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Background CXCL4 (platelet factor4, PF4) is a pleiotropic α-granules chemokine, produced by platelets, dendritic cells, macrophages and neutrophils. Recently, CXCL4 levels have been found to increase under treatment with infliximab, in unresponsive Rheumatoid Arthritis (RA) patients [1]. At present, the influence of non TNFalfa blockers biological DMARDs (Abatacept and Tocilizumab) on CXCL4 serum levels not have been investigated in RA. Objectives To investigate CXCL4 serum levels in RA patients naive for biological disease-modifying anti-rheumatic drugs (bDMARDs) at baseline and after 1-year treatment and look for associations with disease features at baseline and distinct biological therapy overtime. Methods Fifty-six RA were enrolled in the study at the time of initiating a treatment with a biological DMARD. CXCL4 serum levels were investigated by a multiplex suspension immunoassay at baseline and after 1 year and compared with those detected in 30 healthy controls. Epidemiological, clinical and serological features of RA patients at baseline are showed in table 1. Results Out 56 RA patients, 39 (70%) were assigned to an anti-TNF-α treatment (3 patients to Infliximab, 12 to Etanercept, 24 to Adalimumab); 10 to Abatacept and 7 to Tocilizumab. At baseline, CXCL4 serum levels were found to be increased in RA patients as compared to controls but the difference was not significant (1,538 ng/ml vs 1.26 ng/ml, p=0.082). Moreover, CXCL4 levels were not found to be related to any disease feature. At baseline, 6 (10%) RA patients presented serum CXCL4 levels exceeding the 95th percentile of the values detected in controls (4.569 ng/ml). These patients did not differed from the remaining RA patients in any feature. After 1 year of treatment, CXCL4 levels increased with respect to basal values in the whole series (3.280 ng/ml vs 1.538 ng/ml, p Conclusions Similarly to previous studies (1) we found an increase in CXCL4 serum levels in patients undergoing an antiTNFa agent, but we failed to find differences between responsive and unresponsive patients. Moreover, we found that treatment with Abatacept and Tocilizumab did not modify CXCL4 serum level. Finally, we failed to detect any correlation between CXCL4 levels and any disease feature both at baseline and during follow-up. References Trocme C., et al. Ann Rheum Dis. Aug 2009; 68(8): 1328–1333 Disclosure of Interest None declared

Objectives: To investigate the role of microRNA-193b-3p (miR-193b) in the vascular pathophysiology of systemic sclerosis (SSc). Methods: Expression of miR-193b in skin biopsies and fibroblasts from patients with SSc and normal healthy (NH) controls were determined by real-time PCR. Transfection with miR-193b precursor and inhibitor were used to confirm targets of miR-193b. Proliferative effects of urokinase-type plasminogen activator (uPA) were determined by water-soluble tetrazolium salt-1 assay and by analysis of proliferating cell nuclear antigen expression. Fluorescence activated cell sorting analysis was performed to investigate the effect of uPA on apoptosis. For inhibition of the uPA-cellular receptor for uPA (uPAR) pathway, uPAR neutralising antibodies and low molecular weight uPA were used. Results: We found that miR-193b was downregulated in SSc fibroblasts and skin sections as compared with NH controls. The expression of miR-193b was not affected by major profibrotic cytokines and hypoxia. Induction of miR-193b in SSc fibroblasts suppressed, and accordingly, knockdown of miR-193b increased the levels of messenger RNA and protein for uPA. uPA was found to be upregulated in SSc as compared with NH controls in a transforming growth factor-β dependent manner, and uPA was strongly expressed in vascular smooth muscle cells in SSc skin section. Interestingly, uPA induced cell proliferation and inhibited apoptosis of human pulmonary artery smooth muscle cells, and these effects were independent of uPAR signalling. Conclusions: In SSc, the downregulation of miR-193b induces the expression of uPA, which increases the number of vascular smooth muscle cells in an uPAR-independent manner and thereby contributes to the proliferative vasculopathy with intimal hyperplasia characteristic for SSc.

