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)

The genotoxic potency of the hepatocarcinogen N-nitrosodiethylamine (NDEA) was tested in various Reuber hepatoma cell lines, which differ in their expression of differentiated liver specific functions including hepatic cytochrome P450 forms, and in rat hepatocytes, with the aim of characterizing the enzymes involved in activation. DNA single-strand breaks assessed by alkaline elution served as an indicator of genetic damage. Aldrin epoxidase activity was used as a marker for various hepatic cytochrome P450 forms. The poorly differentiated cell lines RH35 and H4IIEC3/T were apparently not affected by NDEA; moderate effects were observed in the well-differentiated lines H4IIEC3/G- and 2sFou, and major effects in two other well-differentiated lines, Fao and C2Rev7, and in hepatocytes. The degree of DNA damage in the cell lines correlated positively with the expression of aldrin epoxidase. Furthermore, DNA damage induced by N-nitrosodimethylamine (NDMA) was determined in C2Rev7 cells and in rat hepatocytes in order to assess a possible involvement of the NDMA-metabolizing cytochrome P45oIIE1 in the activation of NDEA by comparing the genotoxic potencies of the two compounds. NDMA was distinctly less effective than NDEA in C2Rev7 cells at all concentrations tested. In hepatocytes, NDMA induced more DNA damage than NDEA at low concentrations, but was slightly less active at high concentrations. The results suggest that NDEA is preferentially metabolized to genotoxic products by one or several cytochrome P450 forms different from P450IIE1.
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PMID:Induction of DNA damage by N-nitrosodiethylamine in rat hepatoma cells: correlation with cytochrome P450-mediated aldrin epoxidase activity. 205 11

Transcripts of the murine CYP1A1 (cytochrome P1450) mRNA are markedly elevated in mutant hepatoma cell lines that contain missense mutations in the Cyp1a-1 structural gene. This putative derepression extends to other genes in the [Ah] battery. To test whether the Cyp1a-1 gene product is involved in a mechanism of feedback regulation of transcription, we introduced expression plasmids carrying the murine wild-type Cyp1a-1 cDNA into the mutant hepatoma cells. Measurements of steady-state mRNA levels and of transcriptional rates in the transfectants reveal that expression of a functional, exogenous CYP1A1 protein is sufficient to restore the repression of the endogenous gene, as well as restore the inducibility by dioxin, and that this effect takes place primarily at the level of transcription. Similar experiments with expression plasmids that carry the human CYP1A2 cDNA indicate that the CYP1A2 protein (cytochrome P3450) can also function as a transcriptional repressor. In addition, we find that expression of the Nmo-1 [NAD(P)H:menadione oxidoreductase] gene, a third member of the [Ah] gene battery, is also repressed by the exogenous expression of either Cyp1a-1 or CYP1A2 cDNA. These results indicate that the gene product of either member of the mammalian CYP1 family has a previously unrecognized transcriptional regulatory function, which is likely to be exerted by modification of preexisting trans-acting factors. This function may help bring about a fast reprogramming of gene expression, as might be needed during detoxification of toxic foreign chemicals.
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PMID:The murine Cyp1a-1 gene negatively regulates its own transcription and that of other members of the aromatic hydrocarbon-responsive [Ah] gene battery. 208 80

Dimethyl fumarate and dimethyl maleate are potent inducers of cytosolic NAD(P)H:(quinone acceptor) oxidoreductase (here designated quinone reductase) activity in Hepa 1c1c7 murine hepatoma cells in culture, whereas fumaric and maleic acids are much less potent, in agreement with the much greater reactivity of the esters as Michael reaction acceptors (P. Talalay, M. J. De Long, and H. J. Prochaska, Proc. Natl. Acad. Sci. USA, 85:8261-8265, 1988). Dimethyl fumarate also induced quinone reductase in mutants of the Hepa 1c1c7 cell line that were either defective in the Ah receptor or in cytochrome P1-450 activity, thereby establishing that this compound is a monofunctional inducer (H. J. Prochaska and P. Talalay, Cancer Res., 48: 4776-4782, 1988). Addition of dimethyl fumarate to the diet of female CD-1 mice and female Sprague-Dawley rats at 0.2-0.5% concentrations elevated cytosolic glutathione transferases and quinone reductase activities in a variety of organs, whereas much higher concentrations of fumaric acid were only marginally active. The widespread induction of such detoxication enzymes by dimethyl fumarate suggests the potential value of this compound as a protective agent against chemical carcinogenesis and other forms of electrophile toxicity. This proposal is supported by the finding that the concentrations of dimethyl fumarate required to obtain substantial enzyme inductions were well tolerated by rodents. Furthermore, the parent fumaric acid has low chronic toxicity and is a naturally occurring metabolic intermediate that is already in the food chain as an additive, and fumarate salts and esters are used for therapeutic purposes in man.
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PMID:Induction of glutathione transferases and NAD(P)H:quinone reductase by fumaric acid derivatives in rodent cells and tissues. 212 43

