Phenobarbital-Elicited Activation of Nuclear Receptor CAR in Induction of Cytochrome P450 Genes

Zelko, Igor N.; Negishi, Masahiko

doi:10.1006/bbrc.2000.3557

Cited by 105 publications

(71 citation statements)

References 34 publications

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“…Similar to its rodent counterparts, hCAR spontaneously translocates to the nucleus in hepatoma cell lines such as HepG2 and Huh7 independently of ligand binding and/or activation. However, in contrast to its rodent counterparts, hCAR is not inhibited by mCAR repressors, such as androgens, androstanol, progesterone, and calmodulin kinase inhibitors (21). Furthermore, specific ligands identified for mCAR, such as PB and TCPOBOP, have less selectivity for the human receptor.…”

Section: Phy Activates and Translocates Hcar In Vivo Inmentioning

confidence: 91%

“…Hepatocytes-Translocation of CAR from the cytoplasm to the nucleus is the initial step during its activation by xenobiotics (21). The ability to identify efficacious CAR activators using cell-based transfection assays is complicated by the spontaneous nuclear accumulation of CAR in transformed cell lines such as HepG2 and Huh7 (18,22).…”

Section: Phy Translocates Hcar From Cytosol To Nucleus In Primary Humanmentioning

confidence: 99%

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Human Constitutive Androstane Receptor Mediates Induction of CYP2B6 Gene Expression by Phenytoin

Wang

Faucette

Moore

et al. 2004

Journal of Biological Chemistry

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139

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Compared with its rodent orthologs, little is known about the chemical specificity of human constitutive androstane receptor (hCAR) and its regulation of hepatic enzyme expression. Phenytoin (PHY), a widely used antiepileptic drug, is a potent inducer of CYP2B6 in primary human hepatocytes, but does not activate human pregnane X receptor (PXR) significantly in cellbased transfection assays at the same concentrations associated with potent induction of CYP2B6. Based on this observation, we hypothesized that PHY may be a selective activator of hCAR. In primary human hepatocytes, expression of CYP2B6 reporter genes containing phenobarbital-responsive enhancer module (PBREM) or PBREM/xenobiotic-responsive enhancer module (XREM) response elements were activated up to 14-and 28-fold, respectively, by 50 M PHY. By contrast, parallel experiments in HepG2 cell lines co-transfected with an hPXR expression vector did not show increased reporter activity. These results indicated that a PXR-independent pathway, which is retained in primary hepatocytes, is responsible for PHY induction of CYP2B6. Further experiments revealed that PHY effectively translocates hCAR from the cytoplasm into the nucleus in both primary human hepatocytes and CAR ؊/؊ mice. Compared with vehicle controls, PHY administration significantly increased CYP2B6 reporter gene expression, when this reporter construct was delivered together with hCAR expression vector into CAR ؊/؊ mice. However, PHY did not increase reporter gene expression in CAR ؊/؊ mice in the absence of hCAR vector, implying that CAR is essential for mediating PHY induction of CYP2B6 gene expression. Taken together, these observations demonstrate that, in contrast to most of the known CYP2B6 inducers, PHY is a selective activator of CAR in humans.Induction of cytochrome P450 2B (CYP2B) 1 expression by xenobiotics, including clinically used drugs, is mediated by activation of the nuclear receptors constitutive androstane receptor (CAR) and/or pregnane X receptor (PXR) through response elements located in the promoter region of CYP2B genes. In contrast to rodent CYP2B genes, mounting evidence suggests that human PXR (hPXR) may be a key receptor in the regulation of human CYP2B6 gene expression (1-3). hPXR ligands, including rifampicin (RIF), phenobarbital (PB), troglitazone, clotrimazole (CLZ), and SR12813 have been reported to induce CYP2B6 gene expression (1, 4 -7). Recently, fourteen commercially available compounds, including weak, moderate, and strong inducers of CYP2B6 and CYP3A4, were used to compare the potency and magnitude of CYP2B6 and CYP3A4 induction and activation of hPXR in primary cultured hepatocytes and hepatoma cell-based reporter gene assays, respectively. Results from these studies indicate that CYP2B6 induction is highly correlated with the activation of hPXR for most compounds (3). Among the efficacious CYP2B6 inducers evaluated, most activate hPXR but not hCAR, such as RIF and CLZ (CLZ was even reported as a hCAR deactivator by Moore et al. (8)), whereas others can a...

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Section: Phy Activates and Translocates Hcar In Vivo Inmentioning

confidence: 91%

Section: Phy Translocates Hcar From Cytosol To Nucleus In Primary Humanmentioning

confidence: 99%

Human Constitutive Androstane Receptor Mediates Induction of CYP2B6 Gene Expression by Phenytoin

Wang

Faucette

Moore

et al. 2004

Journal of Biological Chemistry

Self Cite

139

100

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“…14,15 The constitutive androstane receptor (CAR) is responsible for phenobarbital-induced expression of many cytochrome P450s and other genes involved in detoxification such as glucuronosyl transferases. 16 CAR is a liver-enriched nuclear receptor that, in the absence of signaling, is retained in the cytoplasm in an inactive state, complexed with HSP90. PB treatment results in translocation of CAR to the nucleus through a phosphorylation-dependent mechanism and subsequent activation, leading to transcriptional regulation of PB-responsive genes.…”

