| Objective: To screen out the core genes and signaling pathways related to the development of glioblastoma(GBM)through bioinformatics analysis,investigate the expression of CEP55 in GBM cells and explore the roles of CEP55 in proliferation,invasion and apoptosis of GBM.Methods:(1)The RNA-seq data of GBM gene expression file were downloaded from TCGA database,GSE4290,GSE50161 dataset and their common GPL570 platform file were downloaded from GEO database.Perl programming language was adopted to convert the gene probe names into gene symbol.The differentially expressed genes(DEGs)between GBM samples and adjacent non-tumor tissues were identified through Limma package in R language.Then we collected the common differentially expressed genes(c DEGs)among three datasets via Venn analysis,GO annotation and KEGG pathway enrichment analysis were performed through online DAVID database to study the function and involved pathways of the c DEGs.Protein-protein interaction(PPI)network was constructed via STRING,and the correspond results were imported to Cytoscape for visualization,core genes in the network were screened out by cyto Hubba plug-in,K-M survival analysis were performed to sift the prognosis related core genes,combining the results of literature research we determined the target gene and calculated the Pearson correlation coefficient to evaluate its correlation with other core genes,then experiments were designed to explore the biological functions of target gene in glioblastoma.(2)U251,U118 cell line were selected for constructing the CEP55-silence stable cell lines through lentivirus technology,q RT-PCR was used to evaluate the efficiency of transfection.EDU cell proliferation assay,Transwell cell invasion assay and flow cytometry were applied to explore the effects of knocking down CEP55 on the cell proliferation,invasion and apoptosis of glioblastoma cells.Results:(1)Differential analysis indicated that there are 2932(1341 up-regulated,1591 down-regulated),2039(816 up-regulated,1223 down-regulated),823(212 upregulated,611 down-regulated)DEGs between tumors and adjacent non-tumor tissues in TCGA,GSE50161,GSE4290 dataset respectively,552 common DEGs from three datasets were finally determined via Venn analysis,including 128 up-regulated genes and 424 down-regulated genes.Up-regulated genes were mostly related to extracellular matrix,cell cycle,cell mitosis,intercellular adhesion,p53 signaling pathway and other relevant biological processes,while down-regulated genes mainly participated in synaptic transmission,transmission of nerve impulse,calcium signaling pathway,Erb B signaling pathway,MAPK signaling pathway and other biological processes.(2)The PPI analysis constructed a complex regulatory network composed of 410 nodes and 1890 edges,where 20 hub genes were screened out through cyto Hubba.Those hub genes were CALB1,CDC20,CDCA8,CDK1,CEP55,DLGAP5,KIF20 A,KIF4A,NDC80,PBK,RRM2,SYN1,SYP,SYT1,TPX2,TTK,VEGFA,BDNF,GNG3 and TOP2 A,they were mainly enriched in cell cycle,cell mitosis and P53 signaling pathway.Survival analysis indicated the expression of CEP55 and RRM2 was associated to the prognosis of GBM patients,high expression of CEP55 or RRM2 usually comes with a poor outcome.Literature research revealed that the explorations about RRM2 in glioblastoma had been reported already,while the CEP55 has not.Hence,we selected the CEP55 for further functional experiments.(3)Data from CCLE and GEPIA database verified that the expression of CEP55 was abnormally high in GBM in the level of both cell and tissue.(P<0.05).(4)After knocking down the expression of CEP55,the proliferation and invasion capacities of GBM cell were decreased importantly,while the proportion of apoptosis were increased markedly.Conclusions:(1)there are genetic and epigenetic differences between GBM tissues and normal tissues,the abnormal regulation of multiple genes may promote the occurrence and development of GBM through different mechanisms.(2)CEP55 is abnormally up-regulated in GBM,and it occupies a pivotal position in the DEGs’ regulation network of GBM.Higher expression of CEP55 is usually associated with shorter survival time.(3)CEP55 participated in the regulation of mitosis,cell cycle and other biological process in GBM along with KIF4 A,KIF20A,CDK1 and other core genes.(4)Knocking down CEP55 can inhibit the proliferation and invasion of GBM cells while promote apoptosis of GBM cells. |
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