Daxx抑制K562细胞向巨核细胞分化  

Role of Daxx in inhibiting megakaryocytic differentiation of K562 cells

作　　者：刘胜兵[1] 潘魏巍 沈忠飞[1] 徐营[1] 郭燕君[1] 王志坚[1] 黄倩[1] 

机构地区：[1]嘉兴学院医学院,浙江嘉兴314001

出　　处：《中国病理生理杂志》2016年第6期1004-1010,共7页Chinese Journal of Pathophysiology

基　　金："十二五"浙江省高校药理学重点学科资助项目;嘉兴市科技局项目(No.2015AY23063);嘉兴学院重点SRT项目(No.851715065)

摘　　要：目的:观察过表达死亡结构域相关蛋白(Daxx)对K562细胞活力和向巨核细胞分化的影响。方法:建立稳定过表达Daxx的K562细胞,荧光显微镜观察、实时荧光定量PCR和Western blot检测Daxx的过表达效果,CCK-8法检测过表达后细胞活力的变化;佛波酯(PMA)诱导K562细胞向巨核细胞系分化,Western blot检测在K562细胞向巨核细胞分化过程中Daxx和p-ERK的表达变化,流式细胞术检测CD41和CD61的表达变化;PMA处理过表达Daxx的K562细胞,NBT还原实验检测细胞分化情况,流式细胞术检测过表达Daxx后CD41和CD61的表达变化,Western blot检测p-ERK的蛋白水平。结果:建立了稳定的过表达Daxx的K562细胞,过表达Daxx抑制K562细胞的活力。PMA诱导K562细胞向巨核细胞分化,CD41和CD61表达水平增高,同时p-ERK的蛋白水平升高,Daxx表达水平下降。过表达Daxx可以抑制K562细胞向巨核细胞分化,CD41和CD61表达降低,同时p-ERK的蛋白水平降低。结论:过表达Daxx可以抑制K562细胞生长及向巨核细胞分化,同时抑制ERK的磷酸化。AIM： To examine the effects of death domain-associated protein（ Daxx） overexpression on the viability and megakaryocytic differentiation of K562 cells. METHODS： Daxx overexpression in the K562 cells was established. The expression of Daxx was detected by fluorescence microscopy,fluorescence quantitative real-time PCR and Western blot after transfection. CCK-8 assay was used to detect the cell viability after overexpression of Daxx. The expression of CD41 and CD61 in phorbol 12-myristate 13-acetate（ PMA） induced K562 cells was detected by flow cytometry. The protein levels of Daxx and p-ERK were determined by Western blot. Nitroblue tetrazolium（ NBT）-reducing test was used to assess leukemia cell differentiation in Daxx-overexpressing K562 cells and control cells. The expression of CD41 and CD61 induced by PMA in Daxx-overexpressing K562 cells was analyzed by flow cytometry. The protein levels of Daxx and p-ERK were also examined by Western blot. RESULTS： The stable overexpression of Daxx in the K562 cells was established. The viability was reduced in Daxx-overexpressing K562 cells. The expression of CD41 and CD61 was significantly increased after PMA induction in the K562 cells（ P〈0. 01）. The protein expression of Daxx was reduced,but the protein level of pERK was increased. The expression of CD41 and CD61 was reduced after PMA induction in Daxx-overexpressing K562cells（ P〈0. 01）. The protein level of p-ERK was also reduced. CONCLUSION： Daxx overexpression inhibits the growth,megakaryocytic differentiation and production of p-ERK in the K562 cells.

关 键 词：死亡结构域相关蛋白 K562细胞 巨核细胞分化 佛波酯 

分 类 号：R329.21[医药卫生—人体解剖和组织胚胎学] R730.23[医药卫生—基础医学]