靶向RNA干扰HOXA7对U937细胞增殖及凋亡的影响

Effects of RNA Interfere Targeting HOXA7 on Proliferation and Apoptosis of U937 Cells

目的运用RNA干扰技术抑制HOXA7表达,研究其对白血病细胞株U937增殖、凋亡的影响,为白血病基因治疗寻找新靶点。方法实验分3组:实验组、空白对照组、阴性对照组。构建靶向HOXA7特异性真核表达载体及阴性对照载体并转染U937细胞,利用RT-PCR和Western blot法分别检测各组细胞中HOXA7 mRNA和蛋白水平的表达情况;MTT法检测转染24、48、72 h后各组细胞增殖能力;流式细胞术检测转染48 h后各组细胞凋亡情况。结果实验组HOXA7表达明显受抑制,在mRNA和蛋白水平抑制率分别为:(47.314±7.394)%和(52.371±9.258)%;MTT结果显示:实验组24、48、72 h细胞增殖抑制率分别为(15.062±5.086)%、(30.052±4.016)%、(52.617±9.292)%,与同时间点空白对照组和阴性对照组相比差异均有统计学意义(P<0.05),且呈时间依赖性;流式结果显示:实验组细胞凋亡率为(24.677±4.161)%,明显高于空白对照组和阴性对照组(P<0.05)。结论靶向HOXA7特异性真核表达载体抑制HOXA7表达后,能有效抑制U937细胞增殖并促进其凋亡,HOXA7有望成为白血病基因治疗的新靶点。

Objective To study the effect of RNA interference on leukemia cell line U937 proliferation and apoptosis by inhibiting HOXA7 expression, to look for a new potential target for gene therapy for leukemia. Methods The experiment was divided into three groups: the experimental group, the blank control group and the negative control group. We created the specificity eukaryotic expression vector of targeting HOXA7 and the negative control vector first, then transfect them into U937 cells. The mRNA and protein expression of HOXA7 were determined by RT-PCR and Western blot. The proliferation of U937 in every group after 24, 48 and 72 h were detected by MTT. The apoptosis of U937 in every group after 48h was detected by flow cytometry. Results The HOXA7 expression was inhibited effectively, with the inhibition rates(47.314±7.394)% and(52.371±9.258)% on mRNA and protein levels in the experimental group,repectively. The MTT results showed the proliferation inhibition rate of 24, 48, 72 h were(15.062±5.086)%,(30.052±4.016)%,(52.617±9.292)% in the experimental group, respectively. Compared with the experimental group, prolferation inhibition rates of other two groups showed significant difference at the same time(P<0.05). Moreover, the inhibition effect of proliferation increased with extended time. The flow cytometry results showed the apoptosis rate of the experimental group was(24.677+4.161) %. And the differences of inhibitory rate and apoptosis rate in the experimental group were statistically significant compared with blank control group and negative control group(P<0.05). Conclusion The eukaryotic expression vector targeting HOXA7 could inhibit proliferation and promote apoptosis of U937 cells effectively after HOXA7 expression suppression. HOXA7 was expected to become the new target of leukemia gene therapy.