Understanding immunity in a tissue‐centric context: Combining novel imaging methods and mathematics to extract new insights into function and dysfunction*

Abstract

A central question in immunology is what features allow the immune system to respond in a timely manner to a variety of pathogens encountered at unanticipated times and diverse body sites. Two decades of advanced and static dynamic imaging methods have now revealed several major principles facilitating host defense. Suborgan spatial prepositioning of distinct cells promotes time‐efficient interactions upon pathogen sensing. Such pre‐organization also provides an effective barrier to movement of pathogens from parenchymal tissues into the blood circulation. Various molecular mechanisms maintain effective intercellular communication among otherwise rapidly moving cells. These and related discoveries have benefited from recent increases in the number of parameters that can be measured simultaneously in a single tissue section and the extension of such multiplex analyses to 3D tissue volumes. The application of new computational methods to such imaging data has provided a quantitative, in vivo context for cell trafficking and signaling pathways traditionally explored in vitro or with dissociated cell preparations. Here, we summarize our efforts to devise and employ diverse imaging tools to probe immune system organization and function, concluding with a commentary on future developments, which we believe will reveal even more about how the immune system operates in health and disease.

Multiplex imaging in immuno-oncology

“… 62 However, the actual nuclear shape as well as position within the cell, the distance from the bounding membrane, and the nucleus-to-cytoplasm ratio all vary among cells, making it technically challenging to determine an accurate way to perform the nuclear expansion for all cells. Recent advances in computer vision now approach segmentation by combining nuclear and membrane staining, then using deep learning methods such as Cellpose, 63 Mesmer, 64 and RAPID 60 to more accurately segment the diverse cell types within a tissue cell. However, pretrained models are limited by their training datasets and their performance can be further enhanced after adaptation to the target dataset.…”

“…Recently, a variety of computational methods have been introduced to address this issue, but to date, these are only approximate solutions to the problem. 60 66 67 This means that careful curation of the segmentation data is needed at an expert level to filter out cell phenotypes that are artifacts of such pixel overlap.…”

“…The detailed statistical consideration can be found in prior reviews. 60 73 In addition, multiple open-source packages and methods are or are becoming available to facilitate this process, such as SPACE, 60 Giotto, 76 ImaCytE, 77 Squidpy, 78 SpatialDE, 79 histoCAT, 80 and SPARK. 81 …”

Locally sourced: site-specific immune barriers to metastasis

“…The past two decades have seen a revolution in the tools available for studying the diversity of immune cells in tissues. Lineage tracing and highly multiplexed immunofluorescence imaging have enabled semiquantitative analyses of the composition and spatial organization of the immune system within tissues, whereas two-photon intravital microscopy has provided insights into immune cell population dynamics, cellcell interactions and functional responses in situ 73 . These advanced imaging modalities, combined with conventional flow cytometry approaches and emerging systems-level immunomics 74 , continue to uncover new phenotype diversity in immune cells across multiple mouse [75][76][77][78][79][80][81][82][83] and human 82,[84][85][86][87][88][89][90][91][92][93] tissues.…”

Spatial Patterning Analysis of Cellular Ensembles (SPACE) discovers complex spatial organization at the cell and tissue levels

“…Whether these interactions occur via direct contact or paracrine signaling, they are determined by the spatial arrangement of the involved entities. Thus discovering new spatial patterns can reveal new mechanisms of tissue (dys)function [Baccin et al 2020, Saviano et al 2020, Lewis et al 2021, Rendeiro et al 2021, Germain et al 2022].…”