Department of Mathematical Sciences, Montana State University, Bozeman, Montana, USA.
Department of Microbiology and Cell Biology, Montana State University, Bozeman, Montana, USA.
Cytometry A. 2022 Dec;101(12):1012-1026. doi: 10.1002/cyto.a.24657. Epub 2022 Jun 6.
Mononuclear phagocytes (MNPs) such as dendritic cells and macrophages perform key sentinel functions in mucosal tissues and are responsible for inducing and maintaining adaptive immune responses to mucosal pathogens. Positioning of MNPs at the epithelial interface facilitates their access to luminally-derived antigens and regulates MNP function through soluble mediators or surface receptor interactions. Therefore, accurately quantifying the distribution of MNPs within mucosal tissues as well as their spatial relationship with other cells is important to infer functional cellular interactions in health and disease. In this study, we developed and validated a MATLAB-based tissue cytometry platform, termed "MNP mapping application" (MNPmApp), that performs high throughput analyses of MNP density and distribution in the gastrointestinal mucosa based on digital multicolor fluorescence microscopy images and that integrates a Monte Carlo modeling feature to assess randomness of MNP distribution. MNPmApp identified MNPs in tissue sections of the human gastric mucosa with 98 ± 2% specificity and 76 ± 15% sensitivity for HLA-DR MNPs and 98 ± 1% specificity and 85 ± 12% sensitivity for CD11c MNPs. Monte Carlo modeling revealed that mean MNP-MNP distances for both HLA-DR and CD11c MNPs were significantly lower than anticipated based on random cell placement, whereas MNP-epithelial distances were similar to randomly placed cells. Surprisingly, H. pylori infection had no significant impact on the number of HLA-DR and CD11c MNPs or their distribution within the gastric lamina propria. However, our study demonstrated that MNPmApp is a reliable and user-friendly tool for unbiased quantitation of MNPs and their distribution at mucosal sites.
单核吞噬细胞（MNP），如树突状细胞和巨噬细胞，在黏膜组织中发挥关键的哨兵功能，负责诱导和维持对黏膜病原体的适应性免疫反应。MNP 位于上皮界面的位置有利于它们获得腔衍生抗原，并通过可溶性介质或表面受体相互作用调节 MNP 功能。因此，准确量化黏膜组织中 MNP 的分布及其与其他细胞的空间关系对于推断健康和疾病中功能性细胞相互作用非常重要。在这项研究中，我们开发并验证了一种基于 MATLAB 的组织细胞计量学平台，称为“MNP 绘图应用程序”（MNPmApp），该平台基于数字多色荧光显微镜图像对胃肠道黏膜中的 MNP 密度和分布进行高通量分析，并集成了蒙特卡罗建模功能来评估 MNP 分布的随机性。MNPmApp 在人胃黏膜组织切片中鉴定 MNPs，对于 HLA-DR MNPs 的特异性为 98±2%，敏感性为 76±15%，对于 CD11c MNPs 的特异性为 98±1%，敏感性为 85±12%。蒙特卡罗建模显示，HLA-DR 和 CD11c MNPs 的平均 MNP-MNP 距离明显低于基于随机细胞放置的预期值，而 MNP-上皮距离与随机放置的细胞相似。令人惊讶的是，H. pylori 感染对胃固有层中 HLA-DR 和 CD11c MNPs 的数量及其分布没有显著影响。然而，我们的研究表明，MNPmApp 是一种可靠且用户友好的工具，可用于在黏膜部位对 MNPs 及其分布进行无偏量化。
