硼替佐米联合三氧化二砷诱导NB4细胞凋亡及相关机制的研究  

NB4 Cell Apoptosis Induced By Bortezomib Combined with As_2O_3 and Its Mechanism

作　　者：陈晓文[1] 夏海龙[1] 夏瑞祥[1] 

机构地区：[1]安徽医科大学第一附属医院血液内科,安徽合肥230022

出　　处：《中国实验血液学杂志》2011年第2期348-352,共5页Journal of Experimental Hematology

摘　　要：本研究旨在探索硼替佐米(bortezomib)联合三氧化二砷(As2O3)诱导急性早幼粒细胞白血病NB4细胞株的凋亡及相关调控机制。用流式细胞术AnnexinⅤ/PI双染法检测细胞凋亡;Hoechst染色法观察凋亡细胞形态;Western blot检测caspase-3,caspase-9的活化以及NOXA蛋白的表达;以小干扰RNA(siRNA)技术特异性"沉默"NOXA基因;用脂质体Lipofectamine2000转染pEGFP-Noxa质粒和空载体pEGFP。结果表明,bortezomib(10 nmol/L)联合As2O3(0.5μmol/L)诱导NB4细胞凋亡率较单药As2O3(0.5μmol/L)诱导时增加,相应的caspase-3,-9的活化明显加强。单药As2O3诱导的NB4细胞中Noxa蛋白水平未发生改变,bortezomib和As2O3联合诱导的NB4中NOXA蛋白水平上调。特异性"沉默"NOXA基因后bortezomib和As2O3联合诱导的NB4细胞中caspase-3的活化程度降低,细胞凋亡率也明显下降。通过转染质粒高表达NOXA蛋白,单药As2O3诱导的NB4细胞中caspase-3的活化程度增加。结论:bortezomib可以增加NB4细胞对As2O3诱导凋亡的敏感性,这可能和促凋亡蛋白NOXA的表达上调有关。This study was aimed to investigate the apoptosis induced by bortezomib combined with As2O3 in APL cell line NB4 and its mechanism.The apoptotic cells were detected by flow cytometry with Annexin V/propidium iodide double staining;the morphology of apoptotic cells was observed by Hoechst staining,Western blot was used to measure activation of caspase-3 and-9 as well as expression of NOXA;the siRNA technique was used to specifically silence NOXA gene;the lipofectamine 2000 was used to transfect pEGFP-Noxa plasmid and pEGFP vacant vector.The results showed that the bortezomib combined with As2O3 could induce significant apoptosis of NB4 cells and activation of caspase 3 and caspase 9,but As2O3（0.5 μmol/L） alone could not cause marked activation of caspase cascade and apoptosis of NB4 cells.The expression level of NOXA in NB4 cells induced by bortezomib combined with As2O3 was up-regulated;the activation level of caspase-3 and apoptotic rate of NB4 cells treated by bortezomib combined with As2O3 decreased after specifically silencing the NOXA gene.The high expression of NOXA induced by transfection of plasmid could enhance the caspase 3 activity induced by As2O3 alone.It is concluded that bortezomib can enhance sensitivity of NB4 cells to apoptosis induced by As2O3 which may be related with up-regulation of proapoptotic protein NOXA.

关 键 词：硼替佐米 三氧化二砷 NB4细胞 Noxa蛋白 

分 类 号：R733.7[医药卫生—肿瘤] R979.1[医药卫生—临床医学]