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Objective: To explore the influence of miR-34a on Notch1 expression in breast cancer cells, and to explore the role of miR-34a in the sensitivity of breast cancer cells to Adriamycin (ADR)., Methods: The expression of miR-34a in MCF-7 and MCF-7/ADR cells were detected by real-time fluorescence quantitative PCR. Cells were transfected with miR-34a mimics or inhibitor using Lipofectamine 2000. Expression of Notch1 in the two groups before and after transfection was detected by real-time fluorescence quantitative PCR and Western blot. MTT test and apoptosis assay were performed to observe the impact of miR-34a expression on the sensitivity of breast cancer cells to ADR. Statistical analysis of the results was conducted using SPSS 16.0 software. The experiment was repeated independently for at least three times, and had a good reproducibility. The data were shown as mean ± standard deviation. The analysis of variance between groups used one-way ANOVA test. Correlation between groups was analyzed using Spearman rank test. P < 0.05 was considered statistically significant., Results: Compared with the control MCF-7 cells, the relative expression level of miR-34a in MCF-7/ADR cells was significantly decreased to 0.02 (P < 0.05). In the MCF-7 ADR-resistant cells, Notch1 mRNA expression was increased compared with that in the MCF-7 ADR-sensitive cells (2.10 ± 0.20-fold, P < 0.05). The expression levels of Notch1 mRNA and protein in MCF-7/ADR cells transfected with miR-34a mimics were significantly up-regulated. On the contrary, the expressions of Notch1 mRNA and protein in MCF-7 cells transfected with miR-34a inhibitor were down-regulated. The IC50 value of ADR in the control MCF-7/ADR cells and cells transfected with miR-34a mimics was (116.33 ± 16.80) and (39.28 ± 4.12) µmol/L, respectively (P < 0.05). The IC50 value of ADR in control MCF-7 cells and cells transfected with miR-34a inhibitor was (0.29 ± 0.21) and (0.51 ± 0.03) µmol/L, respectively (P < 0.05). Further, overexpression of miR-34a significantly enhanced apoptosis in the ADR-treated MCF-7/ADR cells, and miR-34a inhibitor reduced apoptosis in the ADR-treated MCF-7 cells., Conclusions: miR-34a negatively regulates the expression of Notch1 at both mRNA and protein levels. Overexpression of miR-34a may sensitize MCF-7/ADR cells to adriamycin.

Objective: To study the effect of miR-342-3p on the chemotherapy sensitivity in triple-negative breast cancer (TNBC)., Methods: Tissue samples were collected from January 2011 to August 2013 samples in Jiangsu Cancer Hospital from a total of 32 triple-negative breast cancer patients with preoperative chemotherapy, with 5 cases of complete response (CR) and 27 cases of partial response (PR). We detected miR-342-3p expression of TNBC with RT-PCR. We transfected has-miR-342-3p mimic and inhibitor into breast cancer cell lines MDA-MB-231 by lipofection transfection and set up negative control mim-NC and inhi-NC. Group of mimic, mim-NC, inhibitor and inhi-NC were cultivated with 2 μmol /L paclitaxel, cisplatin or 4 μmol/L doxorubicin for 48 h. The cell growth rates were measured by CCK8 reagent kit, and the cell apoptosis rate by flow cytometry., Results: The expressions of miRNA-342-3p in TNBC tissue of CR were higher than those of PR. The cell growth rates of mimic were lower and cell apoptosis rates were higher than those of min- NC after cultivating with paclitaxel or cisplatin for 48 h, with significant difference (P<0.05). The cell growth rates of inhibitor were higher and cell apoptosis rates lower than those of inhi-NC after cultivating with paclitaxel or cisplatin 48 h, with significant difference (P<0.05). The cell growth and cell apoptosis rates of mimic and inhibitor had no difference with those of mim-NC and inhi- NC after cultivating with doxorubicin 48 h (P>0.05)., Conclusion: TNBC with high expression of miR-342-3p are more sensitive to chemotherapy. miRNA-342-3p may regulate the sensitivity of breast cancer cell line MDA-MB-231 to chemotherapy drugs paclitaxel and cisplatin, but can not affect the chemotherapy sensitivity of doxorubicin.

