Monitoring Macrophage Polarization with Gene Expression Reporters and Bioluminescence Phasor Analysis

Giulia Tedeschi, Mariana X. Navarro, Lorenzo Scipioni, Tanvi K. Sondhi, Jennifer A. Prescher* and Michelle A. Digman*, 
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Abstract

Macrophages exhibit a spectrum of behaviors upon activation and are generally classified as one of two types: inflammatory (M1) or anti-inflammatory (M2). Tracking these phenotypes in living cells can provide insight into immune function but remains a challenging pursuit. Existing methods are mostly limited to static readouts or are difficult to employ for multiplexed imaging in complex 3D environments while maintaining cellular resolution. We aimed to fill this void using bioluminescent technologies. Here we report genetically engineered luciferase reporters for the long-term monitoring of macrophage polarization via spectral phasor analysis. M1- and M2-specific promoters were used to drive the expression of bioluminescent enzymes in macrophage cell lines. The readouts were multiplexed and discernible in both 2D and 3D formats with single-cell resolution in living samples. Collectively, this work expands the toolbox of methods for monitoring macrophage polarization and provides a blueprint for monitoring other multifaceted networks in heterogeneous environments.

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利用基因表达报告和生物发光相位分析监测巨噬细胞极化
巨噬细胞在活化后会表现出一系列行为,一般分为两种类型:炎症(M1)或抗炎(M2)。在活细胞中跟踪这些表型可深入了解免疫功能,但这仍是一项具有挑战性的工作。现有的方法大多局限于静态读数,或者难以在复杂的三维环境中进行多重成像,同时保持细胞分辨率。我们的目标是利用生物发光技术填补这一空白。在此,我们报告了基因工程荧光素酶报告器,用于通过光谱相位分析长期监测巨噬细胞极化。我们使用 M1 和 M2 特异性启动子来驱动巨噬细胞系中生物发光酶的表达。在活体样本中,读出的结果是多路复用的,并可在二维和三维格式中以单细胞分辨率辨别。总之,这项工作扩展了监测巨噬细胞极化的方法工具箱,并为监测异质环境中的其他多层面网络提供了蓝图。
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来源期刊
Chemical & Biomedical Imaging
Chemical & Biomedical Imaging 化学与生物成像-
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期刊介绍: Chemical & Biomedical Imaging is a peer-reviewed open access journal devoted to the publication of cutting-edge research papers on all aspects of chemical and biomedical imaging. This interdisciplinary field sits at the intersection of chemistry physics biology materials engineering and medicine. The journal aims to bring together researchers from across these disciplines to address cutting-edge challenges of fundamental research and applications.Topics of particular interest include but are not limited to:Imaging of processes and reactionsImaging of nanoscale microscale and mesoscale materialsImaging of biological interactions and interfacesSingle-molecule and cellular imagingWhole-organ and whole-body imagingMolecular imaging probes and contrast agentsBioluminescence chemiluminescence and electrochemiluminescence imagingNanophotonics and imagingChemical tools for new imaging modalitiesChemical and imaging techniques in diagnosis and therapyImaging-guided drug deliveryAI and machine learning assisted imaging
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