Multiresolution 3D Optical Mapping of Immune Cell Infiltrates in Mouse Asthmatic Lung.
Multiresolution 3D Optical Mapping of Immune Cell Infiltrates in Mouse Asthmatic Lung.
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DOI：
10.1165/rcmb.2022-0353ma
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发表时间：
2023-07
影响因子：
6.4
通讯作者：
中科院分区：
文献类型：
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作者：
哮喘是一种慢性炎症性呼吸道疾病，由多种免疫细胞类型渗入肺部引起。光学显微镜已经被用来研究哮喘肺中的免疫渗透。激光共聚焦扫描显微镜(CLSM)通过使用高倍物镜和多重免疫荧光染色来识别肺组织切片中单个免疫细胞的表型和位置。相比之下，光片荧光显微镜(LSFM)采用光学组织清晰法，可以在三维(3D)上显示整体肺组织的宏观和中观结构。尽管每种显微镜方法都能从组织样本中产生具有独特分辨率的图像数据，但由于组织准备程序的不同，CLSM和LSFM并未一起应用。在这里，我们介绍了一种将LSFM和CLSM结合到序列成像管道中的新方法。我们建立了一种新的光学组织清除流程，在该流程中，浸泡清洗剂可以从有机溶剂切换到糖水溶液，用于小鼠肺的顺序3D LSFM和CLSM。这种顺序组合显微镜在器官、组织和细胞水平上提供了免疫浸润物在同一小鼠哮喘肺组织中分布的定量3D空间分析。这些结果表明，我们的方法促进了多分辨率3D荧光显微镜作为一种新的成像方法，为更好地了解炎症性肺部疾病提供了全面的空间信息。
Asthma is a chronic inflammatory airway disease driven by various infiltrating immune cell types into the lung. Optical microscopy has been used to study immune infiltrates in asthmatic lungs. Confocal laser scanning microscopy (CLSM) identifies the phenotypes and locations of individual immune cells in lung tissue sections by employing high-magnification objectives and multiplex immunofluorescence staining. In contrast, light-sheet fluorescence microscopy (LSFM) can visualize the macroscopic and mesoscopic architecture of whole-mount lung tissues in three dimensions (3D) by adopting an optical tissue-clearing method. Despite each microscopy method producing image data with unique resolution from a tissue sample, CLSM and LSFM have not been applied together because of different tissue-preparation procedures. Here, we introduce a new approach combining LSFM and CLSM into a sequential imaging pipeline. We built a new optical tissue clearing workflow in which the immersion clearing agent can be switched from an organic solvent to an aqueous sugar solution for sequential 3D LSFM and CLSM of mouse lungs. This sequential combination microscopy offered quantitative 3D spatial analyses of the distribution of immune infiltrates in the same mouse asthmatic lung tissue at the organ, tissue, and cell levels. These results show that our method facilitates multiresolution 3D fluorescence microscopy as a new imaging approach providing comprehensive spatial information for a better understanding of inflammatory lung diseases.
DOI：
10.4049/jimmunol.2000885
发表时间：
2021-01-15
期刊：
Journal of immunology (Baltimore, Md. : 1950)
影响因子：
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作者：
Royer DJ;Cook DN
通讯作者：
Cook DN