Effects of CAAP1 on Proliferation, Migration and Invasion of Hepatoma Cell Line SMMC-7721

  Objective  To investigate the effect of caspase activity and apoptosis inhibitor 1 (CAAP1) on the proliferation, migration and invasion of hepatoma cell SMMC-7721.

  Methods  pcDNA3/CAAP1, the overexpression vector of CAAP1, and pSilencer 2.1-U6 neo/shR-CAAP1, the knockdown vector, were constructed and examined. The experiment included 4 groups of SMMC-7721 cells, pcDNA3/CAAP1 group, pcDNA3 control group, shR-CAAP1 group and pSilencer control group. After the SMMC-7721 cells were cultured, the overexpression vector pcDNA3/CAAP1 (the pcDNA3/CAAP1 group), knockdown vector shR-CAAP1 (the shR-CAAP1 group) and their controls (pcDNA3 control group and pSilencer control group) were transfected into SMMC-7721 cells respectively, and the follow-up experiments were carried out 48 h later. The mRNA expression of CAAP1 in each group was examined with qRT-PCR. The protein expression level of CAAP1 and cleaved Caspase-3 were checked with Western blot. The proliferation of cells was examined with CCK-8. The colony formation ability and the motility of cells in each group were assessed with colony formation assay and wound-healing assay, respectively. The migration and invasion of cells were examined with Transwell cell chamber and the apoptosis of cells was examined with flow cytometry. The data of 75 patients with low expression of CAAP1 and 295 patients with high expression of CAAP1 were downloaded from TCGA database and the data of 48 months follow-up were analyzed. Kaplan-Meier survival curve was used to compare the correlation between different levels of CAAP1 expression and overall survival (OS) of hepatocellular carcinoma (HCC) patients.

  Results  Double enzyme digestion analysis showed that the overexpression vector pcDNA3/CAAP1 and knockdown vector shR-CAAP1 were constructed successfully. qRT-PCR and Western blot results showed that pcDNA3/CAAP1 increased the mRNA and protein expression level of CAAP1 in SMMC-7721 cells (in comparison with the pcDNA3 control group, P<0.05), while shR-CAAP1 decreased the mRNA and protein expression of CAAP1 (in comparison with the pSilencer control group, P<0.05). Compared with pcDNA3 control group, the proliferation, colony formation ability, motility, migration and invasion of SMMC-7721 cells in the pcDNA3/CAAP1 group were increased, while the apoptosis of SMMC-7721 cells was inhibited (all P<0.05). Compared with the pSilencer control group, the proliferation, colony formation ability, motility, migration and invasion ability of SMMC-7721 cells in the shR-CAAP1 group decreased, while the apoptosis increased (all P<0.05). TCGA database analysis showed that HCC patients with low CAAP1 expression had better OS than that of HCC patients with high CAAP1 expression.

  Conclusion  CAAP1 can promote the proliferation, migration and invasion of SMMC-7721 cells while it inhibit their apoptosis.