The Expression Of CCNB2 In Gallbladder Cancer And Its Effect On Cell Proliferation And Migration

BackgroundGallbladder Cancer(GBC)is the sixth most common gastrointestinal malignancy worldwide,characterized by rapid progression,poor prognosis,and high mortality.Surgery is the first choice for clinical treatment,but with occessive onset and atypical clinical manifestations,most of the patients with GBC are in the advanced stage and have lost the opportunity of surgical excision.Therefore,in addition to screening early diagnosis,still need to actively explore the biological characteristics of gallbladder cancer,to find other treatment means.PurposeTo investigate the expression of Cyclin B2(CCNB2)in gallbladder carcinoma and its relationship with clinicopathology,as well as the effect of CCNB2 on the proliferation and migration of gallbladder carcinoma cells.Methods1.Gene Expression Profiling Interactive Analysis(GEPIA)database was used to analyze the differential expression of biliary tract carcinoma and paracancer genes.Immunohistochemistry(IHC)was used to detect CCNB2 expression in gallbladder cancer and negative incisal margin tissues.The relevant mRNA was detected by real-time quantitative PCR(qRT-PCR),and CCNB2 protein was detected by Western Blot(WB)for statistical analysis;2.From 2017 to 2020,31 clinical samples after radical resection of gallbladder cancer were collected in Jiangxi Provincial People’s Hospital,and the clinicopathological data of the corresponding patients were statistically analyzed;3.The results of qRT-PCR and WB showed that the expression level of CCNB2 mRNA and protein in NOZ and SGC cell lines was significantly higher than that in GBC and EHGB-1 cell lines,and that sh RNA-CCNB2 stable cell lines of gallbladder carcinoma were successfully constructed.;4.Through in vivo and in vitro experiments,such as CCK8,plate colony formation,and subcutaneous tumor formation in nude mice to detect cell proliferation activity,Transwell chamber assay to detect cell migration ability,flow cytometry to detect cell cycle changes,and Western blot experiments to verify the changes in biological function-related proteins.Results1: Bioinformatics predicted that CCNB2 was upregulated in biliary tract carcinoma.The expression level of CCNB2 in GBC tissues was higher than that in paracancer tissues(p<0.05).The high expression level of CCNB2 was positively correlated with the number of lymph node metastasis,but had no significant relationship with tumor size,nerve invasion and TNM stage;2: The results of qRT-PCR and WB showed that the expression level of CCNB2 mRNA and protein in NOZ and SGC cell lines was significantly higher than that in GBC and EHGB-1 cell lines,and that sh RNA-CCNB2 stable cell lines of gallbladder carcinoma were successfully constructed;3: Experiments in vivo and in vitro proved that the proliferation and migration of gallbladder cancer cells were inhibited after knocking down CCNB2,and the cell cycle arrest was blocked.The expression of two cell cycle arrest proteins,p21 and p27,was significantly increased,but the expression of Cyclin D1 and CDK4 was reduced.,EMT-related proteins did not change significantly.ConclusionsCCNB2 was highly expressed in gallbladder carcinoma,which was positively correlated with the number of metastatic lymph nodes.CCNB2 can promote the proliferation and migration of gallbladder carcinoma cells,and the related mechanism is that CCNB2 can affect the Cyclin D1/CDK4/p21/p27 pathway,resulting in the stagnation of gallbladder carcinoma cells in G2/M phase,which is expected to be a new gene target for the diagnosis and treatment of gallbladder carcinoma.