Objective: To explore whether docetaxel-resistant cells (MCF-7/Doc) and doxorubicin-resistant cells (MCF-7/ADM) can secrete Exosomes and their potential role in cell-cell drug-resistance transfer., Methods: Exosomes were extracted from the cell culture supernatants of MCF-7/Doc and MCF-7/ADM cells by fractionation ultracentrifugation, and were identified by transmission electron microscopy and Western blot analysis. GFP-MCF-7/S, a breast cancer parental sensitive cell line stably expressing green fluorescent protein (GFP), was constructed by recombinant lentiviral vector with GFP. Then the resistance experiment of cells and the experiment of resistance transfer by exosomes were designed to observe the phenomenon of cell-to-cell drug-resistance transfer., Results: Similar to the breast cancer parental sensitive cells (MCF-7/S), the breast cancer resistant sublines could secrete exosomes, which exhibited round or elliptic shape ranging from 30 to 100 nm in diameter with intact membrane, and only expressed the protein marker of exosomes, Tsg101, did not express the endoplasmic reticulum marker calnexin. After MCF-7/S, MCF-7/DOC and MCF-7/ADM cells we cocultured with GFP-MCF-7/S cells for 72 h, there were no significant differences in the expression of fluorescence-labeled cells among the four groups. When treated by the drug ADM or DOC for 24 hours, the MCF-7/DOC+GFP-MCF-7/S group was in favor of a significant higher survival rate of fluorescence-labeled cells compared with the MCF-7/S+GFP-MCF-7/S group (65.5% vs. 25.5%, P < 0.001), and so did the MCF-7/ADM+GFP-MCF-7/S group (53.6% vs. 25.4%, P < 0.001). The exosomes extracted from MCF-7/S, MCF-7/DOC and MCF-7/ADM cells were cultured with the GFP-MCF-7/S cells for 48 h. Among these groups, no significant differences in the expression of fluorescence-labeled cells were found. After treated by the drug ADM or DOC for 24 hours, the exosomes extracted from MCF-7/DOC+GFP-MCF-7/S group was associated with a significant higher survival rate of fluorescence-labeled cells compared with the exosomes extracted from MCF-7/S+GFP-MCF-7/S group (59.9% vs. 32.4%, P < 0.001), and so did the exosomes extracted from the MCF-7/ADM)+GFP-MCF-7/S group (58.3% vs. 27.2%, P < 0.001)., Conclusion: Our results suggest that drug-resistance can be transferred between breast cancer cells, and exosomes are probably the transporter of the drug resistance.

Objective: To investigate the possibility and short-term effect of high dose chemotherapy with peripheral blood stem cell support in the preoperative therapy of breast cancer, and-its influence on the following operation and would healing., Methods: Three patients with T(3)N(1)M(0) (III(a)), T(4)N(1)M(0) (III(b)), T(4)N(1)M(1) (IV) of breast cancer were diagnosed histopathologically. After receiving HDC/APBSCT, the 3 patients were operated on. HDC/APBSCT process included 2 cycles of FEC induction chemotherapy; PBSC mobilization, APBSC collection and cryopreservation and PBSC infusion; and high-dose chemotherapy, APBSC infusion and supportive therapy. The therapy consisted of CTX2.5 g/m(2), VP-16 600 mg/m(2), and cerboplatin 600 mg/m(2) delivered on day 1, APBSC infusion 48 h later, rhG-CSF (150 microg, BID) was administered 4 h after infusion of APBSC until WBC was higher than 10 x 10(9)/L. During HDC/APBSCT, the patients were protected in the air laminar flow room with supportive therapy of antibiotics, anti-virus and anti-fungus drugs. They left the air laminar flow room after their WBC was greater than 2 x 10(9)/L. Case 1 was treated by radical mastectomy, Case 2 by improved radical mastectomy, Case 3 by improved radical mastectomy and transplantation of skin for the large area., Results: Rapid recovery of bone marrow function was observed in all 3 patients. Operation was performed 4 weeks after HDC/PBSCT in Cases 1, 2 and 33 days in Case 3. No influence was seen on operative procedure and would healing, especially in Case 3 with a large area of skin transplantation. Two patients with stage III(a) and III(b) have been alive since the treatment for 30 months and the other with stage IV died of brain metastasis 16 months later., Conclusions: HDC/APBSCT as a preoperative therapy for breast cancer has no influence on the coming surgery and would healing, even on skin transplantation for a large area. It has a practical response in stage III(a) and III(b), but it is still controversial in stage IV. This method as a salvage therapy for patients with breast cancer of intemuediate or stage.