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confidence: 99%

Phenobarbital regulates nuclear expression of HNF-4α in mouse and rat hepatocytes independent of CAR and PXR

Bell

Michalopoulos

2006

Hepatology

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Phenobarbital is a lipophilic molecule used as a sedative and antiepileptic drug that elicits a multitude of effects in the liver, including gross liver enlargement, hepatocyte hypertrophy, and induced expression of drug-metabolizing enzymes and other liver-specific genes. The constitutive androstane receptor (CAR; NR1I3) and to a lesser extent the pregnane X receptor (PXR; NR1I2) are responsible for mediating induction of many phenobarbital-responsive genes. However, CAR-mediated transcriptional control of some genes is critically dependent on hepatocyte nuclear factor 4 alpha (HNF-4␣; NR2A1), which itself regulates multiple liver-specific genes involved in hepatic growth, metabolism, and differentiation. We studied the effects of phenobarbital on HNF-4␣ expression in hepatocytes and provide evidence that HNF-4␣ nuclear expression is regulated in response to phenobarbital. Realtime polymerase chain reaction analyses revealed that HNF-4␣ mRNA is modestly upregulated by phenobarbital. In addition, nuclear expression of HNF-4␣ protein is significantly elevated 3 hours after the administration of phenobarbital in wild-type, CAR ؊/؊ , and CAR ؊/؊ /PXR ؊/؊ mice. In vitro analysis revealed that phenobarbital-induced HNF-4␣ expression is both time-and dose dependent. In addition, the phosphatase inhibitor okadaic acid and the Ca 2؉ /calmodulin-dependent protein kinase II inhibitor KN62 block nuclear induction of HNF-4␣ by phenobarbital. Furthermore, HNF-4␣ nuclear expression is enhanced by inhibition of cyclic AMP-dependent protein kinase A. In conclusion, induced nuclear expression of HNF-4␣ and CAR is an integral part of the phenobarbital response, aimed at coordinated regulation of genes involved in drug metabolism and detoxification as well as maintenance of liver function. (HEPATOLOGY 2006;44: 186-194.) P henobarbital (PB) is an antiepileptic drug and is the prototype of a large family of lipophilic PB-like compounds that have profound effects in the liver. Its effects on liver physiology are typified by hepatic hypertrophy, hyperproliferation of the smooth endoplasmic reticulum, and induction or repression of numerous genes, especially the genes of cytochrome P450 enzymes. 1-3 Hepatic hypertrophy induced by PB is mediated by a moderate increase in hepatocyte DNA synthesis and dramatic enlargement of individual hepatocytes. 4,5 Hepatic enlargement subsides after PB withdrawal and this decrease in liver size is mediated by hepatocyte apoptosis. 6 Liver regeneration in response to partial hepatectomy is markedly diminished in PB-treated rat and mouse livers. 7,8 PB is also one of the most effective liver tumor promoters in both rats and mice. The mechanism of liver tumor promotion by PB is not understood; however, there is evidence that PB affects EGF signaling. 9 . Furthermore, TGF␤1 receptors I, II, and III are specifically down-regulated in early neoplastic lesions following treatment with PB. 10,11 It has been hypothesized that the inhibition of DNA synthesis in normal hepatocytes gives a proliferative advant...

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“…CARmediated transcription is further enhanced by the steroid coactivator 1 (SRC-1) and the ubiquitous transcription factor Sp1 (Muangmoonchai et al, 2001). CAR is also know to exhibit constitutive activation in the absence of any known ligand (Zelko and Negishi, 2000).…”

Section: Discussionmentioning

confidence: 99%

Regional Induction of CYP1A1 in Rat Liver Following Treatment with Mixtures of PCB 126 and PCB 153

et al. 2004

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Liver enzyme induction has been shown previously to be regional with clear borders between induced and uninduced regions in vivo, and cells either fully induced or not induced in vitro. The current study examined this phenomenon in vivo by evaluating enzyme induction after exposure to PCB 126 and PCB 153 in female Fisher 344 (F344) and male Sprague-Dawley (SD) rats. IHC revealed a regional induction of CYP1A1 after exposure to PCB 126, apparent in the centrilobular region at lower doses and progressing to panlobular with higher doses. PCB 153 exposure induced CYP2B1/2 in the centrilobular region, which spread to the midzonal region as the dose increased, but never became panlobular even at the highest dosage tested. In rats treated with PCB 126 in combination with high doses of PCB 153, induction of CYP1A1 occurred preferentially in the periportal region, a reversal from the pattern seen with PCB 126 alone. This CYP1A1 induction pattern reversal is a unique example of complex biological interactions between coplanar (PCB 126) and noncoplanar (PCB 153) halogenated aromatic hydrocarbons.

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Phenobarbital-Elicited Activation of Nuclear Receptor CAR in Induction of Cytochrome P450 Genes

Cited by 105 publications

References 34 publications

Human Constitutive Androstane Receptor Mediates Induction of CYP2B6 Gene Expression by Phenytoin

Human Constitutive Androstane Receptor Mediates Induction of CYP2B6 Gene Expression by Phenytoin

Phenobarbital regulates nuclear expression of HNF-4α in mouse and rat hepatocytes independent of CAR and PXR

Regional Induction of CYP1A1 in Rat Liver Following Treatment with Mixtures of PCB 126 and PCB 153

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