• Environmental Mutagens and Carcinogens
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• v.23 no.1
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• pp.35-40
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• 2003

Polycyclic aromatic hydrocarbons (PAH) are frequently detected in food, water, soil, and sediment and are widespread environmental pollutants formed by the incomplete combustion of fossil fuels, woods and other organic matter. PAHs are considered to be probable human carcinogens. The mechanism of action of PAHs has been studied extensively, however it is not clear how PAHs turn on CYP1A1 in human breast cancer. Our laboratory have been studied the effect of PAHs in the human breast cancer cell MCF7. In this study, we examined the ZR-75-1 human breast cancer cells as a new system to evaluate bioactivity of PAHs. ZR-75-1 human breast cancer cell line responses to estrogen and progesteron. We have been able to estbilish long term culture system of this cells then used for the study to observe the effect of PAHs. We demonstrate that PAHs induced the CYP1A1 promoter and 7-ethoxyresolufin O-deethylase (EROD) activity in a concentration-dependant manner. RT-PCR analysis indicated that PAHs significantly up-regulate the level of CYP1A1 mRNA. Some of PAHs showed stronger stimulatory effect on CYP1 gene expression than TCDD. Apparently, ZR-75-1 cells have Aryl hydrocarbon receptors, therefore it would be good experimental tool to study the cross-talk between PAHs and steroid actions.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1995.04a
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• pp.128-128
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• 1995

To investigate the mechanism of the regulation of cytochrome P450IA1 gene expression, ethoxyresorufin deethylase(EROD) and benzo(a)pyrene hydroxylase in B6 mouse liver, in isolated perfused rat liver system. and in B6 mouse hepatocyte Hepa-I cells were examined. In C57BL/6N mouse, 3-methylcholan- throne( 3MC ) treatment have resulted in the stimulation of EROD activity based on fluorometry by 2.79 fold comparirng with that of control. Measurement of mRNA of cytochrome P450 was carried out by either nothern blot or dot blot analysis. Findings are similar to that of studies with enzymes. Furhtermore, when RTPCR method was applied to detect mRNA in Hepa I cell and liver tissues the results were more clear. Cytochrome P450IA1 upstream DNA containing CAT construct was transfected into Hepa-1 cells. After transfection of CAT construct, 3MC and flavonoids, such as, chrysin, hesperetin, kaempferol, morin, myricetin and aminoyrine were treated. 48 Hours after treatments, cells were harvested and assayed for CAT mRNA by RTPCR. 3MC treatment to hepa I cells transfected with trout P450IA1-CAT construct increased CAT mRNA by 2.81 fold when it was compared with that of control. This increase CAT mRNA was decreased by concomitantly treated flavonoids and aminopyrine. The level of CAT protein was 29.2-58.0% of 3MC stimulated CAT protein. Results of this study suggested that RTPCR seems to be a very good method to study regulation of gene expression in liver tissue or Hepa cells.

• BMB Reports
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• v.39 no.4
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• pp.448-451
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• 2006

Chemopreventive and cytotoxic effect of genistein against human breast cancer cell lines was investigated. Genistein inhibited cell proliferation in estrogen receptor-positive (MCF-7) and estrogen receptor-negative (MDA-MB-231) human breast carcinoma cell lines. Cytochrome P450 (CYP) 1A1-mediated ethoxyresorufin O-deethylase (EROD) activity was inhibited by genistein in a concentrationdependent manner. Genistein significantly inhibited 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced cyclooxy-genase-2 activity and protein expression at the concentrations of 10 (p < 0.05), 25 (p < 0.05) and 50 mM (p < 0.01). In addition, ornithine decarboxylase (ODC) activity was reduced to 53.8 % of the control after 6 h treatment with 50 mM genistein in MCF-7 breast cancer cells. These results suggest that genistein could be of therapeutic value in preventing human breast cancer.

