RESUMEN

Spatial heterogeneity and plasticity of the mammalian liver are critical for systemic metabolic homeostasis in response to fluctuating nutritional conditions. Here, a spatially resolved transcriptomic landscape of mouse livers across fed, fasted and refed states using spatial transcriptomics is generated. This approach elucidated dynamic temporal-spatial gene cascades and how liver zonation-both expression levels and patterns-adapts to shifts in nutritional status. Importantly, the pericentral nuclear receptor Nr1i3 (CAR) as a pivotal regulator of triglyceride metabolism is pinpointed. It is showed that the activation of CAR in the pericentral region is transcriptionally governed by Pparα. During fasting, CAR activation enhances lipolysis by upregulating carboxylesterase 2a, playing a crucial role in maintaining triglyceride homeostasis. These findings lay the foundation for future mechanistic studies of liver metabolic heterogeneity and plasticity in response to nutritional status changes, offering insights into the zonated pathology that emerge during liver disease progression linked to nutritional imbalances.

RESUMEN

Hawke's Bay in New Zealand was impacted by Cyclone Gabrielle in 2023, experiencing intense weather conditions and rainfall. Rivers and streams surged beyond their banks, displacing large amounts of sediment. The sewage treatment plant and industries in the Waitangi catchment, south of the city of Napier, were heavily impacted, making them potential sources of contaminants. The aim of this study was to investigate the risk of displaced sediments deposited south of Napier City, using bioassays and chemical analysis methods. Sediment samples were collected across a gradient between the coastline and the Waitangi Stream. The toxicity of chemically extracted or elutriate samples was assessed by Microtox®, mussel embryo-larval development, and aryl hydrocarbon and constitutive androstane receptor yeast two-hybrid assays. Targeted chemical analysis and automated identification and quantification system (AIQS-GC) methods were used to identify contaminants. The elutriates showed low toxicity and the yeast assays showed levels of activity like those previously reported. Chemical methods confirmed historical contamination by DDT and its metabolites DDE and DDD, as well as by plant sterols. Overall, the toxicity and chemicals detected are what would be expected from a typical agricultural soil. The risk posed by the displaced sediment in the Waitangi catchment can be considered low. Combining chemical and bioanalytical methods was an effective approach to investigate the potential risks of post-disaster contamination.

RESUMEN

OBJECTIVE: The prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) continues to rise with the increasing obesity epidemic. Rezdiffra as an activator of a thyroid hormone receptor-beta is the only Food and Drug Administration approved therapy. As such, there is a critical need to improve our understanding of gene expression regulation and signaling transduction in MASLD to develop new therapies. Matrin-3 is a DNA- and RNA-binding protein involved in the pathogenesis of human diseases. Here we examined its previously uncharacterized role in limiting hepatic steatosis and stress response via the constitutive androstane receptor (CAR). METHODS: Matrin-3 floxed and liver-specific knockout mice were fed either a chow diet or 60 kcal% high-fat diet (HFD) for up to 16 weeks. The mice were euthanized for different analysis including liver histology, lipid levels, and gene expression. Bulk RNA-seq, bulk ATAC-seq, and single-nucleus Multiome were used to examine changes of transcriptome and chromatin accessibility in the liver. Integrative bioinformatics analysis of our data and publicly available datasets and different biochemical assays were performed to identify underlying the molecular mechanisms mediating matrin-3's effects. Liver-tropic adeno-associated virus was used to restore the expression of CAR for lipid, acute phase genes, and histological analysis. RESULTS: Matrin-3 expression is induced in the steatotic livers of mice. Liver-specific matrin-3 deletion exacerbated HFD-induced steatosis, acute phase response, and inflammation in the liver of female mice. The transcriptome and chromatin accessibility were re-programmed in the liver of these mice with signatures indicating that CAR signaling is dysregulated. Mechanistically, matrin-3 interacts with CAR mRNA, and matrin-3 deficiency promotes CAR mRNA degradation. Consequently, matrin-3 deletion impaired CAR signaling by reducing CAR expression. Matrin-3 levels positively correlate with CAR expression in human livers. Ces2a and Il1r1 were identified as new target genes of CAR. Interestingly, we found that CAR discords with the expression of its target genes including Cyp2b10 and Ces2a in response to HFD, indicating CAR signaling is dysregulated by HFD despite increased CAR expression. Dysregulated CAR signaling upon matrin-3 deficiency reduced Ces2a and de-repressed Il1r1 expression. CAR restoration partially abrogated the dysregulated gene expression, exacerbated hepatic steatosis, acute phase response, and inflammation in liver-specific matrin-3 knockout mice fed a HFD. CONCLUSIONS: Our findings demonstrate that matrin-3 is a key upstream regulator maintaining CAR signaling upon metabolic stress, and the matrin-3-CAR axis limits hepatic steatosis and stress response signaling that may give insights for therapeutic intervention.

