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)

In C57BL/6 mouse liver, both murine Cypla-1 (cytochrome P1(450] and Cypla-2 (P3(450] genes are inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin), and Cypla-2 is constitutively expressed at high levels. Although the Cypla-1 gene is constitutively expressed and TCDD-inducible in mouse hepatoma Hepa-1 cell cultures, Cypla-2 gene expression is absent in these cultures. We show here that the 5' flanking region of Cyp1a-2 from - 1843 to +52 (base pairs relative to the Transcription initiation site) linked to the chloramphenicol acetyltransferase (CAT) reporter gene in stable Hepa-1 transformants produces no basal or TCDD- or cycloheximide-inducible CAT activity. On the other hand, the Cyp1a-2 promoter from -63 to +52 driving the CAT gene is inducible by cycloheximide. A chimeric plasmid containing the Cyp1a-1 TCDD-responsive enhancer (-1646 to -245) ligated to a Cyp1a-2 promoter region (-129 to +52) supports TCDD-inducible CAT expression in Hepa-1 cells and in rat 7777 cells. These data suggest that, although sequences between - 1843 and +52 +52 are not sufficient for Cyp1a-2 gene expression, the murine Cyp1a-2 promoter is functional in cell cultures.
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PMID:Expression of the chloramphenicol acetyltransferase (CAT) reporter gene by the murine Cyp1a-2 (cytochrome P3(450)) promoter in hepatoma cell cultures. 217 15

Three nuclear factors, the Ah receptor, XF1, and XF2, bind sequence specifically to the Ah response elements or xenobiotic response elements (XREs) of the cytochrome P450IA1 (P450c) gene. The interactions of these factors with the Ah response element XRE1 were compared by three independent methods, methylation interference footprinting, orthophenanthroline-Cu+ footprinting, and mobility shift competition experiments, using a series of synthetic oligonucleotides with systematic alterations in the XRE core sequence. These studies established the following (i) all three factors interact sequence specifically with the core sequence of XRE1; (ii) the pattern of contacts made with this sequence by the Ah receptor are different from those made by XF1 and XF2; and (iii) although XF1 and XF2 can be distinguished by the mobility shift assay, the sequence specificities of their interactions with XRE1 are indistinguishable. Further characterization revealed the following additional differences among these three factors: (i) XF1 and XF2 could be extracted from nuclei under conditions quite different from those required for extraction of the Ah receptor; (ii) XF1 and XF2 were present in the nuclei of untreated cells and did not respond to polycyclic compounds, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and beta-napthoflavone, while nuclear Ah receptor was undetectable in untreated cells and rapidly increased in response to TCDD; (iii) inhibition of protein synthesis did not affect the TCDD-induced appearance of the Ah receptor but substantially decreased the constitutive activities of XF1 and XF2, suggesting that the Ah receptor must be present in untreated cells in an inactive form that can be rapidly activated by polycyclic compounds, while the constitutive expression of XF1 and XF2 depends on the continued synthesis of a relatively unstable protein; (iv) the receptor-deficient and nuclear translocation-defective mutants of the hepatoma cell line Hepa1, which are known to lack nuclear Ah receptor, expressed normal levels of XF1 and XF2, suggesting that the former factor is genetically distinct from the latter two; and (v) a divalent metal ion, probably Zn2+, is known to be an essential cofactor for the Ah receptor but was not required for the DNA-binding activities of XF1 and XF2. Together, these findings indicate that the Ah receptor is distinct from XF1 and XF2, while the latter two activities may be related. Because the DNA-binding domains of these three factors overlap substantially, their binding to XREs is probably mutually exclusive, which suggests that the interplay of these factors at Ah response elements may be important to the regulation of CYP1A1 gene transcription. The results of preliminary transfection experiments with constructs harboring XREs upstream of the chloramphenicol acetyltransferase gene driven by a minimal simian virus 40 promoter are presented that are consistent with this hypothesis.
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PMID:Multiple DNA-binding factors interact with overlapping specificities at the aryl hydrocarbon response element of the cytochrome P450IA1 gene. 217 7

