KPC1对肝癌细胞增殖和迁移能力的影响

Effect of KPC1 on cell proliferation and migration of hepatocellular carcinoma cell lines

目的探讨Kip1泛素化促进复合体蛋白1(KPC1)对肝癌细胞系Bel-7402和SMMC-7721细胞增殖与迁移能力的影响以及可能的分子机制。方法通过慢病毒包装的方法将KPC1过表达重组质粒转染进肝癌细胞系Bel-7402和SMMC-7721,筛选得到稳定过表达KPC1的肝癌细胞系;采用小干扰RNA(siRNA)瞬转敲低肝癌细胞系中的KPC1;蛋白质免疫印迹法(WB)检测肝癌细胞系中KPC1蛋白的表达水平,验证过表达和敲低KPC1的效果;CCK-8法检测KPC1对肝癌细胞系增殖能力的影响;Transwell法验证KPC1对肝癌细胞系迁移能力的影响;WB法检测扰动KPC1后p105蛋白表达水平的影响;Log-rank检验比较KPC1高表达组和低表达组肝癌患者的生存期差异。结果过表达KPC1可显著抑制Bel-7402和SMMC-7721细胞的增殖和迁移能力;而敲低KPC1显著促进Bel-7402和SMMC-7721细胞的增殖和迁移能力。过表达KPC1抑制p105的表达水平、提升p50的表达水平;而敲低KPC1则促进p105的表达水平、抑制p50的表达水平。临床相关性分析显示KPC1在肝癌癌组织中下调表达,且与肝癌患者的不良预后显著相关。结论 KPC1通过抑制肝癌细胞系的增殖和迁移能力而发挥抑癌基因的功能。

Objective To investigate the effects of Kip1 ubiquitination-promoting complex protein 1（KPC1） on proliferation and migration of hepatocellular carcinoma（HCC） cells Bel-7402 and SMMC-7721,and the underlying molecular mechanism. Methods The KPC1 overexpression recombinant plasmid was transfected into the Bel-7402 and SMMC-7721 cells by lentivirus packaging. The small interfering RNA（siRNA） was used to knockdown endogenous KPC1 in HCC cells. Western blotting（WB） was used to detect the protein levels of KPC1 in HCC cells and verify the effects of its overexpression and knockdown. Cell Counting Kit-8（CCK-8） assays were used to assess the HCC cell proliferation,Transwell assays were used to assess the HCC cell migration,and Western blotting assay was used to detect the effect of perturbation on the expression of p105 protein in KPC1. Log-rank test was used to assess the difference between the HCC patients with high expression levels of KPC1 and those with low expression levels of KPC1. Results Overexpression of KPC1 significantly inhibited the proliferation and migration Bel-7402 and SMMC-7721 cells,whereas knockdown of KPC1 significantly promoted of their proliferation and migration. Overexpression of KPC1 decreased the p105 levels and elevated the p50 levels,while knockdown of KPC1 led to the upregulation of p105 and downregulation of p50. Of note,compared to the matched non-tumor liver tissues,KPC1 was downregulated in HCC tissues,which was associated with poor outcomes of HCC patients. Conclusion KPC1 functions as a tumor suppressor in HCC cells by inhibiting HCC cell proliferation and migration.