CREB3 Inhibits Proliferation,migration And Invasion Of Glioma Cells By Down-regulating FAK Phosphorylation In Vitro

Objective:To investigate the effect of CREB3(c AMP-response element binding protein 3)on the expression of p-FAK(Phosphorylation-Focal Adhesion Kinase)protein,regulate the migration and invasion of glioma U87 and U251 and explore the role of CERB3 as a therapeutic target in the malignant process of glioma,providing a basis for glioma treatment.Methods:The m RNA expression levels of CREB3 and FAK and their survival rates were obtained in the Chinese Glioma Genome Atlas(CGGA)database.Co-immunoprecipitation(Co-IP)was used to verify the interaction between FAK and CREB3,and immunohistochemistry(IHC)was used to verify the expression levels of CREB3 and p-FAK(397)proteins in different grades of human glioma tissues.CREB3 lentiviral knockdown models of U87 and U251 were constructed.The optical density(OD value)of the cells at 450 nm was measured using a cell proliferation assay(Cell Counting Kit-8,CCK-8)and the plate cloning assay was used to verify the proliferation ability of the cells after knockdown of CREB3 by knockdown lentivirus;the migration and invasion ability of the knockdown cell lines were verified using WB,cell scratch,and transwell assay.Western blotting(WB)was used to verify the expression of p-FAK(397)and related proteins on its pathway as well as related apoptotic proteins.Results:In the CGGA database,CREB3 expression was found to be up-regulated,while high FAK and CREB3 expression indicated decreased patient survival.The experimental results of Co-IP indicate an interaction between CREB3 and FAK.The results of IHC indicated that the expression of CREB3 and p-FAK(397)also increased with increasing glioma grade.Thus,cell models of U87 and U251 with stable knockdown of CREB3 by lentivirus were constructed,and a series of cell function experiments such as transwell,cell scratch assay,CCK-8 plotting cell proliferation curve,and plate cloning showed that knockdown of CREB3 in U87 and U251 cell lines could inhibit glioma proliferation,migration,and invasion in vitro.WB results suggested that the expression levels of p-FAK(397)and p-AKT(308)could be down-regulated after CREB3 knockdown relative to the lentiviral unloaded group,and increased Bcl-2 anti-apoptotic protein expression as well as increased BAX pro-apoptotic protein expression as metastasis-related E-cadherin protein expression,and decreased vimentin protein expression.CONCLUSION:In this study,CREB3 may be involved in glioma formation and tumor cell growth by regulating the expression of p-FAK through in vitro experiments,and CERB3 can be a potential target to provide a basis for the diagnosis and treatment of glioma therapy.