicity of 1,4-dioxane remains unclear. Importantly, it is unknown if 1,4-dioxane is genotoxic, a key consideration for risk assessment. To determine the in vivo mutagenicity of 1,4-dioxane, gpt delta transgenic F344 rats were administered 1,4-dioxane at various doses in the drinking water for 16weeks. The overall mutation frequency (MF) and A:T- to -G:C transitions and A:T- to -T:A transversions in the gpt transgene were significantly increased by administration of 5000ppm 1,4-dioxane. A:T- to -T:A transversions were also significantly increased by administration of 1000ppm 1,4-dioxane. Furthermore, the DNA repair enzyme MGMT was significantly induced at 5000ppm 1,4-dioxane, implying that extensive genetic damage exceeded the repair capacity of the cells in the liver and consequently led to liver carcinogenesis. No evidence supporting other MOAs, including induction of oxidative stress, cytotoxicity, or nuclear receptor activation, that could contribute to the carcinogenic effects of 1,4-dioxane were found. These findings demonstrate that 1,4-dioxane is a genotoxic hepatocarcinogen and induces hepatocarcinogenesis through a mutagenic MOA in rats. Because our data indicate that 1,4-dioxane is a genotoxic carcinogen, we estimated the point of departure of the mutagenicity and carcinogenicity of 1,4-dioxane using the no-observed effect-level approach and the Benchmark dose approach to characterize its dose-response relationship at low doses.

Adverse Outcome Pathway-Driven Analysis of Liver Steatosis in Vitro: A Case Study with Cyproconazole

Adverse outcome pathways (AOPs) describe causal relationships between molecular perturbation and adverse cellular effects and are being increasingly adopted for linking in vitro mechanistic toxicology to in vivo data from regulatory toxicity studies. In this work, a case study was performed by developing a bioassay toolbox to assess key events in the recently proposed AOP for chemically induced liver steatosis. The toolbox is comprised of in vitro assays to measure nuclear receptor activation, gene and protein expression, lipid accumulation, mitochondrial respiration, and formation of fatty liver cells. Assay evaluation was performed in human HepaRG hepatocarcinoma cells exposed to the model compound cyproconazole, a fungicide inducing steatosis in rodents. Cyproconazole dose-dependently activated RARα and PXR, two molecular initiating events in the steatosis AOP. Moreover, cyproconazole provoked a disruption of mitochondrial functions and induced triglyceride accumulation and the formation of fatty liver cells as described in the AOP. Gene and protein expression analysis, however, showed expression changes different from those proposed in the AOP, thus suggesting that the current version of the AOP might not fully reflect the complex mechanisms linking nuclear receptor activation and liver steatosis. Our study shows that cyproconazole induces steatosis in human liver cells in vitro and demonstrates the utility of systems-based approaches in the mechanistic assessment of molecular and cellular key events in an AOP. AOP-driven in vitro testing as demonstrated can further improve existing AOPs, provide insight regarding molecular mechanisms of toxicity, and inform predictive risk assessment.

Estrogen-related receptors alpha, beta and gamma expression and function is associated with transcriptional repressor EZH2 in breast carcinoma

