Analysis And Detection Of KPNA2 Gene Expression And Its Correlation With Hepatocellular Carcinoma

Chapter 1 This chapter elaborated the detection methods,principle,advantages and disadvantages and the experimental optimization of nucleic acids and proteins and the significance in biomedical research.The structure,KPNA2 functions and research progress of the nuclear transport protein family are reviewed.Chapter 2 Nuclear transporter KPNA2 regulates some macromolecular substances for nuclear and cytoplasmic transport,and maladjustment of cell transport system is often observed in the process of tumorigenesis.Immunohistochemical detection was used to study the tissue of 100 cases of hepatocellular carcinoma(HCC)and 20 cases of adjacent tissues,and it was found that KPNA2 expression in hepatocellular carcinoma tissues was significantly higher than that in normal tissues.Further application of polymerase chain reaction(policy real-timepolymerase chain reaction,q RT-PCR)technology for 4 kinds of hepatocellular carcinoma cell line(SMMC-7721 and Hep G2,Hep3 B and Huh-7)m RNA KPNA2 gene in quantity detection,the results showed KPNA2 in 4 seedlings are highly expressed in liver cancer,suggests KPNA2 may have a close relationship with the incidence of hepatocellular carcinoma.Chapter 3 Gene silencing technology can be used to to study the gene function of the organism.RNA interference has high efficiency and specificity of gene expression suppression.Here,we chose a lentivirus-mediated short-hairpin RNA(sh RNA)strategy to inhibit human KPNA2 expression in two human hepatocellular carcinoma cell lines,Hep G2 and SMMC-7721,to explore the impact of KPNA2 on cell growth and survival in vitro.MTT assay was used to detect cell proliferation in the two groups.Cell proliferation was significantly inhibited in the experimental group after the silencing of KPNA2 compared with the control group.Flow cytometry was used to detect the cell cycle and apoptosis of the two groups of HCC cells.After the silencing of KPNA2 gene,hepatocellular carcinoma cells were blocked in the G2/M phase and S phase of the cell cycle,and mitosis could not be carried out normally.After silencing the expression of KPNA2 gene in hepatocellular carcinoma cells,the apoptosis rate was significantly higher than that in the control group,suggesting that KPNA2 may play a role in hepatocellular carcinoma cells by affecting cell cycle and apoptosis.Chapter 4 In order to further confirm the biological function of KPNA2,Hep G2 cells were used as tumor forming cells,and the experimental group of hepatocellular carcinoma cells with KPNA2 gene knockdown and the control group of normal hepatocellular carcinoma cells were respectively inoculated under the skin of nude mice to establish the model of transplanted hepatocellular carcinoma.Tumorigenesis was observed every day,tumor size was measured,and tumor weight was measured after the nude mice were killed.Results the tumor size of nude mice in the experimental group was significantly smaller than that in the control group,indicating that targeted silencing of KPNA2 gene significantly inhibited the tumorigenic ability of nude mice in hepatocellular carcinoma cells.In vivo experiments also proved that KPNA2 was closely related to the occurrence and development of hepatocellular carcinoma.Chapter 5 Gene bioinformatics analysis and KPNA2 biological functional mechanism analysis,the KPNA2 gene in hepatocellular carcinoma.cells were introduced into Affymetrix human gene expression profile chip for detection,and the differentially expressed genes were analyzed.Compared with the control group,there were 647 up-regulated genes and 938 down-regulated genes in the experimental group.IPA bioinformatics analysis software was used to analyze the enrichment of differentially expressed genes,and the results suggested gene enrichment of the cell cycle pathway at the G2/M phase and control of chromosomal replication(S phase).Qrt-pcr and Western blot were used to test and verify the expression of key genes,and the results were consistent with the results of IPA differentially expressed genes classic pathway analysis.Finally,it is speculated that KPNA2 gene may play a role in inhibiting tumor cell proliferation and inducing apoptosis by affecting the cell cycle signal pathway of G2/M and S phases.