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目的观察蛋白酶体抑制剂MG132对诱导体外培养激活的肝星状细胞(HSC)凋亡的影响。方法大鼠肝星状细胞分离采用胶原酶原位灌注法,用流式细胞仪和琼脂糖凝胶电泳法检测MG132对诱导激活的HSC凋亡的影响。结果1、2、3μmol/LMG132培养HSC24h后,细胞周期分析发现S期细胞减少,G2/M期细胞显著增加(P<0.01),呈现一个剂量依赖性的关系;流式细胞术检测到明显的亚G1峰,各组的凋亡指数(%)分别是12.70±1.7、17.52±2.3、22.60±3.4,与对照组(1.9±0.6)相比,差异有显著性(P<0.01);3μmol/LMG132作用12、24、36、48h,凋亡指数(%)分别是16.43±2.2、22.60±2.7、29.80±1.7和36.30±1.4,与对照组相比,差异有显著性(P<0.01),呈现一个时间依赖性的关系;琼脂糖凝胶电泳可以看到明显的DNA梯带的形成。结论MG132能够诱导激活的HSC发生凋亡,且在发生凋亡之前有一个明显的G2/M期的阻滞。
Objective To investigate the effect of proteasome inhibitor MG132 on inducing apoptosis of hepatic stellate cells (HSC) activated in vitro. Methods Rat hepatic stellate cells were isolated by collagenase in situ perfusion method. The effects of MG132 on the apoptosis of activated HSC were detected by flow cytometry and agarose gel electrophoresis. Results After cultured with 1,2,3μmol / LMG132 for 24 hours, the number of cells in S phase decreased and the cells in G2 / M phase increased significantly (P <0.01) by cell cycle analysis, showing a dose-dependent relationship. Flow cytometry showed significant (P <0.01). The apoptosis index (%) of each group was 12.70 ± 1.7,17.52 ± 2.3,22.60 ± 3.4, which was significantly different from that of the control group (1.9 ± 0.6) (P <0.01) The apoptotic index (%) of LMG132 at 12,24,36,48h were 16.43 ± 2.2,22.60 ± 2.7,29.80 ± 1.7 and 36.30 ± 1.4, respectively, which was significantly different from the control group (P <0.01) Showing a time-dependent relationship; agarose gel electrophoresis can see significant DNA ladder formation. Conclusion MG132 can induce apoptosis of activated HSC, and there is a significant G2 / M arrest before apoptosis occurs.