Asunto(s)

Receptor de Androstano Constitutivo , Hígado Graso , Hígado , Ratones Noqueados , Animales , Masculino , Ratones , Dieta Alta en Grasa/efectos adversos , Hígado Graso/metabolismo , Hígado Graso/genética , Hígado/metabolismo , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/genética , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética , Transducción de Señal , Estrés Fisiológico

RESUMEN

Triclosan (TCS) is an antimicrobial toxicant found in a myriad of consumer products and has been detected in human tissues, including breastmilk. We have evaluated the impact of lactational TCS on UDP-glucuronosyltransferase 1A1 (UGT1A1) expression and bilirubin metabolism in humanized UGT1 (hUGT1) neonatal mice. In hUGT1 mice, expression of the hepatic UGT1A1 gene is developmentally delayed resulting in elevated total serum bilirubin (TSB) levels. We found that newborn hUGT1 mice breastfed or orally treated with TCS presented lower TSB levels along with induction of hepatic UGT1A1. Lactational and oral treatment by gavage with TCS leads to the activation of hepatic nuclear receptors constitutive androstane receptor (CAR), peroxisome proliferator-activated receptor alpha (PPARα), and stress sensor, activating transcription factor 4 (ATF4). When CAR-deficient hUGT1 mice (hUGT1/Car-/-) were treated with TCS, TSB levels were reduced with a robust induction of hepatic UGT1A1, leaving us to conclude that CAR is not tied to UGT1A1 induction. Alternatively, when PPARα-deficient hUGT1 mice (hUGT1/Pparα-/-) were treated with TCS, hepatic UGT1A1 was not induced. Additionally, we had previously demonstrated that TCS is a potent inducer of ATF4, a transcriptional factor linked to the integrated stress response. When ATF4 was deleted in liver of hUGT1 mice (hUGT1/Atf4ΔHep) and these mice treated with TCS, we observed superinduction of hepatic UGT1A1. Oxidative stress genes in livers of hUGT1/Atf4ΔHep treated with TCS were increased, suggesting that ATF4 protects liver from excessive oxidative stress. The increase oxidative stress may be associated with superinduction of UGT1A1. The expression of ATF4 in neonatal hUGT1 hepatic tissue may play a role in the developmental repression of UGT1A1.

Asunto(s)

Factor de Transcripción Activador 4 , Animales Recién Nacidos , Bilirrubina , Glucuronosiltransferasa , Hígado , PPAR alfa , Triclosán , Animales , Glucuronosiltransferasa/metabolismo , Glucuronosiltransferasa/genética , PPAR alfa/metabolismo , PPAR alfa/genética , Ratones , Factor de Transcripción Activador 4/metabolismo , Factor de Transcripción Activador 4/genética , Triclosán/farmacología , Humanos , Bilirrubina/farmacología , Bilirrubina/metabolismo , Hígado/metabolismo , Hígado/efectos de los fármacos , Ratones Noqueados , Femenino , Receptor de Androstano Constitutivo , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética

