RESUMEN
Recent findings have revealed that exposure to environmental contaminants may result in obesity and pose a health threat to the general public. As the activity of transient receptor potential channels (TRPs) plays a permissive role in adipogenesis, the interactions between TRPs and some food pollutants, i.e. bisphenol A, di (2-ethylhexyl) phthalate, zearalenone, and zeranol at 10 µM were investigated in the present study. TRP-V1,-V3, -C4 and -C6 are reported to be differentially expressed in the adipocyte differentiation, and immunoblotting was performed to quantify changes in these TRPs affected by the pollutants. Our result indicated that the mycoestrogen zeranol or α-zearalanol suppressed the expression of the V1 and C6 isoforms. Subsequently, confocal microscopy was used to measure the calcium inflow repressed by zeranol from 0.1 µM to 10 µM. Oil Red O staining was used to determine the differentiation of 3T3 L1 preadipocytes. Zeranol could suppress the expression of TRP-V1 and -C6 protein and inhibit the associated flow of calcium into the cytosol of 3T3 L1 cells. Its IC50 value for inhibiting calcium inflow stimulated by 40 µM capsaicin or 10 µM GSK1702934A was estimated to be around 6 µM. Reduced TRP-V1 or -C6 activity might result in promoting adipogenesis. In conclusion, this study demonstrated that zeranol could potentiate fat cell differentiation through antagonizing TRP-V1 and -C6 activities.
Asunto(s)
Estrógenos no Esteroides/toxicidad , Canales de Potencial de Receptor Transitorio/antagonistas & inhibidores , Zeranol/toxicidad , Células 3T3-L1 , Animales , Transporte Biológico/efectos de los fármacos , Calcio/metabolismo , Capsaicina/farmacología , Reducción Gradual de Medicamentos , Estradiol/farmacología , Estrógenos no Esteroides/administración & dosificación , Regulación de la Expresión Génica/efectos de los fármacos , Concentración 50 Inhibidora , Ratones , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Canales de Potencial de Receptor Transitorio/agonistas , Canales de Potencial de Receptor Transitorio/metabolismo , Zeranol/administración & dosificaciónRESUMEN
The environmental polycyclic aromatic hydrocarbons (PAH) and dioxins are carcinogens and their adverse effects have been largely attributed to the activation of AhR. Hesperetin is a flavonone found abundantly in citrus fruits and has been shown to be a biologically active agent. In the present study, the effect of hesperetin on the nuclear translocation of AhR and the downstream gene expression was investigated in MCF-7 cells. Confocal microscopy indicated that 7, 12-dimethylbenz[α]anthracene (DMBA) or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) -induced nuclear translocation of AhR was deterred by hesperetin treatment. The reduced nuclear translocation could also be observed in Western analysis. Reporter-gene assay further illustrated that the induced XRE transactivation was weakened by the treatment of hesperetin. Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) assay demonstrated that the gene expressions of CYP1A1, 1A2, and 1B1 followed the same pattern of AhR translocation. These results suggested that hesperetin counteracted AhR transactivation and suppressed the downstream gene expression.
Asunto(s)
Antineoplásicos Fitogénicos/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/antagonistas & inhibidores , Neoplasias de la Mama/metabolismo , Regulación hacia Abajo , Regulación Neoplásica de la Expresión Génica , Hesperidina/metabolismo , Proteínas de Neoplasias/antagonistas & inhibidores , Receptores de Hidrocarburo de Aril/antagonistas & inhibidores , 9,10-Dimetil-1,2-benzantraceno/antagonistas & inhibidores , 9,10-Dimetil-1,2-benzantraceno/toxicidad , Transporte Activo de Núcleo Celular/efectos de los fármacos , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neoplasias de la Mama/inducido químicamente , Neoplasias de la Mama/patología , Neoplasias de la Mama/prevención & control , Carcinógenos Ambientales/química , Carcinógenos Ambientales/toxicidad , Citocromo P-450 CYP1A1/antagonistas & inhibidores , Citocromo P-450 CYP1A1/química , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1A2/química , Citocromo P-450 CYP1A2/genética , Citocromo P-450 CYP1A2/metabolismo , Citocromo P-450 CYP1B1/antagonistas & inhibidores , Citocromo P-450 CYP1B1/química , Citocromo P-450 CYP1B1/genética , Citocromo P-450 CYP1B1/metabolismo , Suplementos Dietéticos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genes Reporteros/efectos de los fármacos , Humanos , Células MCF-7 , Microscopía Confocal , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Dibenzodioxinas Policloradas/antagonistas & inhibidores , Dibenzodioxinas Policloradas/química , Receptores de Hidrocarburo de Aril/metabolismoRESUMEN