• Korean Journal of Environmental Biology
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• v.26 no.2
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• pp.87-93
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• 2008

To assess effects of contaminants on fish in Nakdong river, feral yellowfin goby Acanthogobius flavimanus were caugt in two different sites and its hepatic monooxygenase enzyme, including cytochrome P450 (CYP), NADPH-cytochrome P450 reductase (P450R), NADH-cytochrome b5 reductase (b5R), ethoxyresorufin deethylase (EROD), glutathione S-transferase (GST) were quantitatively determined. Gonadosomatic index (GSI), hepatosomatic index (HSI) and three sex steroid hormone (17$\beta$-estradiol, E2; testosterone, T; 11-ketotestosterone, 11-KT) levels of the fish were also investigated. HSI of fish from polluted site (site 1) were significantly higher than that of unpolluted site (site 2), but GSI levels were significantly lower in polluted site. No significant differences in plasma 11-KT and T levels were observed in two sites surveyed. E2 level was, however, significantly (p<0.05) higher in female fish from site 1 than site 2. In addition, hepatic EROD activity and CYP level of site 1 fish were lower than those of site 2 fish, whereas relatively high levels of P450R, b5R and GST activities were found in site 1. The results imply that yellowfin goby, especially female fish in Nakdong river estuary are affected from contaminants disrupting sex steroid hormone system.

• Korean Journal of Environmental Biology
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• v.26 no.2
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• pp.94-101
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• 2008

The aim of this study was to assess the responses of mixed function oxygenase (MFO) and antioxidative systems of feral Javelin goby, Acanthogobius hasta, caught in two sites of different pollution level in Shihwa lake, which has been a highly polluted lake by organic pollutants from nearby industrial complexes and sites. Enzymes analyzed in phase I of MFO system are cytochrome P450 (CYP), NADPH-cytochrome P450 reductase (P450R), NADH-cytochrome b5 reductase (b5R), and ethoxyresorufin deethylase (EROD). Phase II enzyme of glutathione S-transferase (GST) in MFO system was also investigated. Moreover, oxidative-enzyme system including catalase (CAT), glutathione reductase (GR) and total-glutathione peroxidase (GPX) activities and glutathione concentration in both of oxidized (GSSG) and reduced form (GSH) were determined. P450R, b5R, and GST activities of fish are relatively high in the polluted area, whereas hepatic EROD activity levels of fish in polluted area were lower than those of unpolluted area. CYP concentrations are not different between areas. These results indicated that feral Acanthogobius hasta were adaptive to highly polluted environment and exposed to oxidative stress in Shihwa lake.

• Toxicological Research
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• v.23 no.2
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• pp.135-142
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• 2007

Formononetin is an isoflavonoid phytoestrogen found in certain foodstuffs such as soy and red clover. In this study, we examined the action of formononetin with the carcinogen activation pathway mediated through the aryl hydrocarbon receptor (AhR) in MCF-7 breast carcinoma cells. Treating the cells with formononetin alone caused the accumulation of CYP1A1 mRNA as well as elevation in CYP1A1-specific 7-ethoxyresorufin O-deethylase (EROD) activity in a dose dependent manner. However, a concomitant treatment with 7,12-dimethylbenz[a]anthracene (DMBA) and formononetin markedly reduced both the DMBA-inducible EROD activity and CYP1A1 mRNA level. Under the same conditions, formononetin inhibited the DMBA-induced AhR transactivation, as shown by reporter gene analysis using a xenobiotic responsive element (XRE). Additionally, formononetin inhibited both DMBA-inducible nuclear localization of the aryl hydrocarbon receptor (AhR) and metabolic activation of DMBA, as measured by the formation of the DMBA-DNA adducts. Furthermore, formononetin competed with the prototypical AhR ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), for binding to the AhR in an isolated rat cytosol. These results suggest that formononetin might be considered as a natural ligand to bind on AhR and consequently produces a potent protective effect against DMBA-induced genotoxicity. Therefore, that's the potential to act as a chemopreventive agent that is related to its effect on AhR pathway as antagonist/agonist.

• Archives of Pharmacal Research
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• v.29 no.7
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• pp.570-576
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• 2006

