Effect Of TWIST2 Gene On Invasion And Migration Of SHG44 Maligant Glioma Cell Line

Objective:To detect the expression of Twist2 in human glioma cell lines,observe whether Twist2 affects the invasion and migration of malignant glioma cells,and explore whether Twist2 plays a role through Epithelial-mesenchymal transition(EMT).Methods:1.The expression of Twist2 in four malignant human glioma cell lines(U87,U118,H4,SHG44)was evaluated by q-PCR and Western blot.2.The SHG44 cell line with the highest Twist2 gene and protein expression level in the above cell line was selected as the research object.Malignant glioma cell,SHG44,was infected with Twist2-slienced plasmid via lentivirus.The experiment was divided into three groups: the first group was untreated(blank),the second group was negative control of silence(NC-sh),the third group was the silence of Twist2 gene(Twist2-sh).The expression of Twist2 gene and protein was evaluated by q-PCR and Western blot.3.Transwell invasion assay and migration assay were used to detect the invasion and migration ability of cells in each group.The CCK8 experiment,was to investgate the proliferation ability of different groups.Apoptosis assays by flow cytometry was investgate the cell cycles and the ability of apotosis.4.Immunofluorescence was used to detect the nuclear transfer of NF-kappa B-p65 protein in each group under confocal laser microscope.5.Western blotting detection of EMT-cadherin,N-cadherin,vimentin and fibronectin protein expression.Results:1.Twist2 mRNA and protein were expressed in human malignant glioma cell lines U87,U118,H4,and SHG44,and the expression level was highest in humanmalignant glioma SHG44 cell line.2.Silent Twist2 plasmid was successfully established.The level of mRNA and protein of Twist2 reached the requirements of each groups.3.The invasion and migration of the malignant glioma cells with a down-regulation of Twist2 by shRNA humanly were significantly reduced.4.The nuclear transfer of NF-kappa B-p65 protein with a down-regulation of Twist2 by shRNA humanly were no significantly changed.5.The influence of down-regulation the Twist2 gene take effects on the EMT-related protein.After silencing Twist2 gene,the protein expression level of E-cadherin was increased,while N-cadherin,Vimentin and Fibronectin was decreased.Conclusions:1.Twist2 mRNA and protein is expressed in a variety of human malignant glioma cell lines,such as U87,U118,H4 and SHG44.2.Twist2 can promote the invasion and migration of SHG44 malignant glioma cell line and may affect the invasion and migration of malignant glioma cells through EMT.3.Twist2 can enhance the anti apoptosis ability of malignant glioma cells.