SCL／TAL-1基因沉默对EPO诱导的K562细胞红系分化的影响  

The effect of knockdown of transcription factor SCL/TAL-1 gene on the erythroid differentiation in EPO.induced K562 cell line

作　　者：周睿卿[1] 龚玉萍[1] 郭勇[1] 单卿卿[1] 杨曦[1] 

机构地区：[1]四川大学华西医院血液科、生物治疗国家重点实验室,成都610041

出　　处：《中华血液学杂志》2012年第6期453-456,共4页Chinese Journal of Hematology

基　　金：国家自然科学基金（30770912）;四川省科技厅社会公益项目（2008SZ0017）;教育部回国留学人员启动基金（20071108-184）

摘　　要：目的通过RNA干扰技术降低SCL／TAL-1mRNA的表达，探讨SCL／TAL-1在K562细胞红系分化中的作用。方法以EPO诱导白血病细胞系K562细胞向红系分化为模型，将靶向SCL／TAL-1基因的特异性短发夹RNA（shRNA）慢病毒质粒pTRIP—dU3-RNAiTALh—EF1a—GFP转染K562细胞，RT—PCR检测其SCL／TAL—l和红系相关基因RhD、血型糖蛋白A（GPA，即CD253a）、CD47mRNA表达水平变化，流式细胞术分析红系分化抗原CD71、CD253a的表达，并以空载体质粒pTRIP—dU3-RNAi—luc—EF1-GFP转染的K652细胞为对照。结果①通过慢病毒转染系统将含SCL／TAL．1shRNA的质粒及空载体质粒转染K562细胞48h，绿色荧光蛋白（GFP）＋细胞占总细胞的95％以上，表明感染率达到95％以上。②RT—PCR结果显示转染特异性shRNA的K562细胞SCL／TAL-1mRNA表达水平比空载体对照明显下调（P〈0．05），红系抗原CIM7和RhDmRNA水平也比对照组明显下调（P〈0．05），GPA有所下降，但没有CIM7和RhD下降程度大。③流式细胞术检测到SCL／TAL-1低表达组红系分化抗原CD71、CD235a表达降低，表达率分别是10．4％、76．5％，而在对照组的表达率分别为94．3％、83．6％。结论研究结果提示转录因子SCL／TAL．1参与了红系定向分化。Objective To investigate the role of transcript factor SCL/TAL-1 gene in the erythroid differentiation through the knockdown of SCL/TAL-1 mRNA by RNA interference. Methods The plasmid of pTRIP-dU3-RNAiTALh-EFla-GFP with SCL/TAL1 shRNA was transfected into EPO-induced K562 cell line with erythroid differentiation via lentiviral vector system and the expression of SCL/TAL-1 mRNA decreased. The plasmid pTRIP-dU3- RNAiluc-EF1-GFP expressing EGFP gene was as control. The mRNA levels of SCL/TAL-1 and erythroid related RhD, GPA, CD47 in the cell lines were detected by RT-PCR, and erythroid antigen CD71, CD235a were examined by flow cytometry. Results （1） After 48 h of transfect, more than 95% of K562 cells were GFP positive, indicating the infection rate of the plasmids in the K562 cells more than 95%. （2）The results of RT-PCR showed SCL/TAL-1 mRNA expression in the K562 cell line of knockdown of SCL/TAL-1 was significantly lower than that in the control （ P 〈 0.05 ）. The mRNA levels of CD47 and RhD were also significantly lower, however, GPA decreased slightly in comparison with the control. （3）The expressions of CD71 and CD235a markedly reduced in the K562 cell line of knockdown of SCL/ TAL-1 with positive rates as 10.4% and 76.5% , while the positive rates in the control as 94.3% and 83.6%. Conclusion Our findings suggested that transcription factor SCL/TAL-1 might play an positive role in erythroid differentiation.

关 键 词：基因 SCL/TAL-1 细胞系 K562 细胞分化 RNA干扰 

分 类 号：R363[医药卫生—病理学]