野生型PTEN基因增强青蒿琥酯抑制K562细胞的作用  

Wild-type PTEN gene enhances the inhibitory effect of artesunate on K562 cell growth

作　　者：成志勇[1] 徐倩[1,2] 赵亚玲[1,2] 付建珠 谷蕾[1] 李密[1] 梁丽青[1] 刘芳[1] 

机构地区：[1]保定市第一医院血液内科,河北保定071000 [2]承德医学院,河北承德067000

出　　处：《中国病理生理杂志》2016年第1期112-117,共6页Chinese Journal of Pathophysiology

摘　　要：目的:探讨野生型PTEN转染人白血病K562细胞系对青蒿琥酯敏感性的影响及其分子作用机制。方法:将野生型PTEN以腺病毒为载体转染(感染复数为200)人白血病K562细胞(Ad-WT-PTEN),同时以转染空载体腺病毒(Ad)及未转染细胞为对照组,与青蒿琥酯(ART)联合作用,观察野生型PTEN增强青蒿琥酯抑制K562细胞的作用。根据IC50计算PTEN对青蒿琥酯的增敏倍数。以四甲基偶氮唑蓝(MTT)法检测细胞活力,流式细胞术检测细胞凋亡率,real-time PCR检测PTEN的mRNA水平,Western blot检测PTEN、蛋白激酶B(Akt)及磷酸化Akt(p-Akt)的蛋白水平;caspase活性检测试剂盒检测caspase-3/7的活性。结果:Ad-WT-PTEN转染K562细胞后,对青蒿琥酯敏感性明显增加,依据IC50计算增敏倍数为2.25倍。至第3天,Ad-WT-PTEN+ART组较Ad+ART组细胞活力下降、凋亡率升高。Ad-WT-PTEN转染K562细胞后PTEN的mRNA及蛋白表达明显增加,p-Akt水平及caspase-3/7活性下调,以PTEN及青蒿琥酯联合作用组下调尤为明显。结论:野生型PTEN可能通过降低K562细胞Akt磷酸化的水平,并增加caspase-3/7活性,增强细胞对青蒿琥酯的敏感性。AIM： To investigate the effect of wild-type PTEN transfection on the sensitivity of human leukemia K562 cells to artesunate（ ART） and its molecular mechanism. METHODS： The adenovirus containing wild-type PTEN（ Ad-WT-PTEN） or empty vectors（ Ad） were transfected into K562 cells [with multiplicity of infection（ MOI） = 200].The untransfected cells served as normal control. The effect of wild-type PTEN on the inhibition of K562 cell growth by ART was observed. The sensitizing ratio of PTEN combined with ART based on IC50 was calculated. The viability of K562 cells was detected by MTT assay. The apoptosis was analyzed by flow cytometry. The mRNA level of PTEN was assessed by real-time PCR. The protein expression of PTEN,p-Akt and Akt was detected by Western blot. The activity of caspase-3 /7was measured by caspase activity kits. RESULTS： The sensitivity of K562 cells to ART was significantly increased by2. 25 folds after transfected with PTEN based on the IC50. The cell viability in Ad-WT-PTEN ＋ ART group was significantly lower than that in Ad ＋ ART group after transfection for 3 d（ P 0. 01）. The apoptotic rate in Ad-WT-PTEN ＋ ART group was significantly higher than that in Ad ＋ ART group（ P 0. 01）. The expression of PTEN at mRNA and protein levels in the K562 cells after transfection with PTEN was significantly increased,and the protein level of p-Akt and caspase-3 /7 activity were down-regulated,particularly in PTEN combined with ART group. CONCLUSION： The wild-type PTEN gene enhances the sensitivity of the K562 cells to ART by down-regulating the level of p-Akt and up-regulating the caspase-3 /7activity.

关 键 词：PTEN基因 青蒿琥酯 AKT Caspase-3/7 

分 类 号：R730.23[医药卫生—肿瘤] R733[医药卫生—临床医学]