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RNA干扰抑制HOXA7表达能逆转白血病U937细胞多药耐药 被引量：4

The reversal effect of RNA interference targeting HOXA 7 gene on multi-drug resistance of leukemia cell line U937
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摘要 目的:运用RNA干扰(RNA interference,RNAi)技术抑制同源盒基因A7(homeobox geneA7,HOXA7)表达,并探讨其对白血病U937细胞多药耐药的逆转作用。方法:将靶向HOXA7基因的特异性真核表达载体(pGPU6/GFP/Neo-shHOXA7)及阴性对照载体(pGPU6/GFP/Neo-shNC)分别转染U937细胞,G418稳定筛选。实验分3组:实验组(稳定表达pGPU6/GFP/Neo-shHOXA7的U937细胞)、阴性对照组(稳定表达pGPU6/GFP/Neo-shNC的U937细胞)和空白对照组(U937细胞)。RT-PCR和蛋白质印迹法分别在mRNA和蛋白质水平检测各组细胞中HOXA7的表达情况。MTT法检测各组细胞对阿糖胞苷(cytarabine,Ara-C)和高三尖杉酯碱(homo harringtonine,HHT)的敏感性。FCM法分别检测Ara-C(10μg/mL)和HHT(0.5μg/mL)作用下各组细胞的凋亡情况。结果:实验组HOXA7在mRNA和蛋白质水平的相对表达量明显低于阴性对照组和空白对照组(P<0.05)。实验组Ara-C和HHT的半数抑制浓度(half-inhibitory concentration,IC50)较阴性对照组和空白对照组的IC50分别降低了3.5倍和8.5倍(P<0.05),提示实验组细胞对Ara-C和HHT的敏感性增加。实验组细胞分别经Ara-C(10μg/mL)和HHT(0.5μg/mL)诱导后,细胞凋亡率明显高于相同药物作用下阴性对照组和空白对照组的细胞凋亡率(P<0.05)。结论:RNAi抑制HOXA7表达能增强Ara-C和HHT对U937细胞的增殖抑制及凋亡诱导作用,在一定程度上可逆转白血病细胞的多药耐药性。 Objective： To investigate the effect of RNA interference targeting HOXA7 （homeobox A7） gene on the multi-drug resistance of leukemia U937 cells. Methods： U937 cells were transfected with the specific eukaryotic expression vectors carrying shRNA （small hair RNA） targeting HOXA7 （pGPU6/GFP/Neo-shHOXA7） and the negative control vectors （pGPU6/GFP/Neo-shNC）, respectively. The stably-transfected cells were selected by G418. There were three groups in the design, including the U937 cells stably expressing pGPU6/GFP/Neo-shHOXA7 or pGPU6/GFP/Neo-shNC and the U937 cells without transfection as a blank control. The downregulation of HOXA7 expression in U937 cells transfected with pGPU6/GFP/Neo-shHOXA7 was confirmed by RT-PCR （reverse transcription PCR） and Western blotting. The sensitivities of U937 cells to Ara-C （cytarabine） and HHT （homoharringtonine） were detected by MTT method. The apoptosis of U937 cells treated with Ara-C （10 μg/mL） or HHT （0.5 μg/mL） was determined by FCM （flow cytometry）. Results： The expressions of HOXA7 gene at mRNA and protein levels were both significantly lower in the U937 cells transfected with pGPU6/GFP/Neo-shHOXA7 than those in the blank control cells （P 〈 0.05）. The ICs50 （half-inhibitory concentrations） of Ara-C and HHT in U937 cells transfected with pGPU6/GFP/Neo-shHOXA7 were decreased by 3.5- and 8.5-fold comparing with those in the blank control cells, respectively （P 〈 0.05）. The apoptotic rate of the U937 cells transfected with pGPU6/GFP/Neo-shHOXA7 after treatment with Ara-C （10 μg/mL） or HHT （0.5 μg/mL） was higher than those of the control cells （P 〈 0.05）. Conclusion： The inhibition of HOXA7 gene expression by RNA interference can enhance the effects of Ara-C and HHT on promoting apoptosis and inhibiting proliferation of the U937 cells, which may partly reverse the multidrug resistance of leukemia cells.
作者 尹宝慧 贾秀红 李建厂
机构地区 滨州医学院附属医院儿科
出处 《肿瘤》 CAS CSCD 北大核心 2013年第7期585-591,共7页 Tumor
基金 山东省科学技术发展计划项目(编号:2010GSF10264)
关键词 白血病 实验性 基因 同源盒 RNA干扰 抗药性 多药 U937细胞 Leukemia, experimental Genes, homeobox RNA interference Drug resistance, multiple U937 cells
分类号 R733.7 [医药卫生—肿瘤]
  • 相关文献

参考文献11

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共引文献2

同被引文献32

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肿瘤
2013年 第7期

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