The role of the Ah receptor in mediating the toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) was investigated in 5L rat hepatoma cells containing TCDD-inducible cytochrome P450IA1 activity and in variants lacking cytochrome P450IA1 and Ah receptor. TCDD inhibited growth of the wild-type 5L cells, but not of the Ah receptor deficient variants. The two strong Ah receptor ligands 3,3',4,4'-tetrachlorobiphenyl (3,3',4,4'-TCB) and benz[a]anthracene (BA) exerted toxic effects in 5L cells that resembled those of TCDD. The poor Ah receptor ligand 2,2',4,4'-tetrachlorobiphenyl was not toxic in 5L cells. The concentrations of TCDD, 3,3',4,4'-TCB or BA required for the toxic response were similar to those that elicited P450IA1 induction. The present results suggest strongly that interaction with the Ah receptor is a necessary link in the chain of events leading to toxic effects in 5L cells upon exposure to TCDD.
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PMID:Inhibition of growth by 2,3,7,8-tetrachlorodibenzo-p-dioxin in 5L rat hepatoma cells is associated with the presence of Ah receptor. 217 38

Using the mouse hepatoma Hepa-1c1c7 c37 mutant cell line that exhibits negligible benzo[a]pyrene hydroxylase (Cyp1a1) and acetanilide 4-hydroxylase (Cyp1a2) enzyme activities, we developed stable transfectants of plasmids containing the murine Cyp1a1 (cytochrome P(1)450) and the human CYP1A2 (P(3)450) cDNAs. We show that the assay measuring metabolism of ethoxyfluorescein ethyl ester (EFEE) was invaluable in screening large numbers of individual cell lines for high Cyp1a1 enzyme activity. Nine different plasmid constructs containing various combinations of promoter and enhancer sequences were compared, including: the Drosophila heat shock promoter, the mouse mammary tumor virus long terminal repeat (MMTV LTR) carrying the glucocorticoid-responsive element (GRE), enhancer sequences from simian virus 40 (SV40) and herpes simplex virus type 1 (HSV-1), and the aromatic hydrocarbon-responsive domain (AhRD) of the murine Cyp1a1 gene. Interestingly, only those constructs containing the AhRD produced high levels of Cyp1a1 enzyme activity. In contrast, high levels of CYP1A2 activity were obtained with plasmids carrying the HSV-1 enhancer, as well as the AhRD. These studies suggest that the AhRD, which responds to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), provides a post-transcriptional signal necessary for the induction of functional Cyp1a1 enzyme activity. Although untransfected c37 cells exhibit markedly elevated levels of endogenous Cyp1a1 mRNA, the expression of exogenous Cyp1a1 or CYP1A2 enzyme activity in these cells decreases the concentration of this endogenous Cyp1a1 mRNA to negligible levels and restores Cyp1a1 mRNA inducibility by TCDD; these data indicate that the functional product of either the Cyp1a1 gene or the CYP1A2 gene might have a role in an autoregulatory loop controlling the constitutive expression of the Cyp1a1 gene. The cell lines described herein should be valuable in assessing the contribution of these two P450 enzymes to the processes of cytotoxicity, mutagenesis, and carcinogenesis.
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PMID:Stable expression of mouse Cyp1a1 and human CYP1A2 cDNAs transfected into mouse hepatoma cells lacking detectable P450 enzyme activity. 220 99

In this study, the effects of benzo[a]pyrene (BP), 3-methylcholanthrene (3MC) and tetrachlorodibenzofuran (TCDBF) on the expression of the cytochrome P450IA1 gene in the rat hepatoma cell line H4IIE were examined. The initial rate of increase in the steady-state concentration of the mRNA for this gene was similar with each of these inducers; however, the elevated level of this mRNA was more sustained after TCDBF treatment. Nuclear run-off assays suggested that the elevated level of the mRNA was caused principally by an effect upon transcription.
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PMID:Induction of cytochrome P450IA1 in rat hepatoma cell by polycyclic hydrocarbons and a dioxin. 240 94

