Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UMLS:C0019204 (hepatocellular carcinoma)
71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

alpha-Naphthoflavone (alpha NF) is a weak aryl hydrocarbon (Ah) receptor agonist and inhibits the induction of CYP1A1 gene expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin. It has been suggested that the Ah receptor antagonist activity is due to the formation of alpha NF-cytosolic Ah receptor complexes that fail to undergo transformation. This hypothesis is consistent with data obtained in this and other studies using alpha NF concentrations from 10 to 1000 nM. However, 10 microM alpha NF exhibited Ah receptor agonist activity in several assays. Incubation of rat hepatic cytosol with 10 microM alpha NF caused transformation of the Ah receptor, as determined in a gel retardation assay using a 32P-labeled oligonucleotide containing a single dioxin-responsive element (DRE). Incubation of rat hepatoma (H-4-II E) cells with 10 microM alpha NF not only resulted in the induction of CYP1A1 mRNA levels but also increased chloramphenicol acetyltransferase activity from a DRE-containing chloramphenicol acetyltransferase reporter plasmid. Moreover, the DRE-transformed cytosolic Ah receptor complex liganded with either alpha NF or 2,3,7,8-tetrachlorodibenzo-p-dioxin did not undergo significant dissociation at 4 degrees. These data confirm that alpha NF is an Ah receptor agonist and, based on the results of previous studies, exhibits partial antagonist activity via competition for receptor binding sites.
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PMID:alpha-Naphthoflavone-induced CYP1A1 gene expression and cytosolic aryl hydrocarbon receptor transformation. 838 8

We have used a ligation-mediated polymerase chain reaction technique to analyze protein-DNA interactions at a dioxin-responsive enhancer upstream of the CYP1A1 gene in intact mouse hepatoma cells. In its inactive state, the enhancer binds few, if any, proteins within the major DNA groove in vivo. Thus, the inactive enhancer is relatively inaccessible to DNA-binding proteins. Exposure of cells to 2,3,7,8-tetrachlorodibenzo-p-dioxin leads to the binding of the liganded Ah receptor at six sites within the major DNA groove of the enhancer. The receptor-enhancer interactions occur rapidly and do not require ongoing transcription, consistent with their role in regulating CYP1A1 gene expression. The liganded receptor, which is a heteromer composed of at least two basic helix-loop-helix proteins, is probably the only DNA-binding transcription factor necessary to activate the enhancer in vivo. The small size and irregular distribution of receptor binding sites suggest that chromatin structure imposes substantial steric constraints upon the function of the receptor-enhancer system in intact cells.
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PMID:Mechanism of dioxin action: receptor-enhancer interactions in intact cells. 838 88

The DNA upstream of the dioxin-inducible CYP1A1 gene contains six distinct sites to which the liganded Ah receptor binds in intact mouse hepatoma cells. Here, we have analyzed these six bona fide receptor-binding sites in order to study the relationships between DNA sequence, receptor binding, and dioxin responsiveness. Gel retardation studies reveal that the sites vary by about 7-fold in their relative affinities for the liganded receptor. Within this range, there is no obvious association between the strength of receptor binding and the degree of dioxin responsiveness, as measured in transfection experiments. In fact, one site binds the receptor well but fails to respond to dioxin. This observation implies that the receptor-DNA binding event per se is not sufficient to confer dioxin responsiveness upon a linked gene. Comparison of the DNA sequences of the six receptor-binding sites permits the derivation of a "functional consensus" recognition sequence, which is more extended in length than the "core"-binding sequence previously described. In corroboration of these results, protein-DNA cross-linking studies indicate that the liganded receptor contacts base pairs beyond the core sequence. Our observations also indicate that the liganded receptor can tolerate limited sequence heterogeneity at its DNA-binding site and still elicit a response to dioxin. This finding might reflect corresponding heterogeneity in the amino acid sequence of the liganded receptor's DNA-binding domain.
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PMID:Protein-DNA interactions at a dioxin-responsive enhancer. Analysis of six bona fide DNA-binding sites for the liganded Ah receptor. 838 16