BackgroundOrphan nuclear receptors ERRα, ERRβ and ERRγ that belong to NR3B or type IV nuclear receptor family are well studied for their role in breast cancer pathophysiology. Their homology with the canonical estrogen receptor dictates their possible contributing role in mammary gland development and disease. Although function and regulation of ERRα, ERRγ and less about ERRβ is reported, role of histone methylation in their altered expression in cancer cells is not studied. Transcriptional activity of nuclear receptors depends on co-regulatory proteins. The present study for the first time gives an insight into regulation of estrogen-related receptors by histone methylation specifically through methyltransferase EZH2 in breast cancer.MethodsExpression of ERRα, ERRβ, ERRγ and EZH2 was assessed by immunohistochemistry in four identical tissue array slides that were prepared as per the protocol. The array slides were stained with ERRα, ERRβ, ERRγ and EZH2 simultaneously. Array data was correlated with expression in MERAV expression dataset. Pearson correlation coeficient r was calculated from the partial matrix expression values available at MERAV database to study the strength of association between EZH2 and three orphan nuclear receptors under study. By western blot and real time PCR, their correlated expression was studied in breast cancer cell lines MCF-7, MDA-MB-231, T47D and MDA-MB-453 including normal breast epithelial MCF-10A cells at both protein and RNA level. Regulation of ERRα, ERRβ, ERRγ by EZH2 was further investigated upon overexpression and silencing of EZH2. The interaction between ERRs and EZH2 was validated in vivo by CHIP-qPCR.ResultsWe found a negative correlation between estrogen-related receptors and Enhancer of Zeste Homolog 2, a global repressor gene. Immunohistochemistry in primary breast tumors of different grades showed a correlated expression of estrogen-related receptors and EZH2. Their correlated expression was further validated using online MERAV expression dataset where a negative correlation of variable strengths was observed in breast cancer. Ectopic expression of EZH2 in low EZH2-expressing normal breast epithelial cells abrogated their expression and at the same time, its silencing enhanced the expression of estrogen-related receptors in cancerous cells. Global occupancy of EZH2 on ERRα and ERRβ was observed in-vivo.ConclusionOur findings identify EZH2 as a relevant coregulator for estrogen-related receptors in breast carcinoma.

Novel and emerging therapies for cholestatic liver diseases.

While bile acids are important for both digestion and signalling, hydrophobic bile acids can be harmful, especially when in high concentrations. Mechanisms for the protection of cholangiocytes against bile acid cytotoxicity include negative feedback loops via farnesoid X nuclear receptor (FXR) activation, the bicarbonate umbrella, cholehepatic shunting and anti-inflammatory signalling, among others. By altering or overwhelming these defence mechanisms, cholestatic diseases such as primary biliary cholangitis (PBC) and primary sclerosing cholangitis (PSC) can further progress to biliary cirrhosis, end-stage liver disease and death or liver transplantation. While PBC is currently treated with ursodeoxycholic acid (UDCA) and obeticholic acid (OCA), many fail treatment, and we have yet to find an effective therapy for PSC. Novel therapies under evaluation target nuclear and surface receptors including FXR, transmembrane G-protein-coupled receptor 5 (TGR5), peroxisome proliferator-activated receptor (PPAR) and pregnane X receptor (PXR). Modulation of these receptors leads to altered bile composition, decreased cytotoxicity, decreased inflammation and improved metabolism. This review summarizes our current understanding of the role of bile acids in the pathophysiology of cholestatic liver diseases, presents the rationale for already approved medical therapies and discusses novel pharmacologic therapies under investigation.

Effects of Perinatal Exposure to Dibutyltin Chloride on Fat and Glucose Metabolism in Mice, and Molecular Mechanisms, in Vitro.

Background:The organotin dibutyltin (DBT) is used in the manufacture of polyvinyl chloride (PVC) plastics, in construction materials, and in medical devices. Previous animal studies showed detrimental effects of DBT during in utero development at relatively high doses, but little was known about the effects of DBT exposure at environmentally relevant doses on endpoints such as obesity and metabolic disease.Objectives:We tested the potential obesogenic effects of DBT using in vitro and in vivo models.Methods.We evaluated the effects of DBT on nuclear receptor activation and adipogenic potential using human and mouse multipotent mesenchymal stromal stem cells (MSCs). We also evaluated the effects of perinatal exposure to environmentally relevant doses of DBT in C57BL/6J mice.Results:DBT activated human and mouse and in transient transfection assays, increased expression of adipogenic genes, promoted adipogenic differentiation and increased lipid accumulation in mouse and human MSCs, in vitro. DBT-induced adipogenic differentiation was abolished by the antagonist T0070907, indicating that DBT was acting primarily through . Perinatal exposure to low doses of DBT led to increased fat storage, decreased glucose tolerance, and increased circulating leptin levels in male, but not female, mice.Conclusions:DBT acted as an obesogen by inducing lipid accumulation in human and mouse MSCs through a pathway. In vivo exposure to biologically relevant doses of DBT during perinatal development led to increased fat storage, elevated leptin levels in plasma, and glucose intolerance in mice. Based on these findings, we posit that monitoring of DBT levels in human samples may aid in understanding and potentially preventing the rising rates of metabolic disorders in human populations. https://doi.org/10.1289/EHP3030

The VLDL receptor regulates membrane progesterone receptor trafficking and non-genomic signaling.

