Go to The Journal of Clinical Investigation
  • About
  • Editors
  • Consulting Editors
  • For authors
  • Publication ethics
  • Transfers
  • Advertising
  • Job board
  • Contact
  • Current issue
  • Past issues
  • By specialty
    • COVID-19
    • Cardiology
    • Immunology
    • Metabolism
    • Nephrology
    • Oncology
    • Pulmonology
    • All ...
  • Videos
  • Collections
    • Resource and Technical Advances
    • Clinical Medicine
    • Reviews
    • Editorials
    • Perspectives
    • Top read articles
  • JCI This Month
    • Current issue
    • Past issues

  • Current issue
  • Past issues
  • Specialties
  • In-Press Preview
  • Editorials
  • Viewpoint
  • Top read articles
  • About
  • Editors
  • Consulting Editors
  • For authors
  • Publication ethics
  • Transfers
  • Advertising
  • Job board
  • Contact
Imaging protective mast cells in living mice during severe contact hypersensitivity
Laurent L. Reber, … , Nicolas Gaudenzio, Stephen J. Galli
Laurent L. Reber, … , Nicolas Gaudenzio, Stephen J. Galli
Published May 4, 2017
Citation Information: JCI Insight. 2017;2(9):e92900. https://doi.org/10.1172/jci.insight.92900.
View: Text | PDF
Research Article Immunology Inflammation

Imaging protective mast cells in living mice during severe contact hypersensitivity

  • Text
  • PDF
Abstract

Contact hypersensitivity (CHS) is a common skin disease induced by epicutaneous sensitization to haptens. Conflicting results have been obtained regarding pathogenic versus protective roles of mast cells (MCs) in CHS, and this has been attributed in part to the limitations of certain models for studying MC functions in vivo. Here we describe a fluorescent imaging approach that enables in vivo selective labeling and tracking of MC secretory granules by real-time intravital 2-photon microscopy in living mice, and permits the identification of such MCs as a potential source of cytokines in different disease models. We show using this method that dermal MCs release their granules progressively into the surrounding microenvironment, but also represent an initial source of the antiinflammatory cytokine IL-10, during the early phase of severe CHS reactions. Finally, using 3 different types of MC-deficient mice, as well as mice in which IL-10 is ablated specifically in MCs, we show that IL-10 production by MCs can significantly limit the inflammation and tissue pathology observed in severe CHS reactions.

Authors

Laurent L. Reber, Riccardo Sibilano, Philipp Starkl, Axel Roers, Michele A. Grimbaldeston, Mindy Tsai, Nicolas Gaudenzio, Stephen J. Galli

×

Figure 1

A single injection of fluorochrome-labeled avidin enables specific labeling of dermal mast cell (MC) intracellular secretory granules.

Options: View larger image (or click on image) Download as PowerPoint
A single injection of fluorochrome-labeled avidin enables specific label...
(A) Protocol: 5 μg of sulforhodamine 101–coupled avidin (Av.SRho) was injected intradermally (i.d.) into the ear pinna of a mouse. One week later the mouse was anesthetized and placed under a 2-photon microscope; 3D high-resolution images were taken and Av.SRho fluorescence signal was assessed. (B) Kit+/+ MC-sufficient (upper panel) or KitW-sh/W-sh MC-deficient (lower panel) mice were treated as described in A except that 250 μg of 70-kDa dextran-FITC was injected retro-orbitally 30 minutes before imaging to label blood vessels. Representative 3D photographs of the ear pinna showing merged fluorescence of Av.SRho (red), dextran-FITC (green), and collagen structures (blue). White lines identify the magnified areas. Far right panel: Av.SRho (red) and dextran-FITC (green) fluorescence was modeled and the image was rotated 90° to better depict the modeled fluorescence signals. (C) Number of Av.SRho+ cells in ear pinnae per field of view (FOV) in Kit+/+ or KitW-sh/W-sh mice. (D) Av.SRho mean fluorescence intensity (MFI) per FOV in Kit+/+ or KitW-sh/W-sh mice. (E) Breeding strategy to obtain Mcpt5-Cre+; R26Y+ (Mcpt5-EYFP) mice. (F) Mcpt5-EYFP mice were treated as described in A. Representative 3D photographs of the ear pinna showing merged fluorescence of Av.SRho (red) and EYFP (green). White lines identify the magnified area, dashed white circles identify hair follicles. (G) Left panel: 3D photographs of a single Av.SRho+ EYFP+ cell. Right panel: spatial localization of the Av.SRho (red) and EYFP (green) fluorescence along the white line depicted in the left panel. (H) Percentage of Av.SRho+ EYFP+ (red and green), Av.SRho–EYFP+ (green) and Av.SRho+EYFP– (red) cells in ear pinnae per field of view (FOV) in Mcpt5-EYFP mice. Mean + SEM, 2-tailed, unpaired t test, **P < 0.01; ***P < 0.001; ****P < 0.0001. Data (n = 3 mice per group) are pooled from the 3 experiments performed (each done with 1 mouse per group), each of which gave similar results; circles show values for individual mice. Scale bars: 100 μm.

Copyright © 2023 American Society for Clinical Investigation
ISSN 2379-3708

Sign up for email alerts