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This study aimed to evaluate in vitro the possible mechanisms underlying the estrogenic potential of benzalkonium chloride (BAC) as a disinfectant emerging contaminant. Effects of BAC at the environmentally-relevant concentrations on estrogen synthesis and estrogen receptor (ER) signaling were assessed using the H295R steroidogenesis assay and the MCF-7 proliferation assay, respectively. Results showed that exposure to BAC at concentrations of 1.0-1.5 mg/L for 48 h significantly increased estradiol production of H295R cells in a concentration-dependent manner. Transcription of steroidogenic genes 3β‐HSD2, 17β‐HSD1, 17β‐HSD4, and CYP19A were significantly enhanced by BAC. In ER-positive MCF-7 cells, exposure to 0.5-1.5 mg/L BAC for 48 h significantly promoted cell proliferation and increased the expressions of ERα and G-protein coupled estrogen receptor 1. Flow cytometry analysis showed that 0.5-1.5 mg/L BAC significantly decreased the percentage of cells in G0/G1 phase, increased the percentage in S phase, and BAC at concentrations of 1.0 and 1.5 mg/L increased the G2/M phase cells. Findings of the study suggested that BAC at environmentally-relevant concentrations might act as a xenoestrogen through its inhibitory effect on steroidogenesis and ER-mediated mechanism.
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Objective To explore the mechanism of microRNA-145 (miR-145) involved in proliferation and apoptosis of breast cancer MCF-7 cells. Methods The immortalized breast cancer cell line MCF-7 cells were cultured in vitro and transfected with miR-145. Real-time PCR and Western blotting were used to detect mRNA and protein levels, respectively. The proliferation level of each group was detected by cell counting kit-8 (CCK-8) method , and the apoptosis level of MCF-7 cells in different treatment groups was detected by flow cytometer. Results Real-time PCR result showed that miR-145 did not affect Caspase-3, proliferating cell nuclear antigen (PCNA) and B cell lymphoma factor 2 (Bcl-2) mRNA levels in MCF-7 cells. Western blotting analysis showed that compared with the control group, transfection of miR-145 for 96 hours significantly increased the expression of Caspase-3 and inhibited the expression of PCNA and Bcl-2. The result of CCK-8 assay showed that the proliferation rate of MCF-7 cells was decreased after overexpression of miR-145 for 72 hours and 96 hours (P<0. 05). The result of flow cytometer showed that the apoptosis rate of MCF-7 cells in overexpression group was significantly higher than that in the control group (P<0. 05). Conclusion MiR-145 can inhibit the proliferation and promote the apoptosis of MCF-7 cells by down-regulating PCNA and Bcl-2 and up-regulating the expression of Caspase-3, which may be a new target for breast cancer treatment.
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MiR-142-3p has been reported to act as a tumor suppressor in breast cancer. However, the regulatory effect of miR-142-3p on drug resistance of breast cancer cells and its underlying mechanism remain unknown. Here, we found that miR-142-3p was significantly downregulated in the doxorubicin (DOX)-resistant MCF-7 cell line (MCF-7/DOX). MiR-142-3p overexpression increased DOX sensitivity and enhanced DOX-induced apoptosis in breast cancer cells. High-mobility group box 1 (HMGB1) is a direct functional target of miR-142-3p in breast cancer cells and miR-142-3p negatively regulated HMGB1 expression. Moreover, overexpression of HMGB1 dramatically reversed the promotion of apoptosis and inhibition of autophagy mediated by miR-142-3p up-regulation. In conclusion, miR-142-3p overexpression may inhibit autophagy and promote the drug sensitivity of breast cancer cells to DOX by targeting HMGB1. The miR-142-3p/HMGB1 axis might be a novel target to regulate the drug resistance of breast cancer patients.
