氨肽酶抑制剂对全反式维甲酸诱导人急性早幼粒白血病细胞株NB4细胞分化作用的影响及其机制的初步探讨  

Effect of Aminopeptidase Inhibitor on Differentiation Induction Activity of All-trans Retinoic Acid in Human Acute Promyelocytic Leukemia NB4 Cells and Its Mechanism

作　　者：林茂芳[1] 钱习军[1] 

机构地区：[1]浙江大学医学院附属第一医院血液科,杭州310003

出　　处：《中国实验血液学杂志》2006年第3期471-476,共6页Journal of Experimental Hematology

摘　　要：为了研究氨肽酶抑制剂乌苯美司(bestatin)对全反式维甲酸(ATRA)诱导人APL细胞株NB4细胞分化作用的影响及其可能机制,NB4细胞经bestatin和ATRA单用或联合应用处理一定时间后,用光学显微镜观察细胞形态,流式细胞仪检测CD11b表达,四氮唑蓝(NBT)还原实验检测分化细胞功能,RT-PCR法检测c-myc和c-EBPεmRNA表达,Westernblot检测c-Myc蛋白表达。结果显示:NB4细胞经100μg/mlbestatin和10nmol/LATRA联合处理72小时后,其分化的形态更明显;100μg/mlbestatin与不同浓度ATRA联合处理72小时,能增强NB4细胞的NBT还原能力,与单用相应浓度ATRA组相比,差异显著(P<0.01);从48-96小时,100μg/mlbestatin能增强10nmol/LATRA诱导NB4细胞的NBT还原能力,且呈时间依赖性,与相应时间点ATRA组相比,差异明显(P<0.01);与单用10nmol/LATRA组相比,ATRA(10mmol/L)和bestatin(100μg/ml)联用处理72小时,NB4细胞CD11b阳性表达率明显增加(P<0.01);与单用10nmol/LATRA组相比,联用100μg/mlbestatin处理4小时后,NB4细胞c-mycmRNA表达的降低更明显(P<0.05);50、75、100μg/mlbestatin与10nmol/LATRA联合处理8小时后,NB4细胞c-Myc蛋白表达水平明显降低,与单用ATRA组相比差异显著(P<0.05);药物联用各组NB4细胞c-Myc蛋白表达水平与NBT还原能力之间呈负相关(r=-0.940,p=0.017);与100μg/mlbestatin联用不能增强10nmol/LATRA上调c-EBPεmRNA的表达;50、75、100μg/mlbestatin单独处理对NB4细胞的分化无影响。结论:氨肽酶抑制剂bestatin能够增强ATRA对NB4细胞的诱导分化作用,这可能与bestatin协同ATRA下调c-myc的表达有关。This study was purposed to investigate whether aminopeptidase inhibitor, bestafin, can potentiate all-trans retinoic acid （ATRA） -inducing differentiation in NB4 cells, and to explore its mechanism. The NB4 cells were exposed to either bestatin and ATRA alone or in combination, the morphological changes of NB4 cells were observed by optical microscopy, the CDllb expression was measured by flow cytometry, the function of defferentiation cells was analyzed by nitroblue-tetrazolium （NBT） reduction assay, the mRNA expressions of c-myc and c-EBPe in NB4 cells were detected by RT-PCR, the c-Myc protein expression was determined by Western blot. The results showed that treatment with bestatin alone induced no significant changes in morphology, NBT reduction activity and CDllb expression in NB4 cells. NB4 cells incubated with 10 nmol/L ATRA plus 100μg/ml bestatin showed more morphologic feature of metamyelocyte and band neutrophil than ATRA alone treated cells. 100μg/ml bestatin enhanced the NBT reduction activity in NB4 cells induced by various concentrations of ATRA （ 10, 20, 40 nmol/L）. The effects of various concentrations of ATRA in combination with 100μg/ml bestatin were statistically different from the effect of ATRA alone （ P 〈 0. 01 ）. From 48 to 96 hours, 100μg/ml bestatin time-dependendy increased NBT reduction in NB4 cells induced by 10 nmol/L ATRA （ P 〈0.01 ）. 10 nmol/L ATRA plus 100 μg/ml bestadn for 72 hours prominendy elevated CDllb expression in NB4 cells as compared with ATRA alone treated NB4 cells （ P 〈 0.01 ）. There was a substantial decrease in c-myc mRNA levels when 100μg/ml bestatin was added to 10 nmol/L ATRA （ P 〈 0.05 ）. Various concentrations （50, 75, 100 i^g/ml） of bestatin combined with 10 nmol/L ATRA down-regulated the expression of c-Myc protein, which was negatively correlated with the NBT reduction activity of NB4 cells induced by 10 nmol/L ATRA alone or plus bestatin at various concentrations （r= -0.940, P = 0. 017 ）. However, 100μg/ml

关 键 词：BESTATIN ATRA 细胞株 NB4 细胞分化 

分 类 号：R733.71[医药卫生—肿瘤] R979.1[医药卫生—临床医学]