Based On TGF-β/Smads Signaling Pathway GPR34 Promotes The Proliferation And Malignant Phenotype Of Glioma

ObjectiveThis topic mainly studies that GPR34 activates the TGF-β/Smad signal pathway and promotes the proliferation and malignant phenotype in order to provide a new basis for the clinical treatment of glioma.MethodsThe TCGA and GEO databases were used to analyze the expression of GPR34 in glioma tissues and the survival time of patients.Select 60 glioma samples for immunohistochemical test to detect the difference of GPR34 expression in different grades of glioma tissues,and analyze the relationship between the clinicopathological parameters of glioma and GPR34 expression.The expression of GPR34 protein in glioma tissue was detected by Western blot in 30 cases of tumor tissue and 10 cases of brain trauma decompression tissue.Cultured glioma U251,TG905,PT2,SF295,A172 cell lines,Western blot verified the expression of GPR34 in each cell line,used lentivirus particles to transfected U251 glioma cells with the highest expression,Western blot verified the expression of GPR34 protein at the cell level after knockout,CCK-8,cell scratch,Transwell experiment verified the effect of GPR34 on the proliferation,migration,invasion and other behaviors of glioma cells.The relationship between the expression of GPR34 and the prognosis of patients with glioma was analyzed by drawing the survival analysis curve of patients with glioma.The KEGG gene enrichment analysis was carried out using bioinformatics software to predict the molecular mechanism and related signal pathway that GPR34 protein may participate in.Meanwhile,the effect of GPR34 on the EMT process of tumor cells was also studied.Results1.The bioinformatics analysis of the TCGA database indicated that there was significant difference in the expression of GPR34 in brain tissue and glioma tissue.The expression level of GPR34 in glioma was much higher than that in normal tissue,and the expression of GPR34 was up-regulated with the grade of glioma.The OS and DFS of patients with high GPR34 expression were shorter than those of patients with low expression(P<0.001).2.Immunohistochemical experiments showed that the number of GPR34-positive cells in high-grade glioma tissue was significantly higher than that in low-grade glioma tissue and normal brain tissue,and was significantly correlated with the proliferation index mutant P53(P=0.022).Western blot results showed that the expression of GPR34 in glioma was up-regulated with the increase of WHO grade(P=0.005).3.High expression of GPR34 was significantly correlated with proliferation index mutant P53(P=0.061)and WHO grade(P=0.012).Regression analysis results showed that WHO grade,P53 and GPR34 were independent prognostic indicators(P<0.05).Kaplan-Meier survival analysis showed that the survival time of patients with low GPR34 expression was significantly higher than that of patients with high expression(P<0.001).4.Western blot verified the expression of GPR34 in each cell line,and the highest expression was found in U251.After transfection of U251 using lentiviral particles,Western blot results showed that silencing GPR34 expression significantly inhibited the proliferation,migration and invasive ability of glioma cell lines.5.The results of gene enrichment analysis predicted 12 signaling pathways that GPR34 may be implicated in the regulation of in glioma,focusing on the regulation of DNA repair,TGF-β/Smads signaling pathway.It provides a theoretical basis for the molecular mechanism of GPR34 in glioma.6.GPR34 promotes EMT、the TGF-β/Smads signaling pathway and malignant phenotype of glioma.ConclusionThe expression of GPR34 protein in glioma tissue is up-regulated,which is closely related to the WHO grade of glioma tissue.The overall survival time of glioma patients with high expression of GPR34 is shortened,and its expression level can be used as a marker to judge the prognosis of glioma.After down-regulation of GPR34,the proliferation,migration and invasion of glioma cell lines were significantly inhibited,and the signal pathway or target of GPR34 involved in regulation in glioma was predicted by gene enrichment analysis.Meanwhile,GPR34 could promote EMT in glioma cells.