RESUMEN

The fungicide fluxapyroxad (BAS 700 F) has been shown to significantly increase the incidence of liver tumours in male Wistar rats at dietary levels of 1500 and 3000 ppm and in female rats at a dietary level of 3000 ppm via a non-genotoxic mechanism. In order to elucidate the mode of action (MOA) for fluxapyroxad-induced rat liver tumour formation a series of in vivo and in vitro investigative studies were undertaken. The treatment of male and female Wistar rats with diets containing 0 (control), 50, 250, 1500 and 3000 ppm fluxapyroxad for 1, 3, 7 and 14 days resulted in a dose-dependent increases in relative weight at 1500 and 3000 ppm from day 3 onwards in both sexes, with an increase in relative liver weight being also observed in male rats given 250 ppm fluxapyroxad for 14 days. Examination of liver sections revealed a centrilobular hepatocyte hypertrophy in some fluxapyroxad treated male and female rats. Hepatocyte replicative DNA synthesis (RDS) was significantly increased in male rats given 1500 and 3000 ppm fluxapyroxad for 3 and 7 days and in female rats given 50-3000 ppm fluxapyroxad for 7 days and 250-3000 ppm fluxapyroxad for 3 and 14 days; the maximal increases in RDS in both sexes being observed after 7 days treatment. The treatment of male and female Wistar rats with 250-3000 ppm fluxapyroxad for 14 days resulted in significant increases in hepatic microsomal total cytochrome P450 (CYP) content and CYP2B subfamily-dependent enzyme activities. Male Wistar rat hepatocytes were treated with control medium and medium containing 1-100 µM fluxapyroxad or 500 µM sodium phenobarbital (NaPB) for 4 days. Treatment with fluxapyroxad and NaPB increased CYP2B and CYP3A enzyme activities and mRNA levels but had little effect on markers of CYP1A and CYP4A subfamily enzymes and of the peroxisomal fatty acid ß-oxidation cycle. Hepatocyte RDS was significantly increased by treatment with fluxapyroxad, NaPB and 25 ng/ml epidermal growth factor (EGF). The treatment of hepatocytes from two male human donors with 1-100 µM fluxapyroxad or 500 µM NaPB for 4 days resulted in some increases in CYP2B and CYP3A enzyme activities and CYP mRNA levels but had no effect on hepatocyte RDS, whereas treatment with EGF resulted in significant increase in RDS in both human hepatocyte preparations. Hepatocytes from male Sprague-Dawley wild type (WT) and constitutive androstane receptor (CAR) knockout (CAR KO) rats were treated with control medium and medium containing 1-16 µM fluxapyroxad or 500 µM NaPB for 4 days. While both fluxapyroxad and NaPB increased CYP2B enzyme activities and mRNA levels in WT hepatocytes, only minor effects were observed in CAR KO rat hepatocytes. Treatment with both fluxapyroxad and NaPB only increased RDS in WT and not in CAR KO rat hepatocytes, whereas treatment with EGF increased RDS in both WT and CAR KO rat hepatocytes. In conclusion, a series of in vivo and in vitro investigative studies have demonstrated that fluxapyroxad is a CAR activator in rat liver, with similar properties to the prototypical CAR activator phenobarbital. A robust MOA for fluxapyroxad-induced rat liver tumour formation has been established. Based on the lack of effect of fluxapyroxad on RDS in human hepatocytes, it is considered that the MOA for fluxapyroxad-induced liver tumour formation is qualitatively not plausible for humans.

Asunto(s)

Receptor de Androstano Constitutivo , Fungicidas Industriales , Hepatocitos , Ratas Wistar , Receptores Citoplasmáticos y Nucleares , Animales , Masculino , Femenino , Ratas , Fungicidas Industriales/toxicidad , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Humanos , Hepatocitos/efectos de los fármacos , Hepatocitos/metabolismo , Hepatocitos/patología , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Relación Dosis-Respuesta a Droga , Tamaño de los Órganos/efectos de los fármacos , Neoplasias Hepáticas Experimentales/inducido químicamente , Neoplasias Hepáticas Experimentales/patología , Neoplasias Hepáticas Experimentales/metabolismo , Replicación del ADN/efectos de los fármacos , Sistema Enzimático del Citocromo P-450/metabolismo , Sistema Enzimático del Citocromo P-450/genética , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/metabolismo , Neoplasias Hepáticas/inducido químicamente , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología

RESUMEN

4-Nonylphenol (4-NP), a para-substituted phenolic compound with a straight or branched carbon chain, is a ubiquitous environmental pollutant and food contaminant. 4-NP, particularly the branched form, has been identified as an endocrine disruptor (ED) with potent activities on estrogen receptors. Constitutive Androstane Receptor (CAR) is another crucial nuclear receptor that regulates hepatic lipid, glucose, and steroid metabolism and is involved in the ED mechanism of action. An NP mixture has been described as an extremely potent activator of both human and rodent CAR. However, detailed mechanistic aspects of CAR activation by 4-NP are enigmatic, and it is not known if 4-NP can directly interact with the CAR ligand binding domain (LBD). Here, we examined interactions of individual branched (22NP, 33NP, and 353NP) and linear 4-NPs with CAR variants using molecular dynamics (MD) simulations, cellular experiments with various CAR expression constructs, recombinant CAR LBD in a TR-FRET assay, or a differentiated HepaRG hepatocyte cellular model. Our results demonstrate that branched 4-NPs display more stable poses to activate both wild-type CAR1 and CAR3 variant LBDs in MD simulations. Consistently, branched 4-NPs activated CAR3 and CAR1 LBD more efficiently than linear 4-NP. Furthermore, in HepaRG cells, we observed that all 4-NPs upregulated CYP2B6 mRNA, a relevant hallmark for CAR activation. This is the first study to provide detailed insights into the direct interaction between individual 4-NPs and human CAR-LBD, as well as its dominant variant CAR3. The work could contribute to the safer use of individual 4-NPs in many areas of industry.

