Luker Kathryn E, Hutchens Martha, Schultz Tracey, Pekosz Andrew, Luker Gary D
Department of Radiology, University of Michigan Medical School, Ann Arbor, MI 48109-0648, USA.
Virology. 2005 Oct 25;341(2):284-300. doi: 10.1016/j.virol.2005.06.049. Epub 2005 Aug 10.
Whole animal imaging allows viral replication and localization to be monitored in intact animals, which provides significant advantages for determining viral and host factors that determine pathogenesis. To investigate effects of interferons on spatial and temporal progression of vaccinia infection, we generated recombinant viruses that express firefly luciferase or a monomeric orange fluorescent protein. These viruses allow vaccinia infection to be monitored with bioluminescence or fluorescence imaging, respectively. The recombinant viruses were not attenuated in vitro or in vivo relative to a control WR virus. In cell culture, reporters could be detected readily by 4 h post-infection, showing that these viruses can be used as early markers of infection. The magnitude of firefly luciferase activity measured with bioluminescence imaging in vitro and in vivo correlated directly with increasing titers of vaccinia virus, validating imaging data as a marker of viral infection. Replication of vaccinia was significantly greater in mice lacking receptors for type I interferons (IFN I R-/-) compared with wild-type mice, although both genotypes of mice developed focal infections in lungs and brain after intranasal inoculation. IFN I R-/- mice had greater dissemination of virus to liver and spleen than wild-type animals even when mortality occurred at the same time point after infection. Protective effects of type I interferons were mediated primarily through parenchymal cells rather than hematopoietic cells as analyzed by bone marrow transplant experiments. Collectively, our data define a new function for type I interferon signaling in systemic dissemination of vaccinia and validate these reporter viruses for studies of pathogenesis.
整体动物成像可在完整动物体内监测病毒复制和定位，这为确定决定发病机制的病毒和宿主因素提供了显著优势。为了研究干扰素对牛痘感染时空进程的影响，我们构建了表达萤火虫荧光素酶或单体橙色荧光蛋白的重组病毒。这些病毒分别允许通过生物发光或荧光成像监测牛痘感染。相对于对照WR病毒，重组病毒在体外和体内均未减毒。在细胞培养中，感染后4小时即可轻松检测到报告基因，表明这些病毒可作为感染的早期标志物。体外和体内通过生物发光成像测量的萤火虫荧光素酶活性强度与牛痘病毒滴度的增加直接相关，证实成像数据可作为病毒感染的标志物。与野生型小鼠相比，缺乏I型干扰素受体（IFN I R-/-）的小鼠中牛痘的复制明显更多，尽管两种基因型的小鼠在鼻内接种后肺部和脑部均出现局灶性感染。即使在感染后相同时间点出现死亡，IFN I R-/-小鼠的病毒向肝脏和脾脏的扩散也比野生型动物更大。通过骨髓移植实验分析，I型干扰素的保护作用主要通过实质细胞而非造血细胞介导。总体而言，我们的数据确定了I型干扰素信号在牛痘全身扩散中的新功能，并验证了这些报告病毒在发病机制研究中的作用。
