S-98 1 Department of Medicine, Surgery and Neurosciences; 2 Department of Life Sciences, University of Siena, Italy; 3 Research Laboratory and Academic Division of Clinical Rheumatology, Department of Internal Medicine, University of Genova, Genova, Italy. Claudio Corallo, PhD Luana Paulesu, PhD, Prof. Maurizio Cutolo, MD, Prof. Francesca Ietta, PhD, Assoc. Prof. Claudio Carotenuto, PhD Chiara Mannelli, PhD Roberta Romagnoli, PhD Ranuccio Nuti, MD, Prof. Nicola Giordano, MD, Assoc. Prof. Please address correspondence to: Prof. Nicola Giordano, Department of Medicine, Surgery and Neurosciences, University of Siena, Italy. E-mail: nicola.giordano@unisi.it Received on September 29, 2014; accepted in revised form on January 13, 2015. Clin Exp Rheumatol 2015; 33 (Suppl. 91): S98-S105. © Copyright CliniCal and ExpErimEntal rhEumatology 2015. Key words: systemic sclerosis, macrophage migration inhibitory factor MIF, fbroblasts, microvascular endothelial cells, fbrosis Competing interests: none declared. ABSTRACT Objective. To investigate serum lev- els, tissue/cellular expression of mac- rophage migration inhibitory factor (MIF) in patients with limited (lSSc) and diffuse (dSSc) systemic sclerosis. Methods. 10 lSSc-patients, 10 dSSc- patients and 10 controls were enrolled. MIF serum levels were assayed by ELISA. MIF and its receptors CD74/ CD44 were evaluated by immunohis- tochemistry on skin biopsies from pa- tients with dSSc, lSSc (affected and not- affected skin) and controls. MIF levels were assessed (ELISA) in supernatants of healthy dermal microvascular en- dothelial cells (MVECs) and in control (CTR), non-affected SSc (NA) and af- fected (SSc) fbroblasts treated for 48h with 10% control serum and 10% SSc- serum. MIF supernatant (ELISA) and mRNA (quantitative real-time PCR) lev- els were determined in SSc dermal fbro- blasts and in control dermal fbroblasts untreated or stimulated at 6h-24h-48h with bleomycin (50mU/ml). Results. Serum MIF was signifcant- ly higher in dSSc (18.7±4.1 ng/ml, p<0.001) and in lSSc (10.4±4.4 ng/ml, p<0.001) patients respect to controls (2.6±1.4 ng/ml). Enhanced MIF immu- noreactivity was found in keratinocytes, fbroblasts, endothelium, sebaceous/ sweat glands from lSSc/dSSc affected skin. Faint MIF immunoreactivity was found in control skin and not-affected skin of lSSc patients. No differences were found in CD74/CD44 receptors’ analysis among control and dSSc/lSSc affected and non-affected skin. MVECs and fbroblasts (CTR, NA and SSc) pro- duced signifcantly more MIF, when stimulated with SSc serum respect to control-serum (p<0.001). Finally, MIF mRNA levels signifcantly increased at 6h (p<0.001) and decreased at 48h (p<0.001) in control fbroblasts treated with bleomycin compared to control un- treated. Simultaneously, MIF superna- tant protein levels increased after 48h (p<0.01) in bleomycin-treated fbro- blasts respect to untreated ones. Conclusion. These results suggest that MIF could be implicated in the patho- genesis of SSc, probably acting as pro- tective factor against the SSc stressful conditions. Introduction Systemic sclerosis (SSc) is an autoim- mune connective tissue disease charac- terised by microvascular damage and progressive skin and internal organ fbrosis (1). Its pathogenesis remains incompletely understood (1). Immune activation, vascular impairment, and excessive synthesis of extracellular ma- trix are all known to be important in the development of this illness (2). Despite SSc is considered as a fbrotic disease with relatively poor infammatory com- ponent (3), recent data demonstrated the contribution of some pro-infammatory cytokines in the development of the dis- order (4). Macrophage migration inhibi- tory factor (MIF) is a multi-functional protein that operates as a cytokine and an enzyme (5). As a cytokine, MIF acts as a major regulator of infammation and a central upstream mediator of in- nate immune response (5); moreover, it functions as a key mediator to counter- regulate the immune-inhibitory effects of glucocorticoids (6). MIF was initially identifed as the protein secreted by acti- vated T lymphocytes, capable of inhibit- ing random migration of macrophages, concentrating macrophages at infam- mation loci, and enhancing their ability to kill intracellular parasites and tumour- al cells (7, 8). Nowadays, several types of cells (macrophages, endothelial cells, Serum levels, tissue expression and cellular secretion of macrophage migration inhibitory factor in limited and diffuse systemic sclerosis C. Corallo 1 , L. Paulesu 2 , M. Cutolo 3 , F. Ietta 2 , C. Carotenuto 2 , C. Mannelli 2 , R. Romagnoli 2 , R. Nuti 1 , N. Giordano 1