In the presence of halogenated and polycyclic aromatic hydrocarbons, the CYP1A1 gene is regulated through induction after ligand binding to the cytosolic Ah receptor (AhR). Ligand-dependent AhR activation leads to nuclear translocation and binding of the receptor to dioxin-responsive element (DRE) sequences, an event that initiates transcriptional activation of the CYP1A1 gene. We recently established a human hepatoma cell line stably integrated with the human CYP1A1 promoter and 5'-flanking enhancer sequences fused to the firefly luciferase gene. This cell line, 101L, was used to determine whether the induction of CYP1A1 by omeprazole, a gastric proton pump inhibitor, is AhR mediated. Treatment of 101L cells with either 50 microM omeprazole or 5 nM 2,3,7,8-tetrachlorodibenzo-p-dioxin for 12-72 hr resulted in maximal activity at 24 hr for both inducers. A dose-response curve for omeprazole induction at 24 hr was determined and the EC50 for omeprazole induction of the human CYP1A1 gene was estimated to be 100 microM. The induction of the CYP1A1 gene by omeprazole corresponds to increases in CYP1A1 mRNA. To examine whether omeprazole-initiated transcriptional activation of the CYP1A1 gene correlates with nuclear accumulation of the AhR, binding of nuclear proteins to the DRE was examined. When gel mobility shift assays were performed using nuclear extracts isolated from 101L cells treated with omeprazole or 2,3,7,8-tetrachlorodibenzo-p-dioxin, specific binding of the AhR to the DRE was observed. These studies demonstrate that omeprazole initiates AhR activation and that induction of the human CYP1A1 gene by omeprazole is AhR dependent.
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PMID:Nuclear uptake of the Ah (dioxin) receptor in response to omeprazole: transcriptional activation of the human CYP1A1 gene. 838 5

Immunoprecipitation experiments performed on cytosolic extracts of the mouse hepatoma cell line Hepa-1c1c7 (Hepa-1) confirm that the 9-S, unliganded, cytosolic aryl hydrocarbon (Ah) receptor complex contains the 90-kDa heat shock protein and the Ah receptor protein but reveal that it does not contain the Ah receptor nuclear translocator (ARNT) protein. These experiments confirm that the 6-S liganded form of the receptor identified in nuclear extracts of cells treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) contains the Ah receptor protein and ARNT but not the 90-kDa heat shock protein. The 6-S liganded Ah receptor complex activates transcription of the CYP1A1 gene via its binding to upstream xenobiotic-responsive elements (XREs). Treatment of cytosolic extracts of Hepa-1 cells with TCDD in vitro transforms the Ah receptor complex to the XRE-binding state. No such transformation occurs in a C- mutant deficient in ARNT activity. When in vitro synthesized ARNT was added concomitantly with TCDD to C- cytosolic extracts, it associated with the Ah receptor and restored Ah receptor-dependent XRE-binding activity to the extracts. Covalent cross-linking experiments in nuclear extracts of Hepa-1 and human LS180 cells treated with TCDD in vivo demonstrate that both ARNT and the Ah receptor bind directly to the XRE core sequence.
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PMID:Role of the aryl hydrocarbon receptor nuclear translocator protein in aryl hydrocarbon (dioxin) receptor action. 839 13

Rat CYP1A1 promoter activity was suppressed by the presence of a cis negative regulatory element (NRE) at position -843 to -746 in transiently transfected rat H4IIE and human HepG2 hepatoma cells. Removal of the NRE from the promoter-fusion gene constructs caused an increase in the basal promoter activity of 2-6-fold. Co-transfection of the NRE-containing or non-NRE-containing CYP1A1 promoter-fusion gene constructs with a cloned rat NRE, i.e., pNRE, into HepG2 cells caused a 2-fold or greater reduction in constitutive and induced promoter activities. 2,3,7,8-Tetrachlorodibenzo-p-dioxin-induced expression of the endogenous human CYPA1 was also inhibited by transfection of pNRE into HepG2 cells. Deletion of the sequence from base pairs (bp) -658 to -269 in the NRE-containing construct caused a dramatic decrease of constitutive expression in transiently transfected HepG2 cells, compared with an identical construct that lacked the NRE. Deletion of the sequences between bp -658 and -158 in the CYP1A1 promoter did not affect reporter gene activity, indicating a second site of interaction. At least three different rat liver nuclear proteins bound to the rat NRE, as determined by gel mobility shift and DNase I footprinting assays. A 32-bp sequence within the rat NRE, with significant sequence identity to the 26-bp c-myc, fos/jun-octamer-binding, NRE, was protected from DNAse I cleavage by rat liver nuclear extracts. These data suggested a role for this region in the negative regulation of rat CYP1A1.
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PMID:Rat CYP1A1 negative regulatory element: biological activity and interaction with a protein from liver and hepatoma cells. 839 16