Asunto(s)

Fenoles , Humanos , Fenoles/química , Fenoles/metabolismo , Receptor de Androstano Constitutivo/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Disruptores Endocrinos/química , Simulación de Dinámica Molecular

RESUMEN

Scoparone (6, 7 dimethylesculetin) is a biologically active compound derived from the herb Artemisia capillaris having anti-inflammatory, anti-lipemic, and anti-allergic roles. Activation of the constitutive androstane receptor (CAR) in primary hepatocytes of both wild-type and humanized CAR mice by scoparone, accelerates bilirubin and cholesterol clearance in vivo. This can prevent gallstones which is a dreaded gastrointestinal disease. To date, surgery is regarded as the gold standard for treating gallstones. The molecular interactions between scoparone and CAR leading to gallstone prevention are not yet explored. In this study, we have analyzed these interactions through an insilico approach. After extracting the CAR structures (mice and human) from the protein databank and 6, 7-dimethylesuletin from PubChem, energy minimization of both the receptors was done to make them stable followed by docking. Next, a simulation was performed to stabilize the docked complexes. Through docking, H-bonds and pi-pi interactions were found in the complexes, which imply a stable interaction, thus activating the CAR. A similarity search for scoparone was performed and the selected compounds were docked with the CAR receptors. Esculentin acetate and scopoletin acetate interacted with human CAR through pi-alkyl and H-bond respectively. While Fraxidin methyl ether, fraxinol methyl ether, and 6, 7 diethoxycoumarin interacted with mice CAR through H-bond and Pi-Pi T-shaped bonds. The selected complexes were simulated further. Our results are in accordance with the hypothesis in the literature. We have also analyzed the drug likeliness, absorption, non-carcinogenicity, and other properties of scoparone which can support further in vivo studies.Communicated by Ramaswamy H. Sarma.

Asunto(s)

Cumarinas , Cálculos Biliares , Éteres Metílicos , Ratones , Humanos , Animales , Receptor de Androstano Constitutivo , Receptores Citoplasmáticos y Nucleares , Acetatos

RESUMEN

This study aimed to determine the regulatory mechanism of dietary zinc lactate (ZL) supplementation on intestinal oxidative stress damage in a paraquat (PQ)-induced piglet model. Twenty-eight piglets (mean body weight 9.51 ± 0.23 kg) weaned at 28 d of age were randomly divided into control, ZL, PQ, and ZL + PQ groups (n = 7 in each group). The ZL-supplemented diet had little effect on growth performance under normal physiological conditions. However, under PQ challenge, ZL supplementation significantly improved average daily gain (P < 0.05) and reduced the frequency of diarrhea. ZL improved intestinal morphology and ultrastructure by significantly increasing the expression level of the jejunal tight junction protein, zonula occludens-1 (ZO-1) (P < 0.05), and intestinal zinc transport and absorption in PQ-induced piglets, which reduced intestinal permeability. ZL supplementation also enhanced the expression of antioxidant and anti-inflammatory factor-related genes and decreased inflammatory cytokine expression and secretion in PQ-induced piglets. Furthermore, ZL treatment significantly inhibited the activation of constitutive androstane receptor (CAR) signaling (P < 0.01) in PQ-induced piglets and altered the structure of the gut microbiota, especially by significantly increasing the abundance of beneficial gut microbes, including UCG_002, Ruminococcus, Rikenellaceae_RC9_gut_group, Christensenellaceae_R_7_group, Treponema, unclassified_Christensenellaceae, and unclassified_Erysipelotrichaceae (P < 0.05). These data reveal that pre-administration of ZL to piglets can suppress intestinal oxidative stress by improving antioxidant and anti-inflammatory capacity and regulating the crosstalk between CAR signaling and gut microbiota.

RESUMEN

Traditionally, xenobiotic receptors are known for their role in chemical sensing and detoxification, as receptor activation regulates the expression of various key enzymes and receptors. However, recent studies have highlighted that xenobiotic receptors also play a key role in the regulation of lipid metabolism and therefore function also as metabolic sensors. Since dyslipidemia is a major risk factor for various cardiometabolic diseases, like atherosclerosis and non-alcoholic fatty liver disease, it is of major importance to understand the molecular mechanisms that are regulated by xenobiotic receptors. In this review, three major xenobiotic receptors will be discussed, being the aryl hydrocarbon receptor (AhR), pregnane X receptor (PXR) and the constitutive androstane receptor (CAR). Specifically, this review will focus on recent insights into the metabolic functions of these receptors, especially in the field of lipid metabolism and the associated dyslipidemia.