Progesterone mediates its physiological functions through activation of both transcription-coupled nuclear receptors and seven-pass-transmembrane progesterone receptors (mPRs), which transduce the rapid non-genomic actions of progesterone by coupling to various signaling modules. However, the immediate mechanisms of action downstream of mPRs remain in question. Herein, we use an untargeted quantitative proteomics approach to identify mPR interactors to better define progesterone non-genomic signaling. Surprisingly, we identify the very-low-density lipoprotein receptor (VLDLR) as an mPRβ (PAQR8) partner that is required for mPRβ plasma membrane localization. Knocking down VLDLR abolishes non-genomic progesterone signaling, which is rescued by overexpressing VLDLR. Mechanistically, we show that VLDLR is required for mPR trafficking from the endoplasmic reticulum to the Golgi. Taken together, our data define a novel function for the VLDLR as a trafficking chaperone required for the mPR subcellular localization and, as such, non-genomic progesterone-dependent signaling.This article has an associated First Person interview with the first author of the paper.

Conserved function of mediator in regulating nuclear hormone receptor activation between plants and animals

ABSTRACTPerception of the plant hormone jasmonoyl-isoleucine (JA-Ile) involves the formation of a co-receptor complex between COI1, the F-box subunit of a SCF-type E3 ubiquitin ligase, and its substrates, a group of jasmonate ZIM-domain (JAZ) transcriptional repressors. In recent studies, we show that MED25, a subunit of the Arabidopsis Mediator, physically and functionally interacts with COI1 and the master transcription factor MYC2 on MYC2 target promoters. Here we provide evidence that MED25 also physically interacts with a subset of JAZ proteins. Therefore, in term of their interaction with Mediator, the JA-Ile co-receptor complex SCFCOI1-JAZs together with the master transcription factor MYC2 resembles the nuclear hormone receptor system of metazoans. In addition, we show that the plant MED25 and its animal counterpart also cooperates with similar epigenetic regulators in distinct signaling pathways. Our study reveals a scenario that plants and animals have evolved distinct, yet largely similar, mechanism for nuclear hormone receptor activation at the level of transcriptional regulation.

Agonistic and Antagonistic Effects of Progesterone Derivatives on the Transcriptional Activity of Nuclear Progesterone Receptor B in Yeast Model System.

Identification of progesterone selective agonists and antagonists that act through one of the nuclear progesterone receptor isoforms is of particular importance for the development of tissue-specific drugs in gynecology and anticancer therapy. Fourteen pregna-D'6- and pregna-D'3-pentarane progesterone derivatives with 16α,17α-cycloalkane groups and two progesterone 3-deoxyderivatives were examined for their ability to regulate transcriptional activity of human nuclear progesterone receptor isoform B (nPR-B) expressed in Saccharomyces cerevisiae yeast. Transcriptional activity of nPR-B was measured from the expression of the β-galactosidase reporter gene with a hormone-responsible element in the promoter. Among the compounds tested, two were full progesterone agonists, four were partial agonists, one compound possessed both agonistic and antagonistic activity, one compound displayed only partial antagonistic activity, and eight compounds did not show any activity. Modifications of the pentarane structure, precisely, introduction of an additional double bound in the A or B rings and/or modification at the 6th position of progesterone, lead to a switch from the complete agonistic activity to partial agonistic or mixed activities. These modifications enable progestins to act as selective modulators of progesterone receptor. Steroids with reduced A-ring and 3-ketogroups lose their ability to regulate PR-B activity. Both 3-deoxycompounds, being selective ligands of progesterone membrane receptors, do not affect PR-B activity.

Profiling of 3696 Nuclear Receptor-Coregulator Interactions: A Resource for Biological and Clinical Discovery.

