Dermal fibroblasts respond to IL-4 and IL-13 and promote T cell recruitment in atopic dermatitis
Tomofumi Numata, Michael Shia, Yoshiyuki Nakamura, Fengwu Li, Hung Chan, Teruaki Nakatsuji, Kellen J. Cavagnero, Jared Simmons, Henry Li, Aaroh Anand Joshi, Marta Palomo-Irigoyen, Richard L. Gallo
Tomofumi Numata, Michael Shia, Yoshiyuki Nakamura, Fengwu Li, Hung Chan, Teruaki Nakatsuji, Kellen J. Cavagnero, Jared Simmons, Henry Li, Aaroh Anand Joshi, Marta Palomo-Irigoyen, Richard L. Gallo
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Research Article Dermatology Immunology

Dermal fibroblasts respond to IL-4 and IL-13 and promote T cell recruitment in atopic dermatitis

Abstract

Atopic dermatitis (AD) is a chronic inflammatory skin condition characterized by a type 2 immune response that is not fully understood. Single-cell RNA-seq of human AD skin and murine models of type 2 inflammation identified transcriptionally distinct fibroblast clusters, revealing IL-4Rα–dependent populations of immune-acting fibroblasts (IAFs). These unbiased findings prompted further investigation into the role of dermal fibroblasts during allergic inflammation. These studies demonstrated that, in an inflammatory environment including TNF-α, IL-1β, and IL-17A, the cytokines IL-4 and IL-13 stimulated both mouse and human fibroblasts to produce multiple chemokines, including CCL8, which activated CCR3 to attract T cells. In the skin, fibroblasts were the primary source of many of these chemokines, and targeted deletion of IL-4Rα in mouse fibroblasts reduced T cell infiltration in a mouse model of AD. Additionally, pharmacologic inhibition of CCR3, the receptor shared by many chemokines produced by fibroblasts, decreased T cell infiltration and skin inflammation in mouse models of AD. These findings demonstrate that dermal fibroblasts are more than passive structural cells; they actively participate in the type 2 immune response and contribute to AD by producing chemokines that increase inflammation. Targeting the functions of IAFs could offer an alternative therapeutic approach for AD.

Authors

Tomofumi Numata, Michael Shia, Yoshiyuki Nakamura, Fengwu Li, Hung Chan, Teruaki Nakatsuji, Kellen J. Cavagnero, Jared Simmons, Henry Li, Aaroh Anand Joshi, Marta Palomo-Irigoyen, Richard L. Gallo

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Figure 3

Fibroblasts express multiple chemokines in response to IL-4/13.

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Fibroblasts express multiple chemokines in response to IL-4/13. (A) Sche...
(A) Schematic of the assessment of the response of 3T3-L1 fibroblasts to inflammatory cytokines. (B) PCA of the normalized bulk RNA-seq data from fibroblasts 24 hours after culturing under control conditions, with addition of IL-4 (50 ng/mL) and IL-13 (50 ng/mL) (IL-4/13), with the addition of a mixture of inflammatory cytokines TNF-α (20 ng/mL), IL-17A (50 ng/mL), and IL-1β (0.5 ng/mL) (Inf), or with IL-4/13 + Inf. (C) Heatmap illustrating changes in selected gene expression, scaled by row. (D) Gene enrichment analysis for 3T3-L1 fibroblasts treated with IL-4/13 + Inf and Inf alone. (E) Venn diagram showing the overlap of genes induced in cultured fibroblasts and mouse dermal fibroblasts. Shown is the expression of genes induced in cultured fibroblasts following the addition of Inf or IL-4/13 + Inf compared with control culture conditions, and fibroblast gene expression in mice determined by scRNA-seq of mouse skin exposed to SA alone or MC + SA compared with control mouse skin. Numbers indicate the numbers and percentages of upregulated genes with significant differences. The red dashed area highlights the 28 genes commonly upregulated across all 4 conditions. (F) List of 28 genes upregulated in fibroblasts in culture or in mice following exposure to type 2 cytokines. (G) qRT-PCR analysis of Ccl8, Ccl11, and Ccl17 mRNA expression in 3T3-L1 fibroblasts treated with Inf and IL-4/13 as in B (n = 3). (H) Measurement of CCL8 protein concentration in 3T3-L1 fibroblast supernatants by ELISA 72 hours after treatment with Inf or IL-4/13 as in B (n = 3). (I) Heatmap of protein expression measured by LEGENDPLEX in culture supernatants of the human fibroblast cell line HPAd. Statistical significance was determined by 1-way ANOVA followed by Tukey’s multiple-comparison test (G and H) (n = 3). Experiments in G–I are representative of 2–3 independent experiments. Data are presented as the mean ± SD.