Consumption of fruits and vegetables is generally regarded as beneficial to plasma lipid profile. The mechanism by which the plant foods induce desirable lipid changes remains unclear. SREBP-2 is crucial in cholesterol metabolism, and it is a major regulator of the cholesterol biosynthesis enzyme HMGCR. Our lab has previously illustrated that apigenin and luteolin could attenuate the nuclear translocation of SREBP-2 through an AMPK-dependent pathway. In the present study, these two flavones were studied for their ability to deter the same in an AMPK-independent signaling route. The processing of SREBP-2 protein was promoted by phorbol 12-myristate 13-acetate (PMA) in the hepatic cells WRL and HepG2, and the increased processing was reversed by apigenin or luteolin co-administration. EMSA results demonstrated that the PMA-induced DNA-binding activity was weakened by the flavones. The increased amount of nuclear SREBP-2 in cells was attenuated by the flavonoid as shown by immunocytochemical imaging. Quantitative reverse transcriptase-polymerase chain reaction assay demonstrated that the transcription of HMGCR under both flavone treatments was reduced. However, apigenin appeared to be stronger than luteolin in restraining PMA-induced HMGCR mRNA expression. Since PMA is a diacylglycerol analog, these findings might have some physiological implications.
Asunto(s)
Apigenina/farmacología , Suplementos Dietéticos , Hígado/metabolismo , Luteolina/farmacología , Proteína 2 de Unión a Elementos Reguladores de Esteroles/metabolismo , Acetato de Tetradecanoilforbol/toxicidad , Proteínas Quinasas Activadas por AMP/metabolismo , Células Hep G2 , Humanos , Hidroximetilglutaril-CoA Reductasas/metabolismoRESUMEN
Cyclooxygenase (COX) is the rate-limiting enzyme for the conversion of prostaglandins from arachidonic acid. Upregulation of COX-2 has been well documented during tumorigenesis and metastasis of breast cancer. Isoliquiritigenin (ILN), a flavonoid isolated from licorice (the rhizomes of GLYCYRRHIZA GLABRA, a member of the bean plant family), is known to be a potential suppressor of COX-2 expression. This study focuses on phorbol ester-induced COX-2 expression in the non-tumorigenic MCF-10A cells. Real-time PCR and Western blotting indicated that ILN at 5 microM or above significantly inhibited phorbol 12-myristate 13-acetate (PMA)-induced COX-2 expression in the breast cells. The activated PKC alpha appeared to be not affected, whereas its downstream mitogen-activated protein kinase (MAPK) ERK-1/2 was deactivated. ERK can activate the transcriptional factor binding of AP-1 or CRE, which can be located at the COX-2 promoter region (- 72/- 53). Electrophoretic mobility shift assays illustrated that ILN suppressed DNA binding at this region. The shifted bands could be competed off with consensus sequences of AP-1 and CRE, and the supershift assay demonstrated that CREB-1 instead of c-Jun was responsible for the binding. This study showed that ILN downregulated PMA-induced COX-2 expression by modulating ERK-1/2 signaling, a finding that may be relevant to the disease prevention properties of licorice.
Asunto(s)
Neoplasias de la Mama/patología , Chalconas/farmacología , Ciclooxigenasa 2/metabolismo , Glycyrrhiza/química , Acetato de Tetradecanoilforbol/farmacología , Secuencia de Bases , Western Blotting , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Cartilla de ADN , Dinoprostona/metabolismo , Ensayo de Cambio de Movilidad Electroforética , Femenino , Humanos , Reacción en Cadena de la PolimerasaRESUMEN
Genistein is a phytoestrogen isolated from soyabean, and is a potential nutraceutical gearing for women suffering from perimenopausal symptoms. Because of its differential binding affinity to estrogen receptor (ER) isoforms, genistein is described as a selective estrogen receptor modulator (SERM). The ligand-receptor interaction is established, but the potential confounding factors have not been fully addressed. Alteration in estrogen metabolism is an important issue when determining the downstream effect of ER. Aromatase or CYP19 catalyzes the rate-limiting reaction of estrogen synthesis, and is highly expressed in the ovary. This organ is the source of estrogen in females. After menopause the ovaries cease to produce the hormone, and localized estrogen synthesis in extragonadal tissues could become physiologically significant. In the present study, effect of genistein on CYP19 regulation was investigated in the hepatic cells HepG2. The phytoestrogen induced aromatase activity in the cells. Increased mRNA expression with concurrent elevation in the usage of promoters I.3/II was also demonstrated. Luciferase reporter gene assays verified the transcriptional control dictated by the specific promoters under genistein treatment. Several protein kinases were examined, and PKC?, P38, ERK-1/2 appeared to be activated. Subsequent inhibition and expression experiments demonstrated the involvement of these kinases. The transcriptional factor CREB was ultimately activated in the gene regulation. The present study illustrated an extragonadal pathway by which genistein might increase estrogen synthesis.
