Mastakov M Y, Baer K, Xu R, Fitzsimons H, During M J
Functional Genomics and Translational Neuroscience Laboratory, Division of Molecular Medicine, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand.
Mol Ther. 2001 Feb;3(2):225-32. doi: 10.1006/mthe.2001.0246.
Recombinant adeno-associated viruses (rAAV) are highly efficient vectors for gene transfer into the central nervous system (CNS). However, a major hurdle for gene delivery to the mammalian brain is to achieve high-level transduction in target cells beyond the immediate injection site. Therefore, in addition to improvements in expression cassettes and viral titers, optimal injection parameters need to be defined. Here, we show that previous studies of somatic cell gene transfer to the mammalian brain have used suboptimal injection parameters, with even the lowest reported perfusion rates still excessively fast. Moreover, we evaluated the effect of local administration of mannitol to further enhance transgene expression and vector spread. Ultraslow microperfusion of rAAV, i.e., <33 nl/min, resulted in significantly higher gene expression and less injury of surrounding tissue than the previously reported rates of 100 nl/min or faster. Co-infusion of mannitol facilitated gene transfer to neurons, increasing both the total number and the distribution of transduced cells by 200-300%. Gene transfer studies in the CNS using rAAV should use very slow infusion rates and combined injection with mannitol to maximize transduction efficiency and spread.
重组腺相关病毒（rAAV）是将基因导入中枢神经系统（CNS）的高效载体。然而，将基因递送至哺乳动物大脑的一个主要障碍是要在紧邻注射部位以外的靶细胞中实现高水平转导。因此，除了改进表达盒和病毒滴度外，还需要确定最佳注射参数。在这里，我们表明，以往对哺乳动物大脑进行体细胞基因转移的研究使用的注射参数并不理想，即使是报道的最低灌注速率仍然过快。此外，我们评估了局部施用甘露醇以进一步增强转基因表达和载体扩散的效果。rAAV的超慢速微灌注，即<33 nl/分钟，与先前报道的100 nl/分钟或更快的速率相比，导致基因表达显著更高，周围组织损伤更小。甘露醇的共同输注促进了基因向神经元的转移，使转导细胞的总数和分布增加了200-300%。使用rAAV在CNS中进行基因转移研究应采用非常缓慢的输注速率，并与甘露醇联合注射，以最大限度地提高转导效率和扩散。
