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John Christian Larsen - One of the best experts on this subject based on the ideXlab platform.
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detection of weak estrogenic flavonoids using a recombinant yeast strain and a modified mcf7 Cell Proliferation Assay
Chemical Research in Toxicology, 1998Co-Authors: V Breinholt, John Christian LarsenAbstract:A newly developed recombinant yeast strain, in which the human estrogen receptor has been stably integrated into the genome of the yeast, was used to gain information on the estrogenic activity of a large series of dietary flavonoids. Among 23 flavonoids investigated, 8 were found to markedly stimulate the transcriptional activity of the human estrogen receptor in the yeast Assay increasing transcriptional activity 5−13-fold above background level, corresponding to EC50 values between 0.1 and 25 μM. Five compounds increased the transcriptional activity 2−5-fold over the control, with EC50 values ranging from 84 to 102 μM, whereas the remaining flavonoids were devoid of activity. The most potent flavonoid estrogens tested were naringenin, apigenin, kaempferol, phloretin, and the four isoflavonoids equol, genistein, daidzein, and biochanin A. With the exception of biochanin A, the main feature required to confer estrogenicity was the presence of a single hydroxyl group in the 4‘-position of the B-ring of th...
Rosalee M Mamilton - One of the best experts on this subject based on the ideXlab platform.
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use of a tetrazolium based Cell Proliferation Assay to measure effects of in vitro conditions on perkinsus marinus apicomplexa Proliferation
Journal of Eukaryotic Microbiology, 1995Co-Authors: Christopher F Dungan, Rosalee M MamiltonAbstract:ABSTRACT. Because the in vitro Cell cycle of the apicomplexan oyster pathogen Perkinsus marinus generates Cell populations heterogeneous for size and typified by aggregation, both turbidimetric and counting methods for determining population densities and Proliferation rates are inaccurate or cumbersome. We show that a commercial, tetrazolium-based Cell Proliferation Assay yields a soluble formazan chromophore upon intraCellular reduction by P. marinus. at a rate proportional to Cell population biovolume. Using this Assay system, we have 1) defined selected culture system parameters which maximize P. marinus in vitro Proliferation, 2) assessed selected chemosensitivities, and 3) standardized the Assay system for quantification of densities and doubling times of populations propagated with our optimized system. Growth was supported by four tested base media and was maximized in 1:1 DME/Ham's F-12. Temperatures of 10–40° C permitted growth, which was maximized at 35° C. pH 6.0–8.5 permitted growth, which was maximized at 7.0–7.5. Osmolalities of 340–1,930 mOsm supported growth, which was maximized at 790 mOsm. Serum supplements from 1–10% (v/v) did not enhance log phase growth, but enhanced stationary phase metabolic activity in proportion to concentration. Our isolate (ATCC 50439) has a 13 h log phase doubling time when propagated under optimized conditions: 28° C, 800 mOsm, pH 7.0, 1:1 DME/Ham's F-12 medium, 5% (v/v) FBS. It is tolerant of antibacterial agents at concentrations commonly used in vertebrate tissue culture, but is inhibited by several antimycotics at similar concentrations.
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Use of a Tetrazolium‐based Cell Proliferation Assay to Measure Effects of In Vitro Conditions on Perkinsus marinus (Apicomplexa) Proliferation
Journal of Eukaryotic Microbiology, 1995Co-Authors: Christopher F Dungan, Rosalee M MamiltonAbstract:ABSTRACT. Because the in vitro Cell cycle of the apicomplexan oyster pathogen Perkinsus marinus generates Cell populations heterogeneous for size and typified by aggregation, both turbidimetric and counting methods for determining population densities and Proliferation rates are inaccurate or cumbersome. We show that a commercial, tetrazolium-based Cell Proliferation Assay yields a soluble formazan chromophore upon intraCellular reduction by P. marinus. at a rate proportional to Cell population biovolume. Using this Assay system, we have 1) defined selected culture system parameters which maximize P. marinus in vitro Proliferation, 2) assessed selected chemosensitivities, and 3) standardized the Assay system for quantification of densities and doubling times of populations propagated with our optimized system. Growth was supported by four tested base media and was maximized in 1:1 DME/Ham's F-12. Temperatures of 10–40° C permitted growth, which was maximized at 35° C. pH 6.0–8.5 permitted growth, which was maximized at 7.0–7.5. Osmolalities of 340–1,930 mOsm supported growth, which was maximized at 790 mOsm. Serum supplements from 1–10% (v/v) did not enhance log phase growth, but enhanced stationary phase metabolic activity in proportion to concentration. Our isolate (ATCC 50439) has a 13 h log phase doubling time when propagated under optimized conditions: 28° C, 800 mOsm, pH 7.0, 1:1 DME/Ham's F-12 medium, 5% (v/v) FBS. It is tolerant of antibacterial agents at concentrations commonly used in vertebrate tissue culture, but is inhibited by several antimycotics at similar concentrations.
