Biomedical subjects

David D Moore

Publications and source records attributed to David D Moore.

At least 37 records · Page 2Linked to original sources

PubMed2004

Role of the constitutive androstane receptor in xenobiotic-induced thyroid hormone metabolism.

The induction of hepatic drug metabolizing enzymes alters not only the metabolism of the xenobiotic substances that induce them but also the metabolism of various endogenous hormones. The xenobiotic receptor constitutive androstane receptor (CAR) (NR1I3) mediates the well-studied induction of CYP2B genes and other drug-metabolizing enzymes by phenobarbital (PB), an antiepileptic drug that has been shown to alter thyroid hormone (TH) levels. Here we show that CAR is required for PB-mediated disruption of TH homeostasis and the induction of thyroid follicular cell proliferation. Treatment with PB or the more potent and more effective CAR ligand 1, 4-bis-[2-(3, 5,-dichloropyridyloxy)] benzene resulted in universal induction of thyroid hormone glucuronidation and sulfation pathways in a CAR-dependent manner. This resulted in a decrease in serum T4 concentration and a concomitant increase in serum TSH levels. CAR activation also decreased serum T3 levels in mice in which T3 production was blocked. The increase in serum TSH levels resulted in the stimulation of thyroid-follicular cell proliferation. These results highlight the central role of the xenosensor CAR in drug-hormone interactions.

Animals↗

PubMed2004

The constitutive androstane receptor and pregnane X receptor function coordinately to prevent bile acid-induced hepatotoxicity.

A double null mouse line (2XENKO) lacking the xenobiotic receptors CAR (constitutive androstane receptor) (NR1I3) and PXR (pregnane X receptor) (NR1I2) was generated to study their functions in response to potentially toxic xenobiotic and endobiotic stimuli. Like the single knockouts, the 2XENKO mice are viable and fertile and show no overt phenotypes under normal conditions. As expected, they are completely insensitive to broad range xenobiotic inducers able to activate both receptors, such as clotrimazole and dieldrin. Comparisons of the single and double knockouts reveal specific roles for the two receptors. Thus, PXR does not contribute to the process of acetaminophen hepatotoxicity mediated by CAR, but both receptors contribute to the protective response to the hydrophobic bile acid lithocholic acid (LCA). As previously observed with PXR (Xie, W., Radominska-Pandya, A., Shi, Y., Simon, C. M., Nelson, M. C., Ong, E. S., Waxman, D. J., and Evans, R. M. (2001) Proc. Natl. Acad. Sci. U. S. A. 98, 3375-3380), pharmacologic activation of CAR induces multiple LCA detoxifying enzymes and provides strong protection against LCA toxicity. Comparison of their responses to LCA treatment demonstrates that CAR predominantly mediates induction of the cytochrome p450 CYP3A11 and the multidrug resistance-associated protein 3 transporter, whereas PXR is the major regulator of the Na+-dependent organic anion transporter 2. These differential responses may account for the significant sensitivity of the CAR knockouts, but not the PXR knockouts, to an acute LCA dose. Because this sensitivity is not further increased in the 2XENKO mice, CAR may play a primary role in acute responses to this toxic endobiotic. These results define a central role for CAR in LCA detoxification and show that CAR and PXR function coordinately to regulate both xenobiotic and bile acid metabolism.

Acetaminophen↗

PubMed2004

Meclizine is an agonist ligand for mouse constitutive androstane receptor (CAR) and an inverse agonist for human CAR.

The constitutive androstane receptor (CAR, NR1I3) is a key regulator of xenobiotic and endobiotic metabolism. The ligand-binding domains of murine (m) and human (h) CAR are divergent relative to other nuclear hormone receptors, resulting in species-specific differences in xenobiotic responses. Here we identify the widely used antiemetic meclizine (Antivert; Bonine) as both an agonist ligand for mCAR and an inverse agonist for hCAR. Meclizine increases mCAR transactivation in a dose-dependent manner. Like the mCAR agonist 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene, meclizine stimulates binding of steroid receptor coactivator 1 to the murine receptor in vitro. Meclizine administration to mice increases expression of CAR target genes in a CAR-dependent manner. In contrast, meclizine suppresses hCAR transactivation and inhibits the phenobarbital-induced expression of the CAR target genes, cytochrome p450 monooxygenase (CYP)2B10, CYP3A11, and CYP1A2, in primary hepatocytes derived from mice expressing hCAR, but not mCAR. The inhibitory effect of meclizine also suppresses acetaminophen-induced liver toxicity in humanized CAR mice. These results demonstrate that a single compound can induce opposite xenobiotic responses via orthologous receptors in rodents and humans.

