Light-Sheet Scattering Microscopy to Visualize Long-Term Interactions Between Cells and Extracellular Matrix.

Xiangda Zhou,Bin Qu,Archana K Yanamandra,Markus Hoth,Renping Zhao

doi:10.3389/fimmu.2022.828634

Xiangda Zhou, Bin Qu + Show 3 more

Open Access

https://doi.org/10.3389/fimmu.2022.828634

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Abstract

Visualizing interactions between cells and the extracellular matrix (ECM) mesh is important to understand cell behavior and regulatory mechanisms by the extracellular environment. However, long term visualization of three-dimensional (3D) matrix structures remains challenging mainly due to photobleaching or blind spots perpendicular to the imaging plane. Here, we combine label-free light-sheet scattering microcopy (LSSM) and fluorescence microscopy to solve these problems. We verified that LSSM can reliably visualize structures of collagen matrices from different origin including bovine, human and rat tail. The quality and intensity of collagen structure images acquired by LSSM did not decline with time. LSSM offers abundant wavelength choice to visualize matrix structures, maximizing combination possibilities with fluorescently-labelled cells, allowing visualizing of long-term ECM-cell interactions in 3D. Interestingly, we observed ultrathin thread-like structures between cells and matrix using LSSM, which were not observed by normal fluorescence microscopy. Transient local alignment of matrix by cell-applied forces can be observed. In summary, LSSM provides a powerful and robust approach to investigate the complex interplay between cells and ECM.

Highlights

Under physiological conditions, immune cells and tumor cells encounter complex and dynamic three-dimensional (3D) environments
In order to visualize long-term cell-extracellular matrix (ECM) interactions in larger 3D structures, we looked for a label-free method based on lightsheet microscopy
The results show that the fibrous structures and reconstructed 3D structures visualized by light-sheet scattering microscopy (LSSM) and fluorescence microscopy are very similar (Figure 1A)

Summary

Introduction

Immune cells and tumor cells encounter complex and dynamic three-dimensional (3D) environments. To maintain 3D environments, extracellular matrix (ECM) composed of fibrous mesh networks serves as one main structural component. It is reported that in vivo macrophages structurally form vascular mimicry channels, which are ultrastructurally different from an endothelial vasculature [2]. In vitro evidence shows that fast migrating primary human CD8+ T cells preferably migrate into and expand the existing channels in 3D collagen matrices, facilitating migration of subsequent immune cells [3]. These channels could presumably contribute to metastasis of tumor cells [4].

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