| Breast cancer incidence was significantly increased in recent years. The treatment including operation, chemotherapy, radiotherapy and endocrine therapy. Chemotherapy is a better treatment to alleviate and therapy of breast cancer. Various structures of different anticancer drugs have different mechanisms of action, can be in short-term inside of remission in breast cancer, multidrug resistance which makes the treatment of breast cancer is very difficult. The occurrence of multidrug resistance is the main reason for the failure of chemotherapy, is the biggest obstacle in cancer treatment, is the result of multiple mechanisms. There is still no method of reverse tumor, therefore the treatment of breast cancer depends on the in-depth studying of its pathogenesis,treatment method and explore new methods of drug resistance reversal. The subject on the basis of previous studies, study on reversal effect of Gambogic acid on multidrug resistant human breast cancer cells and its related mechanism, as Gambogic acid is developed as a kind of reversal of multidrug resistance drug provides the corresponding theoretical basis.Objective: To study on reversal effect of Gambogenic acid on multidrug resistant human breast cancer cells and its related mechanism.Methods: The cell growth and proliferation of Gambogenic acid and Adriamycin on MCF-7/ADR cells and MCF-7 cells were measured by MTT assay; Adriamycin combined with low dose of Gambogenic acid(<IC10) effect on the proliferation of MCF-7/ADR cells; MCF-7/ADR cell and MCF-7 cell intracellular accumulation were detected by flow cytometry; Flow cytometry was used to detecting the effect of Gambogenic acid on MCF-7/ADR intracellular adriamycin and Rho123 accumulation,and Rho123 efflux effect; flow cytometry on MCF-7/ADR cell cycle effects of Gambogenic acid; flow cytometry on P-g P protein in MCF-7/ADR cells;Western blot to detect the express of P-g P protein, PTEN protein, P-PI3 K protein,p-AKT protein.Results:(1) When ADR combined with low concentration Gambogenic acid(below IC10) in MCF-7/ADR cells, ADR MCF-7/ADR cells of IC50 was obviously decreased,with significant difference(P<0.05). When Gambogenic acid concentration reached 1μM, IC50 for 48 h was 50.97 μM. Reversal coefficient was 1.57; When Gambogenic acid concentration reached 4 μM, reverse effect is very obvious, the role of IC50 for 48 h was 27.29 μM. Reversal coefficient was 2.94; when Gambogenic acid concentration reached 4 μM, reverse effect is very obvious, the role of IC50 for 48 h was 12.54 μM.Reversal coefficient was 6.40.(2) ADR were added in MCF-7 and MCF-7/ADR cells, the results showed that the content of ADR in MCF-7 cells was higher than MCF-7/ADR cells(P<0.01).(3) After treated by Gambogenic acid, intracellular fluorescence enhanced obviously,suggesting Gambogenic acid may increase intracellular MCF-7/ADR accumulation of ADR and Rho123.(4) In the Rho123 efflux experiments, no drug before and after 30 min, 60 min, 2 h in MCF-7/ADR cells, Rho123 reduced the level of trend; effect of Gambogenic acid,30 min, 60 min, 2 h, MCF-7/ADR cells in Rho123 efflux was inhibited, the results show Gambogenic acid could inhibit the P-g P pump out effect.(5) PI single staining results show that with the increase of Gambogenic acid concentration, the G0/G1 phase of the cell number also increased with the increase of cell number changes of S phase is small, the number of cells in G2/M phase decreased significantly.(6) Different concentrations of Gambogenic acid effect on MCF-7/ADR cells, the expression of P-g P decreased with the increase of Gambogenic acid concentration, and showed dose dependent.(7) Western blot test results showed, Gambogenic acid increased the expression of the PTEN protein, the expression of p-PI3 K and p-Akt reduced, and reduced the expression of P-g P protein.Conclusion: Gambogenic acid could reverse P-g P-mediated human breast MCF-7/ADR cells via inhibiting PTEN/PI3K/Akt signal pathway. |
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