Animals↗

PubMed2004

Interactions between hepatic Mrp4 and Sult2a as revealed by the constitutive androstane receptor and Mrp4 knockout mice.

The ABC transporter, Mrp4, transports the sulfated steroid DHEA-s, and sulfated bile acids interact with Mrp4 with high affinity. Hepatic Mrp4 levels are low, but increase under cholestatic conditions. We therefore inferred that up-regulation of Mrp4 during cholestasis is a compensatory mechanism to protect the liver from accumulation of hydrophobic bile acids. We determined that the nuclear receptor CAR is required to coordinately up-regulate hepatic expression of Mrp4 and an enzyme known to sulfate hydroxy-bile acids and steroids, Sult2a1. CAR activators increased Mrp4 and Sult2a1 expression in primary human hepatocytes and HepG2, a human liver cell line. Sult2a1 was down-regulated in Mrp4-null mice, further indicating an inter-relation between Mrp4 and Sult2a1 gene expression. Based on the hydrophilic nature of sulfated bile acids and the Mrp4 capability to transport sulfated steroids, our findings suggest that Mrp4 and Sult2a1 participate in an integrated pathway mediating elimination of sulfated steroid and bile acid metabolites from the liver.

Animals↗

PubMed2004

Alterations in the distribution and orexigenic effects of dexamethasone in CAR-null mice.

The constitutive androstane receptor (CAR, NR1I3) has emerged as an important regulator of drug metabolism. CAR responds to a wide spectrum of xenobiotics by inducing expression of cytochrome P450 (CYP) enzymes and a number of other proteins responsible for drug metabolism in the liver. The xenosensor function of CAR overlaps with that of the pregnane X receptor (PXR), another xenobiotic receptor that belongs to the nuclear hormone superfamily. We observed that injection of dexamethasone (Dex), a ligand for the glucocorticoid receptor (GR) and PXR but not CAR, results in an unexpected twofold increase in the stomach weight of CAR-null animals relative to wild-type animals. Here, we show that CAR knockout mice have elevated levels of Dex in the brain, resulting in a more rapid and robust increase in the hypothalamic expression of the GR-responsive target genes encoding neuropeptide Y (NPY) and neuropeptide Y receptor subtype 1 (NPY-R1). As expected, this is accompanied by a higher increase in the food intake of the CAR-null animals. The data described here highlight the complexity of the overlapping functions of CAR and PXR.

Animals↗

PubMed2004

A novel constitutive androstane receptor-mediated and CYP3A-independent pathway of bile acid detoxification.

Cytosolic sulfotransferase (SULT)-mediated sulfation plays an essential role in the detoxification of bile acids and is necessary to avoid pathological conditions, such as cholestasis, liver damage, and colon cancer. In this study, using transgenic mice bearing conditional expression of the activated constitutive androstane receptor (CAR), we demonstrate that activation of CAR is both necessary and sufficient to confer resistance to the hepatotoxicity of lithocholic acid (LCA). Surprisingly, the CAR-mediated protection is not attributable to the expected and previously characterized CYP3A pathway; rather, it is associated with a robust induction of SULT gene expression and increased LCA sulfation. We have also provided direct evidence that CAR regulates SULT expression by binding to the CAR response elements found within the SULT gene promoters. Interestingly, activation of CAR was also associated with an increased expression of the 3'-phosphoadenosine 5'-phosphosulfate synthetase 2 (PAPSS2), an enzyme responsible for generating the sulfate donor 3'-phosphoadenosine-5'-phosphosulfate. Analysis of gene knockout mice revealed that CAR is also indispensable for ligand-dependent activation of SULT and PAPSS2 in vivo. Therefore, we establish an essential and unique role of CAR in controlling the mammalian sulfation system and its implication in the detoxification of bile acids.

Animals↗

PubMed2004

A traditional herbal medicine enhances bilirubin clearance by activating the nuclear receptor CAR.

Yin Zhi Huang, a decoction of Yin Chin (Artemisia capillaris) and three other herbs, is widely used in Asia to prevent and treat neonatal jaundice. We recently identified the constitutive androstane receptor (CAR, NR1I3) as a key regulator of bilirubin clearance in the liver. Here we show that treatment of WT and humanized CAR transgenic mice with Yin Zhi Huang for 3 days accelerates the clearance of intravenously infused bilirubin. This effect is absent in CAR knockout animals. Expression of bilirubin glucuronyl transferase and other components of the bilirubin metabolism pathway is induced by Yin Zhi Huang treatment of WT mice or mice expressing only human CAR, but not CAR knockout animals. 6,7-Dimethylesculetin, a compound present in Yin Chin, activates CAR in primary hepatocytes from both WT and humanized CAR mice and accelerates bilirubin clearance in vivo. We conclude that CAR mediates the effects of Yin Zhi Huang on bilirubin clearance and that 6,7-dimethylesculetin is an active component of this herbal medicine. CAR is a potential target for the development of new drugs to treat neonatal, genetic, or acquired forms of jaundice.

