The mouse cortical meninges are the site of immune responses to many different pathogens, and are accessible to intravital imaging
The mouse cortical meninges are the site of immune responses to many different pathogens, and are accessible to intravital imaging
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DOI：
10.1016/j.ymeth.2017.03.020
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发表时间：
2017-08-15
期刊：
影响因子：
4.8
通讯作者：
Brewer, James M.
中科院分区：
文献类型：
--
作者：
Coles, Jonathan A.;Stewart-Hutchinson, Phillip J.;Brewer, James M.
已知多种病毒和微生物感染可导致脑膜炎，有证据表明，脑膜是病理性入侵脑实质的门户。因此，对这些区域的观察对于了解寄主-病原菌的相互作用具有广泛的应用。病原体和免疫反应细胞之间的相互作用可以通过它们环境的变化来改变，例如血液和淋巴流动的抑制，特别是在脑膜没有支撑的情况下，侵入性解剖可以改变组织结构。出于这些原因，通过未穿孔的头骨进行活体成像是首选的方法。我们给出了一种通过麻醉小鼠变薄的皮质头骨进行双光子显微镜的简单方法的方案，以实现通过脑膜进入浅层脑实质的亚微米分辨率的实时成像。在报告小鼠中，选定的细胞类型表达荧光蛋白，感染荧光病原体(淋巴细胞性脉络膜脑膜炎病毒、布氏锥虫或伯氏疟原虫)后的成像显示，免疫细胞的皮质脑膜强烈募集，包括中性粒细胞、T细胞以及可能的树突状细胞和巨噬细胞。在没有特殊标记的情况下，硬脑膜、软脑膜和实质之间的边界不能在活体双光子显微镜中直接显示出来，但我们所说明的其他标志和特征允许研究人员识别正在成像的隔室。虽然大多数感染性脑膜炎主要局限于硬脑膜，但也有一些涉及免疫细胞向软脑膜募集。(C)2017年作者。由爱思唯尔公司出版。
A wide range of viral and microbial infections are known to cause meningitis, and there is evidence that the meninges are the gateway to pathogenic invasion of the brain parenchyma. Hence observation of these regions has wide application to understanding host-pathogen interactions. Interactions between pathogens and cells of the immune response can be modified by changes in their environment, such as suppression of the flow of blood and lymph, and, particularly in the case of the meninges, with their unsupported membranes, invasive dissection can alter the tissue architecture. For these reasons, intravital imaging through the unperforated skull is the method of choice. We give a protocol for a simple method of two-photon microscopy through the thinned cortical skull of the anesthetized mouse to enable real-time imaging with sub-micron resolution through the meninges and into the superficial brain parenchyma. In reporter mice in which selected cell types express fluorescent proteins, imaging after infection with fluorescent pathogens (lymphocytic choriomeningitis virus, Trypanosome brucei or Plasmodium berghei) has shown strong recruitment to the cortical meninges of immune cells, including neutrophils, T cells, and putative dendritic cells and macrophages. Without special labeling, the boundaries between the dura mater, the leptomeninx, and the parenchyma are not directly visualized in intravital two-photon microscopy, but other landmarks and characteristics, which we illustrate, allow the researcher to identify the compartment being imaged. While most infectious meningitides are localized mainly in the dura mater, others involve recruitment of immune cells to the leptomeninx. (C) 2017 The Authors. Published by Elsevier Inc.