Most chemical carcinogens require activation to reactive electrophilic forms by Phase 1 enzymes (cytochromes P-450) in order to exert their toxic and neoplastic effects. The resultant electrophiles are susceptible to metabolic conjugation and other types of detoxications by Phase 2 enzymes (glutathione transferases, NAD(P)H: quinone reductase, glucuronosyltransferases). The balance between Phase 1 and Phase 2 enzymes is an important determinant of whether exposure to carcinogens will result in toxicity and neoplasia. Measurements of the activity of quinone reductase (QR) provide an efficient method for studying the potency and mechanism of Phase 2 enzyme induction. QR can be measured easily in murine hepatoma cells (Hepa lclc7) grown in microtiter plate wells, and the inductive response of these cells closely parallels the behavior of rodent tissues in vivo. Some inducers (such as large planar aromatics) are bifunctional; they induce both Phase 1 and Phase 2 enzymes and require binding to the Ah receptor and enhanced transcription of the cytochrome P1-450 system. Other inducers (e.g., phenolic antioxidants, 1, 2-dithiole-3-thiones, coumarins, thiocarbamates) are monofunctional and are independent of Ah receptor function. Monofunctional enzyme induction protects against carcinogens. The induction of Phase 2 enzymes by monofunctional inducers depends on the presence, or acquisition by metabolism, of electrophilic centers, and many of these inducers are Michael reaction acceptors. Our search for chemoprotective enzyme inducers for potential use as chemoprotectors in man is currently focused on fumarate derivatives, as well as on the identification of other monofunctional inducers in extracts of vegetables.
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PMID:Regulation of enzymes that detoxify the electrophilic forms of chemical carcinogens. 213 77

The cytochrome P450 in the transformable C3H/10T1/2 (10T1/2) cell line has been characterized and compared to the major polycyclic aromatic hydrocarbon (PAH)-inducible hepatic form, cytochrome P450IA1 (P450IA1). The mouse hepatoma cell line, Hepa-1, was used as an in vitro model for P450IA1 expression and regulation by PAH. Microsomes from uninduced and benz[a]anthracene (BA)-induced 10T1/2 cells provided PAH mono-oxygenated product profiles that were totally different from metabolite profiles produced by microsomes from uninduced and BA-induced Hepa-1 cells even though total activities were similar. The proximate carcinogen, 7,12-dimethylbenz[a]anthracene-3,4-diol (DMBA-3,4-diol) was a major product for the 10T1/2 microsomes, while Hepa-1 formed less than 2% of this metabolite. Hepa-1 converted benzo[a]pyrene (BP) to BP-4,5-diol and DMBA to 7-hydroxymethyl-12-methyl-BA, while 10T1/2 did not produce either product. Polyclonal antibody to rat hepatic P450IA1 did not inhibit metabolism of either PAH substrate by 10T1/2 microsomes, but totally inhibited such metabolism by Hepa-1 microsomes. Western immunoblot analysis of BA-induced 10T1/2 microsomes showed that less than 1% of total P450 was P450IA1. The PAH-metabolizing activity of 10T1/2 microsomes was highly inducible (14-fold) by pre-treatment of non-confluent intact cells with BA, but was only half as inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin. In contrast, the P450IA1 activity of Hepa-1 cells was highly inducible by both compounds. The distinct metabolite profiles, antibody inhibition data and lack of immunoreactivity all indicate that PAH metabolism in 10T1/2 cells is catalyzed by a form of P450 distinct from P450IA1. The anomalous induction patterns suggest that this novel isozyme is predominantly regulated by a mechanism other than the Ah receptor.
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PMID:Characterization of a novel cytochrome P450 from the transformable cell line, C3H/10T1/2. 215 39