Animals↗

PubMed2004

Bile acids lower triglyceride levels via a pathway involving FXR, SHP, and SREBP-1c.

We explored the effects of bile acids on triglyceride (TG) homeostasis using a combination of molecular, cellular, and animal models. Cholic acid (CA) prevents hepatic TG accumulation, VLDL secretion, and elevated serum TG in mouse models of hypertriglyceridemia. At the molecular level, CA decreases hepatic expression of SREBP-1c and its lipogenic target genes. Through the use of mouse mutants for the short heterodimer partner (SHP) and liver X receptor (LXR) alpha and beta, we demonstrate the critical dependence of the reduction of SREBP-1c expression by either natural or synthetic farnesoid X receptor (FXR) agonists on both SHP and LXR alpha and LXR beta. These results suggest that strategies aimed at increasing FXR activity and the repressive effects of SHP should be explored to correct hypertriglyceridemia.

Animals↗

PubMed2003

Resistance of SHP-null mice to bile acid-induced liver damage.

The orphan nuclear hormone receptor SHP (gene designation NROB2) is an important component of a negative regulatory cascade by which high levels of bile acids repress bile acid biosynthesis. Short term studies in SHP null animals confirm this function and also reveal the existence of additional pathways for bile acid negative feedback regulation. We have used long term dietary treatments to test the role of SHP in response to chronic elevation of bile acids, cholesterol, or both. In contrast to the increased sensitivity predicted from the loss of negative feedback regulation, the SHP null mice were relatively resistant to the hepatotoxicity associated with a diet containing 0.5% cholic acid and the much more severe effects of a diet containing both 0.5% cholic acid and 2% cholesterol. This was associated with decreased hepatic accumulation of cholesterol and triglycerides in the SHP null mice. There were also alterations in the expression of a number of genes involved in cholesterol and bile acid homeostasis, notably cholesterol 12alpha-hydroxylase (CYP8B1), which was strongly reexpressed in the SHP null mice, but not the wild type mice fed either bile acid containing diet. This contrasts with the strong repression of CYP8B1 observed with short term bile acid feeding, as well as the effects of long term feeding on other bile acid biosynthetic enzymes such as cholesterol 7alpha-hydroxylase (CYP7A1). CYP8B1 expression could contribute to the decreased toxicity of the chronic bile acid treatment by increasing the hydrophilicity of the bile acid pool. These results identify an unexpected role for SHP in hepatotoxicity and suggest new approaches to modulating effects of chronically elevated bile acids in cholestasis.

Animals↗

PubMed2003

Complex effects of rexinoids on ligand dependent activation or inhibition of the xenobiotic receptor, CAR.

BACKGROUND: CAR/RXR heterodimers bind a variety of hormone response elements and activate transcription in the absence of added ligands. This constitutive activity of murine CAR can be inhibited by the inverse agonist ligand androstanol or increased by the agonist TCPOBOP. RXR agonists activate some RXR heterodimer complexes, which are termed permissive, while other non-permissive complexes are not responsive to such ligands. RESULTS: Direct protein-protein interaction studies demonstrate that the RXR agonist 9-cis-RA increases interaction of CAR/RXR heterodimers with the coactivator SRC-3, but also inhibits the ability of TCPOBOP to increase and androstanol to decrease coactivator binding. CAR transactivation of a response element with a five nucleotide spacer (DR-5) is unaffected by 9-cis-RA or the synthetic RXR agonist LG1069. In agreement with the inhibitory effect observed in vitro, these rexinoids block both the TCPOBOP mediated transactivation of this element and the androstanol dependent inhibition. In contrast, CAR transactivation of other response elements is increased by rexinoids. Stable expression of CAR in a HepG2 derived cell line increases expression of the endogenous CAR target CYP2B6. This expression is further increased by TCPOBOP but decreased by either androstanol or LG1069, and LG1069 blocks the stimulatory effect of TCPOBOP but not the inhibitory effect of androstanol. CONCLUSION: We conclude that CAR/RXR heterodimers are neither strictly permissive nor non-permissive for RXR signaling. Instead, rexinoids have distinct effects in different contexts. These results expand the potential regulatory mechanisms of rexinoids and suggest that such compounds may have complex and variable effects on xenobiotic responses.

