The Effect And Mechanism Of MicroRNA-129 In Breast Cancer

Background: Breast cancer is the most common malignant tumor and one of the leading causes of death in female worldwide.Micro RNAs(miRNAs)have been identified to play a crucial role during the tumorigenesis and progression,but the specific mechanism of most miRNAs in breast cancer have not been elucidated.miR-129-5p is a member of the micro RNA family.Many studies have indicated that miR-129-5p function as tumor suppressor in a variety of solid tumors,including nervous system tumors,gastric cancer,hepatocellular carcinoma,and urinary system tumors.At present,the role and mechanism of miR-129-5p in the breast cancer tumorigenesis and progression have not been widely reported.Objective: Our aim is to reveal the specific role and mechanism of miR-129-5p about breast cancer,and to provide theoretical basis to seeking for new therapeutic targets in breast cancer.Method: 1.Samples of tumor tissue and mammary tissue of 30 patients with primary breast cancer were collected,RT-q PCR was performed to detect the expression of miR-129-5p in tumor tissue and mammary tissue.2.We collected pathological paraffin samples of 200 patients with primary breast cancer in Tianjin medical university cancer institute and hospital.We detected the expression of miR-129-5p in the paraffin samples by RT-q PCR.We analyzed the correlation between the clinicopathological factors and the expression level of miR-129-5p.We analyzed 5-year DFS and OS by Kaplan-Meier survival curve;And we also analyzed the risk factors of recurrence and metastasis in breast cancer by Cox proportional risk regression mode.3.We detected the expression levels of miR-129-5p in breast cancer cell lines and normal breast epithelial cell line by RT-q PCR.miR-125-5p mimics or inhibitors were transiently transfected to MDA-MB-231 cell line and MCF7 cell line,and we detected the proliferation of breast cancer cells by MTT and plate colony formation assays.We detected the number of miR-129-5p overexpressed MDA-MB-231 cell apoptosis by flow cytometry.And we also detected the changes in migration and invasion ability of breast cancer cells by Transwell migration assay.4.Epithelial mesenchymal transformation(EMT)in MCF10 A cells was induced by TGF-β1.We detected the expression level of miR-129-5p by RT-q PCR.And we also observed the cell morphology by microscopy.5.We detected the expression levels of EMT-related markers and relevant transcription factors in miR-129-5p overexpressed MDA-MB-231 cell line and down-expressed MCF7 cell line by RT-q PCR.We observed the localization of EMT-related makers by immunofluorescence staining assay.6.Target prediction programs were applied to identify the putative targets of miR-129-5p.We determined the interaction between miR-129-5p and the target genes by dual luciferase assay.We detected the expression level of target genes by RT-q PCR and Western Blot.7.We interfered the target genes in both miR-129-5p overexpressed and downexpressed cell lines.We evaluated the changes in cellular biological behavior of breast cancer cell lines by MTT,plate colony formation assay,Transwell migration assay and flow cytometry.Results: 1.Compared to mammary tissue,the expression level of miR-129-5p in breast cancer tissue decreases.2.Patients with low miR-129-5p expression correlates to advanced clinical staging,higher histological grade,more lymph node metastasis,and higher PR positive rate.The 5-year DFS and OS with low expression of miR-129-5p are significantly lower than those with high expression of miR-129-5p.Low miR-129-5p expression is an independent risk factor for recurrence and metastasis in breast cancer.3.The expression of miR-129-5p decreases in breast cancer cell lines.Overexpression of miR-129-5p inhibits the proliferation,migration and invasion of breast cancer cells and increases the number of apoptosis cells.4.The expression of miR-129-5p in breast cells reduces during EMT.5.miR-129-5p overexpression inhibits the expression levels of EMT-related markers and transcription factors in breast cancer cells.6.Twist1 and CBX4 are direct targets of miR-129-5p.7.miR-129-5p inhibits the proliferation,migration and invasion of breast cancer cells by targeting Twist1 and CBX4.Conclusions: The expression of miR-129-5p is down-regulated in breast cancer and predicts the prognosis in patients with breast cancer.miR-129-5p inhibits the proliferation,migration and invasion by targeting Twist1 and CBX4.Therefore,we uncover the role and mechanism of miR-129-5p in breast cancer and expression of miR-129-5p can be used as a biomarker for prediction of patients with breast cancer.