Asunto(s)

Dislipidemias , Receptores de Esteroides , Humanos , Receptor X de Pregnano/metabolismo , Receptor de Androstano Constitutivo , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores de Esteroides/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Xenobióticos/metabolismo , Proteínas Portadoras

RESUMEN

The constitutive androstane receptor (CAR) and the pregnane X receptor (PXR) are nuclear receptors that are involved in the regulation of gene transcription of enzymes that are responsible for biotransformation and excretion of endo- and xenobiotics. The goal of the work was to study the effect of DL-butyonine sulfoximine (BSO, gamma-glutamylcysteine synthetase inhibitor) on the relative amounts of CAR and PXR in Caco-2 cells and to clarify its mechanisms. BSO was used at concentrations of 1-500 µM for 24 and 72 h. The generation of reactive oxygen species (ROS) has been evaluated using the MitoTracker Red CM-H2 XRos fluorescent probes. Cytotoxicity was analyzed by the MTT test. The relative amount of CAR and PXR was assessed by the Western blot method. It has been shown that BSO caused an increase in ROS formation at concentrations of 10, 50, and 100 µM for 24 h and at concentrations of 50 and 100 µM for 72 h. However, 500 µM BSO reduced the viability of cells during all periods of exposure. The relative amount of CAR increased in 24 h at the BSO concentrations of 50 and 100 µM and in 72 h at its concentrations of 10 and 50 µM. The amount of PXR increased in 72 h during incubation with BSO at the concentration of 50 µM and in 24 and 72 h at its concentrations of 100 and 500 µM. The combined use of BSO (50 µM, 24 h; 10 and 50 µM, 72 h) and glutathione inhibited CAR induction, whereas 50 and 100 µM BSO inhibited PXR formation for 72 h. The addition of 1 mM glutathione to the nutrient medium with BSO (100 and 500 µM, 24 h; 500 µM, 72 h) did not affect the relative amount of PXR. No effect on CAR was observed when 1 mM glutathione was used together with BSO (100 µM, 24 h; 50 and 100 µM, 72 h). Thus, BSO can induce CAR and PXR formation by both increasing the production of free radicals, thus developing oxidative stress, and by acting independently as a xenobiotic.

RESUMEN

Activation of the constitutive androstane receptor (CAR, NR1I3) by chemical compounds induces liver hyperplasia in rodents. 1,4-Bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP), a mouse CAR agonist, is most often used to study chemically induced liver hyperplasia and hepatocyte proliferation in vivo. TCPOBOP is a potent murine liver chemical mitogen, which induces rapid liver hyperplasia in mice independently of liver injury. In recent years, great amount of data has been accumulated on the transcription program that characterizes the TCPOBOP-induced hepatocyte proliferation. However, there are only few data about the metabolic requirements of hepatocytes that divide upon exposure to xenobiotics. In the present study, we have employed liquid chromatography - mass spectrometry technology combined with statistical analysis to investigate metabolite profile of small biomolecules, in order to identify key metabolic changes in the male mouse liver tissue after TCPOBOP administration. Analysis of biochemical pathways of the differentially affected metabolites in the mouse liver demonstrated significant TCPOBOP-mediated enrichment of several processes including those associated with nucleotide metabolism, amino acid metabolism, and energy substrate metabolism. Our findings provide evidence to support the conclusion that the CAR agonist, TCPOBOP, initiates an intracellular program that promotes global coordinated metabolic activities required for hepatocyte proliferation. Our metabolic data might provide novel insight into the biological mechanisms that occur during the TCPOBOP-induced hepatocyte proliferation in mice.

Asunto(s)

Receptor de Androstano Constitutivo , Receptores Citoplasmáticos y Nucleares , Animales , Masculino , Ratones , Proliferación Celular , Receptor de Androstano Constitutivo/agonistas , Hepatocitos/metabolismo , Hiperplasia/metabolismo , Hiperplasia/patología , Hígado/metabolismo , Ratones Endogámicos C57BL , Receptores Citoplasmáticos y Nucleares/metabolismo