Journal Article↗

PubMed2003

Induction of bilirubin clearance by the constitutive androstane receptor (CAR).

Bilirubin clearance is one of the numerous important functions of the liver. Defects in this process result in jaundice, which is particularly common in neonates. Elevated bilirubin levels can be decreased by treatment with phenobarbital. Because the nuclear hormone receptor constitutive androstane receptor (CAR) mediates hepatic effects of this xenobiotic inducer, we hypothesized that CAR could be a regulator of bilirubin clearance. Activation of the nuclear hormone receptor CAR increases hepatic expression of each of five components of the bilirubin-clearance pathway. This induction is absent in homozygous CAR null mice but is observed in mice expressing human CAR instead of mouse CAR. Pretreatment with xenobiotic inducers markedly increases the rate of clearance of an exogenous bilirubin load in wild-type but not CAR knockout animals. Bilirubin itself can also activate CAR, and mice lacking CAR are defective in clearing chronically elevated bilirubin levels. Unexpectedly, CAR expression is very low in livers of neonatal mice and humans. We conclude that CAR directs a protective response to elevated bilirubin levels and suggest that a functional deficit of CAR activity may contribute to neonatal jaundice.

Actins↗

PubMed2003

GUGULIPID: a natural cholesterol-lowering agent.

The resin of the Commiphora mukul tree has been used in Ayurvedic medicine for more than 2000 years to treat a variety of ailments. Studies in both animal models and humans have shown that this resin, termed gum guggul, can decrease elevated lipid levels. The stereoisomers E- and Z-guggulsterone have been identified as the active agents in this resin. Recent studies have shown that these compounds are antagonist ligands for the bile acid receptor farnesoid X receptor (FXR), which is an important regulator of cholesterol homeostasis. It is likely that this effect accounts for the hypolipidemic activity of these phytosteroids.

Animals↗

PubMed2003

The orphan receptor SHP and the three-hybrid interference assay.

The three-hybrid interference method described here is similar to previous interference assays based on the two-hybrid system. In these approaches the presence of a third protein disrupts the interaction between the two different hybrid proteins. The three-hybrid interference assay circumvents a potential problem that arises when the third protein is itself a direct transcriptional repressor by preventing the decrease in transcriptional readout that could occur if the third protein is recruited indirectly to the two-hybrid protein complex. This three-hybrid method should be applicable to any third protein that is a transcriptional repressor. To use this method to explore the effects of such repressors on protein-protein interactions, however, it is important to confirm that the VP16-repressor fusion does not retain active repressor function. It is also appropriate to compare the effects of the VP16-repressor fusion to those with the repressor alone. Finally, it is essential to confirm any conclusions from this or any other single method to study protein-protein interactions with alternative independent approaches.

Animals↗

PubMed2003

Induction of multidrug resistance protein 3 (mrp3) in vivo is independent of constitutive androstane receptor.

We previously demonstrated that multidrug resistance protein 3 (Mrp3/ABCC3) is induced in rat liver by phenobarbital (PB) and several other microsomal enzyme inducers that induce cytochrome P450 2B (CYP2B). CYP2B is induced by constitutive androstane receptor (CAR)-retinoid X receptor (RXR) heterodimer binding to a phenobarbital-responsive promoter element in the CYP2B promoter. Hepatic mRNA levels of CYP2B and Mrp3 were measured in three models of altered CAR activity to determine whether CAR is also involved in the induction of Mrp3. In Wistar Kyoto rats, where males express higher CAR protein levels than females, the induction of CYP2B1/2 was significantly higher in males than in females by PB, diallyl sulfide, and trans-stilbene oxide but not oltipraz. Mrp3 was induced by each of these treatments, but in contrast to CYP2B1/2, to a similar magnitude in males and females. In male hepatocyte-specific RXRalpha-/- mice, CYP2B10 was not induced by diallyl sulfide or oltipraz but remained inducible by PB and trans-stilbene oxide after considering the decrease in basal CYP2B10 expression. Mrp3, however, was induced by PB, diallyl sulfide, trans-stilbene oxide and oltipraz in both wild-type and RXRalpha-/- mice. Additionally, constitutive expression of Mrp3 was significantly reduced in RXRalpha-/- mice. In CAR-/- mice, the robust induction of CYP2B10 by PB was completely absent. However, Mrp3 was equally induced both in wild-type and CAR-/- mice by PB. These data clearly demonstrate that induction of hepatic Mrp3 by PB and other microsomal enzyme inducers is CAR-independent and implies a role for RXRalpha in the constitutive expression of Mrp3.

