A novel adeno-associated viral variant for efficient and selective intravitreal transduction of rat Müller cells.
A novel adeno-associated viral variant for efficient and selective intravitreal transduction of rat Müller cells.
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DOI：
10.1371/journal.pone.0007467
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发表时间：
2009-10-14
期刊：
影响因子：
3.7
通讯作者：
Schaffer DV
中科院分区：
文献类型：
--
作者：
Klimczak RR;Koerber JT;Dalkara D;Flannery JG;Schaffer DV
许多视网膜退行性疾病的病理可归因于多种遗传因素，而针对患者的个体化治疗选择有限且成本低。鉴于这些疾病具有共同的神经退行性表型，需要一种安全和广泛的神经保护方法来克服这些障碍。因此，将可分泌的神经保护因子基因输送到Müller细胞，这是一种接触所有类型视网膜神经元的视网膜胶质细胞，通过简单而无害的眼内或玻璃体内注射有效的载体，如腺相关病毒载体(AAV)，代表了一种理想的中介保护整个视网膜的方法。虽然已经分离出了几种自然产生的AAV变异体，它们具有不同的趋向性或细胞特异性，但这些载体通过玻璃体内注射低效地感染Müler细胞。我们之前已经应用定向进化创造了几个新的AAV变体，通过迭代选择一组高度多样化的AAV文库，能够有效地感染大鼠和人类星形胶质细胞。在此，这些分离的变异株的体内和体外特征确定了一个以前未报道的AAV变异株ShH10，它与AAV6血清类型(AAV6)密切相关，能够通过玻璃体内注射有效、选择性地感染Müler细胞。重要的是，与AAV2(>；60%)和AAV6相比，这种新的变种表现出显著的转导改进。我们的发现表明，AAV是一种高度通用的载体，能够从微小的序列变化中强烈地改变趋向性。这种分离的突变体代表了一种新的治疗载体，通过从Müler细胞分泌神经保护因子来治疗视网膜退行性疾病，并为研究它们在视网膜中的生物学功能提供了新的机会。
The pathologies of numerous retinal degenerative diseases can be attributed to a multitude of genetic factors, and individualized treatment options for afflicted patients are limited and cost-inefficient. In light of the shared neurodegenerative phenotype among these disorders, a safe and broad-based neuroprotective approach would be desirable to overcome these obstacles. As a result, gene delivery of secretable-neuroprotective factors to Müller cells, a type of retinal glia that contacts all classes of retinal neurons, represents an ideal approach to mediate protection of the entire retina through a simple and innocuous intraocular, or intravitreal, injection of an efficient vehicle such as an adeno-associated viral vector (AAV). Although several naturally occurring AAV variants have been isolated with a variety of tropisms, or cellular specificities, these vectors inefficiently infect Müller cells via intravitreal injection. We have previously applied directed evolution to create several novel AAV variants capable of efficient infection of both rat and human astrocytes through iterative selection of a panel of highly diverse AAV libraries. Here, in vivo and in vitro characterization of these isolated variants identifies a previously unreported AAV variant ShH10, closely related to AAV serotype 6 (AAV6), capable of efficient, selective Müller cell infection through intravitreal injection. Importantly, this new variant shows significantly improved transduction relative to AAV2 (>60%) and AAV6. Our findings demonstrate that AAV is a highly versatile vector capable of powerful shifts in tropism from minor sequence changes. This isolated variant represents a new therapeutic vector to treat retinal degenerative diseases through secretion of neuroprotective factors from Müller cells as well as provides new opportunities to study their biological functions in the retina.