白藜芦醇通过促进Apaf-1/caspase-9复合物的形成增强5-氟尿嘧啶对骨肉瘤CD133^+细胞亚群凋亡的诱导  

Resveratrol enhances 5-fluorouracil-induced apoptosis of osteosarcoma CD133^+ cell subset by promoting formation of Apaf-1/caspase-9 complex

作　　者：张红萍[1] 袁梅[1] 

机构地区：[1]嘉兴市中医院检验科,浙江嘉兴314001

出　　处：《中国病理生理杂志》2017年第7期1196-1202,共7页Chinese Journal of Pathophysiology

摘　　要：目的:探讨白藜芦醇联合化疗药物5-氟尿嘧啶对骨肉瘤CD133^+细胞亚群的协同杀伤效应及机制。方法:将人骨肉瘤细胞系MG-63 CD133^+细胞亚群及相应CD133^-细胞亚群用5-氟尿嘧啶及白藜芦醇进行体外处理。MG-63细胞的相对细胞活力用MTT法进行检测;细胞凋亡率用流式细胞术进行检测;用Western blot实验检测caspase-9和caspase-3的活化、Apaf-1的表达水平及细胞色素C的释放;用免疫共沉淀法检测Apaf-1与caspase-9前体的相互作用。结果:5-氟尿嘧啶对MG-63 CD133^+细胞亚群的杀伤活性和凋亡诱导活性均显著低于MG-63 CD133^-细胞亚群。但联用白藜芦醇能显著提高5-氟尿嘧啶对MG-63 CD133^+细胞亚群的细胞活力抑制率。白藜芦醇处理能显著上调MG-63 CD133^+细胞亚群中Apaf-1的表达水平,在MG-63 CD133^+细胞亚群中转染Apaf-1siRNA后,5-氟尿嘧啶联用白藜芦醇的协同效应受到显著抑制。另外,免疫共沉淀实验结果表明白藜芦醇联合5-氟尿嘧啶能显著诱导MG-63 CD133^+细胞亚群中Apaf-1/caspase-9复合物的形成,从而诱导caspase-9发生活化。结论:白藜芦醇通过促进Apaf-1/caspase-9复合物的形成,增强5-氟尿嘧啶对骨肉瘤CD133^+细胞亚群凋亡的诱导。AIM: To investigate the synergistic effect of resveratrol and 5-fluorouracil on osteosarcoma CD133^+ cell subset. METHODS: Human osteosarcoma cell line MG-63 CD133^+ cell subset and the corresponding CD133^- cell subset were treated with resveratrol and 5-fluorouracil. After treatment,the viability of MG-63 cells was measured by MTT assay. The apoptosis of MG-63 cells was analyzed by flow cytometry. Activation of caspase-9 and caspase-3,the expression of Apaf-1,and the release of cytochrome C were evaluated by Western blot. The interaction between Apaf-1 and procaspase-9 was detected by co-immunoprecipitation. RESULTS: The cell death and apoptosis of MG-63 CD133^+ cell subset induced by 5-fluorouracil were significantly weaker than those in the corresponding MG-63 CD133^- cell subset. However,co-treatment with resveratrol significantly enhanced the effect of 5-fluorouracil on inhibiting the viability of MG-63CD133^+ cell subset. Mechanically,treatment with resveratrol upregulated the expression of Apaf-1. Transfection with Apaf-1 siRNA abolished the synergistic effect of resveratrol and 5-fluorouracil in MG-63 CD133^+ cell subset. In addition,the results of co-immunoprecipitation indicated that the combination of resveratrol and 5-fluorouracil significantly induced the formation of Apaf-1/pro-caspase-9 complex,leading to the activation of caspase-9 in MG-63 CD133^+ cell subset. CONCLUSION: Resveratrol enhances 5-fluorouracil-induced apoptosis of osteosarcoma CD133^+ cell subset by promoting the formation of Apaf-1/caspase-9 complex.

关 键 词：白藜芦醇 5-氟尿嘧啶 骨肉瘤 CD133+细胞亚群 APAF-1 CASPASE-9 

分 类 号：R735.7[医药卫生—肿瘤] R965[医药卫生—临床医学]