Asunto(s)
Estrógenos/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Genisteína/farmacología , Glycine max/química , Fitoestrógenos/farmacología , Aromatasa/metabolismo , Western Blotting , Línea Celular Tumoral , Humanos , Hígado/citología , Regiones Promotoras Genéticas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de SeñalRESUMEN
Licorice is the sweet-tasting rhizomes of a bean plant and is quite commonly used in Western countries for culinary purposes, while it is a medicinal herb in China. Many flavonoids have been isolated from licorice, and their pharmacological properties may be applicable in preventive medicine. Overexposure to estrogen has been implicated in the etiology of breast cancer, and cytochrome P450 (CYP) 19 enzyme, or aromatase, catalyzes the rate-limiting reaction. Phytocompounds that are able to inhibit this enzyme may potentially suppress breast cancer development. In the present study the licorice flavonoid isoliquiritigenin (ILN) was shown to be an aromatase inhibitor in recombinant protein and MCF-7 cells stably transfected with CYP19 (MCF-7aro). ILN displayed a K(i) value of around 3 muM, and it also blocked the MCF-7aro cell growth pertaining to the enzyme activity in vitro. Subsequently, the compound administered in diet was given to ovariectomized athymic mice transplanted with MCF-7aro cells. This mouse model is widely accepted for studying postmenopausal breast cancer. The phytochemical significantly deterred the xenograft growth without affecting the body weight. Subsequently, the flavonoid's effect on CYP19 transcriptional control in vitro was also investigated. At the mRNA level, ILN could also suppress the expression in wild-type MCF-7 cells. Reporter gene assay and real-time PCR verified that the transactivity of CYP19 driven by promoters I.3 and II was suppressed in these cells. Deactivation of C/EBP could be the underlying molecular mechanism. Our study demonstrated that ILN was an inhibitor of aromatase and a potential chemopreventive agent against breast cancer.
Asunto(s)
Inhibidores de la Aromatasa/administración & dosificación , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/prevención & control , Chalconas/administración & dosificación , Glycyrrhiza/química , Animales , Aromatasa/genética , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Humanos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Ratones , Ratones Desnudos , Ovariectomía , Regiones Promotoras Genéticas , ARN Mensajero/análisis , Testosterona/farmacologíaRESUMEN
Studies have shown that soya consumption has been associated with low incidence of CVD. Because the chemical structures of soya isoflavones are similar to oestrogen, the beneficial outcome may be attributed to the oestrogenicity of these compounds. In this study, effect of the soya isoflavone genistein on the mRNA expression of apoA-1 in the human hepatoma HepG2 cell was investigated. Without oestrogen receptor (ER) alpha transfection, soya isoflavones in the physiological range had no effect on the apoA-1 transcription. Once ERalpha was ectopically expressed in these cells, soya isoflavone dramatically increased the apoA-1 mRNA abundance quantified by real-time PCR. ApoA-1-reporter assays with plasmid constructed from the 5'-flanking segment upstream to the coding region revealed that the transactivation of the apoA-1 promoter was induced by the soya isoflavone in HepG2 cells expressing ERalpha. This induction was reduced by the anti-oestrogen ICI 182780, but not the inhibitors of protein kinase (PK) C, PKA, or mitogen-activated PK. Based on the previously identified response elements on the promoter, a series of truncated promoter reporter plasmids were then constructed. An induction profile of genistein was built and insulin response core element at -411 to -404 appeared to be a potential site of interaction. This study illustrated that soya isoflavones at physiological concentrations could up regulate apoA-1 mRNA expression in ERalpha-transfected HepG2 cells.
