Martin Keith R G, Klein Ronald L, Quigley Harry A
Wilmer Eye Institute, Wilmer 122, Johns Hopkins Hospital, 600 North Wolfe Street, Baltimore, MD 21287, USA.
Methods. 2002 Oct;28(2):267-75. doi: 10.1016/s1046-2023(02)00232-3.
Adeno-associated virus (AAV) vectors provide a useful way to deliver genes to the eye. They have a number of important properties which make them suitable for this purpose, not least their lack of significant pathogenicity and the potential for long-term transfection of retinal cells. The optimal methods for AAV-mediated gene delivery are determined by the location and characteristics of the target cell type. Efficient gene delivery to photoreceptors and pigment epithelial cells following subretinal injection of AAV has been achieved in various animal models. AAV-mediated gene therapy has been shown to slow photoreceptor loss in rodent models of primary photoreceptor diseases and in dogs with a naturally occurring disease similar to human Leber's congenital amaurosis (LCA). Efficient gene delivery to other cell types such as retinal ganglion cells (RGCs), however, has been more problematic. In this article, we review the potential uses of AAV-mediated gene delivery to the eye. We describe the selection of an appropriate AAV vector for ocular gene transfer studies and discuss the techniques used to deliver the virus to the eye and to assess ocular transfection. We emphasize our techniques for successful gene transfer to retinal ganglion cells, which have often proven challenging to transfect with high efficiency. Using a modified AAV incorporating a chicken beta-actin (CBA) promoter and the woodchuck hepatitis posttranscriptional regulatory element, we describe how our techniques allow approximately 85% of rat retinal ganglion cells to be transfected within 2 weeks of a single intravitreal virus injection. Our techniques facilitate the study of the pathogenesis of RGC diseases such as glaucoma and the development of novel new treatments based on gene therapy.
腺相关病毒（AAV）载体为将基因递送至眼部提供了一种有用的方法。它们具有许多重要特性，使其适用于此目的，尤其是它们缺乏显著的致病性以及对视网膜细胞进行长期转染的潜力。AAV介导的基因递送的最佳方法取决于靶细胞类型的位置和特征。在各种动物模型中，通过视网膜下注射AAV已实现向光感受器和色素上皮细胞的高效基因递送。在原发性光感受器疾病的啮齿动物模型以及患有与人类莱伯先天性黑蒙（LCA）相似的自然发生疾病的犬类中，AAV介导的基因治疗已被证明可减缓光感受器的丧失。然而，向其他细胞类型如视网膜神经节细胞（RGCs）的高效基因递送则更具问题。在本文中，我们综述了AAV介导的基因递送至眼部的潜在用途。我们描述了用于眼部基因转移研究的合适AAV载体的选择，并讨论了将病毒递送至眼部以及评估眼部转染的技术。我们强调了我们成功将基因转移至视网膜神经节细胞的技术，而视网膜神经节细胞的高效转染常常被证明具有挑战性。使用一种包含鸡β-肌动蛋白（CBA）启动子和土拨鼠肝炎转录后调控元件的改良AAV，我们描述了我们的技术如何在单次玻璃体内注射病毒后2周内使约85%的大鼠视网膜神经节细胞被转染。我们的技术有助于青光眼等视网膜神经节细胞疾病发病机制的研究以及基于基因治疗的新型治疗方法的开发。
