TIMP-3对肝癌细胞增殖、凋亡及侵袭转移的影响

The effects of TIMP-3 on the proliferation,apoptosis,invasiveness and migration of human hepatocarcinoma cell line

目的建立以真核表达载体介导的过表达TIMP-3基因的肝癌细胞株(SMMC-7721和HepG2),探讨TIMP-3基因对两细胞增殖、凋亡、周期、侵袭等功能的影响。方法将TIMP-3质粒pCMV6-AC-GFP/TIMP-3(TIMP-3组)和空载对照质粒pCMV6-AC-GFP(对照组)稳定转染入SMMC-7721和HepG2细胞株,用Western blot技术检测细胞内TIMP-3蛋白的表达,通过MTS实验、流式细胞术、侵袭实验测定细胞的各种生物学行为。结果 Western blot检测结果显示,TIMP-3组的SMMC-7721和HepG2细胞高表达TIMP-3蛋白,而对照组细胞没有TIMP-3蛋白表达。MTS实验和流式细胞术检测结果显示,与对照组比较,TIMP-3组两种细胞的增殖能力显著减弱(第5天时P=0.003,P=0.009),凋亡率显著升高(P=0.001,P=0.038),细胞周期停滞于G2/M期(P=0.030,P=0.034);体外侵袭实验结果显示,TIMP-3组两种细胞的侵袭能力较对照组显著下降(P=0.033,P=0.015)。结论 TIMP-3能显著抑制SMMC-7721及HepG2细胞的增殖和侵袭能力,并诱导细胞发生凋亡和产生G2/M期阻滞。

Objective To construct a stable TIMP- 3 over-expression hepatocellular carcinoma （HCC） cell lines （SMMC -7721 and HepG2） for investigating the effects of TIMP -3 on proliferation, apoptosis, cell cycle and inva- siveness of the two cell lines. Methods The pCMV6 - AC - GFP/TIMP - 3 and empty plasmid pCMV6 - AC - GFP （control） were both transfected into SMMC -7721 and HepG2 cells. The TIMP- 3 protein was assessed by Western blot. The effects of TIMP - 3 on the biological behaviors of SMMC - 7721 and HepG2 were tested by MTS assay, flow cytometry, and invasiveness assay. Results Over - expression of TIMP - 3 was observed in SMMC - 7721 and HepG2 cells transfected with TIMP -3, while no expression of TIMP -3 was revealed in controls. The proliferation and invasiveness of SMMC-7721 and HepG2 cells were significantly suppressed by TIMP- 3 overexpression, by which significantly increased apoptosis rate and G2/M cell cycle arrest were also observed （ P 〈 0. 05 ）. Conclusion TIMP - 3 significantly inhibits the capabilities of proliferation and invasiveness of SMMC -7721 and HepG2 cells, induces HCC cells apoptosis and caused cell cycle arrest at G2/M phases.