The Ah receptor, a soluble cytoplasmic receptor that regulates induction of cytochrome P450IA1 and mediates toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), was detected and characterized in the continuous human liver cell line Hep G2. The mean concentration of specific binding sites for TCDD was 112 +/- 26 (SEM) fmol/mg cytosol protein as determined in eight separate cytosol preparations in the presence of sodium molybdate. This is equivalent to 14,000 binding sites per cell, approximately 40% of the sites per cell found in the mouse hepatoma line Hepa-1. The cytosolic Ah receptor from Hep G2 cells sedimented at 9 S and was specific for those halogenated and nonhalogenated aromatic compounds known to be agonists for the Ah receptor in rodent tissues and cells. Specific binding in the 9 S region was detected with both [3H]TCDD and 3-[3H]methylcholanthrene. 3-[3H]Methylcholanthrene did not bind to any component besides that at approximately 9 S. Phenobarbital, dexamethasone, and estradiol did not compete with [3H]TCDD for binding to the Hep G2 Ah receptor. Specific binding of [3H]triamcinolone acetonide to glucocorticoid receptor could also be demonstrated in Hep G2 cytosol. The apparent equilibrium dissociation constant (Kd) for binding of [3H]TCDD to Hep G2 Ah receptor was 9 nM by Woolf plot analysis, about an order of magnitude weaker than the affinity of [3H]TCDD for the mouse Hepa-1 Ah receptor or for the C57BL/6 murine hepatic Ah receptor. [3H]TCDD.Ah receptor complex, which was extracted from nuclei of Hep G2 cells incubated with [3H]TCDD at 37 degrees C in culture, sedimented at approximately 6 S under conditions of high ionic strength. Aryl hydrocarbon hydroxylase (AHH) activity was significantly induced after 24 h of incubation with polycyclic aromatic hydrocarbons: the EC50 for AHH induction was 5.3 microM for benz(a)anthracene and 1.3 microM for 3-methylcholanthrene. Modification of the preparative technique for cell cytosol, especially inclusion of 20 mM sodium molybdate in homogenizing and other buffers, was necessary to detect cytosolic Hep G2 Ah receptor. Hep G2 cells appear to conserve drug-metabolizing activity associated with cytochrome P450IA1 as well as the receptor mechanism which regulates its induction.
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PMID:Characterization of the Ah receptor mediating aryl hydrocarbon hydroxylase induction in the human liver cell line Hep G2. 215 49

Treatment of rat hepatoma H-4-II E cells with alpha-naphthoflavone (alpha NF) (10(-8), 10(-7), 10(-6)M) resulted in only minimum induction of ethoxyresorufin O-deethylase (EROD) activity and cytochrome P4501A1 mRNA levels only at 10(-6)M. In contrast, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) caused maximum or near maximum induction responses at 10(-8) and 10(-9)M. In a time-course study with TCDD (10(-9)M), and TCDD plus alpha NF (cotreated), alpha NF significantly inhibited the induction of EROD activity and cytochrome P4501A1 mRNA levels by TCDD for 6-24 h after initial exposure of the cells to the chemicals. In addition, treatment of the cells with 10(-9)M TCDD in the presence or absence of 10(-8), 10(-7), and 10(-9)M alpha NF showed that the latter compound inhibited the induction effects by TCDD in a concentration-dependent manner and these inhibitory effects could be overcome, in part, by a higher concentration of TCDD (10(-8)M). Treatment of the rat hepatoma H-4-II E cells with [3H]TCDD showed that within 60 min, there was an initial rapid increase in nuclear [3H]TCDD receptor complex levels (38 fmol/mg protein) which decreased to less than 10 fmol/mg protein within 4 h and remained relatively constant for up to 24 h. However, in cells treated with [3H]TCDD (10(-9)M) plus alpha NF (10(-6)M) the levels of the nuclear [3H]TCDD receptor complex were less than 5 fmol/mg protein throughout the 24-h time course. These data, coupled with the results which indicate that the alpha NF competitively inhibits the binding of [3H]-TCDD to the cytosolic aryl hydrocarbon (Ah) receptor, suggest that alpha NF inhibits the TCDD-mediated induction of CYP1A1 gene transcription and translation by direct competition for cytosolic Ah receptor binding sites.
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PMID:The mechanism of action of alpha-naphthoflavone as an inhibitor of 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced CYP1A1 gene expression. 216 79

