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Ultraspecific live imaging of the dynamics of zebrafish neutrophil granules by a histopermeable fluorogenic benzochalcone probe

Abstract : Neutrophil granules (NGs) are key components of the innate immune response and mark the development of the hematopoietic system in mammals. However, no specific fluorescent vital stain existed up to now to monitor their dynamics within a whole live organism. We rationally designed a benzochalcone fluorescent probe (HAB) featuring high tissue permeability and optimal photophysics such as elevated quantum yield, pronounced solvatochromism and target-induced fluorogenesis. Phenotypic screening identified HAB as the first cell-and organelle-specific small-molecule tracer of NGs in live zebrafish larvae, with no labeling of any other cell types or organelles. HAB staining was independent of the state of neutrophil activation, labeling NGs of both resting and phagocytically-active cells with equal specificity. By high-resolution live imaging, we documented the dynamics of HAB-stained NGs during phagocytosis. Upon zymosan injection, labeled NGs were rapidly recruited to the forming phagosome in live zebrafish phagocytosing neutrophils. Despite being a reversible ligand, HAB could not be displaced by high concentrations of pharmacologically-relevant competing chalcones, indicating that this specific labeling was the result of HAB precise physicochemical signature rather than a general feature of chalcones. However, one of the competitors was discovered as a promising interstitial fluorescent tracer illuminating zebrafish histology, similarly to BODIPY-ceramide. As a yellow-emitting histopermeable vital stain, HAB functionally and spectrally complements most genetically-incorporated fluorescent tags commonly used in live zebrafish biology, holding promise for the study of neutrophil-dependent responses relevant to human physiopathology such as developmental defects, inflammation and infection. Furthermore, HAB intensely labeled isolated live human neutrophils at the level of granulated subcellular structures consistent with human NGs, suggesting that the labeling of NGs by HAB is not restricted to the zebrafish model but also relevant to mammalian systems.
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https://hal.archives-ouvertes.fr/hal-02359369
Contributor : Philippe Herbomel <>
Submitted on : Tuesday, November 12, 2019 - 1:32:08 PM
Last modification on : Monday, May 18, 2020 - 4:50:34 PM

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Emma Colucci-Guyon, Ariane Batista, Suellen Oliveira, Magali Blaud, Ismael Bellettini, et al.. Ultraspecific live imaging of the dynamics of zebrafish neutrophil granules by a histopermeable fluorogenic benzochalcone probe. Chemical Science , The Royal Society of Chemistry, 2019, 10 (12), pp.3654-3670. ⟨10.1039/c8sc05593a⟩. ⟨hal-02359369⟩

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