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目的探讨泛素连接酶Pirh2对人肝细胞肝癌(HCC)细胞增殖的影响。方法流式细胞仪测定血清饥饿释放过程中SMMC-7721和Hep3B细胞的周期分布情况,并用Western blot检测增殖过程中Pirh2及p27Kip1蛋白表达。在SMMC-7721和Hep3B细胞中转染Pirh2shRNA,cellcounting kit-8(CCK-8)和流式细胞分析检测转染后细胞增殖的能力,Western blot检测细胞周期调控因子的表达。结果细胞饥饿72h,两种细胞周期进程停滞在G1/G0期,血清释放后增殖到S期。在此过程中,Pirh2表达上调,p27Kip1表达下调。转染Pirh2shRNA组相较于对照组和转染空载体质粒组,CCK-8和流式细胞分析显示细胞的增殖能力下降及细胞的周期进程明显受到抑制(P<0.05)。结论 Pirh2参与HCC细胞的周期进程调控,有望成为HCC治疗的新靶点。
Objective To investigate the effect of ubiquitin ligase Pirh2 on the proliferation of human hepatocellular carcinoma (HCC) cells. Methods The cell cycle distributions of SMMC-7721 and Hep3B cells were detected by flow cytometry during serum starvation, and the expression of Pirh2 and p27Kip1 protein was detected by Western blot. The Pirh2 shRNA was transfected into SMMC-7721 and Hep3B cells, cell counting kit-8 (CCK-8) and flow cytometry were used to detect the cell proliferation after transfection. Western blot was used to detect the expression of cell cycle regulators. The results of cell starvation for 72 h, the two cell cycle progression arrested in the G1/G0 phase, serum release to the S phase. During this process, Pirh2 expression was up-regulated and p27Kip1 expression was down-regulated. Compared with the control group and the transfected vector plasmid group, CCK-8 and flow cytometry analysis showed that the cell proliferation and cell cycle progression were significantly inhibited (P<0.05) in the Pirh2 shRNA transfected group. Conclusion Pirh2 is involved in the regulation of the cycle progression of HCC cells and is expected to become a new target for HCC treatment.