Wuhan National Laboratory for Optoelectronics, Huazhong University of Science and Technology, Wuhan, 430074, China.
Center for Brain Science, State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Key Laboratory of Magnetic Resonance in Biological Systems, Wuhan Center for Magnetic Resonance, Wuhan Institute of Physics and Mathematics, Innovation Academy for Precision Measurement Science and Technology, Chinese Academy of Sciences, Wuhan, 430071, China.
Mol Brain. 2020 Jan 13;13(1):5. doi: 10.1186/s13041-020-0544-2.
Neurotropic viral transsynaptic tracing is an increasingly powerful technique for dissecting the structure and function of neural circuits. Herpes simplex virus type 1 strain H129 has been widely used as an anterograde tracer. However, HSV tracers still have several shortcomings, including high toxicity, low sensitivity and non-specific retrograde labeling. Here, we aimed to construct high-brightness HSV anterograde tracers by increasing the expression of exogenous genes carried by H129 viruses. Using a Trojan horse-like strategy, a HSV/AAV (adeno-associated virus) chimaera termed H8 was generated to enhance the expression of a fluorescent marker. In vitro and in vivo assays showed that the exogenous gene was efficiently replicated and amplified by the synergism of the HSV vector and introduced AAV replication system. H8 reporting fluorescence was brighter than that of currently available H129 tracers, and H8 could be used for fast and effective anterograde tracing without additional immunostaining. These results indicated that foreign gene expression in HSV tracers could be enhanced by integrating HSV with AAV replication system. This approach may be useful as a general enhanced expression strategy for HSV-based tracing tools or gene delivery vectors.
神经嗜性病毒转导示踪技术是一种日益强大的技术,可用于剖析神经回路的结构和功能。单纯疱疹病毒 1 型 H129 株已被广泛用作顺行示踪剂。然而,HSV 示踪剂仍存在一些缺点,包括高毒性、低灵敏度和非特异性逆行标记。在这里,我们旨在通过增加 H129 病毒携带的外源基因的表达来构建高亮度 HSV 顺行示踪剂。利用木马样策略,生成了一种 HSV/AAV(腺相关病毒)嵌合体,称为 H8,以增强荧光标记物的表达。体外和体内实验表明,外源基因通过 HSV 载体和引入的 AAV 复制系统的协同作用得到有效复制和扩增。H8 的报告荧光比目前可用的 H129 示踪剂更亮,并且 H8 可以用于快速有效的顺行示踪,而无需额外的免疫染色。这些结果表明,通过将 HSV 与 AAV 复制系统整合,可以增强 HSV 示踪剂中外源基因的表达。这种方法可能作为基于 HSV 的示踪工具或基因传递载体的通用增强表达策略有用。
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