The cytochrome P4501 gene family consists of two members, CYP1A1 and CYP1A2, that are induced by halogenated hydrocarbons and polycyclic aromatic hydrocarbons. The human CYP1 promoters and 5'-flanking sequences were cloned into luciferase expression vectors to develop cell lines that stably express luciferase activity in response to CYP1 gene induction. Plasmids were initially tested in transient transfection assays. Transient transfections resulted in high-level expression of luciferase by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) from both the CYP1 expression vectors, pLUC1A1 and pLUC1A2. In dose-response experiments, 10 nM TCDD caused a maximal induction of pLUC1A2-directed luciferase activity that was 10-fold over control. Maximal pLUC1A1-directed luciferase activity was 65-fold over control in cells treated with 10 nM TCDD. Stable integration of CYP1-luciferase-neo plasmids, pL1A1N and pL1A2N, into the human hepatoma cell line, HepG2, was achieved by selection with G418. G418-resistant colonies were isolated for both pL1A1N and pL1A2N plasmids. The pL1A2N transfectants showed basal-level luciferase activity, but were nonresponsive to treatment with 10 nM TCDD. These results are in contrast to the observed induction of pLUC1A2-mediated luciferase expression in transient transfection experiments. Stable integration of the human CYP1A2 gene sequences appears to silence the transcriptional activation by TCDD. The pL1A1N transfectants showed inducible luciferase activity and one cell line, referred to as 101L, was used to establish dose-response relationships for TCDD and various polycyclic aromatic hydrocarbons. Maximal induction occurred after treatment with 100 nM TCDD, 10 microM 3-methylcholanthrene, 50 microM benz[a]anthracene, and 50 microM benzo[a]pyrene. These studies illustrate the use of the CYP1A1-luciferase cell line for the study of structure-activity relationships.
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PMID:Response of human CYP1-luciferase plasmids to 2,3,7,8-tetrachlorodibenzo-p-dioxin and polycyclic aromatic hydrocarbons. 844 4

Polychlorinated biphenyls (PCBs) are industrial chemicals which have been detected in fish, birds and humans. They are known to exert marked effects on the liver. They induce hepatocellular carcinoma in rats and birds, and are suspected of being carcinogenic to humans. To better understand the genotoxic effects of PCBs, we used 32P-postlabelling to investigate DNA adduct formation, after exposure to PCBs (Aroclor 1254 and 3,3',4,4'-tetrachlorobiphenyl), in primary cultures of fetal hepatocytes from two animal species and in a human cell line (Hep G2). We also studied the induction of 7-ethoxyresorufin-O-deethylase (EROD) in these PCB-treated cells. The three cell types used are known to express different cytochrome P450 families. The aim was to see whether a correlation could be established between EROD activity (a CYP1A1-related activity) and DNA adduct formation. DNA adducts were found in all three models after exposure to 50 microM 3,3',4,4'-tetrachlorobiphenyl. The number of adducts was higher in quail hepatocytes (37 adducts per 10(9) nucleotides) than in rat hepatocytes or Hep G2 cells (20 adducts per 10(9) nucleotides in both cases). The major adduct was the same in all three cell types, but some adducts were found in only one or two species. These inter-species differences probably reflect metabolic differences leading to different ultimate carcinogens. Exposure to Aroclor 1254 failed to produce significant levels of DNA adducts, suggesting that pre-treated cells are required to magnify Aroclor 1254 metabolism. No correlation was found between adduct formation and the level of EROD induction.
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PMID:DNA adducts and P450 induction in human, rat and avian liver cells after exposure to polychlorobiphenyls. 855 39

