Chinese Clinical Oncology

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Reversal effect of miR-490-3p on the resistance of MCF-7/ADM cells to adriamycin

WANG Xiaoxian, KONG Fanjun, LI Fei, ZHANG Shuo   

  1. Department of Pharmacy and Equipment, 456 Hospital of PLA, Ji'nan 250031, China
  • Received:2015-07-18 Revised:2015-08-05 Online:2015-09-30 Published:2015-09-30

Abstract: 【Abstract】Objective To investigate the expression of miR-490-3p in adriamycin(ADM)resistant human breast cancer cell line MCF-7/ADM and its effect on the cell proliferation,apoptosis and resistance to ADM of MCF-7/ADM cell line. Methods Cultured wild-type MCF-7 human breast cancer cells and its resistant cells MCF-7/ADM were selected as research objectives. The realtime fluorescent quantitative PCR(qPCR)was used to compare the level of miR-490-3p in both cell lines. The recombinant plasmid pcDNA3.1(+)miR-490-3p to over-express miR-490-3p was transiently transfected into MCF-7/ADM cells(overexpression group)and then the transfection effect was verified by qPCR. Meanwhile,the pcDNA3.1(-)empty control group and blank control group were set up. MTT method was used to determine the effect of pcDNA3.1(+)miR-490-3p on the proliferation of each group. The inhibitory rate of ADM on the proliferation of MCF-7/ADM cells was determined, and the inhibition concentration (IC50) and the reversal factor were calculated to evaluate the sensitivity to ADM. The cells in each group were collected at 48 h post-transfection for the detection of apoptotic rate by flow cytometry with Annexin FITC/PI double staining. Meanwhile, the expression of resistance proteins including P-glycoprotein(P-gp),breast cancer resistance protein (BCRP)and multidrug resistance associated protein 1(MRP1)were tested by Western blotting,intracellular ADM concentration was detected by fluorescence photometer and Pgp activity by flow cytometry. Results The level of miR-490-3p in MCF-7/ADM cells was 0.24±0.07,significantly lower than 1.02±0.03 of wild type MCF-7 cells(P<005). The levels of miR-490-3p in the overexpression group were higher than those in the other two groups during 24-96 h post-transfection(P<005). The proliferation inhibition rates were(17.52 ±1.87)%,(31.67±2.79)%,(45.09±1.88)% and(61.82±2.52)% at 48,24,72 and 96 h post-transfection, all higher than those of empty control group(P<0.05). IC50 value of ADM for MCF-7/ADM cells was(11.27±2.34)μg/ml,lower than that of blank control group and empty control group(P<0.05). Moreover,reversal index was 3.35and 3.39fold of blank control group and empty control group for the overexpression group. Compared with the other two groups,there were increased apoptotic rates and intracellular drug concentration but decreased expression of P-gp,BCRP and MRP1 as well as P-gp activity in over-expression group (P<0.05).Conclusion Upregulation of miR-490-3p exhibits the reversal effect of resistance to ADM in MCF-7/ADM cell,possibly by reducing the resistance gene protein expression and inhibition of P-gp activity. Elevated level of miR-490-3p can inhibit proliferation and induce apoptosis.

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