A Study On The In Vivo Experiment And Molecular Mechanism Of The Effect Of TMEM106B On Proliferation And Angiogenesis Of Malignant Meningioma

Objective:This experiment aims to verify the effect of TMEM106 B on the proliferation and angiogenesis of malignant meningioma cells through in vivo experiments,and to explore the possible molecular mechanism.Methods:1.Interference with TMEM106 B gene in two cell lines of malignant meningioma,IOMM-Lee and CH157.Each cell line was divided into three groups:interference with TMEM106 B was the experimental group(TMEM106B-sh),transfection with unrelated lentivirus was the negative control group(NC-sh),and Blank control group(Blank).2.The tumor growth time curves of two malignant meningioma cell lines,IOMM-Lee and CH157,were drawn in vivo,and the tumor inhibition rate was calculated.The expressions of ki-67 and VEGF in the tumors of each group were detected by immunohistochemistry,and the microvascular density(MVD)was calculated by labeling CD34,so as to compare the cell proliferation and angiogenesis of each group.3.General software predicted and analyzed the binding sites between TMEM106 B gene promoter and transcription factor JUN,and constructed overexpressed plasmids of TMEM106 B reporter gene wild-type,mutant plasmids and transcription factor JUN,and verified whether transcription factor JUN could specifically bind to TMEM106 B promoter through dual-luciferase reporter gene experiment.4.Chromatin immunoprecipitation(CHIP)assay further verified the specific binding between TMEM106 B gene promoter and JUN transcription factor.Results:1.After lentivirus transfection interfered with the expression of TMEM106 Bgene,q-PCR and Western blot detection showed that the protein and mRNA expression levels of TMEM106 B in malignant meningioma cell lines decreased significantly,meeting the grouping requirements.2.In the tumorigenesis experiment of nude mice,in the TMEM106 B interference group,the tumorigenesis ability of nude mice was weakened and the tumor proliferation slowed down.Immunohistochemical staining results showed that the expression levels of TMEM106 B,ki-67 and VEGF were significantly decreased,with statistically significant differences.The expression level of CD34 in each group was labeled and the microvascular density(MVD)of each group was analyzed.The results showed that MVD decreased after TMEM106 B was down-regulated.3.The software predictive analysis results showed that there were 7 JUN binding sites of transcription factors in the promoter region of TMEM106 B gene,and the results of dual-luciferase reporter gene experiment showed that JUN could specifically bind to the promoter region of TMEM106 B.4.The results of chromatin immunoprecipitation showed that TMEM106 B promoter could bind to the transcription factor JUN.Conclusion:1.TMEM106 B can inhibit the proliferation and angiogenesis of malignant meninges;2.The promoter region of TMEM106 B has a binding site with JUN,a transcription factor.