| Background:Breast cancer i5 the second most common malignant tumor in the world,which seriously threatens the health of women around the world.How to improve the diagnosis and treatment of breast cancer is a difficult problem facing the whole world.Adriamycin has a high proportion of tumor drug resistance in clinical chemotherapy of breast cancer.Therefore,to explore the mechanism of resistance of breast cancer cells to adriamycin and how to improve the drug resistance will greatly improve the effect of breast cancer chemotherapy.In the early stage of our research group,it has been verified that inhibiting the expression of CDCA4 can inhibit the proliferation,growth and promote apoptosis of breast cancer cells.The relationship and mechanism between miR-15a and adriamycin resistance in breast cancer have been rarely reported.Therefore,it is necessary to explore the mechanism by which miR-1sa may target CDCA4 to regulate the chemical sensitivity of breast cancer to adriamycin.Objective:Our study was to explore miR-15a in breast cancer MCF-7/ADR adriamycin resistant cells of tumor biology behavior(proliferation,migration,invasion,apoptosis and cell cycle),and the possible target genes CDCA4 expression level of the relationship,that we can improved about the pathogenesis of breast cancer cells to adriamycin resistance,for miR-15 a/CDCA4 as a target for targeted therapy provides a new research idea of breast cancer.Methods:In our study,the expression levels of miR-15a-5p in MCF-7/ADR cells,normal breast cell line Hs578Bst and MCF-7 were detected by RT-qPCR.Then,human breast cancer adriamycin-resistant cell line MCF-7/ADR was selected as the research object,and exogenous miR-15a-5p mimics and miR-15a-5p inhibitor were transfected into MCF7/ADR cell line.RT-qPCR was used to detect and compare the expression levels of miR-15a-5p in the miR-15a-5p mimics transfection group,the blank group and miR-15a-5p inhibitor,and on this basis,we tested CDCA4 mRNA expression level by RT-qPCR,cell proliferation and growth by CCK-8,cell migration and invasion ability by Transwell assay,cell cycle and apoptosis rate by flow cytometry.Results:1.RT-qPCR showed that miR-15a-5p in MCF-7/ADR cells was significantly lower than that in normal breast cells and breast cancer cells.2.After transfection with exogenous miR-15a-5p mimics and miR-15a-5p inhibitor respectively in MCF-7/ADR cells,the expression levels of miR-15a-5p in each group were detected and compared by RT-qPCR.It was found that miR-15a-5p expression level in miR-15a-5p mimics transfected group was significantly higher than that in MCF-7/ADR cells in the blank group,and miR-15a-5p expression level was significantly lower after transfection with miR-15a-5p inhibitor than that in the blank group.The differences were statistically significant(P<0.05).3.RT-qPCR was used to detect the expression levels of CDCA4 mRNA in the miR-15a-5p mimics transfection group,the blank group and the miR-15a-5p inhibitor transfection group.The results showed that the expression level of CDCA4 was negatively correlated with miR-15a-5p,and the difference was statistically significant(P<0.05).4.CCK-8 assay was used to detect the proliferation capacity of miR-15a-5p mimics transfection group,blank group and miR-15a-5p inhibitor transfection group.The results showed that compared with blank group and miR-15a-5p inhibitor transfection group,the inhibition rate of adriamycin in MCF-7/ADR cells stably transfected with miR-15a-5p mimics was significantly higher,and the difference was statistically significant(P<0.05).5.Flow cytometry was used to detect the apoptosis rate of the stably transfected miR-15a-5p mimics group,the blank group and the miR-15a-5p inhibitor group,and the results showed that the apoptosis rate of breast cancer adriamycin-resistant cell line MCF-7/ADR stably transfected miR-15a-5p mimics was significantly increased(P<0.05).6.Flow cytometry was used to detect stable transfection of miR-15a-5p mimics transfection group,blank group and miR-15a-5p inhibitor transfection group.Before and after the addition of miR-15a-5p mimics,the number of cells in the S phase was significantly different among the three groups.Before the addition of miR-15a-5p mimics,25.885%of the cells in the S phase were in the S phase,and the proportion of the cells in the S phase was significantly higher than 23.865%in the blank control group and 21.385%in the mir-15a-5p inhibitor transfection group(P<0.05).After addition of the drug,35.455%of the cells in the miR-15a-5p mimics transfection group were in the S phase,and the proportion of the cells in the S phase was still significantly higher than that in the blank control group(26.600%)and the miR-15a-5p inhibitor transfection group(22.545%)(P<0.05).There was no significant difference in the proportion of G1 phase and G2 phase between the three groups before and after administration.Conclusion:Overexpression of miR-15a-5p can inhibit the proliferation and promote the apoptosis of adrianycin-resistant breast cancer cells MCF-7/ADR,which may be related to the targeted inhibition of CDCA4,among which the regulation of miR-15a-5p on the MCF-7/ADR cell cycle may be one of the partial mechanisms.These data provide a scientific basis for reversing adriamycin resistance in breast cancer.Nevertheless,other potential regulatory pathways and roles of miR-15a-5p in breast cancer still need to be clarified through further experiments. |
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