Enlargement and dysfunction of the right atrium might be an early sign for pulmonary hypertension in systemic sclerosis (SSc). This is the first study to analyse right atrial morphology and function in SSc patients compared to healthy controls by speckle-tracking two-dimensional strain echocardiography (2DSE) at rest and during exercise. Furthermore, right atrial function was correlated with further clinical findings. Adult patients with SSc for > 3 years (n = 90) and 55 age- and gender-matched healthy controls underwent a panel of non-invasive assessments including transthoracic echocardiography, pulsed Doppler myocardial imaging and 2DSE at rest and during exercise. Furthermore, serological tests and high-resolution chest computed tomography were performed. SSc patients showed significant impairment of right atrial function and the right atrial enlargement, measured by 2DSE at rest and during exercise compared to controls (both p < 0.001). These findings were more evident in SSc patients with pulmonary fibrosis (p < 0.001) and in patients with high pulmonary artery systolic pressures (PAPs) during exercise. In the SSC patients, right atrial lateral strain was significantly associated with PAPs during effort, right atrial area, left ventricle stroke volume and inferior vena cava diameter using multivariable analysis. The findings of this study suggest that a high proportion of SSc patients reveal right atrial dysfunction even without manifest pulmonary hypertension. Impaired right atrial function occurred mostly in patients with pulmonary fibrosis and/or elevated PAPs during exercise, was independently associated with prognostic factors and may therefore be useful for risk stratification. Further studies are needed to analyse if right atrial dysfunction assessed by 2DSE may help to improve early diagnosis of pulmonary hypertension.

ObjectivesInterleukin (IL) 22 mRNA in systemic sclerosis (SSc) skin and Th22 cells in SSc peripheral blood are increased, but the role of IL-22 in fibrosis development remains poorly understood.MethodsBiopsies were obtained from the involved skin of 15 SSc, 4 morphea and 8 healthy donors (HD). The presence of IL-22+ cells in the skin was determined by immunostaining. The in vitro response of HD and SSc fibroblasts to IL-22, IL-22 in conjunction with tumour necrosis factor (TNF) or keratinocyte conditioned medium was assessed by ELISA, radioimmunoassay (RIA), real-time PCR and western blot. The in vivo response in mice was assessed by histomorphometry.ResultsIL-22+ cells were over-represented in the dermis and epidermis of morphea and in the epidermis of SSc compared with HD. The majority of dermal IL-22+ cells were T cells. Dermal fibroblasts expressed both IL-22 receptor subunits IL-10RB and IL-22RA, expression of which was enhanced by TNF and reduced by transforming growth factor (TGF)-β. IL-22 induced rapid phosphorylation of p38 and ERK1/2 in fibroblasts, but failed to induce the synthesis of chemokines and extracellular matrix components. However, IL-22 enhanced the production of monocyte chemotactic protein 1, IL-8 and matrix metalloproteinase 1 induced by TNF. Fibroblast responses were maximal in the presence of conditioned medium from keratinocytes activated by IL-22 in conjunction with TNF. Dermal thickness was maximal in mice injected simultaneously with IL-22 and TNF.ConclusionsIL-22 capacitates fibroblast responses to TNF and promotes a proinflammatory fibroblast phenotype by favouring TNF-induced keratinocyte activation. These results define a novel role for keratinocyte–fibroblast interactions in the context of skin fibrosis.

Summary Human leucocyte antigen (HLA)-G has a tolerogenic function and could play a role in the pathogenesis of immune-mediated diseases, including systemic sclerosis (SSc). The aim of this study was to evaluate HLA-G serum expression (sHLA-G) and the HLA-G gene 14 base pairs (bp) insertion/deletion (del−/del+) polymorphism in patients with Ssc, to search for possible associations with clinical and laboratory variables. sHLA-G was measured by enzyme-linked immunosorbent assay (ELISA) in sera from 77 patients with SSc and 32 healthy donors (HD); the 14 bp del−/del+ polymorphism was evaluated by polymerase chain reaction (PCR) amplification of peripheral blood mononuclear cells (PBMC) genomic DNA. Receiver operating characteristics (ROC) analysis identified the HLA-G cut-off that best discriminated dichotomized clinical and serological variables, that was subsequently employed to subdivide SSc patients into HLA-G high (HLA-G+) and low (HLA-G−) profile groups. sHLA-G were not statistically different between SSc patients and HD, nor between distinct SSc autoantibody subsets. Subdividing SSc patients by HLA-G positivity or negativity yielded significant differences for the modified Rodnan skin score (mRss) (P = 0·032), ‘general’ (P = 0·031) and ‘kidney’ (P = 0·028) Medsger severity scores (MSS) and disease activity index, and especially Δ heart/lung (P = 0·005). A worse ‘general’ MSS (P = 0·002) and Δ heart/lung (P = 0·011) were more frequent in the low sHLA-G group. These two variables and mRss were associated with sHLA-G levels at logistic regression analysis. Treatment had no influence on sHLA-G. Moreover, a higher frequency of scleredema was detected in the del+/del+ than the del-/del+ group (P = 0.04). These data suggest modulatory effects of sHLA-G on SSc. Prospective studies are needed to investigate a role in predicting the disease course.

