Mechanism Study Of CSTF2 In Regulating CCNB1 Expression And Affecting Hepatocellular Carcinoma Cell Proliferation

Objective:To analyze the expression of cleavage stimulation factor subunit 2(CSTF2)in the ICGC and TCGA hepatocellular carcinoma(HCC)databases and the relationship between survival and prognosis by bioinformatics and to detect the expression of CSTF2 in hepatocellular carcinoma tissues and cells.Then to clarify the effects of CSTF2 expression changes on proliferation of hepatocellular carcinoma cells.Finally,to explore the mechanism of CSTF2 regulating Cyclin B1(CCNB1)expression on hepatocellular carcinoma proliferation.Methods:1.The expression of CSTF2 in HCC and its relationship with prognosis of HCC patients were analyzed by using the public database in ICGC and TCGA;2.The m RNA and protein expression of CSTF2 in HCC tissues were detected by real-time quantitative PCR,immunohistochemistry and Western blotting;the m RNA and protein expression levels of CSTF2 in HL7702,HCCLM3,SKHep1,Hep3 B and Hep G2 were detected by real-time quantitative PCR and Western blotting;3.Cell proliferation and cell cycle changes after interference and overexpression of CSTF2 were detected by CCK-8,cloning formation and flow cytometry,respectively;4.To predict pathways or functions that CSTF2 may affect in HCC by GSEA in TCGA.The correlation between CSTF2 and CCNB1 expression in HCC was analyzed by ICGC and TCGA public database,and the m RNA and protein expression changes of CCNB1 after changing the expression of CSTF2 in HCC cells were observed by real-time PCR and Western blotting method;5.Overexpression of CCNB1 in SKHep1 of hepatocellular carcinoma cell line SKHep1 that stably interferes with CSTF2 reach,or overexpression of CSTF2 in HCCLM3 of hepatocellular carcinoma cell line HCCLM3,the protein expression level and cell proliferation capacity of CSTF2 and CCNB1 were observed.Results:1.In the public database of ICGC and TCGA,the expression of CSTF2 in HCC was significantly higher than normal liver tissue,and high expression of CSTF2 is an independent risk factor for poor prognosis in HCC patients;2.The expression of CSTF2 in HCC tissues and cell lines was significantly increased;3.Cell proliferation ability weakened and G2 stage blocked after interference with CSTF2,and cell proliferation ability increased after overexpression of CSTF2,and G2 stage transition accelerated;4.According to KEGG and HALLMARK analysis,CSTF2 is associated with the G2/M phase of the cell cycle.In the public database of ICGC and TCGA,the expression of CSTF2 and CCNB1 was significantly positively correlated;after interfering or overexpressing the expression of CSTF2 in HCC cells,the m RNA and protein expression levels of CCNB1 were also downregulated or upregulated accordingly;5.CCNB1 is a key effector molecule for CSTF2 to regulate the proliferation of hepatocellular carcinoma cells.Conclusion:CSTF2 is upregulated in HCC and is associated with poor prognosis.In HCC cells,CSTF2 positively regulates the expression of CCNB1,which in turn affects the proliferation ability of liver cancer cells.