Phytoestrogen biochanin A is an isoflavone derivative isolated from red clover Trifolium pratense with anticarcinogenic properties. This study examined the action of biochanin A with the carcinogen activation pathway that is mediated by the aryl hydrocarbon receptor (AhR) in MCF-7 breast carcinoma cells. Treating the cells with biochanin A alone caused the accumulation of CYP1A1 mRNA and an increase in CYP1A1-specific 7-ethoxyresorufin O-deethylase (EROD) activity in a dose dependent manner. A concomitant treatment with 7,12-dimethylbenz[a]anthracene (DMBA) and biochanin A markedly reduced the DMBA-inducible EROD activity and CYP1A1 mRNA level. In addition, the biochanin A treatment alone activated the DNA-binding capacity of the AhR for the dioxin-response element (DRE) of CYP1A1, as measured by the electrophoretic-mobility shift assay (EMSA). EMSA revealed that biochanin A reduced the level of the DMBA-inducible AhR-DRE binding complex. Furthermore, biochanin A competed with the prototypical AhR ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), for binding to the AhR in an isolated rat cytosol. The biochanin A competitively inhibited the metabolic activation of DMBA, as measured by the formation of the DMBA-DNA adducts. These results suggest that biochanin A may thus be a natural ligand to bind on AhR. Therefore, biochanin A may be due to act an antagonist/agonist of the AhR pathway.

• Proceedings of the PSK Conference
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• 2003.10b
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• pp.112.2-112.2
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• 2003

CYP1 enzymes, CYP1A1, CYP1A2 and CYP1B1, are known to bioactivate procarcinogens particularly polyaromatic compounds. Flavonoids are a class of natural compounds that are present in edible plants. Structurally, these compounds are polyphenols with two aromatic rings (A, B) and a heterocycyclic ring (C). We observed the differential inhibition of 5,7-dihydroxyflavones which are different in numbers of B-ring-OH, to the activity of ethoxyresorufin O-deethylase (EROD) in human hepatic microsomes with the IC50 values, ie, 0.57 mM, 1.28 mM, and 3.62 mM, chrysin, apigenin, and Luteolin, respectively. (omitted)

• Proceedings of the Korean Society of Applied Pharmacology
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• 2001.11a
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• pp.114-114
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• 2001

Since hypoxia-inducible factor-lalpha (HIF-lalpha) and the arylhydrocarbon receptor (AhR) shared the AhR nuclear translocator (Arnt) for hypoxia- and AhR-mediated signaling, respectively, it was possible to establish the hypothesis that hypoxia could regulate Cyplal expression. In order to understand the mechanism of Cyplal gene expression, we demonstrated here that hypoxic agents such as cobalt chloride, desferrioxarnine, and picolinic acid reduced the TCDD induced Cyplal promoter activity based on the determination of luciferase activity in Hepa I cells transfected with pmCypla1-Luc. Also cobalt chloride inhibited the TCDD stimulated Cyplal mRNA level as well as EROD activities in both Hepa I and MCF-7 cells. Hypoxic agents such as cobalt chloride, picolinic acid, and desferrioxamine showed inhibition of luciferase activity that was induced by lnM TCDD treatment with dose dependent manner. Concomitant treatment of 150${\mu}$M ferrous sulfate with 1∼100${\mu}$M desferrioxarnine or 1∼100${\mu}$M picolinic acid recovered from the hypoxic agents-inhibited luciferase activity that was stimulated by TCDD. Reciprocally, the hypoxic agents down regulated TCDD induced Cyplal mRNA level and CYP1A1 enzyme activity in Hepa I cells.

• Korean Journal of Pharmacognosy
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• v.38 no.1
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• pp.62-66
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• 2007

Ethanol extract from Salvia miltiorrhiza (SME) was tested for breast cancer chemoprevention and metastasis by measuring the activites of cytochrome P45O 1A1, aromatase, ornithine decarboxylase (ODC), and matrix metalloproteinase (MMP)-9. SME significantly inhibited cytochrome P45O 1A1-mediated ethoxyresorufin O-deethylase (EROD) activity in a dose-dependent manner in a concentration range of 100${\sim}$l,200 ${\mu}g/ml$ (p<0.01). Microsomal aromatase (estrogen synthase) activity was suppressed 54.9%${\sim}$77.5% by the SME at concentration of 600${\sim}$l,200 ${\mu}g/ml$. ODC activity induced by 12-O-tet-radecanoylphorbol-13-acetate (TPA) was significantly reduced by the SME 900 and 1,200 ${\mu}g/ml$ (p<0.05) in MCF-7 breast cancer cells. In addition, SME (900 and 1,200 ${\mu}g/ml$) markedly inhibited MMP-9 activity, a key role in cancer metastasis. Therefore, SME is worth further investigation with respect to breast cancer chemoprevention or therapy.