高三尖杉酯碱联合三氧化二砷诱导人多发性骨髓瘤RPMI 8226细胞株凋亡的实验研究  

Homoharringtonine Combined Arsenic Trioxide Induced Apoptosis in Human Multiple Myeloma Cell Line RPMI 8226: an Experimental Research

作　　者：周秀杰[1,2] 周郁鸿[1] 陈小会[3] 钱文斌[4] 

机构地区：[1]浙江中医药大学第一附属医院血液科,杭州310053 [2]浙江省海宁市人民医院血液科,浙江海市314400 [3]浙江省杭州市第二人民医院血液科,杭州310015 [4]浙江大学第一附属医院血液科,杭州310003

出　　处：《中国中西医结合杂志》2013年第6期834-839,共6页Chinese Journal of Integrated Traditional and Western Medicine

基　　金：国家自然科学基金资助项目(No.30871099);杭州市卫生科技计划资助项目(No.2011A016)

摘　　要：目的通过体外实验阐述高三尖杉酯碱(HHT)单药及联合亚砷酸(ATO)用药对人多发性骨髓瘤细胞株RPMI8226作用及其机制。方法应用四甲基偶氮唑(MTT)比色法,检测HHT、ATO单药及两者联合用药对人多发性骨髓瘤细胞株RPMI8226增殖的影响,Hoechst染色法检测细胞凋亡形态学变化,流式细胞仪检测细胞早期凋亡比率。Western blot检测药物处理后凋亡关键蛋白(Caspase-3、9及PARP)、Bcl-2家族蛋白(Bcl-2、Mcl-1、Bcl-xl)及AKT蛋白的表达。结果 HHT、ATO单药可显著抑制RPMI8226细胞的增殖,其作用呈时间及浓度依赖(P<0.05),且两药联合具有协同作用(CI<1)。HHT、ATO单药可诱导RP-MI8226细胞凋亡且呈浓度依赖,出现细胞凋亡形态学改变及早期凋亡比率增高,两药联合可加强诱导凋亡作用。HHT可呈浓度依赖性激活Caspase-3、PARP表达;HHT(40ng/mL)与ATO(8.5μmol/L)联合用药可显著激活Caspase-3、9,下调抗凋亡蛋白Mcl-1及Bcl-xl的表达,呈时间依赖性降低AKT磷酸化水平。结论 HHT、ATO单药或联合用药可诱导RPMI8226细胞凋亡,其机制可能和激活Caspase通路、调节Bcl-2家族蛋白表达、抑制AKT磷酸化等有关。Objective To clarify the effects and mechanisms of homoharringtonine （HHT） mono- mer therapy or combination therapy with arsenic trioxide （ATO） on human multiple myeloma （MM） cell line RPMI 8226 in in vitro researches. Methods Effects of HHT, ATO, and HHT combined ATO on the growth of MM cell line RPMI 8226 were detected using Ml-r assay. The morphological changes of cell apoptosis were detected by Hoechst staining. The early apoptosis rate was detected using flow cytometry. Expressions of Caspase-3, Caspase-9, poly-ADP-ribose polymerase （PARP）, Bcl-2, Mcl-1, Bcl-xl, and AKT protein were detected by Western blot. Results HHT and ATO inhibited the proliferation of RPMI 8226 cell line in a time- and dose-dependent manner （P 〈0.05）. Synergistic effects was shown in the combination group （CI 〈1 ）. HHT and ATO induced the apoptosis of RPMI 8226 in a dose-dependent manner with typical morphological changes of apoptosis and higher early stage apoptosis rate. The enhancement in apoptotic induction was seen when two agents were combined. HHT activated expressions of Caspase-3 and PARP in a dose de- pendent manner at 24 h. HHT at 40 ng/mL and ATO at 8.5 μmol/L could significantly activate expressions of Caspase-3 and Caspase-9, and down-regulate expressions of anti-apoptotic proteins Bcl-xl and Mcl-l. Inaddition, the combination therapy of HHT at 40 ng/mL and ATO at 8.5 μmol/L inhibited phosphorylation of AKT in a time-dependent manner. Conclusion HTT, ATO, and combination therapy of HHT and ATO in- duced the apoptosis of RPMI 8226 cell line possibly through activating Caspase pathways, regulating ex- pressions of Bcl-2 families, and inhibiting phosphorylation of AKT.

关 键 词：多发性骨髓瘤 高三尖杉酯碱 三氧化二砷 细胞凋亡 

分 类 号：R733.3[医药卫生—肿瘤]