In previous studies we have shown that substances associated with particulates collected from urban air and automobile exhaust bind with high affinity to the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) receptor present in rat liver cytosol. In this study we used a rat hepatoma cell line, H4IIE, to investigate the effect of such substances on an enzyme system, aryl hydrocarbon hydroxylase (AHH), which is regulated via the TCDD receptor. The results demonstrate that AHH activity in the H4IIE cell line can be induced by extracts of particulates collected from urban air and automobile exhausts in a dose-dependent manner, and that the AHH activity is inducible by five polycyclic aromatic hydrocarbons (PAHs) including 1-/3-nitrobenzo[a]pyrene and 6-chlorochrysene, all present in extracts of particulates from urban air and automobile exhausts. The induction of AHH activity is correlated to apparent TCDD receptor affinity for investigated PAHs (r = 0.85) and particulate extracts. Biochemically, treatment of the cells with 5,6-benzoflavone significantly increased the level of cytochrome P-450c but not P-450d as shown by immunoblotting and analysis of mRNA levels. The data indicate that substances present in extracts of urban air particulates can interact with the TCDD receptor in intact cells and cause an accumulation of cytochrome P-450c mRNA leading to an increased synthesis of the gene product and thus an increase in enzyme activity.
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PMID:TCDD receptor ligands present in extracts of urban air particulate matter induce aryl hydrocarbon hydroxylase activity and cytochrome P-450c gene expression in rat hepatoma cells. 244 93

Upon binding of 2,3,7,8-tetrachlorodibenzo-p-dioxin (called dioxin or TCDD), the dioxin receptor exhibits increased affinity for the cell nucleus in vivo and for DNA in vitro. To define the recognition sequence of the dioxin receptor and its relationship with that of the glucocorticoid receptor, oligonucleotides derived from dioxin-responsive elements of the rat cytochrome P-450c gene were tested for their ability to form specific protein-DNA complexes in a gel retardation assay. We found that a previously defined sequence motif that is similar to the glucocorticoid-responsive element and exhibits strong enhancer activity in response to dioxin receptor ligands bound a dioxin-inducible factor with high specificity but was not recognized by the DNA-binding domain of the glucocorticoid receptor. Binding to this element was only observed in nuclear extracts of wild-type mouse hepatoma cells in a time- and dose-dependent manner and not in nuclear extracts from a nonresponsive mutant cell line deficient in DNA binding of the dioxin receptor. The specific DNA-binding activity in wild-type nuclear extracts comigrated in a Superose size-exclusion column and cosedimented on sucrose gradients with the in vivo labeled dioxin receptor. These experiments strongly suggest that the dioxin receptor is a sequence-specific DNA-binding protein and is not only biochemically but also functionally similar to the steroid receptor family.
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PMID:Specific protein-DNA interactions at a xenobiotic-responsive element: copurification of dioxin receptor and DNA-binding activity. 253 61

The mouse hepatoma cell line Hepa-1 was studied for aryl hydrocarbon hydroxylase (AHH) inducibility by sixteen compounds known to be inducers of cytochrome P450 of different "classes". Both 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and sodium phenobarbital induced AHH activity. A cytochrome P450IA1-specific (P1-450) mouse cDNA probe was used to quantitate mRNA induction. There was a good correlation between the amount of cytochrome P450IA1 mRNA induced and AHH activity. Immunoblots with monoclonal antibody 1-7-1, which recognizes rat liver P450IA1 and P450IA2 (P450c and P450d, respectively), showed that both phenobarbital and TCDD increase the amount of a P450 isozyme immunorelated to P450IA1 in this cell line. Hepa-1 mutants with no AHH inducibility (no functional P450IA1 structural gene; no Ah receptor; no nuclear translocation of the inducer-receptor complex; and presence of dominant repressor) did not respond to phenobarbital. The cytosolic receptor for TCDD (Ah receptor) was characterized to see if phenobarbital induced cytochrome P450IA1 mRNA and the hydroxylase enzyme through the same mechanism as TCDD. 20 mM Phenobarbital almost completely abolished the binding of 3H-TCDD to the cytosolic receptor. These data indicate that phenobarbital can be a weak ligand for the Ah receptor and thus induce cytochrome P450IA1 and AHH activity. The observation increases the list of different P450 forms inducible by phenobarbital.
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PMID:Induction of cytochrome P450IA1 in mouse hepatoma cells by several chemicals. Phenobarbital and TCDD induce the same form of cytochrome P450. 254 28