Nuclear receptors (NRs) are ligand-inducible transcription factors that play critical roles in metazoan development, reproduction, and physiology and therefore are implicated in a broad range of pathologies. The transcriptional activity of NRs critically depends on their interaction(s) with transcriptional coregulator proteins, including coactivators and corepressors. Short leucine-rich peptide motifs in these proteins (LxxLL in coactivators and LxxxIxxxL in corepressors) are essential and sufficient for NR binding. With 350 different coregulator proteins identified to date and with many coregulators containing multiple interaction motifs, an enormous combinatorial potential is present for selective NR-mediated gene regulation. However, NR-coregulator interactions have often been determined experimentally on a one-to-one basis across diverse experimental conditions. In addition, NR-coregulator interactions are difficult to predict because the molecular determinants that govern specificity are not well established. Therefore, many biologically and clinically relevant NR-coregulator interactions may remain to be discovered. Here, we present a comprehensive overview of 3696 NR-coregulator interactions by systematically characterizing the binding of 24 nuclear receptors with 154 coregulator peptides. We identified unique ligand-dependent NR-coregulator interaction profiles for each NR, confirming many well-established NR-coregulator interactions. Hierarchical clustering based on the NR-coregulator interaction profiles largely recapitulates the classification of NR subfamilies based on the primary amino acid sequences of the ligand-binding domains, indicating that amino acid sequence is an important, although not the only, molecular determinant in directing and fine-tuning NR-coregulator interactions. This NR-coregulator peptide interactome provides an open data resource for future biological and clinical discovery as well as NR-based drug design.

PXR: Structure-specific activation by hepatotoxic pyrrolizidine alkaloids

Pyrrolizidine alkaloids (PAs) comprise a large group of more than 660 secondary metabolites found in more than 6000 plant species worldwide. Acute PA intoxication induces severe liver damage. Chronic exposure to sub-lethal doses may cause cumulative damage or cancer. Nuclear receptor activation often constitutes a molecular event for xenobiotic-induced toxicity. However, so far nothing is known about potential interactions of PAs with nuclear receptors as a toxicological mode of action. Thus, in the present study PA-dependent activation of a comprehensive panel of nuclear receptors (PPARs, LXRα, RARα, RXRα, FXR, CAR, PXR, ERα/β) was investigated using GAL4/UAS-based transactivation reporter gene assays. To cover the most frequently occurring PA structure types (retronecine, heliotridine and otonecine type; as well as monoester, open-chain diester and cyclic diester) different PAs were analyzed for interaction with nuclear receptors.Most of the nuclear receptors investigated were not affected by the tested PAs. However, significant activation was found for PXR, which was exclusively activated by the open-chain diesters, echimidine and lasiocarpine. Induction of the model PXR target gene CYP3A4 by PAs was verified at the mRNA, protein and enzyme activity level.In conclusion, PXR activation and PXR-mediated induction of CYP3A4 expression by PAs seem to be structure-dependent. Data suggest that only open-chain diesters act as PXR agonists. This might imply that a PXR-mediated mode of action may contribute to the hepatotoxicity of PAs that is dependent on PA structure.

Tissue-specific effects of benzo[a]pyrene and DDT on microRNA expression profile in female rats

Many xenobiotics in the human environment, such as benzo[a]pyrene (B(a)P) and dichlorodiphenyltrichloroethane (DDT), may act as non-genotoxic carcinogens through epigenetic mechanisms, including changes in microRNA expression profile. In part, such disorders can be mediated by the activation of nuclear receptors, resulting in the activation of protein coding gene expression and microRNAs involved in malignant transformation of cells. Therefore, the aim of this study was to investigate the chain of events “xenobiotic administration – receptor activation – up-regulating microRNA expression – down-regulation target genes expression” as one of the key factors in the chemically-induced carcinogenesis. Using in silico methods, an analysis of the rat genome was carried out to find microRNAs putatively regulated by AhR (aryl hydrocarbon receptor) and CAR (constitutive androstane receptor), activated by BP and DDT, respectively. In particular, miR-3577 and -193b were selected as potentially regulated CAR, miR-207 was selected as a candidate for miR under AhR regulation. The results of the study showed that the treatment of female rats with DDT and B(a)P caused a tissue-specific changes in the expression of microRNAs and host genes in both acute and chronic administration of xenobiotics. To confirm the effects of xenobiotics on the microRNA expression, we also estimated the mRNA level of PTPN6, EIF3F, Cbx7, and Dicer1 genes potentially targeting miR-193b, -207, and -3577. The study has shown a high correlation between the expression of target genes and microRNAs; however these changes depended on the tissue types, the dose and time after xenobiotic treatment.

Polyphenols: Novel Signaling Pathways.

