Type-1 cytokines regulate MMP-9 production and E-cadherin disruption to promote melanocyte loss in vitiligo
Nesrine Boukhedouni, … , Julien Seneschal, Katia Boniface
Nesrine Boukhedouni, … , Julien Seneschal, Katia Boniface
Published May 5, 2020
Citation Information: JCI Insight. 2020;5(11):e133772. https://doi.org/10.1172/jci.insight.133772.
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Research Article Dermatology Inflammation

Type-1 cytokines regulate MMP-9 production and E-cadherin disruption to promote melanocyte loss in vitiligo

Abstract

Loss of melanocytes is the pathological hallmark of vitiligo, a chronic inflammatory skin depigmenting disorder induced by exaggerated immune response, including autoreactive CD8 T cells producing high levels of type 1 cytokines. However, the interplay between this inflammatory response and melanocyte disappearance remains to be fully characterized. Here, we demonstrate that vitiligo skin contains a significant proportion of suprabasal melanocytes, associated with disruption of E-cadherin expression, a major protein involved in melanocyte adhesion. This phenomenon is also observed in lesional psoriatic skin. Importantly, apoptotic melanocytes were mainly observed once cells were detached from the basal layer of the epidermis, suggesting that additional mechanism(s) could be involved in melanocyte loss. The type 1 cytokines IFN-γ and TNF-α induce melanocyte detachment through E-cadherin disruption and the release of its soluble form, partly due to MMP-9. The levels of MMP-9 are increased in the skin and sera of patients with vitiligo, and MMP-9 is produced by keratinocytes in response to IFN-γ and TNF-α. Inhibition of MMP-9 or the JAK/STAT signaling pathway prevents melanocyte detachment in vitro and in vivo. Therefore, stabilization of melanocytes in the basal layer of the epidermis by preventing E-cadherin disruption appears promising for the prevention of depigmentation occurring in vitiligo and during chronic skin inflammation.

Authors

Nesrine Boukhedouni, Christina Martins, Anne-Sophie Darrigade, Claire Drullion, Jérôme Rambert, Christine Barrault, Julien Garnier, Clément Jacquemin, Denis Thiolat, Fabienne Lucchese, Franck Morel, Khaled Ezzedine, Alain Taieb, François-Xavier Bernard, Julien Seneschal, Katia Boniface

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Figure 2

Combined activity of IFN-γ and TNF-α reproduces human vitiligo features in a 3D model of reconstructed pigmented epidermis in vitro.

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Combined activity of IFN-γ and TNF-α reproduces human vitiligo features ...
(A–C) Reconstructed human pigmented epidermis (RHPE) containing melanocytes were stimulated for 24 hours in the presence or absence of 10 ng/mL of TNF-α and IFN-γ, alone or in combination. (A) Representative immunofluorescence analysis of Melan-A (red) and E-cadherin (green) expression. Alteration of E-cadherin expression around melanocytes. Dashed lines represent dermoepidermal layer. Scale bars: 20 μm (left), 10 μm (right). (B) Proportion of suprabasal melanocytes in the different conditions (n = 4). (C) Representative analysis of epidermal cell death using a TUNEL assay (green); melanocytes were stained with anti-Melan-A antibody (red). Dashed lines represent the dermoepidermal layer. Scale bar: 20 μm. Staining is representative of 3 independent experiments. (D) Primary cultures of normal human epidermal keratinocytes (NHEK, left, n = 10) or melanocytes (NHEM, right, n = 11) were treated for 24 hours with 20 ng/mL of IFN-γ and/or TNF-α. CDH1 gene expression in epidermal cells was analyzed by real-time PCR. Results are shown as the percentage of change compared with the control culture. (E) Dose-response study of CDH1 gene expression in NHEM after 24 hours stimulation with TNF-α and/or IFN-γ. (F) Kinetic analysis of CDH1 gene expression in NHEM in response to 20 ng/mL of TNF-α and/or IFN-γ. Results from 1 experiment are shown in E and F and are representative of 4 independent experiments with 4 independent donors. GAPDH was used as a housekeeping gene. (G) Confocal microscopy analysis of RHPE treated in the presence or absence of the combination of 10 ng/mL of TNF-α and IFN-γ for 24 hours. Sections were stained for E-cadherin (green) and LAMP-1 (a marker for lysosomes and late endosomes, red). Merge shows the presence of E-cadherin molecule into LAMP-1 vesicle structures. Scale bars: 20 μm (top); 10 μm (bottom). Stainings are representative of 3 independent experiments. (H) Assessment by ELISA of soluble E-cadherin levels in cell-free supernatants of RHPE treated for 24 hours in the presence or absence of 10 ng/mL of TNF-α and IFN-γ. Data in B, D, and H show mean ± SEM. *P < 0.05, **P < 0.01; calculated with 2-tailed Mann-Whitney (B) or Wilcoxon (H) tests.