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. 2025 Jun 2;27(1):119.
doi: 10.1186/s13075-025-03570-3.

Serum IL-40 is elevated in systemic sclerosis and is linked to disease activity, gastrointestinal involvement, immune regulation and fibrotic processes

Adéla Navrátilová  1   2 , Sabína Oreská  1   2 , Hana Wünsch  1 , Klára Mocová  1 , Ondřej Kodet  3   4 , Marian Rybar  5 , Glenda Alquicer  1 , Aneta Prokopcová  1   2 , Viktor Bečvář  1 , Radim Bečvář  1   2 , Aneta Jiránková  1 , Anna Bobrová  1 , Kateřina Bobrová  1 , Karel Pavelka  1   2 , Jiří Vencovský  1   2 , Ladislav Šenolt  1   2 , Lucie Andrés Cerezo  6   7 , Michal Tomčík  8   9
Affiliations

Serum IL-40 is elevated in systemic sclerosis and is linked to disease activity, gastrointestinal involvement, immune regulation and fibrotic processes

Adéla Navrátilová et al. Arthritis Res Ther. .

Abstract

Background: Interleukin 40 (IL-40) is a cytokine implicated in malignancies and rheumatic disorders. Its association with fibrotic mediators has been previously described. Since inflammation and fibrosis are hallmarks of systemic sclerosis (SSc), we aimed to analyze the role of IL-40 in SSc.

Methods: IL-40 levels were analyzed in the serum of 90 SSc patients and 75 healthy controls (HCs). IL-40 expression in dermal biopsies from 5 SSc patients and 5 HCs was assessed via immunohistochemistry. IL-40 was analyzed in 39 SSc patients with interstitial lung disease treated with cyclophosphamide (CPA) and in 24 SSc patients with active progressive disease treated with rituximab (RTX). SSc activity was assessed by the European Scleroderma Study Group (ESSG) index. The effect of recombinant IL-40 on peripheral blood mononuclear cells (PBMCs) from 10 SSc patients was determined in vitro. IL-40 was analyzed in 24 individuals at risk of developing SSc (VEDOSS), who were categorized as progressors (n = 11) and nonprogressors (n = 13).

Results: IL-40 expression was elevated in the skin of SSc patients compared to HCs, particularly in fibroblasts and immune infiltrates. Serum IL-40 was increased in SSc compared to HCs (p < 0.0001) and was associated with ESSG (r = 0.372, p = 0.0005) and gastrointestinal involvement (p < 0.05). IL-40 correlated with serum IL-8 (r = 0.270, p = 0.019) and TGF-β1 (r = 0.301, p = 0.024) levels. In the CPA and RTX cohort, no significant changes in the serum IL-40 were observed upon treatment. Baseline and changes in IL-40 levels were associated with changes in several clinical parameters. IL-40 was elevated in patients at risk of SSc compared to HCs (p = 0.0003). No significant changes were observed in progressors vs. nonprogressors; however, IL-40 was associated with capillaroscopy findings (p < 0.05). IL-40 induced the upregulation of IL-6 (p = 0.002), MCP-1 (p = 0.002) and IL-10 (p = 0.002) in PBMCs from SSc patients in vitro.

Conclusions: IL-40 was upregulated in the skin and serum of SSc patients and was associated with disease activity, gastrointestinal involvement and fibrotic mediators. Our in vitro findings indicate that IL-40 might be involved in the immune response and fibrotic processes in SSc.

Keywords: Biomarkers; Interleukin 40; Systemic sclerosis.

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Conflict of interest statement

Declarations. Ethics approval and consent to participate: The study was approved by the local Ethics Committee of the Institute of Rheumatology in Prague, Czech Republic. The reference number for ethics approval is 6485/2020. Written informed consent was obtained from all participants. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
IL-40 in the cross-sectional analysis of SSc Serum IL-40 levels were increased in SSc patients (n = 90) compared to HCs (n = 75) (A). There was no difference between dcSSc (red) and lcSSc (orange). Furthermore, IL-40 levels were associated with disease activity (ESSG) (B) and gastrointestinal involvement (C), particularly constipation (D) and dysphagia (E). Serum IL-40 was associated with circulating levels of TGF-β1 (n = 56) (F) and IL-8 (n = 76) (G). The data are presented as the median IQR. The association of IL-40 with ESSG or cytokines was analyzed using Pearson correlation. Mann-Whitney was used to compare IL-40 levels. dc, diffuse cutaneous SSc; GI, gastrointestinal; HC, healthy control; lcSSc, limited cutaneous SSc; SSc, systemic sclerosis; TGF-β1, transforming growth factor-beta
Fig. 2
Fig. 2
IL-40 was expressed in the skin of SSc patients. IL-40 was expressed in the involved skin of SSc patients (n = 5) (A) and HCs (n = 5) (B). High intensity IL-40 was observed in the perivascular infiltrates of immune cells in SSc (black arrow). SSc fibroblasts showed moderate IL-40 expression (blue arrow) (A). In HCs, a subset of dermal fibroblasts (blue arrow) and immune cells (black arrow) displayed IL-40 positivity (B). Representative images of immunohistochemistry staining are shown at × 100, the detail at × 200 magnification
Fig. 3
Fig. 3
Serum IL-40 levels in VEDOSS individuals. IL-40 levels were elevated in VEDOSS (n = 24) compared to HCs (n = 75). No significant change in serum IL-40 was detected in VEDOSS progressors (red, n = 11) or nonprogressors (blue, n = 13) at baseline vs. follow-up. The data are presented as the median IQR. The horizontal bar represents the median. Mann-Whitney test was used to compare IL-40 levels in VEDOSS vs. HC, and baseline IL-40 levels in progressors vs. nonprogressors. Wilcoxon test was used to compare IL-40 levels at baseline vs. follow-up. HC, healthy control; ns, not significant; VEDOSS, very early diagnosis of systemic sclerosis
Fig. 4
Fig. 4
IL-40 is involved in the immune regulation of SSc PBMCs in vitro. Recombinant IL-40 increased the secretion of IL-6, MCP-1, and IL-10 in PBMCs from SSc patients after 24 h (n = 10) (A-C). IL-40 had no effect on the release of IL-13 and TGF-β (D, E). PBMCs from SSc patients released IL-40 upon treatment with IL-4 and TGF-β (F). Data are presented as the median IQR. The Wilcoxon test was used to compare the analyzed levels. Ctrl, unstimulated cells; LPS, lipopolysaccharide; MCP-1, Monocyte Chemoattractant Protein-1; N.D., not detected; ns, not significant; PBMCs, mononuclear cells; SSc, systemic sclerosis; TGF-β1, transforming growth factor beta 1
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