Genetic and Imaging Approaches Reveal Pro-Inflammatory and Immunoregulatory Roles of Mast Cells in Contact Hypersensitivity

Abstract

Contact hypersensitivity (CHS) is a common T cell-mediated skin disease induced by epicutaneous sensitization to haptens. Mast cells (MCs) are widely deployed in the skin and can be activated during CHS responses to secrete diverse products, including some with pro-inflammatory and anti-inflammatory functions. Conflicting results have been obtained regarding pathogenic versus protective roles of MCs in CHS, and this has been attributed in part to the limitations of certain models for studying MC functions in vivo. This review discusses recent advances in the development and analysis of mouse models to investigate the roles of MCs and MC-associated products in vivo. Notably, fluorescent avidin-based two-photon imaging approaches enable in vivo selective labeling and simultaneous tracking of MC secretory granules (e.g., during MC degranulation) and MC gene activation by real-time longitudinal intravital microscopy in living mice. The combination of such genetic and imaging tools has shed new light on the controversial role played by MCs in mouse models of CHS. On the one hand, they can amplify CHS responses of mild severity while, on the other hand, can limit the inflammation and tissue injury associated with more severe or chronic models, in part by representing an initial source of the anti-inflammatory cytokine IL-10.

Keywords: IgE; avidin; contact hypersensitivity; interleukin-10; mast cells; mouse models; tumor necrosis factor-alpha; two photon microscopy.

Figure 1

Fluorochrome-labeled avidin can be used to probe either exteriorized or intracellular mast cell (MC) granule structures in living mice. (A) 5 µg of sulforhodamine 101-coupled avidin (Av.SRho) is injected intradermally (i.d.) into the ear pinna of a MC-reporter mouse (e.g., Mcpt5-EYFP). 15 min later, the mouse is anesthetized and placed under a two-photon microscope; MC degranulation is induced by i.d. injection of stimulus; 3D high-resolution images of single cells are taken and Av.SRho fluorescence signal is assessed. The photograph shows a single dermal EYFP⁺ MC activated upon i.d. injection of substance P exhibiting Av.SRho⁺ exteriorized small granule structures. Image extracted from Gaudenzio et al. (38). (B) 5 µg of Av.SRho is injected i.d. into the ear pinna of a mouse. 7 days later, the mouse is anesthetized and placed under a two-photon microscope; 3D high-resolution single-cell images are taken and Av.SRho fluorescence signal is assessed. The photograph shows a single EYFP⁺ MC exhibiting Av.SRho⁺ intracellular granule structures. Image extracted from Reber et al. (39).

Figure 2

Roles of mast cells (MCs) in the sensitization and effector phases of contact hypersensitivity (CHS). (A) During the sensitization phase, MCs are activated directly or indirectly by haptens to release of a diverse spectrum of mediators, including histamine and TNF, which induce vasodilatation and recruitment of leukocytes, mainly neutrophils. Multiple lines of evidence suggest that IgE can amplify MC activation in the sensitization phase though antigen-independent cytokinergic effects. Direct cell-to-cell contacts between MCs and dendritic cells (DCs), as well as MC-derived TNF, can amplify DC migration to the draining lymph nodes (DLNs), where these cells prime naïve T cells to become effector cells (Teff cells) via antigen presentation. (B) During “moderate” CHS responses, MCs and MC-derived TNF amplify ear swelling, leukocyte recruitment (mainly neutrophils and CD8⁺ T cells), and epidermal hyperplasia. Evidence suggests that IgE and FcεRI can amplify the pro-inflammatory functions of MCs in moderate CHS (C). During more severe (and chronic) CHS responses, MCs represent an early source of IL-10 in the skin, which amplifies recruitment of regulatory T cells (Treg) and limits ear swelling and epidermal hyperplasia. MC activation in the skin is amplified by the engagement of FcγRIII by hapten-IgG immune complexes. Additionally, MCs can migrate to the DLNs and the spleen in an IgE-dependent manner, where they produce IL-2 which helps in maintaining the Teff:Treg ratio at the site of inflammation, and thereby contributes to limiting the severity of CHS responses.

Figure 3

Skin mast cells (MCs) represent an early source of IL-10 during severe contact hypersensitivity (CHS) responses. We used Av.SRho in IL-10-GFP reporter mice to preform a longitudinal monitoring of both the release of dermal MC granules and activation of Il10 gene transcription (IL-10-GFP, as detected by emission of GFP fluorescent signal) at the site of severe CHS responses using intravital 2-photon microscopy. Images show representative 3D photographs of the ear pinna one at day 1 upon DNFB challenge. Upper panel: merged fluorescence of Av.SRho (red) and IL-10-GFP (green). Middle panel: Av.SRho (red) fluorescence. Lower panel: IL-10-GFP (green) fluorescence. White dashed lines identify the magnified areas and white circles identify hair follicles (Scale bars: 50 µm). Adapted from Reber et al. (39).