A low rate of cell proliferation and reduced DNA uptake limit cationic lipid-mediated gene transfer to primary cultures of ciliated human airway epithelia
A low rate of cell proliferation and reduced DNA uptake limit cationic lipid-mediated gene transfer to primary cultures of ciliated human airway epithelia
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DOI：
10.1038/sj.gt.3300524
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发表时间：
1997-11-01
期刊：
影响因子：
5.1
通讯作者：
Welsh, MJ
中科院分区：
文献类型：
--
作者：
Fasbender, A;Zabner, J;Welsh, MJ
DNA和阳离子脂质的复合物为基因转移到气道上皮细胞提供了潜在的优势。然而，我们发现，应用DNA-脂质（DMRIE-DOPE）复合物的人纤毛气道上皮细胞或兔气管外植体的原代培养只产生低水平的基因转移。相比之下，当我们在接种后不久将DNA-脂质应用于未成熟的人上皮细胞时，转基因表达显著更高。我们发现了两个限制基因转移的障碍。首先，在接种后，DNA-脂质复合物穿过顶膜进入气道细胞的摄取随着时间的推移而迅速下降，并抑制了转基因表达的下降。第二，细胞分裂随着接种后的时间而减少，我们发现有丝分裂中的细胞（用BrdU标记）比BrdU阴性细胞更可能表达转基因。这些数据表明，跨顶膜的进入步骤和细胞分裂的低速率是阳离子脂质介导的基因转移到气道上皮细胞的重要障碍。试图改变这两个过程可能会提高基因转移到囊性纤维化气道的效率。
Complexes of DNA and cationic lipid offer potential advantages for gene transfer to airway epithelia. However, we found that application of DNA-lipid (DMRIE-DOPE) complexes to primary cultures of human ciliated airway epithelia or explants of rabbit trachea generated only low levels of gene transfer. In contrast, when we applied the DNA-lipid to immature human epithelia shortly after seeding, transgene expression was substantially higher. We identified two barriers that limit gene transfer. First, uptake of the DNA-lipid complexes into airway cells across the apical membrane decreased rapidly with time after seeding and paralleled the decrease in transgene expression. Second, cell division decreased with time after seeding, and we found that cells in mitosis (labeled with BrdU) were much more likely to express transgene than BrdU-negative cells. These data suggest that the entry step across the apical membrane and the low rate of cell division are important barriers for cationic lipid-mediated gene transfer to airway epithelia. Attempts to modify these two processes may yield improvements in the efficiency of gene transfer to the airways in cystic fibrosis.