Exploration For Expression Of NPTX2 In Clear Cell Renal Cell Carcinoma And Its Clinical Significance

Objective1.Bioinformatics was used to analyze genes with significant differences in clear cell renal cell carcinoma(ccRCC)and to obtain the target gene.2.To focuse on the preliminary exploration for the expression of neuronal pentraxin 2(NPTX2)gene in ccRCC and its clinical significance.Materials and methods1.Bioinformatics analysis of ccRCC-related differential genes1.1 Download a ccRCC gene chip data fromthe NCBI-GEO database,standardize the pretreatment with RMAfunction by using R language “affy” package,and then obtain differentially expressed genes by using R language“limma”package.Finally,a list of significantly differentially expressed genes(|logFC|>2 and student's t test with a FDR< 0.05)could be obtained.1.2 GO functional analysis(BP,CC,MF)and KEGG pathway enrichment analysis were performed on the list of significantly differentially expressed genes by using R language“clusterProfiler” package.1.3 Obtain the target gene by analyzing correlation and research significance.2.Exploration for Expression of NPTX2 in Clear Cell Renal Cell Carcinoma and Its Clinical SignificanceThe expression of NPTX2 was detected by qPCR,western blot or immunohistochemistry in ccRCC cell lines and tissues.2.1 Cultured cell lines in vitro(human renal tubular normal epithelial cell line HK-2,human renal clear cell carcinoma primary tumor cell line 786-O,and pleural fluid metastasis cell line ACHN): The expression of NPTX2 mRNA and protein were detected by qPCR and western blot in normal and cancer cell lines,and their expression differences in both types of cell lines were analyzed.2.2 Tissue samples:(1)cDNA microarray(15 cases of paired ccRCC cancer and Paracancer): The expression of NPTX2 mRNA was detected by qPCR in both tissues,and their expression differences were analyzed.(2)Fresh tissues(4 cases of paired ccRCC cancer and Paracancer): The expression of NPTX2 protein was detected by western blot in both tissues,and their expression differences were analyzed.(3)Paraffin embedded tissues(70 cases of paired ccRCC cancer and Paracancer): The expression of NPTX2 protein was detected by EnVsioni method of immunohisto chemistry in both tissues,and their expression differences were analyzed,and it was compared with major clinical information and prognoses of those patients.Results1.Bioinformatics analysis of ccRCC-related differential genes1.1 In the GEO analysis,a total of 9550 ccRCC-related differentially expressed genes were obtained,of which 832 were significantly different(up 438,down 394).1.2 GO and KEGG enrichment analysis found that many significantly differentially expressed genes in ccRCC play an important role in controlling and altering tumor-related metabolic activities,cell adhesion,signal transduction,inflammation/immune response.1.3 Obtain the target gene NPTX2.2.Exploration for Expression of NPTX2 in Clear Cell Renal Cell Carcinoma and Its Clinical Significance2.1 There was a statistical significance of difference between normal cell lines/tissues and ccRCC cell lines/tissues in the expression of NPTX2 mRNAand protein(P<0.05).2.2 The expression of NPTX2 protein was correlated with the WHO/ISUP pathological grade of ccRCC tumors(P<0.05),but it could not be considered related to the gender,age,TNMstage and tumor size of patients(P>0.05).2.3 Kaplan-Meier univariate analysis and log-rank test showed that the expression of NPTX2 protein was related to the survival of patients(P=0.04): The high-expressiongroup had lower survival rate than the low-expression group,and the prognosis was poor.ConclusionNPTX2 gene is over-expressed in ccRCC carcinoma tissues,which is associated with the WHO/ISUP pathological grade of ccRCC tumors and the prognosis of ccRCC patients.It can be used for the diagnosis and prognosis of ccRCC as a potential biomarker.