The Relationship Between MiRNA-216b And The Breast Cancer

Objective:This study aims to investigate the effect of miRNA-216 b on the function of various types of BC cells by clarifying its effects on the prognosis of diverse BC patient subtypes,and predicting its target gene to investigate the expression levels in different BC type populations and their effect on prognosis.The research provides a theoretical basis that whether miRNA-216 b may be a potential diagnostic and prognostic marker for BC patients.Methods:We used the TCGA and Kaplan-Meier plotter databases to determine the expression levels of miRNA-216 b in BC patients and their relationship with the survival of different types of BC patients.RT-qPCR technique was used to determine the expression levels of miRNA-216 b in different BC cell lines.The functional importance of miRNA-216 b in MCF7,LCC2,and MDA-MB-231 cells was examined using four different assays:MTS cell proliferation assay,cell scratch assay,ventricle perforation assay,and flow cytometry,respectively.Using R language(4.0.3),we performed Differentially Expressed Genes(DEGs)analysis and we predicted miRNA-216 b target genes through three databases:Target Scan,miRbase,and mi-RDB,which were used to predict the target genes of miRNA-216 b and integrate them with the results of DEGs analysis to identify differentially expressed target genes.We performed GO enrichment analysis and KEGG pathway analysis on the differentially expressed target genes.Genes on the pathways of interest were extracted and analyzed in the TCGA database for differential expression and the association with the expression and survival in different BC subtypes.Results:1.Association between miRNA-216 b and breast cancer prognosisKaplan-Meier plotter database survival analysis results showed that overall survival(OS)time was reduced in those with high miRNA-216 b expression in all BC patients and ER+ BC patients [BC: HR=1.86(1.34-2.59),P=0.00019;ER+ BC: HR=7.45(2.78-19.97),P<0.001].However,miRNA-216 b was not significantly associated with survival in HER2-positive BC patients(P=0.1),while in TNBC patients with high miRNA-216 b expression was linked to a longer OS time(TNBC: HR=0.21(0.07-0.64),P=0.0025)2.miRNA-216 b functions in ER+ and TNBC breast cancer cellsThe results of the cell function assay demonstrated that,in MCF7 cells,after transfection with miRNA-216 b mimic for 24 hours,the functions of proliferation,migration,and invasion in MCF7 cells were significantly increased(P<0.01),while the ability of apoptosis was inhibited(P<0.001)compared to mimic NC group;after transfection with miRNA-216 b inhibitor After transfection for 24 hours,the functions of proliferation,migration,and invasion in MCF7 cells were significantly inhibited(P<0.01),while the apoptosis ability was increased(P<0.01)compared to the inhibitor NC group.In LCC2 and MDA-MB-231 cells transfected with miRNA-216 b mimic for 24 hours,abilities of proliferation,migration,and invasion in LCC2 and MDA-MB-231 cells were significantly inhibited(P<0.05),while the apoptosis ability was increased(P<0.001)compared to the mimic NC group;transfected with miRNA-216 b inhibitor After 24 hours of transfection,abilities of cell proliferation,migration,and invasion were significantly increased(P<0.01),while the apoptosis ability was inhibited(P<0.01)compared with the inhibitor NC group.These results suggest that miRNA-216 b has a cancer-promoting effect in non-TAM-resistant cells and an oncogenic effect in TAM-resistant BC cells and TNBC cells.3.Differential expression of miRNA-216 b in breast cancer patientsTCGA database showed that the level of miRNA-216 b expression was higher in breast cancer tissues than that in non-cancerous tissues.Moreover,the difference was statistically significant.4.Predictive screening of miRNA-216 b target genes We obtained 2913 down-regulated DEGs and 1935 up-regulated DEGs.Using the TCGA database: screening for genes differentially expressed between breast cancer tissues and control sample tissues.Using Taeget Scan,miRbase,and mi-RDB to predict the target genes,we attained 4333,15142,and 593 possible target genes respectively,and identified 242 target genes by intersecting the results.After intersecting with the DEGs,we obtained 233 differentially expressed target genes for the KEGG pathway analysis and GO enrichment analyses.Our findings suggest that the Hippo signaling pathway was significantly enriched,and the main biological roles were metal ion binding and protein phosphorylation in GO enrichment analysis.We further investigated the expression levels of target genes in various types of breast cancer patients and performed survival analyses.SRPK3 was found to be lowly expressed in patients with ER+、PR+ breast cancer and highly expressed in patients with ER+、PR-and TNBC.Therefore,SRPK3 may serve as a protective factor for non-TAM-resistant breast cancer patients and a risk factor for patients with TAM-resistant breast cancer and TNBC,SRPK3 is a possible target gene for miRNA-216 b.Conclusions:1.The higher miR-216 b expression level was associated with poorer prognosis in ER+ breast cancer patients,and better prognosis in TNBC patients.2.miR-216 b expression in breast cancer tissues was higher than that in paraneoplastic tissues.3.miR-216 b was expressed at higher levels in ER+ breast cancer cells than in TNBC and TAM-resistant BC cells.4.In TAM-resistant breast cancer and TNBC,high miR-216 b expression could inhibit the proliferation,invasion,and migration ability of tumor cells and promote apoptosis;in ER+ breast cancer cells,high miR-216 b expression could promote the proliferation,invasion,and migration ability of tumor cells and inhibit apoptosis.5.SRPK3 is lowly expressed in ER+ breast cancer patients,and the higher the expression level,the better the prognosis of patients;it is highly expressed in TAM-resistant breast cancer and TNBC patients,and the higher the expression level,the worse the prognosis of patients.