Asunto(s)
Apolipoproteína A-I/biosíntesis , Receptor alfa de Estrógeno/metabolismo , Genisteína/farmacología , Isoflavonas/farmacología , Fitoestrógenos/farmacología , Activación Transcripcional/efectos de los fármacos , Apolipoproteína A-I/genética , Relación Dosis-Respuesta a Droga , Estradiol/análogos & derivados , Estradiol/farmacología , Antagonistas de Estrógenos/farmacología , Receptor alfa de Estrógeno/fisiología , Fulvestrant , Humanos , Regiones Promotoras Genéticas , Inhibidores de Proteínas Quinasas/farmacología , ARN Mensajero/genética , Células Tumorales Cultivadas , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Biochanin A is an isoflavone isolated from red clover (Trifolium pratense), and is a commercially available nutraceutical for women suffering from postmenopausal symptoms. Isoflavones resemble the structure of oestrogen, and display agonistic and antagonistic interactions with the oestrogen receptor. Overexposure of oestrogen is a major contributing factor in the development of breast cancer, and cytochrome P450 (CYP) 19 enzyme, or aromatase, catalyses the reaction converting androgen to oestrogen. In the present study the effect of biochanin A on the gene regulation and enzyme activity of aromatase was investigated. By assaying MCF-7 cells stably transfected with CYP19, biochanin A inhibited aromatase activity and hampered cell growth attributing to the enzyme activity. In addition, 25 microm-biochanin A significantly reduced CYP19 mRNA abundance in the oestrogen receptor-negative breast cancer cells SK-BR-3. The transcriptional control of the CYP19 gene is exon-specific, and promoter regions I.3 and II have been shown to be responsible for CYP19 expression in SK-BR-3 cells. Luciferase reporter gene assays also revealed that biochanin A could repress the transcriptional control dictated by the promoter regulation. Interestingly, genistein did not inhibit aromatase but it might down regulate promoter I.3 and II transactivation. Since genistein is a major metabolite of biochanin A, it might contribute to biochanin A's suppressive effect on CYP19 expression. The present study illustrated that biochanin A inhibited CYP19 activity and gene expression.
Asunto(s)
Inhibidores de la Aromatasa/farmacología , Aromatasa/metabolismo , Neoplasias de la Mama/enzimología , Genisteína/farmacología , Fitoestrógenos/farmacología , Trifolium/química , Aromatasa/genética , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , División Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Humanos , Regiones Promotoras Genéticas/efectos de los fármacos , ARN Mensajero/genética , Activación Transcripcional/efectos de los fármacos , Células Tumorales CultivadasRESUMEN
Estrogen is crucial in preparing of pregnancy, and its role in the maintenance of pregnancy has yet to be elucidated. During the course of pregnancy, the placenta is responsible for the provision of estrogen. The hormone biosynthesis is catalyzed by cytochrome P450 (CYP) 19 or aromatase. In the present study, we screened several common dietary components and identified the grape polyphenol resveratrol to be a potential inhibitor in the hormone synthesis. In a recombinant protein system resveratrol inhibited the aromatase activity with an IC(50) value of approximately 40 microM. Subsequent analysis was performed in the human placental JEG-3 cells, and 25 microM resveratrol significantly reduced the mRNA abundance in these cells. Since the transcriptional control of CYP19 gene is tissue-specific and the proximal promoter region of exon Ia has previously been shown to be crucial in CYP19 expression in placental cells, we also evaluated the promoter activity of this gene. Reporter gene assays revealed that resveratrol repressed the transcriptional control of promoter Ia. The present study illustrated the possibility that dietary supplementation of resveratrol interfered with the normal functioning of placental cells.
