EXTENDED REPORT IL-17-producing CD4 - CD8 - T cells are expanded in the peripheral blood, infiltrate salivary glands and are resistant to corticosteroids in patients with primary Sjögren’s syndrome Alessia Alunno, 1 Onelia Bistoni, 1 Elena Bartoloni, 1 Sara Caterbi, 1 Barbara Bigerna, 2 Alessia Tabarrini, 2 Roberta Mannucci, 3 Brunangelo Falini, 2 Roberto Gerli 1 ▸ Additional data are published online only. To view these files please visit the journal online (http://dx.doi. org/10.1136/annrheumdis- 2012-201511). 1 Rheumatology Unit, Department of Clinical & Experimental Medicine, University of Perugia, Perugia, Italy 2 Department of Clinical & Experimental Medicine, Institute of Hematology, University of Perugia, Perugia, Italy 3 Laboratory of Confocal Microscopy and Image Analysis, University of Perugia, Perugia, Italy Correspondence to Professor Roberto Gerli, Rheumatology Unit, Department of Clinical & Experimental Medicine, University of Perugia, Via Enrico Dal Pozzo, Perugia I- 06122, Italy; gerlir@unipg.it Received 10 February 2012 Accepted 18 July 2012 Published Online First 17 August 2012 ABSTRACT Objectives It has been recently observed that a T-cell subset, lacking of both CD4 and CD8 molecules and defined as double negative (DN), is expanded in the blood of patients with systemic lupus erythematosus, produces IL-17 and accumulates in the kidney during nephritis. Since IL-17 production is enhanced in salivary gland infiltrates of primary Sjögren’s syndrome (SS) patients, we investigated whether DN T cells may be involved in the pathogenesis of salivary gland damage. Methods Phenotypic characterisation of peripheral blood mononuclear cells from SS patients and controls was performed by flow cytometry in freshly isolated and anti-CD3-stimulated cells. SS minor salivary glands were processed for immunofluorescence staining. Results CD3 + CD4 - CD8 - DN T cells were major producers of IL-17 in SS and expressed ROR-γt. They were expanded in the peripheral blood, spontaneously produced IL-17 and infiltrated salivary glands. In addition, the expansion of αβ-TCR + DN T cells was associated with disease activity. Notably, IL-17-producing DN T cells from SS patients, but not from healthy controls, were strongly resistant to the in vitro effect of dexamethasone. Conclusions These findings appear to be of great interest since the identi fication of a peculiar T-cell subset with pro-inflammatory activity, but resistant to corticosteroids, in an autoimmune disorder such as SS may help to design new specific treatments for the disease. INTRODUCTION Primary Sjögren’s syndrome (SS) is a systemic auto- immune disease primarily characterised by chronic inflammation of exocrine glands that leads to com- promised secretory function and tissue damage. 1 Surprisingly, despite the recognised autoimmune nature of the disease, there is no evidence for the efficacy of corticosteroids (CS) or other traditional immunosuppressive drugs in affecting the natural course of chronic inflammatory infiltration and its destructive potential of exocrine glands. 2 It is thought that T helper 1 (Th1) cells are major players in the induction of glandular damage. However, the discovery of Th17 cells chal- lenged the long-standing Th1–Th2 paradigm and prompted a re-evaluation of glandular infiltrate in SS. Th17 cells, indeed, represent a main pathogenic effector subset involved in inflammation and autoimmunity 3–5 and are actively involved in glan- dular tissue damage of SS. 6–9 Th17 lymphocytes are developmentally and functionally divergent from classic Th1 and Th2 cells. 10–12 They can derive from CD4 T cells acti- vated by interleukin (IL)-6 and transforming growth factor (TGF)-β stimulation. 10–12 However, CD4 expression by Th17 cells is not mandatory, since a small CD3 cell population, lacking both CD4 and CD8, can produce IL-17 following T cell receptor (TCR) stimulation. 13 These cells, known as CD4 - CD8 - double negative (DN), represent less than 5% of the total peripheral blood (PB) T-cell pool and express either αβ- or γδ-TCR. The conflict- ing data that have been published about DN T-cell functional role could be explained by their identifi- cation by exclusion and, therefore, may include T cells with different functions, including suppressive activity. 14–16 Several lines of evidence, however, appear to suggest that DN T cells play a major role in the pathogenesis of autoimmune disorders. DN T-cell accumulation is associated with the develop- ment and severity of autoimmune disease in animal models. 17 In addition, deficiency of Fas or its ligand causes dramatic accumulation of DN T cells in autoimmune lymphoproliferative syndrome. 18 Increased DN T-cell proportions, associated with anti-DNA production and kidney involvement, have been described in systemic lupus erythemato- sus (SLE). 19 20 In these patients, DN T cells can promote immunoglobulin B cell production. 21 22 More importantly, DN T lymphocytes represent a large component of kidney-infiltrating T cells during lupus nephritis and also represent a major source of IL-17 in SLE. 23 The evidence of a pro-inflammatory role of DN T cells in autoimmune disorders and the recogni- tion of a potential pathogenic involvement of IL-17 in SS inflammatory glandular damage prompted us to investigate a possible involvement of DN T lymphocytes in SS pathogenesis and to verify whether these T cells may contribute to increased production of IL-17 in this disorder. 24 METHODS Patients Thirty consecutive patients with SS, classified according to the Euro-American criteria, 25 were 286 Ann Rheum Dis 2013;72:286–292. doi:10.1136/annrheumdis-2012-201511 Basic and translational research group.bmj.com on January 17, 2013 - Published by ard.bmj.com Downloaded from