Eliana Aparecida Varanda - One of the best experts on this subject based on the ideXlab platform.
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evaluation of estrogenic potential of flavonoids using a recombinant yeast strain and mcf7 bus Cell Proliferation Assay
PLOS ONE, 2013Co-Authors: Flavia Aparecida Resende, Ana Paula Siqueira Oliveira, Mariana Santoro De Camargo, Wagner Vilegas, Eliana Aparecida VarandaAbstract:Phytoestrogens are of interest because of their reported beneficial effects on many human maladies including cancer, neurodegeneration, cardiovascular disease and diabetes. Furthermore, there is a search for compounds with estrogenic activity that can replace estrogen in hormone replacement therapy during menopause, without the undesirable effects of estrogen, such as the elevation of breast cancer occurrence. Thus, the principal objective of this study was to assess the estrogenic activity of flavonoids with different hydroxylation patterns: quercetin, kaempferol, luteolin, fisetin, chrysin, galangin, flavone, 3-hydroxyflavone, 5-hydroxyflavone and 7-hydroxyflavone via two different in vitro Assays, the recombinant yeast Assay (RYA) and the MCF-7 Proliferation Assay (E-screen), since the most potent phytoestrogens are members of the flavonoid family. In these Assays, kaempferol was the only compound that showed ERα-dependent transcriptional activation activity by RYA, showing 6.74±1.7 nM EEQ, besides acting as a full agonist for the stimulation of Proliferation of MCF-7/BUS Cells. The other compounds did not show detectable levels of interaction with ER under the conditions used in the RYA. However, in the E-screen Assay, compounds such as galangin, luteolin and fisetin also stimulated the Proliferation of MCF-7/BUS Cells, acting as partial agonists. In the evaluation of antiestrogenicity, the compounds quercetin, chrysin and 3-hydroxyflavone significantly inhibited the Cell Proliferation induced by 17-β-estradiol in the E-screen Assay, indicating that these compounds may act as estrogen receptor antagonists. Overall, it became clear in the Assay results that the estrogenic activity of flavonoids was affected by small structural differences such as the number of hydroxyl groups, especially those on the B ring of the flavonoid.
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Evaluation of estrogenic potential of flavonoids using a recombinant yeast strain and MCF7/BUS Cell Proliferation Assay.
PLOS ONE, 2013Co-Authors: Flavia Aparecida Resende, Ana Paula Siqueira Oliveira, Mariana Santoro De Camargo, Wagner Vilegas, Eliana Aparecida VarandaAbstract:Phytoestrogens are of interest because of their reported beneficial effects on many human maladies including cancer, neurodegeneration, cardiovascular disease and diabetes. Furthermore, there is a search for compounds with estrogenic activity that can replace estrogen in hormone replacement therapy during menopause, without the undesirable effects of estrogen, such as the elevation of breast cancer occurrence. Thus, the principal objective of this study was to assess the estrogenic activity of flavonoids with different hydroxylation patterns: quercetin, kaempferol, luteolin, fisetin, chrysin, galangin, flavone, 3-hydroxyflavone, 5-hydroxyflavone and 7-hydroxyflavone via two different in vitro Assays, the recombinant yeast Assay (RYA) and the MCF-7 Proliferation Assay (E-screen), since the most potent phytoestrogens are members of the flavonoid family. In these Assays, kaempferol was the only compound that showed ERα-dependent transcriptional activation activity by RYA, showing 6.74±1.7 nM EEQ, besides acting as a full agonist for the stimulation of Proliferation of MCF-7/BUS Cells. The other compounds did not show detectable levels of interaction with ER under the conditions used in the RYA. However, in the E-screen Assay, compounds such as galangin, luteolin and fisetin also stimulated the Proliferation of MCF-7/BUS Cells, acting as partial agonists. In the evaluation of antiestrogenicity, the compounds quercetin, chrysin and 3-hydroxyflavone significantly inhibited the Cell Proliferation induced by 17-β-estradiol in the E-screen Assay, indicating that these compounds may act as estrogen receptor antagonists. Overall, it became clear in the Assay results that the estrogenic activity of flavonoids was affected by small structural differences such as the number of hydroxyl groups, especially those on the B ring of the flavonoid.
