Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0406810 (NAME)
 13,345 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hepatocyte nuclear factor 4alpha (HNF4alpha) is an important transcription factor in hepatic gene expression. Here, we have investigated the role of HNF4alpha in the expression of drug-metabolizing enzymes and transporters in human hepatocytes using an adenovirus expressing human HNF4alpha-small interfering RNA (hHNF4alpha-siRNA). The hHNF4alpha-siRNA effectively reduced the mRNA and nuclear protein levels of hHNF4alpha in a concentration-dependent manner. The hHNF4alpha-siRNA also decreased the mRNA levels of CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4, UGT1A1, UGT1A9, SULT2A1, ABCB1, ABCB11, ABCC2, OATP1B1 and OCT1, as well as those of PXR and CAR. To discern the role of these nuclear receptors, we co-infected hepatocytes with hHNF4alpha-siRNA and PXR- or CAR-expressing adenovirus. The hHNF4alpha-siRNA-induced reductions of the enzyme and transporter mRNA levels were not restored except CYP2B6 mRNA levels, which were returned to the control level by overexpressing CAR. Furthermore, although hHNF4alpha-siRNA did not significantly affect the fold-induction of CYP2B6, CYP2C8, CYP2C9, or CYP3A4 mRNA levels following treatment with CYP inducers, the levels in hHNF4alpha-suppressed cells fell significantly compared to the control. These results suggest that HNF4alpha plays a dominant role in the expression of drug-metabolizing enzymes and transporters in human hepatocytes, and that HNF4alpha expression levels is a possible determinant for inter-individual variations in the expression of these enzymes and transporters.
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PMID:Role of human hepatocyte nuclear factor 4alpha in the expression of drug-metabolizing enzymes and transporters in human hepatocytes assessed by use of small interfering RNA. 1782 83

Although regulation of phase I drug metabolism in human liver is relatively well studied, the regulation of phase II enzymes and of drug transporters is incompletely characterized. Therefore, we used human liver slices to investigate the PXR, CAR and AhR-mediated induction of drug transporters and phase I and II metabolic enzymes. Precision-cut human liver slices were incubated for 5 or 24h with prototypical inducers: phenobarbital (PB) (50 microM) for CAR, beta-naphthoflavone (BNF) (25 microM) for AhR, and rifampicin (RIF) (10 microM) for PXR, and gene expression of the phase I enzymes CYP1A1, 1A2, 3A4, 3A5, 2B6, 2A6, the phase II enzymes UGT1A1 and 1A6, and the transporters MRP2, MDR1, BSEP, NTCP and OATP8 was measured. BNF induced CYP1A1, UGT1A1 and UGT1A6 and MRP2, NTCP and MDR1. RIF induced CYP3A4, 3A5, 2B6, 2A6, UGT1A1, UGT1A6 and BSEP, MRP2 and MDR1 and slightly downregulated OATP8. PB induced CYP3A4, 3A5, 2B6 and 2A6, UGT1A1 and all transporters. Large interindividual differences were found with respect to the level of induction. Enzyme activity of CYP3A4, measured by testosterone metabolism, was increased after 24h by RIF. 7-Ethoxycoumarin O-deethylation activity, mediated predominantly by CYP 1A1/1A2 but also by other CYPs, was increased after 24h with PB. We have shown that regulation of all phases of the (in)activation of a drug via the CAR, AhR and the PXR pathways can be studied in human liver slices. The concomitant induction of metabolic enzymes and transporters shows that also in the human liver transporters and metabolic enzymes are regulated coordinately.
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PMID:Coordinated induction of drug transporters and phase I and II metabolism in human liver slices. 1832 80

