目的：研究丹参多酚酸盐(salvianolate)体外诱导人肝癌细胞SMMC-7721凋亡作用及其可能机制。方法：不同质量浓度丹参多酚酸盐(0.5、1、2 mg/ml)与肝癌细胞共培养24 h后，流式细胞仪检测肝癌细胞凋亡，线粒体膜电位试剂盒(JC-1)检测线粒体膜电位变化；比色法测定1.0 mg/ml丹参多酚酸盐作用后肝癌细胞内caspase8、caspase9 及caspase3的活性，流式细胞仪检测培养体系内加入caspase9抑制剂(zLEHDfmk)或caspase3抑制剂(zDEVDfmk)后细胞凋亡率的变化，Western blotting检测肝癌细胞内线粒体凋亡途径相关蛋白Bax、Bcl2表达水平。结果：丹参多酚酸盐显著诱导肝癌细胞SMMC7721凋亡(P<0.05)，同时线粒体膜电位随着药物浓度的升高而加剧下降(P<0.05)。1.0 mg/ml 丹参多酚酸盐处理肝癌细胞24 h后caspase-9与caspase-3的活性明显升高(P<0.05)，而caspase-8的活性无明显变化(P>0.05)；当培养体系内加入caspase-9或caspase3活性抑制剂后，丹参多酚酸盐诱导肿瘤细胞凋亡的作用明显降低(P<0.05)。Western blotting检测显示，丹参多酚酸盐处理组前凋亡蛋白Bax表达明显升高，抗凋亡蛋白Bcl2表达降低。结论：丹参多酚酸盐(0.5~2.0 mg/ml)剂量具有促进肝癌细胞凋亡的作用，且有剂量依赖的趋势，其机制与线粒体凋亡途径有关。

Objective:To explore the apoptosisinducing effect of salvianolate on hepatoma SMMC7721 cells and the underlying mechanism. Methods:SMMC7721 cells were cocultured in vitro with different concentrations (0.5, 1, 2 mg/ml) of salvianolate for 24 h. The apoptotic SMMC7721 cells were examined by flow cytometry, and the changes of mitochondrial transmembrane potential were examined by mitochondrial transmembrane potential JC1 kit. The activities of caspase8, caspase9, and caspase3 were detected by spectrophotometry in the hepatoma SMMC7721 cells after cocultured with 1 mg/ml salvianolate. The changes of apoptotic SMMC7721 cells induced by salvianolate in the presence or absence of caspase9 inhibitor or caspase3 inhibitor were measured by flow cytometry. The expressions of proapoptotic protein Bax and antiapoptotic protein Bcl2 were detected by Western blotting analysis. Results:Salvianolate significantly induced apoptosis of hepatoma SMMC7721 cells (P<0.05), and the decline of mitochondrial membrane potential increased with the increase of salvianolate concentration (P<0.05). The activities of caspase9 and caspase3, but not caspase8, were increased in hepatoma cells after treatment with 1 mg/ml salvianolate for 24 h (P<0.05). The apoptosisinducing effect of salvianolate was significantly decreased in the presence of caspase9 or caspase3 inhibitors (P<005). Western blotting results showed that salvianolate increased proapoptotic protein Bax expression and decreased antiapoptotic protein Bcl2 expression. Conclusion:Salvianolate can induce the apoptosis of human hepatoma SMMC7721 cells in a dosedependent manner, which is probably mediated by mitochondrial apoptosis pathway.

上海市卫生局青年科研项目资助（No. 2008Y085）；卫生部国家科技支撑计划课题资助（No. 2008BAI60B03）