LncRNA AL591178.1 Regulates The Function Of Nasopharyngeal Carcinoma Cells During EMT

Objective:Select the nasopharyngeal carcinoma cell line CNE-1 to construct a cell model of epithelial-mesenchymal transformation of nasopharyngeal carcinoma cells,screen out differentially expressed lncRNA,and determine the key lncRNA in the development of nasopharyngeal carcinoma epithelial mesenchymal transformation And explore its functions.It provides theoretical evidence for exploring the role of lncRNA in the invasion and metastasis of nasopharyngeal carcinoma,and provides new ideas for the diagnosis and treatment of nasopharyngeal carcinoma.Methods:1.Incubate nasopharyngeal carcinoma cell line CNE-1 cells for 24 hours in vitro,treat the experimental group cells with 5ng/mL TGF-β1 reagent for 24 hours to induce CNE-1 cells to undergo epithelial-mesenchymal transformation,and observe with light microscope Changes in cell morphology;2.Extract cellular RNA and protein,and verify the molecular markers of epithelial-mesenchymal transition E-cadherin,N-cadherin,vimentin,and Twist protein in experimental group and control group by qPCR and Western Blot,respectively The difference in the expression level of 3.The addition of TGF-β1 signaling pathway inhibitor LY2109761 and TGF-β1 co-treated CNE-1 cells,further verifying that TGF-β1 is a key factor in inducing EMT in CNE-1 cells;4.Extract the experimental group The RNA of the control group was sequenced,bioinformatics analysis,GO function enrichment analysis,KEGG signal pathway enrichment analysis,differential gene verification,etc.were selected to screen for differentially expressed IncRNA;5.The most up-selected from the screened IncRNA The gene IncRNA AL591178.1 was verified,and its expression was interfered by means of cell transfection.The molecular marker E-cadherin,N-cadherin of CNE-1 cell epithelial-mesenchymal transition was verified by qPCR and Western Blot.Changes in the expression levels of shape protein and Twist protein,etc.6.CNE-1 cells not treated with TGF-β1,CNE-1 cells treated only with TGF-β1,treated with TGF-β1 and transfected with negative controls CNE-1 cells with siRNA and CNE-1 cells treated with TGF-β1 and transfected with si-AL siRNA were divided into four groups.The RNA and protein of the four groups of cells were extracted,and qPCR and Western Blot verification were performed respectively.The expression levels of epithelial-mesenchymal transition molecular markers between the four groups were different.Results:1.100 times magnification,it can be seen that the cells have undergone epithelial interstitial changes,which show that the intercellular adhesion is reduced and the cell morphology becomes spindle-shaped;2.TGF-β1 can induce the expression level of EMT molecular markers in CNE-1 cells Changes occurred,E-cadherin expression increased,N-cadherin,Vimentin and Twist expression decreased;3.After adding LY2109761 and TGF-β1 co-treatment of CNE-1 cells,CNE-1 cells EMT process was inhibited;4.Extraction experiment RNA of the group and the control group was sequenced,bioinformatics analysis,GO function enrichment analysis,KEGG signal pathway enrichment analysis,differential gene verification,etc.,and differentially expressed lncRNAs were selected,and 10 up-regulated lncRNAs and 10 were selected.The down-regulated lncRNA was verified by qPCR,and the qPCR verification results of most genes were consistent with the analysis results;5.After cell transfection,compared with the negative control group si-NC group,MOYM3 of CNE-1 cells in si-AL group The expression level of E-cadherin decreased significantly,while the expression of N-cadherin,vimentin and Twist protein decreased,indicating that the transfected siRNA can effectively inhibit the expression of lncRNA AL591178.1.At the same time,it indicated that after interfering with the expression of lncRNA AL591178.1,the EMT process of CNE-1 cells was reversed;6.The rescue experiment further showed that the expression level of lncRNA AL591178.1 was up-regulated during the EGF process of CNE-1 cells induced by TGF-β1,E-cadherin expression level was significantly reduced,while the expression of N-cadherin,vimentin and Twist increased,promoted EMT of CNE-1 cells,inhibited the expression of lncRNA AL591178.1,E-cadherin expression level was significantly increased,while N-The expression of cadherin,vimentin and Twist is reduced and the EMT process is reversed.Conclusion:TGF-β1 can induce EMT in CNE-1 cells.LncRNA AL591178.1 is a potential key LncRNA gene in this process.Its expression is up-regulated during EMT in CNE-1 cells.Inhibiting its expression can effectively suppress the occurrence of EMT.