| Objectives Long noncoding RNA(lcn RNA)plays a regulatory role in lots of biological processes.It is also involved in cell proliferation,differentiation and carcinogenesis.HOX transcript antisense RNA(HOTAIR)is a common studied RNA in lnc RNA family.It is dysregulated in many kinds of malignant tumors.It has significant effect on the occurrence or metastasis of malignant tumors.This study explored the expression of lnc RNA HOTAIR in human breast cancer tissue,and the relationship to clinical progress indicators of human breast cancer.Then,this study further explored the effects of high expression of HOTAIR on proliferation,apoptosis and invasion of breast cancer cells,and its effect on epithelial mesenchymal transition(EMT),so as to provide a new basis for the diagnosis of patients with advanced breast cancer and the development of targeted therapeutic drugs.Methods(1)A total of 80 cases with breast cancer who did not receive neoadjuvant therapy were selected.Real-time PCR was used to detect the expression of HOTAIR in tumor tissues and surrounding normal tissues of breast cancer,and the correlation between HOTAIR expression and clinical pathological parameters was analyzed.(2)MTT assay was used to analyze the effect of HOTAIR overexpression on MCF-7 cell viability at 24 h,48 h and 72 h after transfection.After 48 h transfection,the changes of apoptosis were detected by flow cytometry,and the cell invasion was detected by Transwell assay.(3)MCF-7 cells were cotransfected with HOTAIR vector and human CDH1 or mouse CDH1(E-cadherin core promoter of)vector individually.After 48 h transfection,the fluorescence activity of CDH1 was analyzed by double luciferase reporter gene assay to verify the relationship between HOTAIR and E-cadherin promoter.The protein expression of E-cadherin,N-cadherin and vimentin related to EMT were further determined by Western blot after cotransfecting.Results(1)The results of quantitative PCR showed that the expression of HOTAIR in human breast cancer tissues was higher than that in surrounding normal tissues,with significant difference(p<0.01).The expression of HOTAIR was not correlated with age and nationality(p>0.05),but was significantly correlated with clinical stage,differentiation degree and lymph node metastasis(all p<0.001);(2)Compared the transfected cells with empty vector,high expression of exogenous HOTAIR significantly increased the proliferation activity of MCF-7 cells(p< 0.05)in a time-dependent manner(increased at 24 h,but p>0.05,p<0.05 at 48 h,p<0.01 at 72h).Overexpression of HOTAIR significantly inhibited the apoptosis of MCF-7 cells,and the difference between the two groups was statistically significant(p<0.01).Transwell assay showed that the invasion of MCF-7 cells was also significantly increased(p<0.05).At the same time,after HOTAIR transfection,the cells became long spindle shape and the pseudopodia elongation increased,which significantly promoted like epithelial mesenchymal transition;(3)Cotransfection of HOTAIR and CDH1 into MCF-7 cells decreased the luciferase activity of human or mouse CDH1 respectively(p<0.01 for human and p<0.05 for mouse).Western blot results showed that after cotransfection,the expression of E-cadherin was decreased(p<0.05),and the expression of N-cadherin and vimentin were incereased(both p<0.01).Conclusion The expression of HOTAIR gene was up regulated in breast cancer tissues,and it was related to clinical stage,tumor differentiation and lymph node metastasis.Overexpression of HOTAIR in MCF-7 cells can promote cell proliferation and invasion,inhibite cell apoptosis.Overexpression of HOTAIR can inhibit the transcription of E-cadherin promoter,then down regulate the epithelial protein marker Ecadherin and up regulate the mesenchymal protein markers N-cadherin and vimentin.These changes indicate that overexpression of HOTAIR can promote the mesenchymal transformation of breast cancer cells,which may be one of the mechanisms of the increase of proliferation,invasion and metastasis of breast cancer. |
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