Asunto(s)
Inhibidores de la Aromatasa/toxicidad , Aromatasa/metabolismo , Estrógenos/metabolismo , Placenta/efectos de los fármacos , Estilbenos/toxicidad , Aromatasa/biosíntesis , Aromatasa/genética , Línea Celular , Relación Dosis-Respuesta a Droga , Represión Enzimática , Genes Reporteros , Humanos , Luciferasas/genética , Placenta/enzimología , Placenta/metabolismo , Regiones Promotoras Genéticas/efectos de los fármacos , ARN Mensajero/biosíntesis , Proteínas Recombinantes/metabolismo , Resveratrol , Transcripción Genética/efectos de los fármacos , TransfecciónRESUMEN
Epidemiological studies have indicated that soya consumption may produce a better plasma lipid profile. The effect may be attributed to the phyto-oestrogens in soya. The red clover (Trifolium pratense) isoflavone biochanin A has a chemical structure similar to those phyto-oestrogens found in soya beans, and is marketed as a nutraceutical for alleviating postmenopausal symptoms. In the present study we investigated the effect of biochanin A on the mRNA expression of ApoA-1 in the hepatic cell line HepG2. Real-time PCR revealed that biochanin A increased ApoA-1 mRNA abundance in cells expressing oestrogen receptor (ER) alpha. Without ERalpha transfection, biochanin A had no effect on mRNA abundance. In order to study the transcriptional control, a fragment of the 5'-flanking region of the ApoA-1 gene was amplified and inserted in a firefly luciferase reporter plasmid. The reporter assay indicated that the transactivation of the ApoA-1 promoter was induced by biochanin A in HepG2 cells transfected with the ERalpha expression plasmid. This induction was reduced by the anti-oestrogen ICI 182,780, whereas the inhibitors of protein kinase (PK) C, PKA, or mitogen-activated kinase (ERK) had no suppressive effect. The present study illustrated that biochanin A might up regulate hepatic apoA-1 mRNA expression through an ER-dependent pathway.
Asunto(s)
Apolipoproteína A-I/genética , Receptor alfa de Estrógeno/genética , Genisteína/farmacología , Fitoestrógenos/farmacología , Receptor beta de Estrógeno/genética , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/genética , Genisteína/análisis , Humanos , Isoflavonas/farmacología , Luciferasas/antagonistas & inhibidores , Fitoestrógenos/análisis , Regiones Promotoras Genéticas/genética , Inhibidores de Proteínas Quinasas/farmacología , ARN Mensajero/análisis , Transfección , Células Tumorales CultivadasRESUMEN
Epidemiological studies indicate that Asian women have a lower incidence of breast cancer compared with their counterparts in the West, and soya consumption has been suggested as a contributory factor. Clinical and animal studies have revealed that cyclooxygenase-2 (COX-2) expression is associated with a risk of breast cancer. In the present study, we investigated the effect of soya isoflavones on the expression of COX-2 in the breast cell line MCF-7. Genistein, daidzein and equol were found to inhibit COX-2 expression induced by phorbol 12-myristate 13-acetate (PMA). Similar findings were observed in the COX-2 protein analysis. In order to study transcriptional control, a fragment of the 5'-flanking region of the hCOX-2 gene was amplified and inserted into a firefly luciferase reporter plasmid. The reporter assay indicated that the transactivation of the hCOX-2 promoter was induced by PMA, and activity was inhibited with the co-administration of genistein, daidzein or equol. An activator protein-1 (AP-1)/cyclic AMP response element binding protein (CREB) binding site (-59/-53) was identified in hCOX-2 promoter, and this could be critical in PMA-induced COX-2 expression. Truncation reporter plasmids with (-70/-36) and without (-51/-36) AP-1/CREB were constructed for subsequent analysis. The results revealed that the hCOX-2 promoter transactivation suppressed by isoflavone could be dependent on AP-1/CREB binding. Nonetheless, this study illustrated that the soya isoflavones reduced COX-2 expression, which could be important in the post-initiation events of breast carcinogenesis.
Asunto(s)
Anticarcinógenos/farmacología , Ciclooxigenasa 2/análisis , Glycine max/química , Isoflavonas/farmacología , Acetato de Tetradecanoilforbol/análogos & derivados , Neoplasias de la Mama/enzimología , Carcinógenos , Línea Celular Tumoral , Ciclooxigenasa 2/genética , Inhibidores de la Ciclooxigenasa 2/farmacología , Equol , Femenino , Genisteína/farmacología , Humanos , Proteínas de la Membrana/análisis , Fitoestrógenos/farmacología , Extractos Vegetales/farmacología , Regiones Promotoras Genéticas/genética , Acetato de Tetradecanoilforbol/farmacologíaRESUMEN
Polycyclic aromatic hydrocarbons (PAHs) are established cancer initiators that can be found in our food and environment. Some dietary plant polyphenols are strong inhibitors to PAH-induced mutagenesis, whereas others may not be as effective. To identify the chemopreventive compounds from a huge volume of dietary components, the development of an efficient screening method is required. In this study, a xenobiotic response element (XRE)-luciferase reporter plasmid was constructed to screen for some potential chemopreventive agents in tea against PAH-induced DNA damage. Tea is one of the most consumed beverages worldwide, and its beneficial effects on health have been documented. Previous studies have claimed that tea polyphenols could be protective against various cancers, and the rich database can be a source for comparison. Among the green and black tea polyphenols, the XRE-luciferase reporter assays suggested that only epigallocatechin gallate (EGCG) was effective in reducing XRE-driven luciferase assay in MCF-7 cells at the concentrations tested. Further study indicated EGCG could reduce CYP1A1 and CYP1B1 mRNA abundances and decrease the DMBA-DNA lesions. The results of DNA covalent binding of all tea polyphenols tested were consistent with the XRE-reporter assays. This study illustrated that the XRE-reporter assay was a viable screening test for dietary chemopreventive agents against PAH-initiated breast mutagenesis. It has the advantages of shorter sample processing time and producing no radioactive waste over directly measuring the CYP1A1/1B1 expressions, DNA lesion, or gel mobility shift assay.
