OAZ1基因诱导慢粒白血病K562细胞向成熟红系方向分化  

Effect of Ornithine Decarboxylase Antizyme 1 on the Erythroid Differentiation in Human Leukemia Cell Line K562

作　　者：吴兵平[1] 王星[2] 马文丽[1] 张思毅[2] 郑文岭[1] 姜立[2] 

机构地区：[1]南方医科大学基因工程研究所,广州510515 [2]广东省医学科学院,广东省人民医院,广州510080

出　　处：《中国生物化学与分子生物学报》2013年第4期361-367,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金项目(No.30901757);广东省自然科学基金(No.10151008004000028);广东省领军人才项目资助~~

摘　　要：近年来,鸟氨酸脱羧酶抗酶(OAZ)作为肿瘤治疗的潜在靶点备受关注.本文研究了OAZ1基因过表达对慢粒白血病K562细胞红系分化的作用.构建框移位点突变的OAZ1过表达慢病毒载体pLVX-Neo-OAZ1-IRES-ZsGreen,包装病毒并感染K562细胞,Western印迹验证其过表达效果.FACS检测细胞分化标志物CD71和GPA,结合联苯胺染色分析细胞红系分化情况.对比氯化高铁血红素(hemin)诱导组,实时RT-PCR检测与K562细胞红系分化、癌变的关键基因(GATA1、BCR/ABL、TGFβ)转录水平,对OAZ1诱导分化的机制进行初步探索.结果表明,慢病毒过表达载体及K562细胞过表达体系构建成功.OAZ1过表达后细胞红系分化标志物CD71+/GPA+为(11.22±2.09)%,与对照组(4.07±1.04)%、空病毒组(1.79±2.36)%相比差异极显著(P<0.01);联苯胺蓝染阳性率为(14.037±0.083)%,与对照组、空病毒组比较,差异也极显著(P<0.01).定量分析结果提示,相对于GATA1、BCR/ABL基因mRNA转录水平的影响,OAZ1对TGFβ基因的作用更为明显.为此推断,OAZ1基因可诱导白血病K562细胞向成熟红系方向分化,其作用机制可能与TGFβ信号转导通路相关.Recently, ornithine decarboxylase antizyme （OAZ1） has been much concerned for its potentials in anti-tumor. To investigate the role of OAZ1 in inducing the erythroid differentiation in human leukemia cell line K562, the lentivirus vector pLVX-Neo-OAZI-IRES-ZsGreen targeting gene OAZ1 modified at the frameshift site was constructed to transfect K562 cells. OAZ1 overexpression efficiency was assessed by Western blotting. The potential effect of OAZ1 in inducing erythroid differentiation were obtained by detecting two surface markers （CD71 and GPA） and the benzidine staining. To explore the induction mechanism, the relevant genes （ GATA1, BCR/ABL and TGF~） related with the differentiation and cancer formation of leukemia were detected by real-time RT-PCR adding the hemin-induced group. The results showed that recombinant lentiviral vector and the overexpression cell line were successfully constructed. The positive rate of double labeling （CD71＋/GPA~） was （11.22 _＋ 2.09）% in cells transfected with OAZ1 lentivirus, which was significantly higher than that in the untreated cells （4.07 ＋ 1.04）% or in the treated cells with empty virus （ 1.79 ＋ 2.36）% （P 〈 0.01 ）. The positive rate of benzidine staining was （ 14. 037 ＋ 0. 083） % , which was also significantly higher than that in other two groups （P 〈0.01 ）. Quantitative results indicated that OAZ1 had more siginificant function on TGFfl than that in GATA1 or BCR/ABL. In conclusion, OAZ1 eould effectively induce the erythroid differentiation in K562 cells and this induction is possiblely related to TGFβ signaling pathway.

关 键 词：鸟氨酸脱羧酶抗酶-1(OAZ1) 慢粒白血病 红系分化 

分 类 号：R34[医药卫生—基础医学]