摘要
目的:研究PI3K/Akt抑制剂渥曼青霉素对白血病细胞增殖和凋亡的影响,并探讨其可能的作用机制。方法:以不同浓度的渥曼青霉素作用于人类髓细胞白血病细胞K562,采用MTT法检测细胞增殖活性,单细胞凝胶电泳技术检测细胞DNA损伤形成的"彗星"样拖尾现象,AnnexinV-FITC/PI双标法检测细胞凋亡,Westernblotting、RT-PCR检测渥曼青霉素作用K562细胞后总Akt和磷酸化Akt以及NF-κB基因及蛋白表达水平的变化。结果:渥曼青霉素以时间-剂量依赖性方式抑制K562细胞的增殖,其24h的IC50是25nmol/L。渥曼青霉素诱导K562细胞发生凋亡,其作用呈明显剂量依赖性增强。渥曼青霉素作用后K562细胞DNA链断裂,呈现"彗星"拖尾现象,其尾长与拖尾率显著高于对照组(P<0.01)。渥曼青霉素能同时在蛋白和基因水平,以剂量依赖性方式抑制磷酸化Akt以及NF-κB表达,但对总Akt蛋白没有明显影响。结论:渥曼青霉素以时间和剂量依赖方式明显抑制K562细胞的增殖及诱导其凋亡,其机制可能与其下调磷酸化PI3K/Akt信号通路以及NF-κB蛋白表达有关。
Objective：To study the effect of wortmannin （WM）,a PI3K/Akt inhibitor,on the proliferation and apoptosis of leukemia cells and the possible mechanism.Methods：Human leukemia cell line K562 was treated with different concentrations of WM.The proliferation of K562 cells was examined by MTT assay.DNA damage in K562 cells was examined by single cell gel electrophoresis assay,and apoptosis of K562 cells was detected by Annexin V-FITC/PI double-staining.The expressions of total Akt,phosphorate-Akt （p-Akt）,and NF-κB p65 mRNA and protein were detected by RT-PCR and Western blotting,respectively.Results：WM inhibited the proliferation of K562 cells in a dose-and time-dependent manner,with the IC50 value of 24 h being 25 nmol/L.WM also induced apoptosis of K562 cells in a dose-dependent manner.DNA damage in K562 cells was demonstrated by appearance of comet tail after treatment with WM,with the rate of DNA tail and the tail length being significantly higher than those in the control group （P〈0.01）.WM dose-dependently inhibited P-Akt and NF-κB p65,but not the total Akt,mRNA and protein expressions.Conclusion：WM can inhibit proliferation and induce apoptosis of K562 cells in a dose-and time-dependent manner,probably through down-regulation of phosphorate PI3K/Akt signal pathway and NF-κB expression.
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2010年第1期82-87,共6页
Chinese Journal of Cancer Biotherapy
基金
湖北教育厅科技基金资助项目(No.D200511008)~~