Cardiovascular disease (CVD) is currently the leading cause of death globally. The metabolic syndrome (MetS), a clustering of risk factors including hypertension, hyperglycemia, elevated low-density lipoprotein (LDL) cholesterol, reduced high-density lipoprotein (HDL) cholesterol and increased visceral adiposity, is a significant risk factor for the development of CVD. Non-alcoholic fatty liver disease (NAFLD), often referred to as the hepatic manifestation of MetS, is a constellation of progressive liver disorders closely linked to obesity, diabetes, and insulin resistance. NAFLD initially presents as relatively benign, non-progressive hepatic steatosis, but it may, in certain individuals, progress to nonalcoholic steatohepatitis, fibrosis, cirrhosis, or hepatocellular carcinoma. Currently, there are no validated treatments for NAFLD. Polyphenols are important bioactive dietary compounds and may represent a natural complementary and integrative therapy for the treatment of CVDassociated risk factors, including elevated serum cholesterol and triglyceride levels, as well as NAFLD. Understanding their molecular mechanisms of action is important in the design of future human intervention studies. Several studies utilizing in vitro and in vivo models have helped to identify underlying molecular mechanisms of action of polyphenols. This review will highlight recent advances regarding the molecular actions of dietary procyanidins, with a special focus on those originating from procyanidin-rich grape seed extracts, with a focus on the signaling pathways utilized to exert beneficial metabolic effects. Modulation of nuclear receptor activity and histone deacetylase inhibition has been identified as underlying mechanisms contributing to procyanidin-mediated amelioration of dyslipidemia and steatosis.

Ubiquitylation Pathways In Insulin Signaling and Organismal Homeostasis.

The insulin/insulin-like growth factor-1 (IGF-1) signaling (IIS) pathway is a pivotal genetic program regulating cell growth, tissue development, metabolic physiology, and longevity of multicellular organisms. IIS integrates a fine-tuned cascade of signaling events induced by insulin/IGF-1, which is precisely controlled by post-translational modifications. The ubiquitin/proteasome-system (UPS) influences the functionality of IIS through inducible ubiquitylation pathways that regulate internalization of the insulin/IGF-1 receptor, the stability of downstream insulin/IGF-1 signaling targets, and activity of nuclear receptors for control of gene expression. An age-related decline in UPS activity is often associated with an impairment of IIS, contributing to pathologies such as cancer, diabetes, cardiovascular, and neurodegenerative disorders. Recent findings identified a key role of diverse ubiquitin modifications in insulin signaling decisions, which governs dynamic adaption upon environmental and physiological changes. In this review, we discuss the mutual crosstalk between ubiquitin and insulin signaling pathways in the context of cellular and organismal homeostasis.

Toxic Effects of Bisphenol A, Propyl Paraben, and Triclosan on Caenorhabditis elegans.

Bisphenol A (BPA) is a ubiquitous plasticizer which is absorbed by ingestion and dermal contact; propyl paraben (PPB) inhibits the microbiome and extends the shelf life of many personal care products, whereas triclosan (TCS) is commonly found in antiseptics, disinfectants, or additives. In this work, Caenorhabditis elegans was used as a biological model to assess the toxic effects of BPA, PPB, and TCS. The wild type strain, Bristol N2, was used in bioassays with the endpoints of lethality, growth, and reproduction; green fluorescent protein (GFP) transgenic strains with the hsp-3, hsp-4, hsp-16.2, hsp-70, sod-1, sod-4, cyp-35A4, cyp-29A2, and skn-1 genes were evaluated for their mRNA expression through fluorescence measurement; and quick Oil Red O (q ORO) was utilized to stain lipid deposits. Lethality was concentration-dependent, while TCS and PPB showed more toxicity than BPA. BPA augmented worm length, while PPB reduced it. All toxicants moderately increased the width and the width–length ratio. BPA and PPB promoted reproduction, in contrast to TCS, which diminished it. All toxicants affected the mRNA expression of genes related to cellular stress, control of reactive oxygen species, and nuclear receptor activation. Lipid accumulation occurred in exposed worms. In conclusion, BPA, PPB, and TCS alter the physiology of growth, lipid accumulation, and reproduction in C. elegans, most likely through oxidative stress mechanisms.