A panel of retinoids and carotenoids was screened as potential inducers of CYP3A4 through the RXR/VDR-mediated signaling pathway. Transient transfection assays revealed that 3 out of 12 retinoids screened transactivated RXRalpha/VDR and induced CYP3A4 reporter activity. These three retinoids are the active metabolites of retinoids, 9-cis-retinal, 9-cis-retinoic acid (9-cis-RA), and all-trans-retinoic acid (all-trans-RA). 9-cis-RA and all-trans-RA preferentially transactivated the RXR/VDR heterodimers and RXR homodimers. Retinoids and VDR agonist 1alpha, 25-dihydroxyvitamin D(3), but not PXR or CAR activator, could induce Cyp3a11 mRNA level in hepatocytes derived from PXR/CAR-double null mouse. Moreover, retinoids induced CYP3A4 enzyme activity in HepG2 human hepatoma and Caco-2 human colorectal adenocarcinoma cells. A direct role of retinoid-mediated CYP3A4 induction through RXRalpha/VDR was proved by the results that 9-cis-retinal, 9-cis-RA, and all-trans-RA recruited RXRalpha and VDR to CYP3A4 regulatory region pER6 (proximal everted repeat with a 6-nucleotide spacer) and dXREM (distal xenobiotic-responsive enhancer module). Thus, using various approaches, we have unequivocally demonstrated that retinoids transactivate RXR/VDR heterodimers and RXR homodimers and induce CYP3A expression at mRNA as well as enzyme activity levels in both liver and intestinal cells. It is possible that retinoids might alter endobiotic metabolism through CYP3A4 induction in vivo.
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PMID:Retinoids induce cytochrome P450 3A4 through RXR/VDR-mediated pathway. 1840 Feb 6

Lentiviral vectors effectively transduce both dividing and non-dividing cells and stably integrate into the genome of the host cell. In this study, we evaluated the usefulness of a lentiviral system for genetic modulation of primary human hepatocyte cultures. Infection with GFP-expressing lentivectors shows that Huh7 and HepG2 cell lines, as well as primary cultures of human hepatocytes, are efficiently transduced by lentiviral vectors. Real-time RT-PCR analyses demonstrate that infection with lentivectors does not alter hepatic hallmarks such as the expression of the nuclear receptors CAR, PXR, RXR alpha, or HNF4 alpha, or expression of the secretory protein, albumin. Additionally, infected hepatocytes retain the capacity for CYP3A4 induction in response to treatment with phenobarbital, a uniquely sensitive indicator of hepatic differentiation status. Lentivectors may be used for both over-expression and knockdown analyses in primary hepatocytes, as demonstrated in this study by >200-fold CAR over-expression and knockdown of CAR to less than 40% of endogenous levels, with corresponding effects on CYP2B6 expression. In summary, lentiviral vectors provide a novel methodology by which primary human hepatocytes may be stably genetically manipulated, with minimal effects on the differentiated hepatic phenotype. These approaches offer considerable advantage over current methodologies, providing a valuable alternative for use in pharmacological and toxicological investigations involving primary human hepatocyte models and potentially for cell-based therapeutics to treat hepatic dysfunction in vivo.
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PMID:Preservation of hepatic phenotype in lentiviral-transduced primary human hepatocytes. 1846 91

The transcript levels of CYP2B22, 3A22, 3A29, 3A46, CAR, PXR and HNF4alpha were investigated in liver, kidney and airways from control and rifampicin-treated male pigs. The presence and induction of CYP genes transcription were studied by RT-PCR, real-time PCR, Western blotting and enzymatic activity whereas the expression of receptors was studied by RT-PCR or real-time PCR. Pretreatment with rifampicin resulted in a transcriptional activation, although to different extents, of all the CYP3A genes in liver but not in kidney, lung, bronchi or trachea. In the hepatic microsomes, the induction of CYP3A genes was accompanied by an increase of CYP3As marker activities and of two protein bands immunoreactive with anti-human CYP3A4. The CYP2B22 transcript was found to be markedly induced only in liver and kidney. In parallel, a protein band immunoreactive with anti-rat CYP2B1 was elevated while enhanced CYP2B marker activities were observed in hepatic and renal microsomes. As expected, based on human data, the basal expression of CAR, PXR and HNF4alpha was found to be high in liver and low in airways and not susceptible to induction by rifampicin. A significant expression of these transcriptional factors was also demonstrated in kidney. Thus, it is likely that rifampicin induced CYP2B22 both in liver and kidney of pig, not via activation of CAR, but via PXR, through a cross-talk mechanism, as previously observed in human liver. Taken together, our results demonstrated a differential expression and regulation of three individual CYP3As, CYP2B22, CAR, PXR and HNF4alpha genes in liver, kidney and airways of pig.
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PMID:Expression and induction by rifampicin of CAR- and PXR-regulated CYP2B and CYP3A in liver, kidney and airways of pig. 1878 98

