Xu L, Ferry A E, Monteiro C, Pace B S
Department of Biology, University of South Alabama, Mobile, AL 36688, USA.
Gene Ther. 2000 Mar;7(5):438-44. doi: 10.1038/sj.gt.3301106.
Sickle cell disease is caused by a mutation in the beta globin gene leading to hemoglobin S (Hb S) production. Several approaches have been explored to prevent Hb S polymerization in red blood cells and the symptoms associated with this disorder. To this end we tested a mammalian expression vector carrying a human beta globin antisense cDNA (pZeobetaAS) fragment in a mouse erythroleukemia cell line expressing the human gamma and beta globin genes. We observed a relative reduction in beta globin mRNA levels compared with gamma mRNA levels in the presence of pZeobetaAS. Moreover, analysis at the protein level showed an average 76% decrease in beta chains and a 517% increase in gamma chain biosynthesis. The inhibitory effect of the antisense vector on globin expression was maintained long term in culture. The expression vector pZeobetaAS was also transfected into primary erythroid progenitors to test its effects on globin genes undergoing normal developmental switching during differentiation. We observed a relative reduction of beta globin mRNA levels compared with gamma mRNA levels. These results support a novel role for antisense cDNA expression vectors as an alternative gene therapy strategy to inhibit betas gene expression in sickle cell disease. Gene Therapy (2000) 7, 438-444.
镰状细胞病是由β珠蛋白基因突变导致血红蛋白S（Hb S）产生引起的。人们探索了多种方法来防止红细胞中Hb S聚合以及预防与这种疾病相关的症状。为此，我们在表达人γ和β珠蛋白基因的小鼠红白血病细胞系中测试了携带人β珠蛋白反义cDNA（pZeobetaAS）片段的哺乳动物表达载体。在存在pZeobetaAS的情况下，我们观察到β珠蛋白mRNA水平相对于γmRNA水平有所降低。此外，蛋白质水平分析显示β链平均减少76%，γ链生物合成增加517%。反义载体对珠蛋白表达的抑制作用在培养中长期维持。表达载体pZeobetaAS也被转染到原代红系祖细胞中，以测试其对分化过程中经历正常发育转换的珠蛋白基因的影响。我们观察到β珠蛋白mRNA水平相对于γmRNA水平有所降低。这些结果支持了反义cDNA表达载体作为一种新型基因治疗策略在镰状细胞病中抑制βs基因表达的作用。《基因治疗》（2000年）7卷，438 - 444页