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ABSTRACT Plants are considered among the main sources of biologically active chemicals. The species Solidago chilensis Meyen, Asteraceae, is native to the southern parts of South America, where the aerial parts of the plant are commonly used for the treatment of inflammatory conditions. However, the effects of S. chilensis on human cancer cells remain to be elucidated. In this study, we evaluated the antiproliferative effects of the hydroalcoholic and dichloromethane extracts of S. chilensis, as well as their chemical constituents quercitrin and solidagenone against the five human tumor cell lines in vitro. The dichloromethane extract showed a promisor antiproliferative effects in vitro, especially against glioma cell line. Besides, the hydroalcoholic extract and quercitrin were inactive. The diterpene solidagenone showed highly potent antiproliferative effects against breast (MCF-7), kidney (786-0), and prostate cancer (PC-3) cells (total growth inhibition: TGI < 6.25 µg/ml). Solidagenone meets the theoretical physico-chemical criteria for bioavailability of drugs, according to the "Rule of Five" and, by theorical studies, the observed biological effects were probably related to the interaction of the molecule with nuclear receptors and as an enzymatic inhibitor. This study contributes to chemical study and to the identification of antiproliferative molecules in S. chilensis.
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Objective@#To investigate the properties of radiation-induced changes of cell cycle and apoptosis in breast cancer cell lines MDA-MB-231 and MCF-7, and to further explore its relationship with radiosensitivity.@*Methods@#The two cell lines MCF-7 and MDA-MB-231 were irradiated with 6 MV X-rays. The cell survival curves were fitted by clonogenic formation assay, then according to the radiosensitivity parameters average lethal dose (D0), quasi-threshold dose (Dq) and survival fraction after 2 Gy irradiation (SF2), the radiosensitivity of the two cell lines was compared. The apoptosis rate and cell cycle distribution of the two cell lines were detected by Hoechst 33342 and PI staining.@*Results@#The survival curve of MDA-MB-231 cell line shifted to the right compared with that of MCF-7 cell line. The values of D0, Dq and SF2 of MDA-MB-231 cell line were higher than those of MCF-7 cell line (1.603±0.023 vs. 1.233±0.027, 1.76±0.04 vs. 1.03±0.10, 0.639 3±0.008 2 vs. 0.398 1±0.018 5, t values were -17.981, -11.745, -20.596, P values were 0.000, 0.003, 0.000). The apoptosis rate of MCF-7 cell line was significantly higher than that of MDA-MB-231 cell line at the doses of 2, 4, 8 Gy irradiated 24 h and at the 12, 48 h after 6 Gy X-irradiation (t values were 4.441, 7.299, 10.499, 6.375, 7.743, P values were 0.011, 0.002, 0.000, 0.003, 0.001). Compared with the non-irradiated group, G2 phase arrest appeared in both cell lines after irradiation. The percentage of the G2/M phase in MDA-MB-231 cell line increased as the time or dosage accumulated. Furthmore, the percentage didn't go down even after 48 hours later. However, the blockage began to gradually release in MCF-7 cell line at the dose of 8 Gy irradiated 24 h and the 48 h after 6 Gy X-irradiation. Followed with that, it turned out the percentage of the G0/G1 phase increased [(65.80±0.56)%, (62.53±0.67)%].@*Conclusions@#6 MV X-irradiation with the doses of 2-8 Gy can induce the cell cycle arrest at G1 and G2 phase in MCF-7 cell line, G2 phase in MDA-MB-231 cell line. Thus more apoptosis appears in MCF-7 cell line, which may cause the difference in radiosensitivity between the two cell lines.
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Objective: To select and identify the endophytic fungi of Rabdosia rubescens as well as study on its antitumor activity in vitro in order to obtain the high yield strains of oridonin. Methods: Using the methods of TLC and HPLC, the oridonin high yield strain was selected from 256 strains of endophytic fungi which were isolated from the previous experiments. Oridonin of fermentation broth was determined by HPLC-MS. The strain was identified by morphological characteristics and ITS sequence methods and its antitumor activity in vitro was detected by MTT assay. Results: An oridonin high yield strain M-J-5 was obtained and identified as Penicillium oxalicum, which can inhibit the activity of human breast cancer cell line MCF-7. Conclusion: a oridonin high yield strain is obtained with antitumor activity and provides scientific basis for microbial production of oridonin and the development of new anticancer drugs.
