Detection Of MiR-21 Targeting MARCKS Gene On The Sensitivity Of Human Breast Cancer Cells To Docetaxel

ObjectiveDocetaxel-resistant breast cancer parental sensitive cells(MDA-MB-231/S)and their homologous docetaxel-resistant cells(Doc;MDA-MB-231/Doc)were selected as the research objects.Objective To investigate the regulation of microRNA-21(microRNA-21,miR-21)on MARCKS gene expression and its potential relationship with docetaxel resistance in breast cancer MDA-MB-231 cell line,and to find a target for reversal therapy of breast cancer resistance.MethodsDifferential expression of microRNAs-21 in parental sensitive cell lines(MDA-MB-231/S)and docetaxel-resistant cell lines(MDA-MB-231/Doc)was screened by microarray and verified by real-time fluorescence quantitative analysis(RT-qPCR),and mimics and inhibitors were synthesized and transfected respectively.Interventional experiments were carried out to observe the expression of microRNAs-21 and the changes of drug sensitivity in these cells after intervention;target genes of microRNAs-21 were predicted by target gene prediction software,and the targeting relationship between microRNAs-21 and MARCKS was analyzed;MTT,flow cytometry,RT-qPCR and Western blotting were used.To verify the effect of microRNAs-21 expression on docetaxel resistance in triple-negative breast cancer cells by targeting the MARCKS gene.Results1.Compared with MDA-MB-231/S,the expression level of miR-21 in MDA-MB-231/Doc was significantly higher,and the difference was(2.03+0.06)times(P<0.05).2.Compared with the negative control group,the expression level of miR-21 in sensitive cells(MDA-MB-231/S)transfected with miR-21 mimics was significantly increased(2.26(+0.07)times(P<0.05),and the expression of IC50(Doc concentration leading to 50%cell death)in the transfected cells was significantly increased,and the apoptosis rate induced by Doc was significantly decreased.After transfection of MDA-MB-231/Doc with the inhibitors of microarray-21,the expression of microarray-21 was significantly down-regulated(0.36 + 0.03 times as high as that of negative control group;P<0.05),and the expression of IC50 was significantly decreased and the apoptosis rate induced by Doc was significantly increased in MDA-MB-231/Doc transfected cells.3.PicTar,TargetScan and miRDB prediction software analysis revealed that MARCKS was a specific target gene for miR-21.Further studies confirmed that the expression of MARCKS gene was negatively correlated with microRNAs-21 mimics:the expression of MARCKS mRNA and protein in MDA-MB-231/S cells transfected with microRNAs was significantly decreased and drug resistance was enhanced;the expression of MARCKS mRNA and protein in MDA-MB-231/Doc cells transfected with microRNAs-21 inhibitors was significantly increased,and drug resistance was reversed.Conclusion1.miR-21 expression in breast cancer drug-resistant cells(MDA-MB-231/Doc)was significantly higher than that in sensitive cells(MDA-MB-231/S).2.Introducing mimics and inhibitors of microRNAs into breast cancer cells in vitro,we found that the expression level of microRNAs was positively correlated with the drug resistance of breast cancer cells to docetaxel.3.Bioinformatics results show that MARCKS is a specific target gene for microRNAs-21,and miR-21 is negatively correlated with the expression of MARCKS gene.4.miR-21 may affect the sensitivity of triple-negative breast cancer cells to docetaxel chemotherapy by targeting the MARCKS gene.