RESUMEN

Non-Alcoholic Fatty Liver Disease (NAFLD) is the most common cause of chronic liver disease worldwide, affecting 70-90% of obese individuals. In humans, a lower NAFLD incidence is reported in pre-menopausal women, although the mechanisms affording this protection remain under-investigated. Here, we tested the hypothesis that the constitutive androstane nuclear receptor (CAR) plays a role in the pathogenesis of experimental NAFLD. Male and female wild-type (WT) and CAR knock-out (CAR-/-) mice were subjected to a high-fat diet (HFD) for 16 weeks. We examined the metabolic phenotype of mice through body weight follow-up, glucose tolerance tests, analysis of plasmatic metabolic markers, hepatic lipid accumulation, and hepatic transcriptome. Finally, we examined the potential impact of HFD and CAR deletion on specific brain regions, focusing on glial cells. HFD-induced weight gain and hepatic steatosis are more pronounced in WT males than females. CAR-/- females present a NASH-like hepatic transcriptomic signature suggesting a potential NAFLD to NASH transition. Transcriptomic correlation analysis highlighted a possible cross-talk between CAR and ERα receptors. The peripheral effects of CAR deletion in female mice were associated with astrogliosis in the hypothalamus. These findings prove that nuclear receptor CAR may be a potential mechanism entry-point and a therapeutic target for treating NAFLD/NASH.

Asunto(s)

Enfermedades Metabólicas , Enfermedad del Hígado Graso no Alcohólico , Femenino , Masculino , Humanos , Animales , Ratones , Enfermedad del Hígado Graso no Alcohólico/etiología , Dieta Alta en Grasa/efectos adversos , Obesidad , Peso Corporal

RESUMEN

The constitutive androstane receptor (CAR) regulates enzyme transcription related to drug metabolism; therefore, natural compound clarification in food that interacts with CAR is significant for drug development. We revealed that 13-epimanool, which is a compound found in the common sage, is bound to hCAR based on differential scanning fluorometry (DSF) measurements using recombinant hCAR protein. Similar labdane diterpenoids were examined, which revealed that manool and sclareol, which were both natural compounds contained in herbs, are bound to hCAR. They exhibited different effects for CAR activity in the luciferase assay despite the structural similarity. Manool was a partial agonist, 13-epimanool was a weak partial agonist, and sclareol was an antagonist. The activity of hCAR may be regulated by slight differences in the bound compound.

Asunto(s)

Receptor de Androstano Constitutivo , Diterpenos , Humanos , Receptores Citoplasmáticos y Nucleares , Diterpenos/farmacología

RESUMEN

Breast cancer resistance protein (BCRP,ABCG2) is an efflux transporter protein that transports various substrates from the cell to the extracellular space or organ cavities. The aim of this study was a complex assessment of the amount of BCRP during pregnancy in rabbits. The amount of BCRP in samples of the rabbit jejunum, liver, kidney, cerebral cortex, and placenta was determined by enzyme immunoassay, and in human hepatocellular carcinoma (HepG2) cells by the Western blot. To study the mechanisms involved in control of the dynamic BCRP levels during pregnancy, serum concentrations of sex hormones were investigated by radioimmunoassay and relative amounts of constitutive androstane receptor (CAR) and pregnane X receptor (PXR) in these organs were evaluated using the Western blot method. The putative role of CAR and PXR in regulation of the BCRP level by progesterone was evaluated in vitro experiments on HepG2 cells. It was found that amount of BCRP in the jejunum of pregnant rabbits was higher than in the placenta, liver, kidneys, and cerebral cortex. An increase in the amount of BCRP in the liver of rabbits was noted on the 21st day of pregnancy and a tendency to the increase was also detected on the 28th day; in the kidney and cerebral cortex increased BCRP levels were detected on the 28th day and 14th day of pregnancy, respectively, as compared with non-pregnant females. In vitro experiments with HepG2 cells have shown that the increase in the BCRP level is determined by the activating effect of progesterone on PXR.

Asunto(s)

Neoplasias de la Mama , Proteínas de Neoplasias , Femenino , Humanos , Embarazo , Animales , Conejos , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Progesterona , Riñón

RESUMEN

Environmental chemicals, which are known to impact offspring health, have become a public concern. Constitutive activated receptor (CAR) is activated by various environmental chemicals and participates in xenobiotic metabolism. Here, we described the effects of maternal exposure to the CAR-specific ligand 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP, TC) on offspring health outcomes. Maternal TC exposure exhibited a stronger inhibition of body weight in 3-week-old and 8-week-old first-generation (F1) offspring female mice compared to controls. Further, maternal TC exposure obtained a strong increase in hepatic drug-metabolizing enzyme expression in 3-week-old female mice that persisted into 8-week-old adulthood. Interestingly, we observed distorted intestinal morphological features in 8-week-old F1 female mice in the TC-exposed group. Moreover, maternal TC exposure triggered a loss of intestinal barrier integrity by reducing the expression of intestinal tight junction proteins. Accordingly, maternal exposure to TC down-regulated serum triglyceride levels as well as decreased the expression of intestinal lipid uptake and transport marker genes. Mechanistically, maternal TC exposure activated the intestinal inflammatory response and disrupted the antioxidant system in the offspring female mice, thereby impeding the intestinal absorption of nutrients and seriously threatening offspring health. Altogether, these findings highlight that the effects of maternal TC exposure on offspring toxicity could not be ignored.