V Breinholt - One of the best experts on this subject based on the ideXlab platform.
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detection of weak estrogenic flavonoids using a recombinant yeast strain and a modified mcf7 Cell Proliferation Assay
Chemical Research in Toxicology, 1998Co-Authors: V Breinholt, John Christian LarsenAbstract:A newly developed recombinant yeast strain, in which the human estrogen receptor has been stably integrated into the genome of the yeast, was used to gain information on the estrogenic activity of a large series of dietary flavonoids. Among 23 flavonoids investigated, 8 were found to markedly stimulate the transcriptional activity of the human estrogen receptor in the yeast Assay increasing transcriptional activity 5−13-fold above background level, corresponding to EC50 values between 0.1 and 25 μM. Five compounds increased the transcriptional activity 2−5-fold over the control, with EC50 values ranging from 84 to 102 μM, whereas the remaining flavonoids were devoid of activity. The most potent flavonoid estrogens tested were naringenin, apigenin, kaempferol, phloretin, and the four isoflavonoids equol, genistein, daidzein, and biochanin A. With the exception of biochanin A, the main feature required to confer estrogenicity was the presence of a single hydroxyl group in the 4‘-position of the B-ring of th...
Toshimasa Nishiyama - One of the best experts on this subject based on the ideXlab platform.
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evaluation of the estrogenic activities of some pesticides and their combinations using mtt se Cell Proliferation Assay
International Journal of Hygiene and Environmental Health, 2006Co-Authors: Mari Manabe, Seiji Kanda, Kenji Fukunaga, Airo Tsubura, Toshimasa NishiyamaAbstract:Abstract A number of pesticides are used in agricultural production with some having estrogenic activities, such as endocrine-disrupting chemicals that may affect wildlife and humans. This study aimed to detect the estrogenic effects of some mixed agricultural chemicals in agricultural production. The Assay to measure estrogenic activity was evaluated by the Cell proliferative activity of MtT/Se Cells, which respond well to estrogen. To evaluate MtT/Se Cells we went down to the molecular level of estrogen receptor (ER)-α and ER-β expression. The proportion of ER-α to ER-β was 3.55:1, as determined by semi-quantitative real-time PCR. These results showed that ER-α was dominant in MtT/Se Cells on the transcriptional level, therefore implying that the estrogenic activity detected by these Cells may be mainly mediated by ER-α. It was found that diazinon, tolclofos–methyl, pyriproxyfen, prothiofos and thiabendazole had estrogenic activity. Several pesticides are often present in agricultural products. Therefore, we evaluated the estrogenic activity of a mixture of two pesticides. The REC10 levels of prothiofos/pyriproxyfen and thiabendazole/orthophenylphenol were increased up to 10-fold. We concluded that those two pesticide combinations showed a significantly higher estrogenic effect in comparison to the results of the respective pesticides when tested individually.
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Evaluation of the estrogenic activities of some pesticides and their combinations using MtT/Se Cell Proliferation Assay.
International Journal of Hygiene and Environmental Health, 2006Co-Authors: Mari Manabe, Seiji Kanda, Kenji Fukunaga, Airo Tsubura, Toshimasa NishiyamaAbstract:Abstract A number of pesticides are used in agricultural production with some having estrogenic activities, such as endocrine-disrupting chemicals that may affect wildlife and humans. This study aimed to detect the estrogenic effects of some mixed agricultural chemicals in agricultural production. The Assay to measure estrogenic activity was evaluated by the Cell proliferative activity of MtT/Se Cells, which respond well to estrogen. To evaluate MtT/Se Cells we went down to the molecular level of estrogen receptor (ER)-α and ER-β expression. The proportion of ER-α to ER-β was 3.55:1, as determined by semi-quantitative real-time PCR. These results showed that ER-α was dominant in MtT/Se Cells on the transcriptional level, therefore implying that the estrogenic activity detected by these Cells may be mainly mediated by ER-α. It was found that diazinon, tolclofos–methyl, pyriproxyfen, prothiofos and thiabendazole had estrogenic activity. Several pesticides are often present in agricultural products. Therefore, we evaluated the estrogenic activity of a mixture of two pesticides. The REC10 levels of prothiofos/pyriproxyfen and thiabendazole/orthophenylphenol were increased up to 10-fold. We concluded that those two pesticide combinations showed a significantly higher estrogenic effect in comparison to the results of the respective pesticides when tested individually.