Drawing Sticky Adeno-Associated Viruses on Surfaces for Spatially Patterned Gene Expression
Drawing Sticky Adeno-Associated Viruses on Surfaces for Spatially Patterned Gene Expression
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DOI：
10.1002/anie.201201495
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发表时间：
2012-01-01
影响因子：
16.6
通讯作者：
Jang, Jae-Hyung
中科院分区：
文献类型：
--
作者：
Kim, Eunmi;Song, In Taek;Jang, Jae-Hyung
在发育中的组织中，细胞外信号的空间控制分泌产生生物界面，其调节分化、增殖和迁移的细胞过程，并提供分子线索以组织组织结构。[1-4]已经开发了控制细胞外分子在基质上的空间分布的系统，以诱导细胞内诱导因子的模式化表达。[1，5，6]或者，空间模式化的基因递送已被用于克服蛋白质递送中发现的限制：短蛋白质半衰期和全身毒性。[7-9]体外模型系统的发展，模仿组织或器官中的基因表达的空间控制是阐明各种生物学机制的关键。然而，大多数基因递送系统依赖于将基因载体直接简单地添加到培养基中，这在其空间控制基因表达的能力方面固有地受到限制。微纳米纤维技术的进步使研究人员能够控制基因载体在表面上的位置。[10现有的方法包括微流体，[8，12]表面涂层，[13]和自组装。[14]通常，这些技术涉及多个费力的程序，通常是基底的化学活化、抗蚀剂沉积、使用光掩模的图案生成和基因载体固定。此外，往往需要昂贵的设备。与上述复杂的过程不同，本研究描述了一种简单，通用的空间模式化基因传递方法，灵感来自海洋贻贝的粘附。在紫贻贝的特殊粘附蛋白中发现的关键粘附部分--儿茶酚胺被用来配制"粘性"病毒。本研究中使用的粘合剂儿茶酚胺聚合物是聚（乙烯亚胺）-儿茶酚（PEI-C），其已用于碳纳米管纤维的材料无关的逐层组装和机械增强。[15腺相关病毒（AAV）是一种安全有效的细小病毒，[17]与PEIC复合。由于PEI-C的水下粘附特性，与PEI-C复合的AAV载体变成可以稳定粘附到表面上的高粘性病毒。最重要的是，通过使用粘性病毒载体，我们能够使用微量移液器作为"笔"在基质上创建病毒模式。这种"基因矢量绘制"技术绕过了繁琐的多个步骤，因此可以成为控制基因表达以建立复杂组织的通用平台。将支链PEI与3-（3，4-二羟基苯基）丙酸（DPA）缀合以产生PEI-C，其提供粘性病毒载体（图1A）。一种新型病毒AAVr3. 45，这是专门为神经元细胞设计的，
In developing tissues, the spatially controlled secretion of extracellular signals creates biointerfaces that modulate cellular processes of differentiation, proliferation, and migration, and provide molecular cues to organize the structure of tissues.[1–4] Systems that control spatial distribution of extracellular molecules on substrates have been developed in order to induce patterned expression of intracellular inductive factors.[1, 5, 6] Alternatively, spatially patterned gene delivery has been employed to overcome limitations found in protein delivery: short protein half-life and systemic toxicity.[7–9] The development of in vitro model systems that mimic the spatial control of gene expression in tissues or organs is critical to elucidate a variety of biological mechanisms. However, the majority of gene delivery systems rely on simple additions of gene carriers directly to media, which are inherently limited in their ability to spatially control gene expression. Advances in micro-and nanofabrication technologies have enabled researchers to control locations of gene vectors on surfaces.[10, 11] Existing methods include microfluidics,[8, 12] surface coating,[13] and self-assembly.[14] In general, the techniques involve multiple laborious procedures, typically chemical activation of substrates, resist depositions, pattern generation using a photomask, and gene vector immobilization. Furthermore, expensive equipment is often necessary. Unlike the aforementioned complex processes, this study describes a simple, versatile approach for spatially patterned gene delivery inspired by adhesion of marine mussels. Catecholamine, the key adhesive moiety found in the specialized adhesive proteins of mytilus edulis, was used to formulate “sticky” viruses. The adhesive catecholamine polymer used in this study is poly (ethylenimine)-catechol (PEI-C), which has been used for material-independent layerby-layer assembly and mechanical reinforcement of carbon nanotube fibers.[15, 16] The adeno-associated virus (AAV), which is a safe and efficient parvovirus,[17] was complexed with the PEIC. Because of the underwater adhesive property of PEI-C, the AAV vector complexed with the PEI-C became a highly sticky virus that can stably adhere onto surfaces. Most importantly, by using the sticky viral vectors, we were able to use a micropipette as a “pen” to create viral patterns on substrates. This “genevector drawing” technique bypasses laborious multiple steps and can thus be a versatile platform to control gene expression for the establishment of complex tissues. Branched PEI was conjugated with 3-(3, 4-dihydroxyphenyl) propionic acid (DPA) to generate PEI-C, which provides sticky viral vectors (Figure1A). A novel virus, AAVr3. 45, which was specifically designed for neuronal cell