CYP3A4 is an important xenobiotic metabolizing enzyme. We previously found that CYP2C55 is highly up-regulated in Cyp3a(-/-) mice. Here, we have further investigated the mechanism of regulation of CYP2C55 and other detoxifying systems in Cyp3a(-/-) mice. Induction studies with prototypical inducers demonstrated an important role for the nuclear receptors PXR and CAR in the up-regulation of CYP2C55. Subsequent diet-switch experiments revealed that food-derived xenobiotics are primarily responsible for the increased induction of CYP2C55, as well as of several other primary detoxifying systems in Cyp3a(-/-) mice. Our data suggest that CYP3A normally metabolizes food-derived activators of PXR and/or CAR, explaining the increased levels of such activators in Cyp3a(-/-) mice and subsequent up-regulation of a range of detoxifying systems. Interestingly, our studies with tissue-specific CYP3A4 transgenic Cyp3a(-/-) mice revealed that not only hepatic but also intestinal expression of CYP3A4 could reduce the hepatic expression of detoxifying systems to near wild-type levels. Apparently, intestinal CYP3A4 can limit the hepatic exposure to food-derived activators of nuclear receptors, thereby regulating the expression of a range of detoxifying systems in the liver. This broad biological effect further emphasizes the importance of intestinal CYP3A activity and could have profound implications for the prediction of drug exposure.
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PMID:Intestinal cytochrome P450 3A plays an important role in the regulation of detoxifying systems in the liver. 1879 35

Phenobarbital (PB) induces or represses a wide spectrum of genes in rodent liver. Much less is known about its effects in human liver. We used pangenomic cDNA microarrays to analyze concentration- and time-dependent gene expression profile changes induced by PB in the well-differentiated human HepaRG cell line. Changes in gene expression profiles clustered at specific concentration ranges and treatment times. The number of correctly annotated genes significantly modulated by at least three different PB concentration ranges (spanning 0.5 to 3.2 mM) at 20 h exposure amounted to 77 and 128 genes (p< or =0.01) at 2- and 1.8-fold filter changes, respectively. At low concentrations (0.5 and 1 mM), PB-responsive genes included the well-recognized CAR- and PXR-dependent responsive cytochromes P450 (CYP2B6, CYP3A4), sulfotransferase 2A1 and plasma transporters (ABCB1, ABCC2), as well as a number of genes critically involved in various metabolic pathways, including lipid (CYP4A11, CYP4F3), vitamin D (CYP24A1) and bile (CYP7A1 and CYP8B1) metabolism. At concentrations of 3.2 mM or higher after 20 h, and especially 48 h, increased cytotoxic effects were associated with disregulation of numerous genes related to oxidative stress, DNA repair and apoptosis. Primary human hepatocyte cultures were also exposed to 1 and 3.2 mM PB for 20 h and the changes were comparable to those found in HepaRG cells treated under the same conditions. Taken altogether, our data provide further evidence that HepaRG cells closely resemble primary human hepatocytes and provide new information on the effects of PB in human liver. These data also emphasize the importance of investigating dose- and time-dependent effects of chemicals when using toxicogenomic approaches.
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PMID:Dose- and time-dependent effects of phenobarbital on gene expression profiling in human hepatoma HepaRG cells. 1908 49