Isothiocyanates Activate the Constitutive Androstane Receptor in an Oxidative Stress Dependent Manner in hUGT1 Mice

Our laboratory has previously developed a humanized UDP‐glucuronosyltransferase 1 (hUGT1) mouse model, creating an in vivo model to examine UGT1A metabolism and gene regulation. Mirroring human development, neonatal hUGT1 mice experience a developmental delay in expression of UGT1A1, the sole enzyme capable of bilirubin glucuronidation, resulting in an increase in unconjugated bilirubin. The accumulation of total serum bilirubin (TSB) is readily quantifiable and creates an observable phenotype linked to the expression of UGT1A1. The UGT1A1 gene is regulated by a host of nuclear receptors allowing drugs to be examined in vivo for their effects on nuclear receptor activity. By examination of the relationship between oxidative stress and UGT1A1 expression, we discovered a novel association between the constitutive androstane receptor (CAR) and oxidative stress. In order to induce oxidative stress, hUGT1 mice were treated with phenylethyl isothiocyanate (PEITC), a natural pesticide found in cruciferous vegetables with known anticancer and antioxidant response properties. It is well established that PEITC reacts with glutathione, depleting a major cellular antioxidant, which then activates antioxidant response pathways in response to the change in the cells redox potential. Neonatal hUGT1 mice experienced a drastic decrease in TSB levels following oral PEITC treatment. Concurrently, a significant increase in hepatic and intestinal UGT1A1 expression was observed. In characterizing a mechanism responsible for UGT1A1 induction, we determined CAR to be the major contributor by examining nuclear receptor target genes. CAR's activation was observed by the significant induction CYP2B10, with the Cyp2b10 gene being a consensus CAR target. This suggested a novel association between CAR activation and oxidative stress. In order to confirm CAR's role in the induction of UGT1A1 and CYP2B10, we treated hUGT1/Car(−/−) and hUGT1/Car(+/−) mice orally with PEITC and demonstrated CAR dependence for intestinal and hepatic CYP2B10 induction. We further identified PEITC induction of UGT1A1 was significantly dependent on CAR in liver. PEITC's ability to directly activate CAR was examined using an in vitro transactivation assay, and determined that PEITC is incapable of directly activating CAR by binding to its ligand binding domain. To probe the role of oxidative stress on UGT1A1 induction/CAR activation, hUGT1 mice were pretreated with N‐acetyl cysteine (NAC), an antioxidant and precursor to glutathione. NAC effectively prevented PEITC induced oxidative stress and a concurrent drop in hepatic UGT1A1 and CYP2B10 induction was observed. This suggests a direct correlation between oxidative stress and CAR activation.Support or Funding InformationU.S. Public Health Service Grants ES010337, GM100481, GM086713 (R.H.T.) and R21‐ES024818 (S.C)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

An innate immune response and altered nuclear receptor activation defines the spinal cord transcriptome during alpha-tocopherol deficiency in Ttpa-null mice.

Mice with deficiency in tocopherol (alpha) transfer protein gene develop peripheral tocopherol deficiency and sensory neurodegeneration. Ttpa-/- mice maintained on diets with deficient α-tocopherol (α-TOH) had proprioceptive deficits by six months of age, axonal degeneration and neuronal chromatolysis within the dorsal column of the spinal cord and its projections into the medulla. Transmission electron microscopy revealed degeneration of dorsal column axons. We addressed the potential pathomechanism of α-TOH deficient neurodegeneration by global transcriptome sequencing within the spinal cord and cerebellum. RNA-sequencing of the spinal cord in Ttpa-/- mice revealed upregulation of genes associated with the innate immune response, indicating a molecular signature of microglial activation as a result of tocopherol deficiency. For the first time, low level Ttpa expression was identified in the murine spinal cord. Further, the transcription factor liver X receptor (LXR) was strongly activated by α-TOH deficiency, triggering dysregulation of cholesterol biosynthesis. The aberrant activation of transcription factor LXR suppressed the normal induction of the transcription factor retinoic-related orphan receptor-α (RORA), which is required for neural homeostasis. Thus we find that α-TOH deficiency induces LXR, which may lead to a molecular signature of microglial activation and contribute to sensory neurodegeneration.