Direct comparison of efficiency and stability of gene transfer into the mammalian heart using adeno-associated virus versus adenovirus vectors
Direct comparison of efficiency and stability of gene transfer into the mammalian heart using adeno-associated virus versus adenovirus vectors
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DOI：
10.1016/s0022-5223(03)00082-5
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发表时间：
2003-09-01
影响因子：
6
通讯作者：
Thistlethwaite, PA
中科院分区：
文献类型：
--
作者：
Chu, D;Sullivan, CC;Thistlethwaite, PA
目的：目前的基因治疗策略依赖于使用腺病毒或质粒作为载体将基因递送到心脏。这些方法受到低转导频率和瞬时转基因表达的限制。我们试图确定是否腺相关病毒产生更稳定，更高效率的基因表达在啮齿动物的心脏比以前的常规methods.Methods：两个重组病毒构建体：腺相关病毒含有lacZ基因的巨细胞病毒启动子（AAV-lacZ）和腺病毒表达lacZ的控制下相同的启动子（腺病毒lacZ）。向20只大鼠注射（心室尖）每种病毒的1 × 10（7-8）个基因组颗粒。动物被置于死亡在一系列的时间点和转基因表达定量β-半乳糖苷酶活性，心肌染色，和Western blot蛋白质analysis.Results：三个月后，腺相关病毒基因转移，动物表现出稳定的β-半乳糖苷酶表达在60%的心肌细胞没有心肌炎症/坏死的证据。3个月时蛋白表达的分布和程度以及阳性细胞的数量与4周时的转基因表达相当。在心脏以外的器官中未检测到腺相关病毒。相比之下，Adeno-lacZ动物在60%的心肌细胞中显示出短暂的β-半乳糖苷酶活性，这在基因转移后4周无法检测到。腺病毒治疗的动物表现出显着的心肌炎症和转基因expression in other organs.Conclusion：直接心肌内注射腺相关病毒载体程序稳定，长期，心脏特异性转基因表达在啮齿动物心脏长达3个月。我们的研究结果表明，与腺病毒载体相比，腺相关病毒在心脏长期转基因表达方面具有显着优势。
Objective: Recent gene therapy strategies have relied on the use of adenovirus or plasmid as vehicles for gene delivery to the heart. These approaches have been limited by low transduction frequencies and transient transgene expression. We sought to determine whether adeno-associated virus produces more stable, higher efficiency gene expression in the rodent heart than did previous conventional methods.Methods: Two recombinant viral constructs were made: an adeno-associated virus containing the lacZ gene under the control of the cytomegalovirus promoter (AAV-lacZ) and an adenovirus expressing lacZ under the control of the same promoter (Adeno-lacZ). Twenty rats were injected (into the ventricular apex) with 1 X 10(7-8) genomic particles of each virus. Animals were put to death at serial time points and transgene expression quantitated by beta-galactosidase activity, myocardial staining, and Western blot protein analysis.Results: Three months after adeno-associated virus gene transfer, animals demonstrated stable beta-galactosidase expression in 60% of cardiomyocytes without evidence of myocardial inflammation/necrosis. The distribution and degree of protein expression and number of positive cells at 3 months were equivalent to transgene expression at 4 weeks. Adeno-associated virus was not detected in organs other than the heart. In contrast, Adeno-lacZ animals displayed transient beta-galactosidase activity in 60% of cardiomyocytes, which was undetectable 4 weeks after gene transfer. Adenovirus-treated animals manifest significant myocardial inflammation and had transgene expression in other organs.Conclusion: Direct intramyocardial injection of an adeno-associated virus vector programs stable, long-term, cardiac-specific transgene expression in the rodent heart for up to 3 months. Our results suggest adeno-associated virus has significant advantages for long-term transgene expression in the heart compared to adenovirus vectors.