Objective To investigate the long-term disease course of patients with recently deteriorated systemic sclerosis (SSC)-interstitial lung disease (ILD) undergoing continuous immunosuppressive treatment with cyclophosphamide (CYC) as induction therapy. Methods A total of 45 consecutive SSc patients were treated with weekly pulses of 500mg of CYC up to 10-g cumulative dose followed by azathioprine (AZA) in those experiencing improvement (>10% increase) or stabilization of both forced vital capacity and diffusion lung capacity for carbon dioxide and by micophenolic acid (MMF) in those experiencing deterioration (>10% decrease of either parameter). The follow-up ranged from 6 to 62 months post-CYC regimen (median = 36 months). Results Overall, 39 patients completed the CYC regimen. Of them, 24 (61.5%) experienced improvement or stabilization of lung function parameters and received AZA; the remaining 15 received MMF. During follow-up, lung function parameters improved in 3 (12.5%), remained stable in 18 (75%), and worsened in 3 (12.5%) AZA-treated patients, whereas they worsened in 8 (67%) and remained stable in 4 (33%) MMF-treated patients. The incidence of improvement or stabilization was significantly higher in AZA-treated than in MMF-treated patients ( p = 0.001). The time to the decline of lung function was significantly shorter in CYC non-responders, and CYC unresponsiveness was predictive of lung function worsening over time in a multivariate analysis (HR = 9.14; 95% CI: 2.28–36.64; p = 0.0018). Conclusion Our study supports the use of low-dose pulse CYC as induction therapy of recently deteriorated SSc-ILD. Moreover, it suggests that AZA should be administered to CYC-responsive patients but does not show any definite effect of MMF in unresponsive patients.

Background Identification of parameters for early diagnosis and treatment response would be beneficial for patients with early rheumatoid arthritis (ERA) to prevent ongoing joint damage. miRNAs have features of potential biomarkers, and an altered expression of miRNAs was shown in established rheumatoid arthritis (RA). Objective To analyse RA associated miRNAs in the sera of patients with ERA to find markers of early disease, clinical activity or predictors of disease outcome. Methods Total RNA was isolated from whole sera in ERA patients (prior to and after 3 and 12 months of therapy with disease modifying antirheumatic drugs), in patients with established RA and in healthy controls (HC) using phenol–chloroform extraction. Expression of miR-146a, miR-155, miR-223, miR-16, miR-203, miR-132 and miR-124a was analysed by TaqMan Real Time PCR. Results From all analysed miRNAs, levels of miR-146a, miR-155 and miR-16 were decreased in the sera of ERA patients in comparison with established RA. A change in circulating miR-16 in the first 3 months of therapy was associated with a decrease in DAS28 in long term follow-up in ERA (p=0.002). Levels of circulating miR-223 in treatment naive ERA correlated with C reactive protein (p=0.008), DAS28 (p=0.031) and change in DAS28 after 3 months (p=0.003) and 12 months (p=0.011) of follow-up. However, neither miR-16 nor miR-223 could distinguish ERA from HC. Conclusions Differential expression of circulating miR-146a, miR-155 and miR-16 in the sera of ERA patients may characterise an early stage of the disease. We suggest miR-223 as a marker of disease activity and miR-16 and miR-223 as possible predictors for disease outcome in ERA.