Two established human hepatoma cell lines, Hep3B and HepG2, were examined for aryl hydrocarbon (benzo[a]pyrene) hydroxylase (AHH) induction and for the presence of the murine-equivalent aromatic hydrocarbon (Ah) receptor. Both cell lines demonstrated polycyclic aromatic hydrocarbon (PAH)-induced AHH activity; however, assay conditions for induction were different than those established for the control mouse hepatoma cell line, Hepa c1-9. When cytosols from either cell line were exposed to tritiated 2,3,7,8-tetrachlorodibenzo-p-dioxin [( 3H]TCDD) and analyzed on sucrose gradients with or without prior charcoal treatment, two peaks were observed at positions corresponding to 4-5 S and 8-9 S. The 8-9 S peak was identified as the probable human Ah receptor equivalent since, like the mouse Ah receptor, this peak: (a) was eliminated only by cytochrome P1-450 inducers; (b) was sensitive to protease digestion; and (c) was thermolabile. Levels of TCDD specifically bound in the 8-9 S peak for HepG2 and Hep3B were 27 and 34 fmol/mg cytosolic protein respectively. The level of TCDD specifically bound was not affected by charcoal treatment or by the addition of sodium molybdate, which is known to stabilize ligand binding to steroid receptors. Incubation of Hep3B or HepG2 cells with [H]TCDD at 37 degrees for 1 hr effected a redistribution of binding from the cytosol 8-9 S peak to a nuclear 6 S peak. The nuclear peaks from both human cell lines demonstrated similar sedimentation properties, temperature-dependence and inducer-specificity, as for the mouse nuclear Ah receptor. Appearance of nuclear 6 S binding is consistent with a temperature-dependent translocation process, supporting the observation that these human hepatoma cell lines contain a binding component which is similar to the mouse Ah receptor in structure and function during AHH induction.
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PMID:Induction of aryl hydrocarbon hydroxylase and demonstration of a specific nuclear receptor for 2,3,7,8-tetrachlorodibenzo-p-dioxin in two human hepatoma cell lines. 254 64

The tissues of hepatocellular carcinoma were operatively resected from six patients. All four components of the systems of microsomal cytochrome P-450-linked monooxygenase of the tissues were investigated and compared to those of normal liver tissue. The concentrations of cytochromes P-450, P-420 and b5 were measured optically and the concentrations of all components except cytochrome P-450 were measured by the Western blotting method followed by immunochemical staining. In microsomes of hepatocellular carcinoma tissues, there was as much cytochrome P-450 and other redox components as in the normal liver tissues, but cytochrome P-450 in liver cancer tissues was unstable and easily converted to cytochrome P-420. The specific activities of NADPH- and NADH-ferricyanide and cytochrome c reductase of each sample were also measured. In the microsomes of the cancer tissues, the specific activities were remarkably reduced compared with those of normal liver tissues. The lipid compositions of the microsomes and the phospholipid/cholesterol ratios (w/w) were 13.1 +/- 3.13 in the cancer tissues and 43.0 +/- 6.74 in normal liver tissues. This difference of the lipid composition elucidates the instability of cytochrome P-450 molecules and the inefficiency of the electron transport of cytochrome P-450-linked monooxygenase systems.
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PMID:Microsomal cytochrome P-450-linked monooxygenase systems and lipid composition of human hepatocellular carcinoma. 254 14


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