人参皂苷单体Rh_2诱导小鼠前胃癌细胞系凋亡的分子机制

Molecular mechanism of G-Rh_2-induced apoptosis in gastric carcinoma cells

目的:观察人参皂苷Rh2(G-Rh2)诱导小鼠前胃癌MFC细胞的凋亡,探讨Ca2+在G-Rh2诱导细胞凋亡中的作用,揭示G-Rh2诱导MFC细胞凋亡的分子机制.方法:分别对无血清培养24h后的MFC正常细胞组、G-Rh2(3、10、30mg/L)给药组、Ca2+螯合剂BAPTA(20μmol)+G-Rh2(10mg/L)给药组及BAPTA(20μmol)组,荧光显微镜(Hoechst33258)观察细胞形态学改变;流式细胞仪Annexin V-FITC双染法检测细胞早期凋亡率;琼脂糖凝胶电泳法观察细胞晚期凋亡;共聚焦显微镜分析G-Rh2给药后细胞内钙和细胞线粒体膜电位,Western blot分析caspase3的表达.结果:10mg/L的G-Rh2能显著抑制无血清培养的MFC细胞增殖和诱导MFC细胞凋亡,可剂量和时间依赖性的提高细胞内钙浓度,降低细胞线粒体膜电位,增加caspase3的表达,但这些作用均可被BAPTA(20μmol)抑制.结论:G-Rh2可激活细胞内Ca2+信号转导途径,从而最终通过线粒体凋亡途径诱导胃癌细胞凋亡.

AIM： To investigate the effect of Ginseng （G） -Rh2 on the proliferation and apoptosis of mouse gastric carcinoma cells （MFCs）, and to explore the activation of the Ca^2＋ transduction pathways mediating G-Rh2-induced MFC cell apoptosis. METHODS： MFCs were divided into a normal free-serum MFC cell group, a G-Rh2 （3, 10, 30 mg/L） treatment group, a Ca^2＋ chelator BAPTA （20 μmol） ＋ G-Rh2 （10 mg/L） group and a BAP- TA （20 μmol） group. Morphologic changes were observed by fluorescence microscopy （Hoechst 33 258）. The early apoptosis rate was detected by Annexin V-FITC using flow cytometry; late apoptosis was analyzed by agarose gel electrophoresis. The mitochondrial membrane potential （△ψm） and Ca^2＋ density were measured by confocal microscopy. The expression of caspase 3 was observed by Western blotting. RESULTS： G-Rh2 （10 mg/L） reinforced cell activity and induced apoptosis of MFC cells. The apoptosis rate in the drug group increased greatly after G-Rh2 treatment. G-Rh2 induced MFC cell apoptosis, markedly enhance calcium ionconcentration, effectively reduced △ψm and increased the protein expression of caspase 3, but all of these effects could be inhibited by BAPTA （20 μmol）. CONCLUSION： G-Rh2 can induce apoptosis of gastric carcinoma cells, possibly via a Ca^2＋ - dependent mitochondrial apoptosis pathway.