OBJECTIVES: In agreement with other autoimmune diseases, systemic sclerosis (SSc) is associated with a strong sex bias. However, unlike lupus, the effects of sex on disease phenotype and prognosis are poorly known. Therefore, we aimed to determine sex effects on outcomes. METHOD: We performed a prospective observational study using the latest 2013 data extract from the EULAR scleroderma trials and research (EUSTAR) cohort. We looked at (i) sex influence on disease characteristics at baseline and (ii) then focused on patients with at least 2 years of follow-up to estimate the effects of sex on disease progression and survival. RESULTS: 9182 patients with SSc were available (1321 men) for the baseline analyses. In multivariate analysis, male sex was independently associated with a higher risk of diffuse cutaneous subtype (OR: 1.68, (1.45 to 1.94); p

BACKGROUND: High interleukin (IL)-17A levels are characteristically found in the skin of systemic sclerosis (SSc) individuals. Our aim was to investigate whether the dermal expression of IL-17A and related IL-17 family members (i.e. IL-17C, IL-17E and IL-17F) could distinguish fibrotic from healthy skin and could show similarities in SSc and morphea, two disorders with presumed distinct pathogenesis, but characterized by skin fibrosis. METHODS: Biopsies were obtained from the involved skin of 14 SSc, 5 morphea and 8 healthy donors (HD) undergoing plastic surgery. Immunohistochemistry/immunofluorescence techniques were coupled to a semi-automated imaging quantification approach to determine the presence of the IL-17 family members in the skin. The in vitro effects induced by the IL-17 family members on fibroblasts from normal and SSc individuals were assessed by ELISA and RIA. RESULTS: Positive cells for each of the IL-17 isoforms investigated were present in the dermis of all the individuals tested, though with variable frequencies. SSc individuals had increased frequency of IL-17A+ (p = 0.0237) and decreased frequency of IL-17F+ (p = 0.0127) and IL-17C+ cells (p = 0.0008) when compared to HD. Similarly, morphea individuals had less frequent IL-17C+ cells (p = 0.0186) in their skin but showed similar number of IL-17A+ and IL-17F+ cells when compared to HD. Finally, IL-17E+ cells were more numerous in morphea (p = 0.0109) and tended to be more frequent in SSc than in HD. Fibroblast production of IL-6, MMP-1 and MCP-1 was enhanced in a dose-dependent manner in the presence of IL-17E and IL-17F, but not in the presence of IL-17C. None of the cytokine tested had significant effect on type I collagen production. Of interest, in SSc the frequency of both IL-17A and IL-17F positive cells increased with disease duration. CONCLUSIONS: The frequency of IL-17A and IL-17F distinguish SSc to morphea individuals while dermal expression of IL-17C (low) and IL-17E (high) identifies a fibrosis-specific motif. The specific IL-17C/IL-17E cytokine combination may thus play a role in the development of fibrosis. PMID: 25136988 PMCID: PMC4138152 DOI: 10.1371/journal.pone.0105008

For many years observational studies and clinical trials on systemic sclerosis (SSc) have been carried out only on patients who met the 1980 American College of Rheumatology-ACR preliminary classification criteria. However, this lead to the exclusion from all those studies of subset patients, particularly those with a limited cutaneous SSc-sine scleroderma subset, because, despite a diagnosis of SSc based on Raynaud's phenomenon (RP) associated with digital pitting scars/ulcers, or by sclerodactyly and telangiectasia and/or typical esophagopathy and/or interstitial fibrosis detected by High Resolution Computed Tomography of the lungs, they did not satisfy the classification criteria. In that setting, LeRoy and Medsger proposed to label as affected by limited SSc (lSSc) or early SSc, cases presenting with RP associated with an SSc-type nailfold capillary pattern and/or SSc-selective autoantibodies. In 2008, Koenig et al. validated these criteria and proposed to name early SSc, or pre-scleroderma, patients with RP and either a scleroderma marker autoantibody or typical capillaroscopy abnormalities or both, who had been clearly shown to have high probabilities but not the certainty to develop definite SSc during a 20-year follow-up. This definition has been recently challenged by the development of the new ACR/EULAR criteria for the classification of SSc. At present, to be labeled as affected by early SSc, or pre-scleroderma, a patient should suffer from RP associated with either scleroderma marker autoantibodies or typical capillaroscopy findings and should not meet the 2013 ACR/EULAR criteria for the classification of SSc nor should he/she meet the criteria for SSc sine scleroderma.

