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目的:研究三氧化二砷(As2O3)对人肝癌细胞SMMC-7721的促凋亡作用及对Smac、caspase-9、caspase-3表达的影响。方法:人肝癌细胞SMMC-7721经As2O3处理,共分为四组,分别为空白对照组、低剂量组、中等剂量组、高剂量组。分别采用MTT、Hoechst 33258染色法、Annexin V-FITC/PI双染法观察其对SMMC-7721细胞增殖的抑制,凋亡细胞核的形态学变化,以及诱导凋亡作用;采用Western blot法检测凋亡相关蛋白Smac、caspase-9、caspase-3表达的变化。结果:MTT显示:As2O3在体外能明显抑制SMMC-7721的生长,具有时间剂量依赖关系,与空白对照组相比,其余三组细胞生存率明显下降,差异均有统计学意义(P<0.05);Hoechst 33258显示细胞呈明显的凋亡细胞形态学特征,具有剂量依赖性;Annexin V-FITC/PI双染法显示:As2O3作用24小时可诱导SMMC-7721细胞凋亡,且呈剂量依赖性,与空白对照组相比(2.69±0.58),其余三组(4.01±0.58)、(5.99±1.69)、(9.26±2.34)差异均有统计学意义(P<0.05);Western blot显示:As2O3作用SMMC-7721细胞24小时,Smac、caspase-9、caspase-3表达上升,呈剂量依赖性,与空白对照组相比,其余三组蛋白表达量明显增加,差异均有统计学意义(P<0.05)。结论:一定量的As2O3能抑制SMMC-7721细胞增殖,促进其凋亡,其机制可能与调控Smac、caspase-9、caspase-3表达有关。
AIM: To investigate the effects of arsenic trioxide (As2O3) on the apoptosis of human hepatocellular carcinoma cell line SMMC-7721 and its effects on the expression of Smac, caspase-9 and caspase-3. Methods: Human hepatocellular carcinoma cell line SMMC-7721 was treated with As 2 O 3 and divided into four groups: blank control group, low dose group, middle dose group and high dose group. The proliferation of SMMC-7721 cells was observed by MTT, Hoechst 33258 staining and Annexin V-FITC / PI staining respectively. The morphological changes of apoptotic nuclei and the apoptosis of SMMC-7721 cells were observed. Apoptosis was detected by Western blot Changes in the expression of Smac, caspase - 9 and caspase - 3 proteins. Results: MTT showed that As2O3 could significantly inhibit the growth of SMMC-7721 in vitro in a dose-dependent manner. Compared with the blank control group, the survival rates of the other three groups were significantly decreased (P <0.05) ; Hoechst 33258 cells showed obvious morphological characteristics of apoptotic cells in a dose-dependent manner; Annexin V-FITC / PI double staining showed that: As2O3 can induce apoptosis of SMMC-7721 cells in a dose- Compared with control group (2.69 ± 0.58), the other three groups (4.01 ± 0.58), (5.99 ± 1.69) and (9.26 ± 2.34) showed statistical significance (P <0.05) The expression of Smac, caspase-9 and caspase-3 in SMMC-7721 cells increased in a dose-dependent manner at 24 hours. Compared with the blank control group, the expression levels of other three groups were significantly increased (P <0.05 ). Conclusion: A certain amount of As2O3 can inhibit the proliferation and promote the apoptosis of SMMC-7721 cells. The mechanism may be related to the regulation of the expression of Smac, caspase-9 and caspase-3.