Ad5F35嵌合腺病毒载体介导的突变性IκBα对白血病细胞凋亡的诱导作用  

Chimeric Ad5F35 Adenoviral Vector-mediated Expression of Mutant IκBα Induces Apoptosis of Leukemia Cells

作　　者：王光平[1] 王凯[2] 信红亚[1] 段朝军[3] 荆照正[3] 谭三勤[1] 齐振华[1] 陈方平[1] 

机构地区：[1]中南大学湘雅医院血液科 [2]海军总医院儿科,北京100037 [3]中南大学医学实验中心,湖南长沙410008

出　　处：《中国实验血液学杂志》2011年第2期332-336,共5页Journal of Experimental Hematology

基　　金：国家自然科学基金资助(编号30270575)

摘　　要：多种不同类型的白血病均存在持续活化的核转录因子κB(NF-κB),而抑制NF-κB活化诱导细胞凋亡有可能成为治疗白血病新的方法。本研究探讨修饰的5型腺病毒载体即Ad5F35嵌合腺病毒载体(Ad5F35 Vec)介导的突变性IκBα(IκBαDN)对白血病细胞凋亡的作用。将携带缺失5'端1-70个氨基酸编码序列的人IκBαAd5F35-IκBαDN Vec转染HL-60细胞。采用流式细胞术、DNA结合试验、实时定量PCR和Western blot技术分别检测细胞凋亡、NF-κB DNA结合活性、IκBα,cIAP-2和xIAP的表达。结果显示,转染48小时后,转染Ad5F35-IκBαDN Vec细胞、空白载体Ad5F35-EGFP Vec细胞以及未转染细胞的凋亡率分别为(22.53±2.999)%,(6.08±2.464)%和(4.86±1.366)%,其差异具有显著意义(Ad5F35-IκBαDN Vec vs未转染:p<0.001;Ad5F35-IκBαDNVec vs Ad5F35-EGFP Vec:p<0.001,p<0.002)。同时,NF-κB DNA结合活性降低,IκBα表达增加,但cIAP-2和xIAP mRNA的表达降低。结论:Ad5F35 Vec能够有效介导IκBαDN cDNA在HL-60细胞的表达,IκBαDN可抑制HL-60细胞NF-κB DNA结合活性并诱导其凋亡。Constitutive activation of nuclear transcription factor-κB（NF-κB） exists in a variety of leukemia,and induction of apoptosis through blocking NF-κB activation may be an alternative strategy for leukemia treatment.The aim of this study was to investigate the inducing effect of modified adenovirus 5-based adenovirus vector（i.e.chimeric Ad5F35 Vec）-mediated expression of mutant IκBα（IκBαDN） on apoptosis of HL-60 cells.The recombinant Ad5F35-IκBαDN Vec carrying IκBαDN cDNA which deleted the first 1-70 amino acids coding sequences at 5′ terminal of human IκBα was transfected into HL-60 cells.The apoptosis,NF-κB DNA binding activity,the expressions of IκBα,cIAP-2 and xIAP in HL-60 cells were detected by DNA binding assay,flow cytometry,real-time quantitative polymerase chain reaction and Western blot respectively.The results showed that apoptosis rates were 22.53±2.999%,6.08±2.464% and 4.86±1.366% for Ad5F35-IκBαDN Vec-infected or blank vector of Ad5F35-EGFP Vec-transfected and untransfected HL-60 cells respectively,which showed a significant difference between Ad5F35-IκBαDN Vec-transfected and untransfected cells（p〈0.001） and between Ad5F35-IκBαDN Vec-transfected and Ad5F35-EGFP Vec-transfected cells（p〈0.001,p〈0.002）,while NF-κB DNA binding activity was decreased,the truncated IκBα was expressed,and IκBα mRNA expression was up-regulated,but the expression of cIAP-2 and xIAP mRNA was down-regulated after transduction for 48 hours.It is concluded that the chimeric Ad5F35 Vec can effectively mediate the expression of IκBαDN cDNA in HL-60 cells,leading to the inhibition of NF-κB DNA binding activity and inducing apoptosis of HL-60 cells.

关 键 词：Ad5F35载体 NF-ΚB IΚBΑ 白血病 

分 类 号：R733.7[医药卫生—肿瘤]