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Objective: To explore the anti-proliferation and apoptosis-inducing effects of decitabine combined with paclitaxel on paclitaxel-resistant MCF-7 cells. Methods:Cell counting kit-8 (CCK-8) assays were used to determine the proliferation inhibition of drugs at different concentrations in 24, 48 and 72h. The group was treated with decitabine, paclitaxel and the combination of the two drugs, respectively. The cell apoptosis rate was determined by flow cytometry after the treatment with the drugs at different concentra-tions in 48h. Results:The results of CCK-8 showed the combination group significantly inhibited the cell proliferation when compared with the single drug use group(P<0. 05), and the anti-proliferation value was in a dose-dependent manner in all groups. The apopto-sis values after the treatment with decitabine, paclitaxel and the combination in 48h was(20. 4 ± 1. 98)%,(21. 8 ± 3. 34)% and(70. 8 ± 8. 23)%,respectively. Conclusion:The proliferation inhibition and apoptosis induction are both increased significantly in the com-bination group. Synergistic effect of decitabine and paclitaxel is found in multidrug resistant breast cancer cell line MCF-7 in vitro.
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The use of gold nanoparticle in drug delivery has emerged as a promising avenue to reduced toxicity and frequency of dosage while maintaining therapeutic effects and biocompatibility. Therefore, the possibility of developing eco-friendly metallic gold nanoparticles is evaluated. To achieve this, aqueous leave extracts of Calotropis procera was used to synthesis gold nanoparticles and its cytotoxic effect was investigated. The gold nanoparticles (AuNPs) produced were characterized using Ultra Violet–Visible spectroscopy, Zeta-sizer nano, High Resolution Scanning Electron Microscopy (HRSEM), Energy-Dispersive X-ray (EDAX) spectroscopy and Fourier Transmission Infrared (FTIR) spectroscopy. The cytotoxic ability of the synthesized gold nanoparticles was evaluated on MCF-7 cell using MTT assay. The result of Ultra Violet–Visible spectroscopy showed development of gold nanoparticle reaction at 550 nm of Surface Plasmon Resonance and average particle size of 45 nm was confirmed using nano Zeta-sizer. EDAX profile result suggested the presence of gold at 2.30ke while FTIR result confirms the presence of biomolecules serving as reducing and capping agents on the synthesized gold nanoparticle with a strong signal at 3426 cm of the hydroxyl group of alcohol or phenol. The cytotoxic effect of the synthesis gold nanoparticles shows cell viability decreased as the concentration of AuNPs increased from 0.156 mg to 5 mg with an IC50 of 0.312 mg/l. In conclusion, this study demonstrated the bioreductive capability of aqueous leaf extract of Calotropis procera to produced gold nanoparticle and its cytotoxicity effect on MCF-7cell line.
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Aims: To evaluate the In vitro cytotoxic effects of the hydro ethanolic extract (HEE) of the Delonix regia flowers against different cell lines. Methodology: The dried Delonix regia flowers are subjected to soxhlet extraction by using 70% ethanol. The dried extract was used to determine the qualitative preliminary phytochemical analysis, total phenolic and flavonoid content. The cytotoxic property of the extract was determined by using MTT assay against breast cancer (MCF-7), cervix (HeLa), brain and colon cancer cells. Tamoxifen is used as a standard for all the cell lines. Results: Qualitative phytochemical tests of extract HEE showed the presence of sugars, flavonoid, tannins, phenolic compounds, steroids and saponins. The percentage of phenolic and flavonoid content was determined as 31.42 mg/g, 29.22 mg/g respectively. The cytotoxic activity of the extract showed, IC50 concentrations (μg/ml) against MCF-7 (breast), carcinoma of cervix HeLa cells, carcinoma of the brain, and carcinoma of colon cells against tamoxifen are 141.6 ± 0.08, 223.7 ± 2.16, 173.9 ± 0.7, 168.33 ± 0.04 respectively. Conclusions: The experimental data clearly demonstrate the hydro ethanolic extract (HEE) showed cytotoxic properties against human cancer cells.