Animals↗

PubMed2002

Modulation of acetaminophen-induced hepatotoxicity by the xenobiotic receptor CAR.

We have identified the xenobiotic receptor CAR (constitutive androstane receptor) as a key regulator of acetaminophen metabolism and hepatotoxicity. Known CAR activators as well as high doses of acetaminophen induced expression of three acetaminophen-metabolizing enzymes in wild-type but not in CAR null mice, and the CAR null mice were resistant to acetaminophen toxicity. Inhibition of CAR activity by administration of the inverse agonist ligand androstanol 1 hour after acetaminophen treatment blocked hepatotoxicity in wild type but not in CAR null mice. These results suggest an innovative therapeutic approach for treating the adverse effects of acetaminophen and potentially other hepatotoxic agents.

Acetaminophen↗

PubMed2002

A natural product that lowers cholesterol as an antagonist ligand for FXR.

Extracts of the resin of the guggul tree (Commiphora mukul) lower LDL (low-density lipoprotein) cholesterol levels in humans. The plant sterol guggulsterone [4,17(20)-pregnadiene-3,16-dione] is the active agent in this extract. We show that guggulsterone is a highly efficacious antagonist of the farnesoid X receptor (FXR), a nuclear hormone receptor that is activated by bile acids. Guggulsterone treatment decreases hepatic cholesterol in wild-type mice fed a high-cholesterol diet but is not effective in FXR-null mice. Thus, we propose that inhibition of FXR activation is the basis for the cholesterol-lowering activity of guggulsterone. Other natural products with specific biologic effects may modulate the activity of FXR or other relatively promiscuous nuclear hormone receptors.

Animals↗

PubMed2002

CREB-binding protein/p300 co-activation of crystallin gene expression.

Although some of the transcription factors that are required for expression of crystallins during lens development have been identified, the molecular interactions that contribute to enhanced crystallin expression are not yet well defined. In this study, we designed experiments to test whether the co-activators CREB-binding protein (CBP) and/or p300 interact with c-Maf, Prox-1, or Sox-1 to enhance transcription of crystallin genes. Promoter regions from the mouse alphaA-, betaB2-, and gammaF-crystallin genes were linked to a luciferase reporter. Expression of c-Maf transactivated each of these promoters. Of particular interest, co-expression of CBP or p300 with c-Maf was found to synergistically co-activate each promoter. CBP and p300 were less effective or ineffective at co-activation with Prox-1 or Sox-1. Co-immunoprecipitation and mammalian two-hybrid experiments revealed that CBP and p300 bind to c-Maf and Prox-1 but not to Sox-1. The co-activation of c-Maf by CBP/p300 requires histone acetyltransferase activity. Our results suggest that c-Maf recruits CBP and/or p300 to crystallin promoters leading to up-regulation of crystallin gene expression through localized histone acetylation and consequent chromatin re-modeling. In a promoter-specific fashion, co-activation can be modulated by Prox-1 and/or Sox-1. This modulation may help to specify the endogenous levels of crystallin gene expression.

Acetyltransferases↗

PubMed2002

FOR, a novel orphan nuclear receptor related to farnesoid X receptor.

We have identified and characterized a new amphibian orphan member of the nuclear receptor superfamily and termed it FOR1 (farnesoid X receptor (FXR)-like Orphan Receptor) because it shares the highest amino acid identity with the mammalian FXR. We also identified a variant of FOR1, called FOR2, which has 15 additional C-terminal amino acids. Both variants include an unusual insertion of 33 amino acids in the helix 7 region of the canonical ligand binding domain sequence, suggesting a unique structure for FOR. Northern blot analysis demonstrates that the FOR gene is highly expressed in adult and tadpole liver, kidney, and tail bud stage of the embryo. Detailed expression analysis using in situ hybridization indicates that FOR expression is first detectable at stage 30/31 in the presumptive liver region lasting until stage 41 with a peak level evident at stage 35/36. FOR forms heterodimeric complexes with retinoid X receptor (RXR) as demonstrated by biochemical and mammalian two-hybrid approaches. Gel mobility shift assays demonstrate that FORs form specific DNA-protein complexes on an FXR binding element consisting of an inverted repeat DNA element with 1 nucleotide spacing (IR1) from the phospholipid transfer protein gene promoter. Finally, although FORs do not exhibit constitutive transcriptional activity, frog gallbladder extract significantly augments the transcriptional activities of FORs.

Amino Acid Sequence↗