论文部分内容阅读
目的:探讨4,5,6,7-四溴苯三唑(TBB)对人结肠癌SW480细胞的促凋亡作用,并初步探讨其可能的作用机制。方法:选取处于对数生长期的人结肠癌SW480细胞,将其分为对照组(给予0μmol·L~(-1)TBB)和实验组(给予1、3、10、30和100μmol·L~(-1) TBB)。采用MTT比色法检测SW480细胞存活率,采用AnnexinⅤ-FITC/PI双染法及流式细胞术检测SW480细胞凋亡率及细胞中活性氧(ROS)水平,采用Western blotting法检测细胞中抗凋亡蛋白p-Akt、Bcl-2和促凋亡蛋白Bad、pro-caspase-9及cleaved-caspase-3表达水平。结果:MTT法检测,与对照组比较,实验组SW480细胞存活率明显降低(P<0.05),且呈浓度依赖性;AnnexinⅤ-FITC/PI双染法及流式细胞术检测,随着处理时间的增加,与对照组比较,TBB作用3、6、12和24h时SW480细胞凋亡率明显高于作用0h时(P<0.05);流式细胞术检测,TBB作用3、6、12和24h时结肠癌SW480细胞中ROS水平高于作用0h时(P<0.05);Western blotting法检测,SW480细胞中抗凋亡蛋白p-Akt和Bcl-2表达水平逐渐降低,促凋亡蛋白Bad和cleaved-caspase-3表达水平逐渐升高,pro-caspase-9蛋白表达水平逐渐降低,与对照组(0h)比较差异有统计学意义(P<0.05)。结论:TBB对体外培养的人结肠癌SW480细胞有明显的杀伤作用,可诱导细胞凋亡,其机制可能与抑制Akt活性或促进细胞中ROS产生有关。
AIM: To investigate the apoptosis-promoting effect of 4,5,6,7-tetrabromobenzotriazole (TBB) on human colon cancer SW480 cells and to explore its possible mechanism. Methods: Human colon cancer SW480 cells in logarithmic growth phase were selected and divided into control group (given 0μmol·L -1 TBB) and experimental group (given 1,3,10,30 and 100μmol·L ~ (-1) TBB). The survival rate of SW480 cells was detected by MTT colorimetric assay. The apoptosis rate and the level of reactive oxygen species (ROS) in SW480 cells were detected by AnnexinⅤ-FITC / PI double staining and flow cytometry. The anti-apoptosis P-Akt, Bcl-2 and pro-apoptotic proteins Bad, pro-caspase-9 and cleaved-caspase-3. Results: Compared with the control group, the survival rate of SW480 cells in the experimental group was significantly decreased (P <0.05) by MTT assay, and the concentration-dependent manner. The results of flow cytometry and AnnexinⅤ-FITC / Compared with the control group, the apoptotic rates of SW480 cells at 3, 6, 12 and 24 hours after TBB treatment were significantly higher than those at 0 hours (P <0.05). Flow cytometry, TBB treatment at 3, 6, 12 and 24h The level of ROS in SW480 cells was higher than that in 0h (P <0.05). Western blotting showed that the expression levels of p-Akt and Bcl-2 in SW480 cells decreased gradually, while the levels of pro-apoptotic proteins Bad and cleaved The expression of caspase-3 gradually increased and the expression of pro-caspase-9 protein gradually decreased. The difference was statistically significant (P <0.05) compared with the control group (0h). CONCLUSION: TBB has obvious killing effect on human colon cancer SW480 cells in vitro and induces apoptosis. Its mechanism may be related to inhibiting Akt activity or promoting ROS production.