Abstract
Real-time in vivo imaging has become an integral tool for the investigation and understanding of cellular processes in health and disease at single-cell resolution. This includes the dynamic and complex cellular interactions that occur during cancer progression and the subsequent metastatic dissemination of tumor cells to sites distant from the primary tumor. Herein we outline the methodology for the establishment and intravital imaging of the pulmonary metastatic niche, a preferred site of metastasis for many cancers, and describe the implementation of a lung window to visualize and dissect the intricate behaviour of multiple cell types within this environment. We also address the advantages and limitations of this high-resolution technology.
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Video 1:
Intravital imaging of B16F10 lung metastasis environment. Time series intravital imaging of the tumor environment in the lung 17Â days after mouse melanoma B16F10-mCherry cells (cyan) were intravenously injected. Neutrophils are shown in magenta (anti-Ly6G, BV421), and lung endothelial (yellow, Alexa fluor 647) cells were labeled with anti-CD31 (WMV 6705 kb)
Video 2:
Intravital imaging of E0771 lung metastasis environment. Time lapse intravital imaging of the tumor environment in the lung 14Â days after the injection of mammary E0771-iRFP720 cells (cyan). Neutrophils are shown in magenta (anti-Ly6G, BV421), and lung endothelial (yellow, Alex fluor 594) cells were labeled with anti-CD31 (WMV 9689 kb)
Video 3:
Intravital imaging of E0771 lung metastasis in 4D. Four-dimensional time lapse imaging of lung mammary E0771-iRFP tumor micro-environment. Images over time were acquired at indicated depth (0 μm–70 μm depth; 1.29-μm-thick sections). E0771-iRFP720 cells are shown in cyan, neutrophils labeled with anti-Ly6G are depicted in magenta (BV421), and lung endothelial cells in yellow (anti-CD31, Alexa fluor 594). Total variation denoising tool in the Leica software was used to process all time lapse images. Scale bar 50 μm (WMV 5236 kb)
Video 4:
Intravital imaging of B16F10 lung metastasis in 4D. Four-dimensional time lapse imaging of melanoma B16F10-mCherry lung metastasis at indicted depth (0 μm–32.76 μm; 1.26-μm-thick sections) over time. B16F10-mCherry cells (cyan), neutrophils labeled with anti-Ly6G (magenta, BV421), and lung endothelial (yellow, Alexa fluor 647) cells labeled with anti-CD31. Total variation denoising tool in the Leica software was used to process all time lapse images. Scale bar 50 μm (WMV 8431 kb)
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Babes, L., Yipp, B.G., Senger, D.L. (2023). Intravital Microscopy of the Metastatic Pulmonary Environment. In: Ursini-Siegel, J. (eds) The Tumor Microenvironment. Methods in Molecular Biology, vol 2614. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2914-7_23
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DOI: https://doi.org/10.1007/978-1-0716-2914-7_23
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Publisher Name: Humana, New York, NY
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Online ISBN: 978-1-0716-2914-7
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