Asunto(s)

Receptor de Androstano Constitutivo , Receptores Citoplasmáticos y Nucleares , Animales , Femenino , Humanos , Ratones , Crecimiento y Desarrollo , Hígado/metabolismo , Exposición Materna , Ratones Endogámicos C57BL , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo

RESUMEN

Phthalates are extensively used in the production of plastics products and have been verified to induce lung injury. Lycopene (LYC) has proved an effective preventive and can be utilized to prevent phthalates-induced toxicity. However, the role of phthalate in pathogenesis of lung injury remain poorly researched, and little work has been devoted whether LYC could alleviate phthalate-induced lung toxicity via modulating nuclear xenobiotic receptors (NXRs) response. Here, di (2-ethylhexyl) phthalate (DEHP) is used as the representative of phthalates for further studies on toxicity of phthalates and the antagonistic role of LYC in phthalates-induced lung injury. We found that DEHP exposure caused alveoli destruction and alveolar epithelial cells type II damage. Mechanistically, DEHP exposure increased nuclear accumulation of aryl hydrocarbon receptor (AHR) and its downstream genes level, including cytochrome P450-dependent monooxygenase (CYP) 1A1 and CYP1B1. Constitutive androstane receptor (CAR) and their downstream gene level, including CYP2E1 are also increased after phthalates exposure. Significantly, LYC supplementation relieves lung injury from DEHP exposure by inhibiting the activation of NXRs. We confirm that NXRs plays a key role in phthalates-induced lung injury. Our study showed that LYC may have a positive role in alleviating the toxicity effects of phthalates, which provides an effective strategy for revising phthalates-induced injury.

Asunto(s)

Dietilhexil Ftalato , Lesión Pulmonar , Ácidos Ftálicos , Humanos , Dietilhexil Ftalato/toxicidad , Lesión Pulmonar/inducido químicamente , Licopeno/farmacología , Ácidos Ftálicos/toxicidad , Receptores Citoplasmáticos y Nucleares/metabolismo , Xenobióticos/toxicidad , Aminoácidos/metabolismo

RESUMEN

BACKGROUND: Breviscapine is a flavonoid extracted from Erigeron breviscapus (Vant.) Hand.-Mazz., and mainly contains scutellarin. Nuclear receptors play important roles in regulating transporter and drug metabolic enzymes. OBJECTIVE: To investigate the regulatory effects of scutellarin on CYP3A4 and 2C19 in HepG2 and Caco-2 cells based on nuclear receptors PXR and CAR. METHODS: The proteins and mRNA levels of CYP3A4 and CYP2C19 treated with scutellarin were detected by Western Blot and RT-qPCR. Using assays of the dual-luciferase reporter, promoter sequences containing hPXR and hCAR protein recognition and binding regulatory elements CYP3A4 and CYP2C19 were inserted upstream of the reporter gene, and the expression vector and the reporter vector were cotransfected into HepG2 and Caco-2 cells. RESULTS: Scutellarin inhibited mRNA of CYP3A4 and PXR, and promoted mRNA expression of CYP2C19 and CAR in RT-qPCR results. Western-blot results showed scutellarin inhibited the expression of CYP3A4 and promoted the expression of CYP2C19. The dual-luciferase reporter genes showed that scutellarin enhanced the expression level of CYP2C19, and when its concentration was 40 and 80µmol/L, CYP3A4 was significantly increased. CONCLUSION: Scutellarin down-regulates CYP3A4 through PXR, and its mechanism may work by up-regulating CAR, binding to PXR to inhibit PXR-mediated expression of CYP3A4. Scutellarin up-regulates CYP2C19 through CAR.

Asunto(s)

Receptor de Androstano Constitutivo , Receptores de Esteroides , Humanos , Receptor X de Pregnano , Citocromo P-450 CYP3A/genética , Citocromo P-450 CYP3A/metabolismo , Citocromo P-450 CYP2C19 , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Células CACO-2 , Receptores Citoplasmáticos y Nucleares/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Luciferasas

