Toyoda K, Ooboshi H, Chu Y, Fasbender A, Davidson B L, Welsh M J, Heistad D D
Departments of Internal Medicine, Physiology and Biophysics, and Pharmacology, Cardiovascular Center and Center on Aging, University of Iowa College of Medicine, Iowa City, Iowa.
Stroke. 1998 Oct;29(10):2181-8. doi: 10.1161/01.str.29.10.2181.
Improvement of efficiency of gene transfer to endothelium could be useful for several applications. We tested the hypothesis that cationic nonviral molecules augment adenovirus-mediated gene transfer to blood vessels, perhaps by alteration of the surface charge of adenovirus and facilitation of binding to endothelium.
Carotid arteries from rabbits were incubated in vitro for 0.5 to 2 hours with an adenoviral vector alone or noncovalent complexes of adenovirus with poly-L-lysine (a cationic polymer) or lipofectin (a cationic lipid). Binding of adenovirus to the vessels was evaluated immediately after incubation with virus, and assay of transgene (ss-galactosidase) activity and histochemistry were performed 24 hours after gene transfer. To determine whether cationic molecules can be used to augment alteration of vascular function by adenovirus-mediated gene transfer, we also examined effects on gene transfer of endothelial nitric oxide synthase.
Assay of ss-galactosidase activity indicated that both cationic molecules increased transgene expression in vessels by approximately 5- to 6-fold. In contrast, when endothelium was removed from the vessels after gene transfer, poly-L-lysine and lipofectin did not significantly increase transgene activity. Histochemistry for ss-galactosidase also suggested that the adenovirus-cationic molecule complexes augmented transgene expression mainly in the endothelium. In addition, we found that complexing adenovirus with cationic molecules increased binding of adenovirus to the vessels. After gene transfer with recombinant adenovirus containing endothelial nitric oxide synthase, calcium ionophore (A23187) produced greater relaxation of vessels treated with adenovirus complexed with poly-L-lysine or lipofectin than those treated with adenovirus alone.
Cationic molecules improve the efficiency of adenovirus-mediated gene transfer to blood vessels.
提高基因向内皮细胞转移的效率在多个应用领域可能会有所帮助。我们检验了这样一个假设，即阳离子非病毒分子可增强腺病毒介导的基因向血管的转移，这可能是通过改变腺病毒的表面电荷并促进其与内皮细胞的结合来实现的。
将兔颈动脉在体外分别与单独的腺病毒载体、腺病毒与聚-L-赖氨酸（一种阳离子聚合物）或脂质体（一种阳离子脂质）的非共价复合物孵育0.5至2小时。在与病毒孵育后立即评估腺病毒与血管的结合情况，并在基因转移24小时后进行转基因（β-半乳糖苷酶）活性测定和组织化学分析。为了确定阳离子分子是否可用于增强腺病毒介导的基因转移对血管功能的改变，我们还研究了其对内皮型一氧化氮合酶基因转移的影响。
β-半乳糖苷酶活性测定表明，两种阳离子分子均使血管中的转基因表达增加了约5至6倍。相比之下，在基因转移后去除血管内皮时，聚-L-赖氨酸和脂质体并未显著增加转基因活性。β-半乳糖苷酶的组织化学分析也表明，腺病毒-阳离子分子复合物主要在内皮细胞中增强了转基因表达。此外，我们发现将腺病毒与阳离子分子复合可增加腺病毒与血管的结合。在用含内皮型一氧化氮合酶的重组腺病毒进行基因转移后，钙离子载体（A23187）对与聚-L-赖氨酸或脂质体复合的腺病毒处理的血管产生的舒张作用比对单独用腺病毒处理的血管更强。
阳离子分子可提高腺病毒介导的基因向血管转移的效率。
