中文摘要：

目的研究脂质体转染反义寡核苷酸（ASON）对重型β-地中海贫血（β-地贫）患者红系细胞α-珠蛋白基因的调控作用，为基因治疗β-地贫提供新的思路。方法将前期实验筛选获得的高效抑制α-珠蛋白基因表达的ASON以最佳浓度作用于体外培养的重型β-地贫患者红系细胞，并设空白对照组，经实时荧光定量RT—PCR分析ASON组和空白对照组中红系细胞α、β、γ-珠蛋白基因表达情况，观察ASON对相应珠蛋白基因表达的影响，通过高效液相色谱法（HPLC）检测ASON作用后血红蛋白水平，并经透射电子显微镜（电镜）观察ASON作用前后红系细胞内α-珠蛋白肽链沉积情况，了解ASON对β-地贫患者红系细胞内过剩α-珠蛋白肽链的影响。结果实时荧光定量RT—PCR结果显示ASON组α-珠蛋白基因mRNA表达（9．04±0．29）较空白对照组（24．23±0．29）减低（P〈0．01），α/（β＋γ）珠蛋白比例失衡得到改善[ASON组、空白对照组分别为（0．79±0．02）、（2．26±0．06），P〈0．01]；HPLC检测表明ASON作用后仅-珠蛋白基闪表达下调，HbA：和HbF表达增加；透射电镜结果证实ASON作用后重型β-地贫患者红系细胞内过剩的α-珠蛋白肽链沉积减少，特别是在早期幼稚红细胞中，在空白对照组中沉积物无明显变化。结论高效ASON可抑制体外培养的重型β-地贫患者红系细胞α-珠蛋白基因表达，有助于改善珠蛋白基因α（β＋γ）比例失衡，并减少过剩α-珠蛋白肽链在红系细胞内的沉积，为基因治疗β-地贫提供新的思路，

英文摘要：

Objective To study the effect of liposomal transfection of antisense oligonucleotide（ASON） on the erythroid cell α-globin gene in the patients with severe β-thalassemia, and provide a new idea for β- thalassemia gene therapy. Methods A highly effective ASON targeting α-globin gene was transfected into severe β-thalassemic erythroid cells cultured in vitro by liposomal at an optimal concentration. The expression level of α,β,γ-globin gene, the level of hemoglobin, and the excess α-globin chains precipitates in ASON group and control group were carefully analyzed by quantitative real-time PCR（ Q-RT-PCR）, high performance liquid chromatography （HPLC） , and electron microscope, respectively. Results The mRNA expression of α-globin gene was significantly lower in ASON group （9.04 ±0.29） than in control group （24.23 ±0.29）（P 〈0.01 ）. Simultaneously, the disqulibrinm between α- and β-, γ-globin gene expression was partly modified by ASON, the ratios of ASON group and control group being 0.79 ± 0. 02 and 2.26 ±0.06 respectively（ P 〈 0.01 ）. HPLC demonstrated that the levels of HbA2 and HbF increased with downregulation of α-globin gene in β-thalassemic erythroid cells, particularly HbF. The precipitates of α-globin chains in ASON group were lessened under electron microseope, particularly in early erythroblast while no change in the control group. Conlusion The high effective ASON contributes to inhibit the α-globin gene expression of severe β-thalassemic erythroid cells, partly modify the disqulibfium between α-, β- and γ-globin gene expression and obviously reduce the precipitates of α- globin chains in elythroid cells. It might provide a new idea for gene therapy of β-thalassemia.