Expression Of MT1H In Triple-negative Breast Cancer Cell Line MDA-MB-231 And Its Effect On The Biological Behavior Of Its Malignant Biology

Objective: 1.To analyze the expression of metallothioneins(MTs)in triple negative breast cancer(TNBC)tissues and their correlation with clinical prognostic information,and to analyze the specific biological significance of metallothionein 1H(MT1H)in triple negative breast cancer;2.To investigate the expression level of MT1 H in triple negative breast cancer cell line(MDA-MB-231);3.To investigate the effect of MT1 H expression on the malignant biological behavior of triple negative breast cancer cells through in vitro cellular assays,and to provide a theoretical basis and clinical rationale for finding potential therapeutic targets for triple negative breast cancer.Methods: 1.Selected transcriptome data from breast cancer patients in The Cancer Genome Atlas database(The Cancer Genome Atlas,TCGA),bioinformatics analysis of MTs expression and prognosis using R language programming package,and single gene analysis of MT1H;2.q RT-PCR technique to detect differences in MT1 H transcriptome levels in different cell lines;3.si RNA transfection technique was constructed to interfere with MDA-MB-231 cell line by MT1 H,grouped as blank control group(Control group);NC si RNA transfection group as negative control group(si-NC group);MT1H si RNA transfection group as MT1 H transfection interference experimental group(si-MT1 H group),and detected by q RT-PCR technique.si RNA interference efficiency;4.Flow cytometry detection of apoptosis in different treatment groups.Cell function assay was performed to detect the effect of malignant behavior(proliferation,migration)of cells in different treatment groups;5.Western blot was performed to detect the effect of MT1 H on the epithelial-mesenchymal transition process(EMT)of MDA-MB-231 cells.Results: 1.Bioinformatics analysis revealed that MTs were differentially expressed in various subtypes of breast cancer,and MTs were found to be associated with poor prognosis in triple negative breast cancer patients;2.MT1 H single gene analysis revealed that its expression was correlated with clinical prognostic information,TMN staging,Tumor mutation burden(TMB)and microsatellite instability(MSI)data.The analysis of differential gene function enrichment revealed that MT1H-related differential genes were mostly enriched in Pathways in cancer,and drug sensitivity analysis revealed that patients with high MT1 H expression might be more sensitive to gemcitabine treatment;3.q RT-PCR showed that the expression level of MT1 H transcriptome was significantly higher in triple-negative breast cancer cell line MDA-MB-231 compared with normal breast epithelial cell line MCF-10A(P<0.05);4.plate cloning assay suggested that the value-added capacity of tumor cells was decreased in si-MT1 H group compared with the control group(P<0.05);5.Flow experiments showed that the apoptosis level was increased in the si-MT1 H group compared with the control group(P<0.05);6.Transwell experiments showed that the cell migration ability was decreased in the si-MT1 H group compared with the control group(P<0.05);7.Westernblot results showed that the expression level of the epithelial marker E calcineurin expression level was increased and the expression levels of mesenchymal markers vimentin and N-cadherin were decreased in the si-MT1 H group compared with the control group(P<0.05).Conclusion:1.Bioinformatics analysis revealed that MTs were differentially expressed in TNBC and mostly associated with poor prognosis in TNBC patients;MT1H may be associated with tumor pathway and highly correlated with gemcitabine,which can be used as a biomarker for breast cancer prognosis;2.The expression level of MT1 H transcriptome in triple-negative breast cancer cell line MDA-MB-231 was significantly higher than that of normal breast epithelial cell line MCF-10A;3.Interfering with MT1 H promotes apoptosis of MDA-MB-231 cells and inhibits their proliferation,migration ability,and epithelial-mesenchymal transition.These suggest that MT1 H is expected to be a new therapeutic target for triple negative breast cancer in the future.