The triazole antifungals myclobutanil, propiconazole and triadimefon cause varying degrees of hepatic toxicity and disrupt steroid hormone homeostasis in rodent in vivo models. To identify biological pathways consistently modulated across multiple timepoints and various study designs, gene expression profiling was conducted on rat livers from three separate studies with triazole treatment groups ranging from 6 h after a single oral gavage exposure, to prenatal to adult exposures via feed. To explore conservation of responses across species, gene expression from the rat liver studies were compared to in vitro data from rat and human primary hepatocytes exposed to the triazoles. Toxicogenomic data on triazoles from 33 different treatment groups and 135 samples (microarrays) identified thousands of probe sets and dozens of pathways differentially expressed across time, dose, and species--many of these were common to all three triazoles, or conserved between rodents and humans. Common and conserved pathways included androgen and estrogen metabolism, xenobiotic metabolism signaling through CAR and PXR, and CYP mediated metabolism. Differentially expressed genes included the Phase I xenobiotic, fatty acid, sterol and steroid metabolism genes Cyp2b2 and CYP2B6, Cyp3a1 and CYP3A4, and Cyp4a22 and CYP4A11; Phase II conjugation enzyme genes Ugt1a1 and UGT1A1; and Phase III ABC transporter genes Abcb1 and ABCB1. Gene expression changes caused by all three triazoles in liver and hepatocytes were concentrated in biological pathways regulating lipid, sterol and steroid homeostasis, identifying a potential common mode of action conserved between rodents and humans. Modulation of hepatic sterol and steroid metabolism is a plausible mode of action for changes in serum testosterone and adverse reproductive outcomes observed in rat studies, and may be relevant to human risk assessment.
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PMID:Toxicogenomic effects common to triazole antifungals and conserved between rats and humans. 1940 4

Retinoids and carotenoids are frequently used as antioxidants to prevent cancer. In this study, a panel of retinoids and carotenoids was examined to determine their effects on activation of RXR/CAR-mediated pathway and regulation of CYP3A gene expression. Transient transfection assays of HepG2 cells revealed that five out of thirteen studied retinoids significantly induced RXRalpha/CAR-mediated activation of luciferase activity that is driven by the thymidine kinase promoter linked with a PXR binding site in the CYP3A4 gene [tk-(3A4)(3)-Luc reporter]. All-trans retinoic acid (RA) and 9-cis RA were more effective than CAR agonist TCBOPOP in induction of the tk-(3A4)(3)-Luc reporter. Addition of retinoid and TCBOPOP further enhanced the inducibility and the induction was preferentially mediated by RXRalpha/CAR and RXRgamma/CAR heterodimer. Chromatin immunoprecipitation assay showed that retinoids recruit RXRalpha and CAR to the proximal ER6 and distal XREM nuclear receptor response elements of the CYP3A4 gene promoter. The experimental data demonstrate that retinoids can effectively regulate CYP3A gene expression through the RXR/CAR-mediated pathway.
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PMID:Retinoids activate RXR/CAR-mediated pathway and induce CYP3A. 1968 1

Weak blood irrigation within solid tumours including hepatocellular carcinomas (HCCs) plays an important role in resistance to anticancer drugs by decreasing accessibility of cytotoxic agents to tumour cells. Reduced oxygen levels, or hypoxia, also contribute to drug resistance because many anticancer drugs require molecular oxygen to be cytotoxic. Our aim was to develop a new in vitro model mimicking hypoxic cells within HCCs in order to further explore the molecular responses to hypoxia, including regulation of drug-metabolising enzymes (DMEs) expression. For this purpose, we used the highly differentiated human hepatoma HepaRG cells cultured under either normoxic or hypoxic (24h at 1% O(2)) conditions. Gene and protein expressions were investigated by quantitative PCR and immunoblotting, respectively. We showed that HepaRG cells adapt to prolonged moderate hypoxia by a switch from aerobic to anaerobic glycolysis and a repression of critical genes involved in amino acid, lipid and ethanol metabolisms. Importantly, expression of several DMEs (particularly cytochromes P450 (CYPs) and phase II enzymes) and xenosensors (CAR, PXR and AhR) was down-regulated and CYPs activities (using testosterone and paclitaxel as substrates) were decreased during hypoxia. In addition, a new role for HIF-1alpha in the repression of CYP3A4 is demonstrated in cells treated with chemical inducers of HIF-1alpha, cobalt chloride or desferrioxamine, and by transfecting untreated HepaRG cells with HIF-1alpha expression vector. In conclusion, HepaRG cells cultured under hypoxia might mimic metabolic changes occurring within poorly irrigated differentiated HCCs. Furthermore, hypoxia down-regulates hepatic DMEs, a phenomenon that might compromise chemotherapy effectiveness in HCC treatment. Thus, HepaRG cells might represent a new in vitro model to test anticancer agents in hypoxic versus normoxic conditions.
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PMID:Drug-metabolising enzymes are down-regulated by hypoxia in differentiated human hepatoma HepaRG cells: HIF-1alpha involvement in CYP3A4 repression. 1969 66


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