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Objective To investigate the effect of Shugan-Yishen formula on Tamoxifen-resistant MCF-7 cell line(LCC9) through HER2-ERK/MAPK-ERα pathway. Methods Four serums were prepared with the method of serum pharmacology:the serum of Shugan-Yishen formula, Shugan-Yishen formula plus Tamoxifen(TAM), blank and TAM alone. After LCC-9 cells were affected in the above-mentioed serums in vitro, the inhibition rate and the level of mRNA as well as the protein expression of HER2, ERα, ERK/MAPK, p-ERK/MAPK were detected. Results MTT showed that TAM alone group、Shugan-Yishen formula group and the Shugan-Yishen formula plus TAM group had higher inhibition rates at 48th hour, the inhibition rates was(6.61±0.129)%, (47.43±2.24)%, (54.19±3.364)%, compared to the blank group, Shugan-Yishen formula group and the Shugan-Yishen formula plus TAM group had a stronger inhibition on LCC9 at 48th hour (P0.05). RT-PCR result showed that compared to the blank group, the expression levels of ESR1 mRNA of group of Shugan Yishen formula plus TAM werehigher by and large at 48th hour (P0.05). Conclusions ①The combination of Shugan-Yishen formula with tamoxifen can inhibit the LCC9 cells. The inhibition effect reaches the peak at 48th hour. ②The mechanism of Shugan-Yishen formula plus TAM combination effects on human breast cancer TAM-resistant cell line LCC9 may improve the sensitivity of ERαby inhibiting the expression of HER2, ERK/MAPK.
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Objective We established a model of human breast cancer in nude mice, to discuss the feature of pathology and biology of breast cancer,and give help to establish tools of pathogen research. Methods Human breast cancer cells MCF-7 were subcutaneously injected into the right armpit of nude mice to establish human breast cancer models.The mice were divided into composite group, estrogen group, leptin group and the blank group (30 in each). In the composite group,estrogen and leptin were injected into peripheral region of the tumor daily.In the estrogen group,estrogen was injected.In the leptin group, leptin was injected.In the blank group, physiological saline was injected.Tumor growth was observed, and the volume of the tumor was recorded.The tumor tissues obtained from the mice were pathologically examined. Results (1)The tumor-taking rate of leptin group and the blank group were 46.7% (14/30)and33.3% (10/30) ,and the mode is failure.In composite group and estrogen group they were 96.7% (29/30) and 70% (28/30).There was not significant difference between them ( P < 0.05).(2) The differences of average diameter and volume were statistically significant between the composite group and the estrogen group (P < 0.05).(3) Pathology diagnosis was invasive ductal carcinoma. Conclusions (1) Establishing human breast cancer model in nude mice need the stimulation of estrogen. The tumor-taking rate of nude mice has no relationship with leptin.(2) In the study in vivo, leptin as same as estrogen has stimulating effect on MCF-7 cells proliferation.(3) The transplanted cancer cell partly have the pathology and biology features of the human breast cancer cells and give help to establish tools of pathogen research.
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Objective To explore the effect of vascular endothelial growth factor-C (VEGF-C) gene transfection on the expression level of VEGF-C in human breast cancer MCF-7 cell. Methods The constructed VEGF-C gene eukaryotic expression vector was transfected into the human breast cancer MCF-7 cell by using lipofectamine transfection reagents, and the positive cell clones were obtained through G418 selection after transfection. The expressions of VEGF-C mRNA and protein were detected by RT-PCR and Western blot respectively. Results Following the transfection of the VEGF-C recombination plasmid, there were significant differences on the expression levels of VEGF-C mRNA and protein between pcDNA3.1-VEGF-C transfection group and pcDNA3.1 transfection group (12.382?2.183 vs 6.039?1.950, P
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Objective: Multidrug resistance (MDR) of malignant tumors to chemotherapeutic agents has played a major role in the failure of cancer chemotherapy. Finding effective and low toxic reversing agents against MDR has been a critical challenge. In this study, we aim to investigate the reversing effects of ginsenoside Rh2 on MDR in vitro its the corresponding mechanism. Methods: MCF-7 or MCF-7/ADM cells were incubated with ginsenoside Rh2 alone or in combination with adriamycin (ADM), respectively. The viabilities of MCF-7 and MCF-7/ADM cells were assessed by MTT method and the in IC50 values were calculated. The inhibitory effects of Rh2 and verapamil on the efflux of rhodamine 123 were assayed by flow cytometric analysis. The expression of mdr1 gene was measured by RT-PCR and P-gp expression was determined by flow cytometry. Results: Ginsenoside Rh2 significantly decreased IC50 value of ADM on MCF/ADM cells (P <0.05), but it had no effect on MCF-7 cells. Rh2 and verapamil significantly increased the accumulation of rhodamine 123 in drug-resistant cells MCF - 7/ADM in a dose-dependent manner (P <0.05), but they had no effect on MCF-7 cells. Ginsenoside Rh2 had stronger effect in inhibition of rhdamine 123 efflux than verapamil. RT-PCR and flow cytometric analysis showed that the expressions of mdr1 gene and P-gp had no significant difference in MCF-7/ADM cells after ginsenoside Rh2 treatment. Conclusion: Ginsenoside Rh2 effectively reverses P-gp-mediated drug resistance of MCF-7/ADM cells.
