【摘要】 目的 探讨隐丹参酮（CPT）对HepG2细胞凋亡的影响及其机制。方法 采用MTT试验检测终浓度为0、1、5、10、20、40、100μmol/L的CPT对HepG2细胞活性的影响，并计算出CPT对HepG2细胞的半数致死浓度(IC₅₀)。将HepG2细胞分为A～D组，分别使用终浓度0、1、5、10μmol/L的CPT培养48 h,分别通过划痕实验、JC-1染色和流式细胞术检测各组HepG2细胞的迁移能力、线粒体膜电位以及细胞凋亡率。将HepG2细胞分为E～H组，分别使用含有0μmol/L CPT+20μmol/L Spautin-1、1μmol/L CPT+20μmol/L Spautin-1、5μmol/L CPT+20μmol/L Spautin-1、10μmol/L CPT+20μmol/L Spautin-1的培养液培养。采用MTT实验检测E～H组处理48 h时的细胞活性，Western blot方法检测A、D、H组处理48 h时HepG2细胞中LC3B-Ⅱ蛋白的表达情况，JC-1染色检测A、D、E、H组处理48 h时的细胞线粒体膜电位，流式细胞术检测A、D、E、H组处理24 h时的细胞凋亡情况。结果 随着CPT浓度的递增，HepG2细胞活性明显下降，处理48 h时的IC₅₀值为3.9μmol/L。随着CPT浓度的升高，抑制HepG2细胞迁移的能力逐渐增强（t=5.96～29.63,P<0.05）;绿色荧光与红色荧光比值也逐渐升高（t=4.24～23.36,P<0.05）;HepG2细胞的细胞凋亡率逐渐增高（t=7.30～18.15,P<0.05）。A～H组间比较，细胞活性均有显著性差异（F=231.15,P<0.05）,E～H与A～D组相比，细胞活性增高（t=3.96～18.80,P<0.05）。H组与D组相比，自噬相关蛋白LC3B-Ⅱ蛋白表达显著降低（t=3.52,P<0.05）。JC-1染色结果显示，E组、H组与D组比较绿色荧光与红色荧光比值均有显著差异（t=3.58、14.76,P<0.05）。流式细胞术检测细胞凋亡的结果显示，E组、H组与D组比较绿色荧光与红色荧光比值均有显著差异（t=12.38、4.99,P<0.05）。结论CPT可能通过诱导自噬从而促进HepG2细胞的凋亡，达到对HepG2细胞活性的抑制作用。更多还原

【Abstract】 Objective To investigate the effect of cryptotanshinone（CPT） on the apoptosis of HepG2 cells and the underlying mechanism. Methods The MTT assay was used to measure the effects of CPT at final concentrations of 0, 1, 5, 10, 20, 40, and 100 μmol/L on the viability of HepG2 cells. The median lethal concentration(IC₅₀) of CPT for HepG2 cells was calculated. HepG2 cells were divided into groups A to D for 48 h culture with CPT at final concentrations of 0, 1, 5, and 10 μmol/L, respectively. The migration ability, mitochondrial membrane potential, and apoptosis rate of HepG2 cells were determined by wound healing assay, JC-1 staining, and flow cytometry, respectively. HepG2 cells were divided into groups E to H for culture with 0 μmol/L CPT+20 μmol/L Spautin-1, 1 μmol/L CPT+20 μmol/L Spautin-1, 5 μmol/L CPT+20 μmol/L Spautin-1, 10 μmol/L CPT+20 μmol/L Spautin-1, respectively. After 48 h culture, cell viability in groups E to H was measured by the MTT assay; LC3B-Ⅱ protein expression in groups A, D, and H was determined by Western blot; the mitochondrial membrane potential in groups A, D, E, and H was measured with JC-1 staining; and cell apoptosis in groups A, D, E, and H was measured by flow cytometry. Results With the increase of CPT concentrations, the viability of HepG2 cells was decreased significantly, with the IC₅₀ value being 3.9 μmol/L after 48 h culture. As CPT concentrations increased, the migration ability of HepG2 cells was decreased significantly（t=5.96-29.63,P<0.05）; the green/red fluorescence ratio was increased significantly（t=4.24-23.36,P<0.05）; the apoptosis rate of HepG2 cells was increased significantly（t=7.30-18.15,P<0.05）. There was a significant difference in cell viability between groups A to H（F=231.15,P<0.05）, with higher cell viability in groups E-H than in groups A-D（t=3.96-18.80,P<0.05）. Group H showed significantly lower expression of the autophagy-related protein LC3B-Ⅱ than group D（t=3.51,P<0.05）. The JC-1 staining results showed that groups E and H significantly differed from group D in the green/red fluorescence ratio（t=3.58,14.76,P<0.05）. The results of apoptosis by flow cytometry showed significant differences between group E and group D as well as between group H and group D（t=12.38,4.99,P<0.05）. Conclusion CPT can effectively inhi-bit the viability of HepG2 cells through autophagy-mediated apoptosis.更多还原