摘要
目的探讨干扰Nrf2对HirsutanolsA抑制人结肠癌细胞SW480、肝癌细胞HepG2细胞增殖的影响。方法采用四甲基偶氮唑蓝比色法检测不同浓度Hirsutanols A对细胞增殖抑制作用。流式细胞仪检测细胞内ROS的含量;Annexin V-FITC/PI双染检测细胞凋亡;Western blot检测siRNA干扰NRF2蛋白表达的效果。结果 HirsutanolsA(1.25、2.5、5、10、20和40μmol/L)对SW480、HepG2细胞增殖有明显的抑制作用,并呈现剂量依赖关系;Hirsutanols A处理SW480(15μmol/L)、HepG2(30μmol/L)细胞后,诱导细胞中过氧化氢迅速增加,并引起细胞凋亡,用自由基清除剂N-乙酰半胱氨酸(5 mmol/L)预处理完全抑制HirsutanolsA诱导的ROS增加及细胞凋亡。siRNA有效地干扰Nrf2表达后可极大增强Hirsutanols A对肿瘤细胞的增殖抑制作用。结论Hirsutanols A通过增加细胞内ROS诱导其凋亡同时激活Nrf2,Nrf2在维持细胞氧化还原稳态中起重要作用,干扰其表达可增强Hirsutanols A的凋亡诱导作用。
Objective To investigate the effect of Nrf2 gene knockdown on hirsutanols A-induced cytotoxity in cancer cells. Methods The changes in the cell viability following treatment with different concentrations of hirsutanols A was detected by MTT assay, and the generation of reactive oxygen species (ROS) was assayed using flow cytometry. AnnexinV-FITC apoptosis kit was used to detect the cell apoptosis. Nrf2 protein expression in HepG2 and SW480 ceils transfected with the siRNA targeting Nrf2 was analyzed with Western blotting. Results At the concentrations of 1.25, 2.5, 5, 10, 20 and 40 μmol/L, hirsutanols A obviously inhibited the cell proliferation of human liver cancer HepG2 and colon cancer SW480 cells in a concentration-dependent manner. The levels of hydrogen peroxide increased rapidly after hirsutanols A treatment in both HepG2 (30 μmol/L) and SW480 (15 μmol/L) cells. Hirsutanols A also induced apoptosis of the two cells. Pretreatment with 5 mmol/L NAC totally inhibited apoptosis and ROS accumulation in the two cells induced by hirsutanols A. Transfection of HepG2 and SW480 cells with the siRNA caused a significant reduction in Nrf2 protein expression, which resulted in an increased sensitivity of the cells to hirsutanols A. Conclusion Hirsutanols A can induce apoptosis in HepG2 and SW480 cells by promoting ROS production and up-regulating Nrf2 expression. Nrf2 knockdown by siRNA can increase the sensitivity of the cancer cells to hirsutanols A in vitro.
出处
《南方医科大学学报》
CAS
CSCD
北大核心
2012年第8期1093-1097,共5页
Journal of Southern Medical University
基金
国家自然科学基金(81001446)~~