POS0177 POTENTIAL INVOLVEMENT OF IL-40 AND IL-40-PRODUCING CELLS IN PRIMARY SJOGREN’S SYNDROME (pSS) AND pSS-ASSOCIATED LYMPHOMA

Abstract

Background:The pathogenesis of pSS relies on a complex interplay between both innate and adaptive immune responses in which B cells play a pivotal role. Their chronic aberrant hyperactivation may drive clonal escape and consequent lymphomagenesis [1]. In the last few years, B cells have emerged as potential effector cells, able to release a wide range of cytokines that actively contribute to shape the microenvironment they act in. Recently, IL-40, a novel B cell associated cytokine encoded by an uncharacterized gene (C17orf99; chromosome 17 open reading frame 99) was described. Naïve B cells can express IL-40 at both tissue and peripheral level and the stimulation of B cells with IL-4 and TGF-β significantly increases IL-40 release. In addition, human B cells lymphomas are able to constitutively produce IL-40 [2]. Taking into account this emerging evidence and considering the well-known role of IL-4 and TGF-β in pSS pathogenesis, as well as the association with lymphomas, we decided to focus our attention on IL-40 in pSS patients.Objectives:The aim of the present study was to investigate IL-40 expression in the salivary glands of patients affected by pSS and pSS-associated non-Hodgkin’s lymphoma (NHL).Methods:Minor salivary gland biopsies were obtained from 22 patients with pSS and 12 patients with non-specific chronic sialoadenitis (nSCS), included as controls. Paraffine-embedded samples of parotid glands from patients with a previous diagnosis of pSS-associated NHL (n=10) were selected from the biopsy bank of the Pathology Unit of the Ospedale Cervello (Palermo, Italy). Quantitative gene expression analysis by TaqMan real-time PCR and immunohistochemistry (IHC) for IL-40, IL-4, TGF-β1 was performed on salivary glands from patients and controls. The cellular sources of IL-40 among infiltrating inflammatory cells were determined by fluorescence-activated cell sorting (FACS) analysis and immunofluorescence (IF). Serum IL-40 levels were measured by ELISA in both patients (n=10) and controls (n=9).Results:IL-40 was significantly increased at both protein and mRNA level in the inflamed salivary glands of patients with pSS where a positive strong correlation between the IL-40 mRNA levels and the focus score (FS) was evidenced. The expression of IL-40 in parotid glands of pSS-associated NHL was also markedly increased (Figure 1). IL-40 expression correlated with the presence of IL-4 and TGF-β; both cytokines were significantly increased in pSS at mRNA and protein level. Among infiltrating immune cells, CD19+ B cells resulted the major source of IL-40. However, we identified CD4+, CD8+ T cells and CD68+ macrophages as additional producers of IL-40 in both FACS and IF analysis. The ELISA test also showed a significant increase of serum IL-40 concentration in pSS patients (p value = 0.0190), compared to controls.Conclusion:Our preliminary results suggest that IL-40 may play a role in the pathogenesis of pSS and pSS-associated NHL. To the best of our knowledge, this is the first demonstration of the overexpression of this cytokine in salivary gland tissue and sera in pSS. Moreover, we demonstrated that IL-40 is produced by several cellular types, such as T cells and macrophages, and is not exclusively released by B cells. Further studies are necessary to clarify IL-40 pathways and functions in order to unravel IL-40 possible role in pSS development.References:[1]Mariette X, Criswell LA. Primary Sjogren’s Syndrome. N Engl J Med. 2018;379(1):97.[2]Catalan-Dibene J, Vazquez MI, Luu VP, Nuccio SP, Karimzadeh A, Kastenschmidt JM, et al. Identification of IL-40, a Novel B Cell-Associated Cytokine. J Immunol. 2017;199(9):3326-35.Figure 1.IL-40 overexpression in pSS and pSS-associated lymphoma A, Salivary gland biopsies stained for IL-40 showing a marked increase in IL-40 expression in presence of higher focus score. B, Parotid gland specimens of pSS-associated lymphoma showing intense staining for IL-40.Disclosure of Interests:Chiara Rizzo: None declared, Marianna Lo Pizzo: None declared, Leila Mohammadnezhad: None declared, Vincenzo Luca Lentini: None declared, Diana Di Liberto: None declared, Giulia Grasso: None declared, Piero Ruscitti Consultant of: Pfizer, Novartis, Celgene, Abbvie, Roche, Lilly, Rorberto Giacomelli Consultant of: Pfizer, Novartis, Celgene, Abbvie, Roche, Lilly, francesco ciccia Consultant of: Pfizer, Novartis, Celgene, Abbvie, Roche, Lilly, Giuliana Guggino Consultant of: Pfizer, Novartis, Celgene, Abbvie, Roche, Lilly

New pro-inflammatory cytokine IL-40 is produced by activated neutrophils and plays a role in the early stages of seropositive rheumatoid arthritis

“…Here, we show that IL-40 is up-regulated in the serum of ERA patients compared with healthy individuals, which is in agreement with studies on patients with RA 11 12 and SS 9 and further highlights the role of IL-40 in the pathogenesis of autoimmune inflammation. As this study was performed simultaneously with the study on RA 11 with sample replicates to assess plate-to-plate consistency, we can state that the levels of IL-40 in patients with ERA are approximately two times lower compared with long-standing RA.…”

“…Interleukin (IL)-40 (encoded by the C17orf99 gene) represents one of the newly described cytokines implicated in autoimmune inflammation. [7][8][9][10][11][12] IL-40 was identified in 2017 as a small (27 kDa) secreted protein with a unique structure distinct from any known cytokine family. 7 Initially, IL-40 was detected in organs associated with the immune system such as the fetal liver, bone marrow or activated B-cells potentiated by IL-4 and transforming growth factor (TGFβ)-1.…”

“… 7 8 13 Recent data show that IL-40 is also expressed by other leucocytes or fibroblasts. 9 11 In mice, IL-40 expression peaks with lactation, and C17orf99 - / - mice exert microbiome alternations, reduced IgA levels and impaired B-cell functions. 7 In addition, a link between IL-40 and malignancy has been proposed based on the differential expression of IL-40 found in B-cell lymphomas.…”

A Comprehensive Review on the Role of Interleukin-40 as a Biomarker for Diagnosing Inflammatory Diseases

“…Supporting this result, researches showed that neutrophils from arthritis patients are one of the important sources of interleukin-40 production during the early stages of the disease [17]. However, another study revealed that CD4+, CD8+ T cells, and CD68+ macrophages are additional producers of IL-40 in pSS patients, supporting the hypothesis that other immune cells may also create IL-40 [18]. Terefore, it is not just secreted by B cells and for the development of appropriate treatments to target this infammatory cytokine in the future, it is crucial to identify the main source from which this cytokine is secreted in various diseases.…”

“…Sjogren's syndrome (pSS). Interestingly, the expression of IL-40 was noticeably enhanced in two types of patients compared with controls [18]. In support of the role of IL-40 in infammation, studies showed that in the presence of the anti-infammatory cytokine like IL-38 in the coculture of human respiratory epithelial cells with macrophages, C17orf99 was downregulated [15].…”