The antiestrogenic effects of black cohosh on BRCA1 and steroid receptors in breast cancer cells

Figures & data

Figure 1 (A) BC downregulates ER-α protein expression in T-47D cells. (B) BC downregulates ER-α in MCF-7 cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with 80–500 µM BC and subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at *P<0.05; **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for Windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviations: BC, black cohosh; ER-α, estrogen receptor alpha.

Figure 2 (A) Black cohosh downregulates PR protein expression in T-47D cells. (B) BC downregulates PR protein expression in MCF-7 cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with 80–500 µM BC and subjected to SDS-PAGE and Western blot analysis as described in Materials and methods section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at *P<0.05; **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviations: BC, black cohosh; PR, progesterone receptor.

Figure 3 (A) BC downregulates BRCA1 protein expression in T-47D cells. (B) BC downregulates BRCA1 protein expression in MCF-7 cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with 80–500 µM BC and subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at *P<0.05; **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviation: BC, black cohosh.

Figure 4 The effects of hormones and antihormones in combination with BC on ER-α expression in T-47D cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with varying combinations of BC, estrogens, and antiestrogens. They were subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at *P<0.05; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviations: BC, black cohosh; ER-α, estrogen receptor alpha.

Figure 5 The effects of hormones and antihormones in combination with BC on ER-α expression in MCF-7 cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with varying combinations of BC, estrogens, and antiestrogens. They were subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at *P<0.05; **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviations: BC, black cohosh; ER-α, estrogen receptor alpha.

Figure 6 The effects of hormones and antihormones in combination with BC on PR expression in T-47D cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with varying combinations of BC, estrogens, and antiestrogens. They were subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviations: BC, black cohosh; PR, progesterone receptor.

Figure 7 The effects of hormones and antihormones in combination with BC on PR expression in MCF-7 cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with varying combinations of BC, estrogens, and antiestrogens. They were subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviations: BC, black cohosh; PR, progesterone receptor.

Figure 8 The effects of hormones and antihormones in combination with BC on BRCA1 expression in T-47D cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with varying combinations of BC, estrogens, and antiestrogens. They were subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviation: BC, black cohosh.

Figure 9 The effects of hormones and antihormones in combination with BC on BRCA1 expression in MCF-7 cells.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 24 hours with varying combinations of BC, estrogens, and antiestrogens. They were subjected to SDS-PAGE and Western blot analysis as described in the “Materials and methods” section. Actin bands represent equivalence in protein loading. The optical densities of each protein band were measured using the computer program Image Studio Lite. Differences are considered significant at ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviation: BC, black cohosh.

Figure 10 (A, B) BC decreases T-47D and MCF-7 cell viability.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 6 days with 80–500 µM BC and cell viability was determined by propidium iodide staining and image cytometry using the Cellometer vision CBA. The samples were compared to the control. Differences were considered significant at *P<0.05; **P<0.01; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviation: BC, black cohosh.

Figure 11 (A, B) The effects of hormones and antihormones in combination with BC on T-47D and MCF-7 cell viability.

Notes: The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. The cells were treated for 6 days with varying combinations of BC, estrogens, and antiestrogens. Cell viability was determined by propidium iodide staining and image cytometry using the Cellometer vision CBA. The samples were compared to the control. Differences were considered significant at *P<0.05; ***P<0.001. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviation: BC, black cohosh.

Figure 12 The effects of BC on the cellular localization of BRCA1.

Notes: Images (magnification ×20) were acquired according to the “Materials and methods” section. The control, “Cs”, represents DMSO, the solvent for BC.
Abbreviations: BC, black cohosh; DMSO, dimethyl sulfoxide.

Figure 13 (A, B) The effects of BC, E₂, and ICI alone and in combination on ESR1 levels in T-47D and MCF-7 breast cancer cells.

Notes: ESR1 mRNA levels determined by reverse transcription quantitative real-time PCR. T-47D cells were treated in the presence or absence of 400 µM BC, E₂, and ICI for 24 hours. The control, “Cs”, represents dimethyl sulfoxide, the solvent for BC. Results are shown as the mean ± SEM of at least three independent experiments with three replicates in each experiment. The samples were compared to the control. Differences were considered significant at **P<0.01. Statistical analyses were carried out using SPSS for windows version 11.5 (SPSS Inc., Chicago, IL, USA).
Abbreviations: BC, black cohosh; SEM, standard error of the mean.