Asunto(s)
Neoplasias de la Mama/prevención & control , Flavonoides/uso terapéutico , Luciferasas , Fenoles/uso terapéutico , Hidrocarburos Policíclicos Aromáticos/toxicidad , Té , Xenobióticos , Análisis de Varianza , Animales , Neoplasias de la Mama/genética , Supervivencia Celular/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Evaluación Preclínica de Medicamentos , Genes Reporteros/fisiología , Polifenoles , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
Several flavonoids have shown their anti-carcinogenic effects in various models. The soyabean isoflavone genistein was demonstrated earlier in our laboratory to be an effective inhibitor of dimethylbenz[a]anthracene (DMBA)-induced DNA damage in MCF-7 cells by curbing cytochrome P450 (CYP) 1 enzymes. The red clover (Trifolium pratense) isoflavone biochanin A is a methylated derivative of genistein, and its anti-mutagenic effect in bacterial cells has been shown previously. Because of its protection against chemical carcinogenesis in an animal model, biochanin A was selected for testing in our established MCF-7 cell system. From the results obtained in the semi-quantitative reverse transcription-polymerase chain reaction and xenobiotic response element (XRE)-luciferase reporter assays, biochanin A could reduce xenobiotic-induced CYP1A1 and -1B1 mRNA abundances through the interference of XRE-dependent transactivation. Enzyme kinetic studies also indicated that biochanin A inhibited both CYP1A1 and -1B1 enzymes with inhibition constant (Ki) values 4.00 and 0.59 microm respectively. Since the biotransformation of DMBA was dependent on CYP1 enzyme activities, biochanin A was able to decrease the DMBA-DNA lesions. The present study illustrated that the red clover isoflavone could protect against polycylic aromatic hydrocarbon-induced DNA damage.
Asunto(s)
9,10-Dimetil-1,2-benzantraceno/metabolismo , Antimutagênicos/uso terapéutico , Hidrocarburo de Aril Hidroxilasas/antagonistas & inhibidores , Carcinógenos/metabolismo , Citocromo P-450 CYP1A1/antagonistas & inhibidores , Genisteína/uso terapéutico , Animales , Hidrocarburo de Aril Hidroxilasas/genética , Biotransformación , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1B1 , Aductos de ADN/metabolismo , Luciferasas/metabolismo , ARN Mensajero/análisis , Elementos de Respuesta , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas/metabolismoRESUMEN
Flavonoids are phenolic compounds isolated from plants, and several of them like genistein and quercetin, have been documented to be effective in preventing cancer. Baicalein, a flavonoid extracted from the root of Scutellaria species, is widely used as a health supplement and herbal medicine in Asian countries. In this study, the chemopreventive effect of baicalein on 7,12-dimethylbenz[a]anthracene (DMBA)-induced DNA damage was evaluated in an established cell culture model. In a preliminary screening, baicalein was identified to be a strong inhibitor to EROD activities induced by DMBA in MCF-7 cells. Subsequent enzyme kinetic analysis revealed that baicalein was a competitive inhibitor to EROD, and CYP1A1 and CYP1B1 gene expressions were also determined. Baicalein could reduce the CYP1A1/1B1 mRNA expressions induced by DMBA, and the mRNA abundance of CYP1A1 appeared to be more responsive than that of CYP1B1. A XRE-luciferase gene reporter assay indicated that AhR transactivation was suppressed. Since CYP1A1/1B1 were responsible for the biotransformation of polycyclic aromatic hydrocarbons, baicalein also demonstrated its ability to reduce DMBA-DNA adduct formation in MCF-7 cells. This study suggested that the natural occurring baicalein could be an agent preventing carcinogen-DNA adduct formation.