Department of Molecular Genetics and Cell Biology, The University of Chicago, Chicago, IL, USA.
Ludwig Center for Metastasis Research, The University of Chicago, Chicago, IL, USA.
Sci Rep. 2017 Dec 5;7(1):17031. doi: 10.1038/s41598-017-16987-x.
Recent developments in optical tissue clearing and microscopic imaging have advanced three-dimensional (3D) visualization of intact tissues and organs at high resolution. However, to expand applications to oncology, critical limitations of current methods must be addressed. Here we describe transparent tissue tomography (T3) as a tool for rapid, three-dimensional, multiplexed immunofluorescent tumor imaging. Cutting tumors into sub-millimeter macrosections enables simple and rapid immunofluorescence staining, optical clearing, and confocal microscope imaging. Registering and fusing macrosection images yields high resolution 3D maps of multiple tumor microenvironment components and biomarkers throughout a tumor. The 3D maps can be quantitatively evaluated by automated image analysis. As an application of T3, 3D mapping and analysis revealed a heterogeneous distribution of programmed death-ligand 1 (PD-L1) in Her2 transgenic mouse mammary tumors, with high expression limited to tumor cells at the periphery and to CD31 vascular endothelium in the core. Also, strong spatial correlation between CD45 immune cell distribution and PD-L1 expression was revealed by T3 analysis of the whole tumors. Our results demonstrate that a tomographic approach offers simple and rapid access to high-resolution three-dimensional maps of the tumor immune microenvironment, offering a new tool to examine tumor heterogeneity.
光学组织透明化和显微镜成像的最新进展提高了完整组织和器官的高分辨率三维（3D）可视化。然而，为了将应用扩展到肿瘤学，必须解决当前方法的关键限制。在这里，我们将透明组织断层扫描（T3）描述为一种用于快速、三维、多重免疫荧光肿瘤成像的工具。将肿瘤切成亚毫米级的大块切片可实现简单快速的免疫荧光染色、光学透明化和共聚焦显微镜成像。注册和融合大切片图像可生成整个肿瘤中多个肿瘤微环境成分和生物标志物的高分辨率 3D 图谱。通过自动图像分析可以对 3D 图谱进行定量评估。作为 T3 的应用，3D 映射和分析显示了 Her2 转基因小鼠乳腺肿瘤中程序性死亡配体 1（PD-L1）的异质性分布，高表达仅限于肿瘤细胞的外周和核心的 CD31 血管内皮细胞。此外，通过对整个肿瘤进行 T3 分析，还揭示了 CD45 免疫细胞分布与 PD-L1 表达之间的强烈空间相关性。我们的结果表明，断层扫描方法提供了一种简单而快速的方法来获取肿瘤免疫微环境的高分辨率三维图谱，为研究肿瘤异质性提供了一种新工具。
