Mechanisms Of PAIP1 In Hepatocarcinogenesis

Objective: To detect the expression of PAIP1 in hepatocellular carcinoma(HCC)and HCC cell lines,and to investigate the relationship between the expression level of PAIP1 and the clinicopathologic parameters of HCC.To investigate the effect of down-regulation of PAIP1 gene expression on hepatocellular carcinoma cell line SMMC-7721 in vitro and in vivo.To explore the mechanism of PAIP1 in hepatocarcinogenesis.Methods: Part I Detection of PAIP1 expression in HCC and HCC cell lines.The expression of PAIP1 protein in hepatocellular carcinoma tissue was detected by immunohistochemical method.The expression level of PAIP1 in hepatocellular carcinoma tissues and adjacent tissues was compared with that in adjacent tissues.The relationship between the expression of PAIP1 and the clinicopathological parameters was analyzed.The expression of PAIP1 mRNA in hepatocellular carcinoma cell lines was detected by real-time fluorescent quantitative PCR(RT-PCR).Part ? Down-regulation of PAIP1 expression may affect the biological behavior of hepatoma cells in vitro and in vivo.The expression of PAIP1 was down-regulated by RNA interference.To study the biological behavior of SMMC-7721 hepatocarcinoma cell line with PAIP1 down-regulated.The cell cycle and apoptosis of SMMC-7721 cells were analyzed by flow cytometry.The clonogenic ability of SMMC-7721 cells was observed by clone formation assay.The growth of SMMC-7721 cells was detected by xenograft model in nude mice.Part ? Screening and verifying the differentially expressed genes after PAIP1 gene expression silencing.The downstream response gene after PAIP1 knockdown were screened by whole-gene expression microarray.Bioinformatics analysis was performed to detect the key genes related to cell cycle regulation by Western blotting.Results: Part I Detection of PAIP1 expression in HCC and HCC cell lines.The expression level of PAIP1 in hepatocellular carcinoma tissue was significantly higher than that in para-cancerous tissue,the expression level of PAIP1 had no significant correlation with sex,age,tumor size and TNM staging of liver cancer,but the pathological grade was significantly correlated with the survival rate of patients.In addition,PAIP1 mRNA was also highly expressed in hepatoma cell lines.Part ? Down-regulation of PAIP1 expression may affect the biological behavior of hepatoma cells in vitro and in vivo.After the expression of PAIP1 was down-regulated,the proliferation ability of hepatoma cells was decreased and the apoptosis rate was increased.In S phase and G1 phase,the cells had no significant changes,but the cells in G2 phase were increased,the ability of clone formation was weakened in vitro,nude mice in vivo tumor formation capacity weakened.Part ? Screening and verify the differentially expressed genes after PAIP1 gene expression silencing.The results of cDNA microarray showed that 333 genes were changed in SMMC-7721 cells after PAIP1 knockdown after ablation with FC> 2.0 or FC <0.5,P<0.05.The up-regulated genes were 134,the down-regulated genes were 199.The KEGG and BIOCARTA pathways were used to identify the 10 signal transduction pathways of the enriched differential genes.Western blotting results showed that the expression of CCND1,CCND2,CDK6 and MDM2 were changed significantly after PAIP1 down-regulated.CONCLUSION:(1)PAIP1 is highly expressed in hepatocellular carcinoma and hepatocarcinoma cell lines,which may play a role in the development and progression of hepatocellular carcinoma.(2)Down-regulation of PAIP1 can significantly inhibit the proliferation of hepatocellular carcinoma cells,promote cell apoptosis,inhibit the formation of in vitro clones,change the cell cycle period,inhibition of tumorigenesis in vivo.(3)PAIP1 plays an important role in the development of hepatocellular carcinoma by affecting the key genes that regulate cell cycle.