Exposure of iron-loaded C57BL/10ScSn mice to the polychlorinated biphenyls (PCBs) mixture Aroclor 1254 in the diet (0.01%) for 5 weeks caused massive hepatic porphyria far greater than occurred with PCBs alone. This regime eventually causes hepatocellular carcinoma. Hepatic microsomal ethoxy-, pentoxy-, and benzyloxyresorufin dealkylase activities (respectively EROD, PROD, and BROD) catalyzed primarily by cytochrome P4501A1 and 2B isoenzymes were markedly induced after 2 weeks of diet (when no porphyria had developed) but showed little effect of iron. EROD activity in the nuclear membrane was also induced by the PCBs as was CYP1A1 protein when shown by immunoblotting. Nuclear dealkylase activities of PCBs-treated mice were considerably less than microsomal activities but were stimulated by iron pretreatment. The mechanism of the iron-enhanced toxicity may be due to oxidative damage associated with chronic induction of CYP1A1 isoforms. Lucigenin-enhanced chemiluminescence (CL) by microsomes and nuclear membranes was used as a method to estimate their potential to form reactive oxygen species. Despite CL being induced by PCBs it was less with microsomes from iron-treated mice. In a comparison of a variety of inducers of microsomal cytochrome P450 there was no correlation between inducer, uroporphyrogenic agent, and intensity of CL. On the other hand, cytosolic glutathione S-transferase (GST) activities with 1-chloro-2,4-dinitrobenzene and 1,2-dichloro-4-nitrobenzene (DCNB) as substrates, were also induced by the PCBs mixture, the induction with DCNB being synergistically potentiated by iron pretreatment. Complementary results were observed by immunocytochemistry using anti alpha-GST antibody. In contrast, total glutathione peroxidase activity and selenium-dependent glutathione peroxidase activity were depressed by PCBs but particularly in mice also administered iron. The results illustrate that PCBs not only induce CYP1A1 in microsomes but also in the nuclear membrane, which may be of significance in the mechanism of the iron-enhanced carcinogenicity of these chemicals. The iron-enhanced induction of GST with accompanying depletion of glutathione peroxidase provides evidence for oxidative processes induced in vivo by the PCBs.
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PMID:Modulation by iron of hepatic microsomal and nuclear cytochrome P450, and cytosolic glutathione S-transferase and peroxidase in C57BL/10ScSn mice induced with polychlorinated biphenyls (Aroclor 1254). 856 Apr 83

We have studied the response of genes in the dioxin-inducible [Ah] battery to three compounds that protect mouse hepatoma cells (Hepa-1c7c7 wild-type, wt) against menadione toxicity. Pretreatment of wt cells with 25 microM 5,10-dihydroindenol[1,2-b]indole (DHII), 25 microM tert-butylhydroquinone (tBHO) or 10 microM menadione itself, generated substantial protection against toxicity produced by subsequent menadione exposure. The gene response was examined in wt cells, and three mutant lines: CYP1A1 metabolism-deficient (c37 or P1-); nuclear translocation-impaired (c4 or nt-); and AHR-deficient (c2 or r-, containing < 10% of normal functional receptor levels). DHII treatment of wt cells for 12 hr markedly elevated the enzyme activities and mRNA levels of genes in the [Ah] battery: aryl hydrocarbon hydroxylase (Cyp1a1), NAD(P)H:menadione oxidoreductase (Nmol), cytosolic aldehyde dehydrogenase class 3 (Ahd4), and UDP-glucuronosyltransferase form 1*06 (Ugt1*06). Treatment of the c4 and c2 cells with DHII failed to induce mRNA levels of the genes, indicating that induction of the [Ah] gene battery by DHII is aromatic hydrocarbon receptor (AHR)-mediated. On the other hand, neither tBHO nor menadione caused increases in CYPlAl mRNA, but tBHQ significantly enhanced the NMO1, AHD4, and UGT1*06 mRNA levels in all three mutant cell lines. In conclusion, we expect one or more putative electrophile response elements (EpRE), previously found in the regulatory regions of the murine Nmol, Ahd4, and ugt1*06 genes, to be functional in responding to phenolic antioxidants.
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PMID:Response of [Ah] battery genes to compounds that protect against menadione toxicity. 861 69


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