Glutathione S-transferase (GST) Ya subunit gene expression is induced in mammalian tissues by two types of chemical agents: (i) planar aromatic compounds (e.g., 3-methylcholanthrene, beta-naphthoflavone, and 2,3,7,8-tetrachlorodibenzo-p- dioxin) and (ii) electrophiles (e.g., trans-4-phenyl-3-buten-2-one and dimethyl fumarate) or compounds easily oxidized to electrophiles (e.g., tert-butylhydroquinone). To study the mechanism of this induction, we have introduced deletions in the 5' flanking region of a mouse GST Ya subunit gene, fused it to the coding sequence for chloramphenicol acetyltransferase (CAT) activity, and transfected the Ya-CAT genes for expression into hepatoma cells. We show that a single cis-regulatory element, between nucleotides -754 and -713 from the start of transcription, is responsible for the induction by both planar aromatic and electrophilic compounds. Using murine hepatoma cell mutants defective in either the Ah-encoded aryl hydrocarbon receptor (BPrc1 mutant) or in cytochrome P1-450 gene (c1 mutant), we show that induction by planar aromatic but not by electrophilic inducers requires a functional Ah receptor and cytochrome P1-450 activity. From this it is concluded that Ya gene activation by planar aromatic compounds involves metabolism of these inducers by the phase I xenobiotic-metabolizing cytochrome P1-450 system into electrophilic compounds, which is consistent with a recently proposed model [Prochaska, H. J. & Talalay, P. (1988) Cancer Res. 48, 4776-4782]. Therefore, the regulatory sequence of the Ya gene should be considered an electrophile-responsive element (EpRE) activated exclusively by inducers containing an electrophilic center. An EpRE-containing 41-bp oligonucleotide ligated at the -187 site of the Ya gene promoter confers upon it an increase in basal activity and xenobiotic inducibility. The basal activity augments with the number of EpRE copies. DNase I protection patterns show the protection of the EpRE domain by a nuclear factor(s) that becomes more abundant upon exposure of Hepa 1c1c7 cells to tert-butylhydroquinone.
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PMID:Xenobiotic-inducible expression of murine glutathione S-transferase Ya subunit gene is controlled by an electrophile-responsive element. 216 52

The synergistic effect of dexamethasone (DEX) and polycyclic aromatic hydrocarbons on the induction of cytochrome P450IA1 (P450IA1) was examined in H4IIEC3/T Reuber hepatoma cells. P450IA1 activity was determined by the hydroxylation of benzo[a]pyrene (AHH) and deethylation of 7-ethoxyresorufin (EROD). The amount of Ah receptor, i.e. the specific cytosolic binding protein of 3-methylcholanthrene or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in H4IIEC3/T cells was characterized and quantitated by high performance gel filtration. Benz[a]anthracene and TCDD induced AHH and EROD activities, respectively, about 20-fold within 4 h. The increase was about 100-fold when cells were pretreated with DEX. The glucocorticoid alone induced P450IA1 activities 3-4 fold. DEX elicited half maximum AHH induction at a concentration of 20 nM in the presence or absence of benz[a]anthracene. Maximal potentiation of AHH induction required treatment with DEX for at least 32 h prior to the exposure to benz[a]anthracene. Treatment of H4IIEC3/T cells with DEX for 20 h caused a 2-3-fold increase in the amount of Ah receptor. The results suggest that the synergistic effect of DEX and polycyclic aromatic hydrocarbons on P450IA1 induction involves a time-consuming process which may consist of the synthesis or modification of a factor, possibly the Ah receptor.
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PMID:Dexamethasone-mediated potentiation of P450IA1 induction in H4IIEC3/T hepatoma cells is dependent on a time-consuming process and associated with induction of the Ah receptor. 217 91

We have developed a homologous in vitro transcription system that requires (i) 2,3,7,8-tetrachlorodibenzo-p-dioxin (called TCDD or dioxin), (ii) the Ah receptor, and (iii) a dioxin-responsive enhancer for activity. Unfractionated nuclear extracts from mouse hepatoma cells contain an inhibitor and fail to direct transcription in vitro. However, following phosphocellulose chromatography and reconstitution, the fractionated nuclear extract directs accurate transcription in vitro, using as a template the promoter/enhancer region from the mouse cytochrome P1-450 gene (Cyp1a1) linked to a "G-free cassette" (which generates a transcript with no guanosine residues). Extracts from TCDD-treated cells exhibit higher activity than extracts from untreated cells when transcribing a template containing both the promoter and enhancer but not when transcribing a template containing the promoter alone. Extracts from Ah receptor-defective cells fail to direct in vitro transcription in a TCDD-inducible fashion. A regulatory element that contains two binding sites for the liganded Ah receptor plus a truncated Cyp1a1 promoter suffices to direct TCDD-inducible, Ah receptor-dependent transcription in vitro. The inducible, receptor-dependent, enhancer-dependent properties of this system make it appropriate for analyzing in vitro the mechanism of dioxin action and the function of the Ah receptor.
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PMID:Dioxin-inducible, Ah receptor-dependent transcription in vitro. 217 90


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