Objective To investigate the effect of low-frequency and low-intensity ultrasound irradiation microbubbles(LILFU-MB)on drug resistance of triple-negative breast cancer MDA-MB-231/DOX cells.Methods The best non-toxic irradiation time of LILFU-MB was screened and the cell survival rate was calculated.The MDA-MB-231/DOX cells were divided into LILFU-MB irradiation group and non LILFU-MB irradiation group.The drug inhibition fitting curve was drawn to obtain the half inhibitory concentration(IC50)of the two groups of drugs,the best dose of doxorubicin was screened.According to different treatment methods,the cells were divided into control group,doxorubicin group,LILFU-MB group,LILFU-MB+doxorubicin group,the cell proliferation activity was observed,and the cell migration rate was calculated.The morphology of the cells were observed by inverted fluorescence microscope,and the fluorescence intensity and karyotype of the cells were recorded.The apoptosis rate was detected by flow cytometry.The protein expression was detected by Western blot.Results The best non-toxic irradiation time of LILFU-MB was 30 s,and the cell survival rate was(92.33±2.58)%under this condition.This condition was used for the duration of LILFU-MB irradiation in subsequent experiments.The IC50 of doxorubicin on drug-resistant cells in the LILFU-MB irradiation group was lower than that in the non LILFU-MB irradiation group(7.51µmol/L vs.35.40µmol/L).The best dose of doxorubicine was 35.40µmol/L.The cell proliferation activity of doxorubicin group and LILFU-MB+doxorubicin group was significantly inhibited,and the proliferation rate was slowed down,and the inhibition effect of LILFU-MB+doxorubicin group was the most obvious.The cell migration rates of the control group,doxorubicin group,LILFU-MB group,LILFU-MB+doxorubicin group were(56.79±1.11)%,(51.34±4.66)%,(46.09±9.42)%,(22.01±6.02)%,respectively,and the cell migration rates of LILFU-MB+doxorubicin group were significantly different from that of other groups(all P<0.01).Inverted fluorescence microscope revealed that most of the nuclei in the LILFU-MB+doxorubicin group were obviously shrunk and fragmented,the blue staining of the nuclei was highlighted,the chromatin was concentrated and marginalized,and the strong apoptotic karyotype change was showed.The results of flow cytometry showed that the apoptosis rates of the control group,doxorubicin group,LILFU-MB group and LILFU-MB+doxorubicin group were(13.30±0.66)%,(22.68±2.85)%,(20.60±3.72)%,(31.84±2.87)%,respectively.The apoptosis rate of LILFU-MB+doxorubicin group was higher than that of the other groups,and the differences were statistically significant(all P<0.05).The expressions of P-gp protein and Bcl-2 in LILFU-MB+doxorubicin group were significantly down-regulated,while the expressions of Cyt-c and Cl-caspase3 protein were significantly up-regulated,and the differences were statistically significant compared with that of other groups(all P<0.05).Conclusion LILFU-MB can significantly inhibit the proliferation and migration of MDA-MB-231/DOX cells,promote cell apoptosis,and reverse the drug resistance of triple negative breast cancer MDA-MB-231/DOX cells.
Ultrasound irradiation,low-frequency,low-intensityMicrobubblesTriple-negative breast cancerMultidrug resistanceDoxorubicin
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