| Objective: Gliomas are one of the most common primary intracranial tumors in adults.Although the comprehensive treatment strategies of surgery,chemotherapy,radiotherapy and molecular targeted therapy have been well developed in recent years,the prognosis of patients with gliomas has not been significantly improved and the median survival time is only 15 months.Research on molecular targeting has become one of the hot topics in the therapeutic study of gliomas.This study aimed to investigate whether micro RNA mi R-150 and RNA binding protein DAZAP1 can regulate the malignant biological behaviors of glioma cells,and further analyze its possible molecular mechanism,namely that mi R-150 inhibits MMP-9 by down-regulating FOXB1 and the expression of XIAP inhibited the malignant biological behavior of glioma cells.DAZAP1 down-regulated the expression of FOXE1 by stabilizing the expression of long-chain non-coding RNA00598(Linc00598)and inhibited the malignant behavior of glioma cells.This study aims to provide a new basis for the development of glioma,and at the same time provide a new target for molecular targeted therapy of glioma.Methods: The human glioma cells U87,U251,human normal astrocytes were cultured.Real-time PCR was used to detect mi R-150,FOXB1,DAZAP1,Linc00598 and FOXE1.Western blot was used to detect the expression levels of FOXB1,DAZAP1 and FOXE1 in normal brain tissue,glioma tissue and human normal astrocytes.CCK-8 was used to detect cell proliferation.Transwell was used to detect changes in cell migration and invasion.Flow cytometry and PI-FITC double staining were used to detect apoptosis.A cell line transfected with mi R-150,FOXB1,DAZAP1,Linc00598,FOXE1 overexpressing and silencing expression was constructed.The expression of XIAP,MMP9,FOXE1,and MARK4 m RNA was detected by Real-time PCR.The expression of XIAP,MMP9,FOXE1,MARK4,STAU1,UPF1,LATS,p-LATS,MST,p-MST,YAP and p-YAP antibody proteins was detected by Western Blot.The dual luciferase gene reporter system was assayed for the targeted binding of mi R-150 and FOXB1,Linc00598 and FOXE1.Actinomycin D-3 induction assay was used to determine the change in half-life of Linc00598.The binding of DAZAP1 to Linc00598,Linc00598 and FOXE1 were verified by RIP and RNA-pulldown experiments.The direct interaction of FOXB1 with MMP9 and XIAP,FOXE1 and MARK4 promoters was detected by immunoprecipitation.The effects of mi R-150 and FOXB1 on the growth of transplanted tumors and the survival time of tumor-bearing nude mice in nude mice were examined.Results: The expression of mi R-150 in glioma tissue was significantly lower than that in the peritumoral and normal brain tissues,and it decreased with the increase of the pathological grade of glioma.The expression of mi R-150 in U87 and U251 cells was also significantly lower than normal human brain glial cells.Overexpression of mi R-150 significantly inhibited the proliferation,migration and invasion of U87 and U251 cells,and promoted apoptosis.Silencing of mi R-150 significantly enhanced the proliferation,migration and invasion of U87 and U251 cells,and inhibited cell apoptosis.The expression of FOXB1 was up-regulated in glioma tissues,and increased with the increase of the pathological grade of gliomas.The expression of FOXB1 in U87 and U251 cells was also significantly higher than that of normal human brain glial cells.Overexpression of FOXB1 significantly enhanced the proliferation,migration and invasion of U87 and U251 cells,and inhibited cell apoptosis.Silencing FOXB1 significantly inhibited the proliferation,migration and invasion of U87 and U251 cells,and promoted apoptosis.There is a targeted binding of mi R-150 to FOXB1.Overexpression of mi R-150 negatively regulates FOXB1,decreases the expression of MMP9 and XIAP,and inhibits the malignant biological behavior of U87 and U251 cells.There is binding of FOXB1 to the promoter regions of MMP9 and XIAP.Overexpression of mi R-150 in combination with silencing of FOXB1 significantly inhibited the growth of xenografts in nude mice and prolonged the survival of tumor-bearing nude mice.DAZAP1 is endogenously expressed in glioma tissues and cells and is significantly down-regulated in glioma tissues and cells.Overexpression of DAZAP1 significantly inhibited the proliferation,migration and invasion of glioma cells and promoted apoptosis.Linc00598 is endogenously expressed in glioma tissues and cells and is significantly down-regulated in glioma tissues and cells.Overexpression of mi R-217 significantly inhibited the proliferation,migration and invasion of glioma cells,and promoted apoptosis.There is a targeted binding between DAZAP1 and Linc00598.Overexpression of DAZAP1 or overexpression of Linc00598 decreased FOXE1 expression.FOXE1 was endogenously expressed in glioma tissues and glioma cells and was significantly up-regulated in glioma tissue and glioma cells.Linc00598 binds to FOXE1 through the SBS site.Overexpression of Linc00598 inhibits the proliferation,migration and invasion of glioma cells by promoting the degradation of FOXE1 m RNA and promotes apoptosis.FOXE1 binds to the promoter region of MARK4,increasing its promoter activity.Silencing FOXE1 negatively regulates the expression of MARK4,inhibits the proliferation,migration and invasion of glioma cells and promotes apoptosis.Overexpression of Linc00598 negatively regulates the expression of FOXE1,decreases the expression of MARK4,regulates the activity of Hippo pathway,and then regulates the malignant biological behavior of glioma cells.Conclusions: 1.mi R-150 is lowly expressed in glioma tissues and U87 and U251 cells.Overexpression of mi R-150 reduces the proliferation,migration and invasion of glioma cells and promotes apoptosis.FOXB1 was highly expressed in glioma tissues and U87 and U251 cells.Silencing of FOXB1 inhibited the proliferation,migration and invasion of glioma cells and promoted apoptosis.2.MMP9 and XIAP are target genes for FOXB1.Over-expression of mi R-150 in U87 and U251 cells negatively regulates the expression of FOXB1,thereby regulating the expression of MMP9 and XIAP and inhibiting the malignant biological behavior of U87 and U251 cells.3.DAZAP1 and Linc00598 were lowly expressed in glioma tissues and U87 and U251 cells.Overexpression of DAZAP1 and overexpression of Linc00598 inhibited the proliferation,migration and invasion of glioma cells and promoted apoptosis.FOXE1 was highly expressed in glioma tissues and U87 and U251 cells,and silenced FOXE1 inhibited the malignant biological behavior of glioma cells.4.Overexpression of DAZAP1 in U87 and U251 cells increased the stability expression of Linc00598.Up-regulated Linc00598 promoted the degradation of FOXE1 m RNA through SMD pathway,decreased the expression of FOXE1,and inhibited the malignant biological behavior of glioma cells.5.FOXE1 binds with MARK4 promoter.Silencing FOXE1 inhibits the malignant biological behavior of glioma cells by downregulating MARK4 expression and promoting the activity of the Hippo pathway. |
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