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OBJECTIVE: We investigated the effects on the cell cycle and p53 expression by the treatment of various concentrations of staurosporine to elucidate the molecular mechanism of staurosporine induced cell cycle arrest in MCF-7 cell line. METHODS: Various concentrations of staurosporine were treated in MCF-7 cells cultured with RPMI 1640 media. The harvested cells were fixed and permeabilized with 1% paraformaldehyde and absolute methanol. Then the cells were stained indirectly with anti-p53 primary antibody and FITC conjugated goat anti-mouse(GAM)-IgG secondary antibody. Sequentially DNA were stained with 0.1% RNase and PI solution. These stained cells were analyzed by the standard FACScan flow cytometer. The obtained results were analyzed further with WinList 3.0, and ModiFit LT software program. RESULTS: MCF-7 cells were arrested mostly in G1 phase of cell cycle at 5-10 nM of staurosporine, however, the cells were arrested in G2 phase at 20-100 nM of staurosporine. The expressions of p53 protein were higher in the MCF-7 cells treated with both concentrations of 10 nM and 100 nM of staurosporine compaired with the control cells. This suggests that the p53 may be involved in the mechanism of G1 and G2M arrest of cell cycle in MCF-7 cell. CONCLUSIONS: The points of arrest in cell cycle differred depending on the concentrations of staurosporine and these cell cycle arrests at G0G1 and G2M pahse were related with p53 protein expression. It suggested that these results could be extended to study for staurosporine to be usefull as a potential anti-tumor agent.
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Cell Cycle Checkpoints , Cell Cycle , DNA , Fluorescein-5-isothiocyanate , G1 Phase , G2 Phase , Goats , MCF-7 Cells , Methanol , Ribonucleases , StaurosporineABSTRACT
Objective To explore the efficacy of calmodulin antagonist berbamine(BBM) in down-regulating survivin mRNA. Methods Human breast cancer cell line MCF7 and its adriamycin-resistant counterpart MCF7/ADR were used in the study. The cells were cultured with different concentration of BBM for 72 hours. The mRNA expression level of survivin gene in both MCF and MCF7/ADR cells was detected by semi-quantitative RT-PCR. Results After treating MCF7 and MCF7/ADR cells by 20?mol/L BBM, the mRNA expression level of survivin gene decreased from 0.43?0.02 to 0.21?0.04 in MCF7 cells, and from 0.57?0.05 to 0.45?0.04 in MCF/ADR cells(P
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Object To study the effects of artemisinin and its analogue artemisunate on the proliferation of human breast cancer MCF-7 cell line, as well as their mechanism comparatively. Methods The inhibition of cell proliferation was determined by SRB method. Cell cycle was determined by flow cytometry (FCM) analysis. Apoptosis was confirmed by sub-G 1 cells content and DAPI method. Results The cell cycle of MCF-7 was changed greatly when treated 24 h with either 10 ?mol/L artemisinin or 1 ?mol/L artemisunate, the distribution of MCF-7 cells among S phase was reduced greatly, while inereased during G 0+G 1. However, artemisinin had weaker effect on the proliferation of MCF-7 cell, while artemisunate effectively inhibited the proliferation of MCF-7, the IC 50 was 0.31 ?mol/L. Apoptosis induced by 1 ?mol/L artemisunate was stronger than that by 10 ?mol/L artemisinin, too. Conclusion The inhibitory effect of artemisunate on the proliferation of tumor cell is stronger than that of artemisinin in vitro.
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8 ?mol/L)was markedly restrained. Zearalenone, like estradiol, could markedly increase the proliferation of MCF-7 cells. The effects were time-dependent and dose-dependent. Conclusion: The tested phytoestrogens are biologically active, and they can differently affect the proliferation of human breast cancer cells in vitro. These data suggest that genistein may have preventive and therapeutic applications against breast tumors.