RESUMEN

Constitutive androstane receptor (CAR) is a nuclear receptor that plays a key role in drug metabolism and disposition and in the development of liver tumors in rodents. CAR is activated by ligands and indirect activators, which do not bind to the receptor but activate it through cellular signaling. In this study, we sought to identify direct and indirect activators of rat CAR (rCAR). Assessment of the influence of mutations on the transcriptional activity of rCAR identified a mutant termed rCAR-3A-G354Q that displays low constitutive activity and high ligand responsiveness. Reporter assays using the mutant were performed with compounds that increased the mRNA levels of Cyp2b1, a CAR target gene, in rat primary hepatocytes. Several compounds activated rCAR-3A-G354Q and were implicated as rCAR ligands. Since indirect CAR activators are considered to display little species differences, we then determined CYP2B6 mRNA levels in human hepatocyte-like HepaRG cells after treatment with compounds that increased Cyp2b1 mRNA levels in rat hepatocytes but did not activate rCAR-3A-G354Q. The results demonstrated six compounds as possible rCAR indirect activators. Taken together, the combined measurement of Cyp2b1 mRNA levels in rat primary hepatocytes and rCAR-3A-G354Q activation in reporter assays can be useful for evaluating rCAR activation by chemicals.

Asunto(s)

Receptor de Androstano Constitutivo , Citocromo P-450 CYP2B1 , Ratas , Humanos , Animales , Citocromo P-450 CYP2B1/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Hepatocitos/metabolismo , Ligandos , ARN Mensajero/genética , ARN Mensajero/metabolismo

RESUMEN

Phenanthrene (Phe) exposure is associated with skin ageing, cardiotoxicity and developmental defects. Here, we investigated the mode of Phe toxicity in human keratinocytes (HaCaT cells) and the attenuation of toxicity on pre-treatment (6 h) with ethanol extract of Hibiscus sabdariffa calyxes (HS). Cell viability, reactive oxygen species (ROS) generation, mitochondrial membrane potential (ΔΨm) alteration, changes in the transcriptional activity of selected genes involved in phase I and II metabolism, antioxidant response and gluconeogenesis, western blot and docking studies were performed to determine the protective effect of HS against Phe. Phe (250 µM) induced cytotoxicity in HaCaT cells through AhR-independent, CAR/PXR/RXR-mediated activation of CYP1A1 and the subsequent alterations in phase I and II metabolism genes. Further, CYP1A1 activation by Phe induced ROS generation, reduced ΔΨm and modulated antioxidant response, phase II metabolism and gluconeogenesis-related gene expression. However, pre-treatment with HS extract restored the pathological changes observed upon Phe exposure through CYP1A1 inhibition. Docking studies showed the site-specific activation of PXR and CAR by Phe and inhibition of CYP1A1 and CYP3A4 by the bioactive compounds of HS similar to that of the positive controls tested. Our results conclude that HS extract can attenuate Phe-induced toxicity in HaCaT cells through CAR/PXR/RXR mediated inhibition of CYP1A1.

Asunto(s)

Hibiscus , Fenantrenos , Extractos Vegetales/farmacología , Receptores de Esteroides , Antioxidantes/farmacología , Receptor de Androstano Constitutivo , Citocromo P-450 CYP1A1 , Citocromo P-450 CYP3A , Etanol , Células HaCaT , Humanos , Receptor X de Pregnano , Especies Reactivas de Oxígeno , Receptores Citoplasmáticos y Nucleares , Receptores de Esteroides/metabolismo

RESUMEN

Bilirubin is a heme catabolite and Ugt1a1 is the only enzyme involved in the biological elimination of bilirubin. Partially functional or non-functional Ugt1a1 may result in neuronal damage and death due to the accumulation of unconjugated bilirubin in the brain. The understanding of the role of alternative bilirubin detoxification mechanisms that can reduce bilirubin toxicity risk is crucial for developing novel therapeutic strategies. To provide a proof-of-principle showing whether activation of alternative detoxification pathways could lead to life-compatible bilirubin levels in the absence of Ugt1a1 activity, we used Ugt1-/- hyperbilirubinemic mice devoid of bilirubin glucuronidation activity. We treated adult Ugt1-/- mice with TCPOBOP, a strong agonist of the constitutive androstane receptor (CAR). TCPOBOP treatment decreased plasma and liver tissue bilirubin levels by about 38%, and resulted in the transcriptional activation of a vast array of genes involved in bilirubin transport and metabolism. However, brain bilirubin level was unaltered. We observed ~40% degradation of bilirubin in the liver microsomes from TCPOBOP treated Ugt1-/- mice. Our findings suggest that, in the absence of Ugt1a1, the activation of alternative bilirubin clearance pathways can partially improve hyperbilirubinemic conditions. This therapeutic approach may only be considered in a combinatorial manner along with other treatments.

Asunto(s)

Bilirrubina , Hiperbilirrubinemia , Animales , Modelos Animales de Enfermedad , Glucuronosiltransferasa/genética , Glucuronosiltransferasa/metabolismo , Hemo/metabolismo , Hígado/metabolismo , Ratones