Expression And Significance Of PRKAR2B In Serous Ovarian Cancer

BackgroundOvarian cancer has a poor prognosis due to difficulties in early diagnosis,chemotherapy resistance,and recurrence and metastasis.It is the gynecological malignant tumor with the highest fatality rate.Further research is urgently needed to find new early sensitive biomarkers or therapeutic targets.Chip technology based on high-throughput sequencing is a reliable method for studying tumors at present,and the integration and re-analysis of massive chip data can provide new and valuable insights for clinical research.The preliminary analysis of the ovarian cancer chipset in the TCGA and GEO databases found that PRKAR2B is closely related to the survival prognosis of ovarian cancer patients,and RRKAR2B can participate in cell differentiation,proliferation,transcription and other processes through the classic cellular pathway of cAMP-PKA.There is no literature report on the correlation between PRKAR2B and ovarian cancer at home and abroad,so this study intends to further collect clinical specimens to detect their expression and explore the effect of PRKAR2B on the biological behavior of ovarian cancer cells through in vitro cell function experiments.The ovarian cancer sensitive prognosis gene provides a theoretical basis.Objective:1.Screen the common differentially expressed genes through 5 sets of ovarian cancer chip data sets in TCGA and GEO databases,and determine the target research gene by comprehensive bioinformatics analysis.2.Detect the expression of PRKAR2B in serous ovarian cancer tissues and normal ovarian tissues from the level of protein and mRNA,and analyze the relationship between its expression level and the clinicopathological characteristics and survival prognosis of patients.3.Observe the effect of PRKAR2B overexpression on ovarian cancer cell proliferation,apoptosis,invasion and metastasis,and explore the effect of PRKAR2B on the biological behavior of ovarian cancer cells.Methods:1.First download the original chip data of ovarian cancer from the TCGA and GEO databases,select differentially expressed genes,and then use bioinformatics methods to conduct integrated analysis of the differential genes.The next five sets of differential genes are taken to obtain the common differentially expressed genes,and further analyzed the common differential genes for survival analysis and protein co-expression analysis to determine the target research genes.2.Collect clinical specimens and use immunohistochemical methods to detect the expression of PRKAR2B protein and PRKAR2B mRNA in serous ovarian cancer tissues and normal ovarian tissues.3.Construct PRKAR2B overexpression plasmid and transfect into SKOV3 ovarian cancer cells,and then use MTT method to detect cell proliferation,flow cytometry to detect apoptosis,and Transwell to detect cell migration and invasion.Results:1.A total of 7,050 DEGs were identified by comprehensive bioinformatics,of which 4043 genes were up-regulated and 3007 genes were down-regulated.Enrichment analysis suggests that DEGS is mainly located in extracellular regions and exosomes,and its molecular functions are mainly related to heparin binding and growth factor activity;biological processes are mainly related to Wnt signaling pathway and retinol metabolism.Protein interaction analysis revealed that PRKAR2B,ALDH1A1,EFEMP1,OGN,and CP are central genes with high connectivity.Survival analysis found that PRKAR2B,ALDH1A1,GFPT2,AQP9,CP are closely related to the survival prognosis of ovarian cancer patients.2.TCGA database analysis of PRKAR2B mRNA expression level in patients with ovarian cancer is closely related to histological grade and 5-year survival rate of ovarian cancer(P<0.05).3.The PRKAR2B protein is expressed in the cytoplasm of normal cells and is pale yellow to brownish yellow.The intensity of PRKAR2B staining in serous ovarian cancer tissue was significantly weaker than that in normal ovarian tissue.The positive expression rates of PRKAR2B protein in normal ovarian tissue and serous ovarian cancer tissue were 90.0%(18/20)and 10.0%(6/60),respectively,and the difference was statistically significant(χ2=45.71,P<0.001).4.The expression level of PRKAR2B mRNA in serous ovarian cancer is lower than that in normal ovarian tissue,and the difference in expression between the two is statistically significant(P=0.028).5.Western Blot and RT-qPCR experiments confirmed that the overexpressed PRKAR2B plasmid was successfully transfected into SKOV3 ovarian cancer cell line.The CCK8 cell proliferation experiment showed that compared with the NC group and the GFP-NC group,the cell proliferation rate of the PRKAR2B group was significantly reduced,and the cell suppression rate was significantly increased,and the differences were statistically significant(P<0.05);flow cytometry detection Apoptosis showed that the percentage of apoptotic cells in the PRKAR2B overexpression group was 8.19%,the percentage of apoptotic cells in the NC group was 3.31%,and the percentage of apoptotic cells in the GFP-NC group was 2.59%;Transwell cell migration and invasion experiments showed that:Compared with the GFP-NC group,PRKAR2B overexpression significantly inhibited the invasion and metastasis ability of SKOV3 ovarian cancer cells,the difference was statistically significant(P<0.05).Conclusion:1.PRKAR2B is a central gene with high connectivity and is closely related to the prognosis of ovarian cancer,and the overall survival of ovarian cancer patients with high PRKAR2B expression level is longer.2.The expression of PRKAR2B at the protein and mRNA levels in serous ovarian cancer tissues is lower than that in normal ovarian tissues.3.Overexpression of PRKAR2B can promote SKOV3 ovarian cancer cell apoptosis and inhibit its proliferation,invasion and metastatic ability.