Background IL-17A has been recently implicated in the pathogenesis of systemic sclerosis (SSc). Objectives Therefore, we explored its expression and effects in peripheral blood mononuclear cells (PBMCs) co-cultured with autologous skin fibroblasts. Methods PBMCs and autologous skin fibroblasts from 5 patients with early (disease duration Results Real-time PCR analysis showed an increased expression of IL17A in co-cultured PBMCs by 11.5 fold (p Lastly, we investigated the effects of co-cultured PBMCs on the expression of pro-fibrotic genes in fibroblasts. We found that COL1A1, COL3A1, and CTGF mRNAs were down-regulated by 0.33 fold, 0.24 fold, and 0.31 fold respectively (p Conclusions Here we show for the first time that PBMCs from early dcSSc co-cultured with autologous skin fibroblasts over-express IL17A and exert anti-fibrotic effects in vitro. The simultaneous up-regulation of IL17RA and IL-17A target genes in the co-cultured fibroblasts suggests that IL-17A pathway is active in early dcSSc fibroblasts. These findings are paralleled by the down-regulation of TGF-beta signalling components. We previously showed that in co-cultures performed with PBMCs and autologous skin fibroblasts from early dcSSc patients T cells are expanded and kill the autologous fibroblasts. Taken together, these novel data support the hypothesis that immune system may be primarily aimed to control fibroblast activation in the early phases of the disease, potentially opening new therapeutic approaches in SSc. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.5910

pURPOSE: To assess the serum profile of factors involved in endothelial, T-cell, and fibroblast interplay in patients with Raynaud's phenomenon (RP) associated with nailfold vodeocapillaroscopy (NVC) scleroderma findings and/or systemic sclerosis (SSc) marker autoantibodies, recently labeled as early SSc patients. METHODS: Serum levels of soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular adhesion molecule-1 (sVCAM-1), CCL2, CXCL8, IL-13, IL-33, and transforming growth factor-β (TGF-β) were measured in 24 early SSc patients, 48 definite SSc patients, and 24 osteoarthritis/fibromyalgia controls by multiplex suspension immunoassay. All SSc patients were investigated for the presence/absence of preclinical and clinical organ involvement, SSc marker autoantibodies, and NVC abnormalities. RESULTS: Serum sICAM-1, CCL2, CXCL8, and IL-13 were increased in all SSc patients as compared to controls, and paralleled the severity of the disease subset (early SSc < limited cutaneous SSc < diffuse cutaneous SSc; p

INTRODUCTION: Early systemic sclerosis (SSc) is characterized by Raynaud's phenomenon together with scleroderma marker autoantibodies and/or a scleroderma pattern at capillaroscopy and no other distinctive feature of SSc. Patients presenting with marker autoantibodies plus a capillaroscopic scleroderma pattern seem to evolve into definite SSc more frequently than patients with either feature. Whether early SSc patients with only marker autoantibodies or capillaroscopic positivity differ in any aspect at presentation is unclear. METHODS: Seventy-one consecutive early SSc patients were investigated for preclinical cardiopulmonary alterations. Out of these, 44 patients and 25 controls affected by osteoarthritis or primary fibromyalgia syndrome were also investigated for serum markers of fibroblast (carboxyterminal propeptide of collagen I), endothelial (soluble E-selectin) and T-cell (soluble IL-2 receptor alpha) activation. RESULTS: Thirty-two of the 71 patients (45.1%) had both a marker autoantibody and a capillaroscopic scleroderma pattern (subset 1), 16 patients (22.5%) had only a marker autoantibody (subset 2), and 23 patients (32.4%) had only a capillaroscopic scleroderma pattern (subset 3). Patients with marker autoantibodies (n = 48, 67.6%) had a higher prevalence of impaired diffusing lung capacity for carbon monoxide (P = 0.0217) and increased serum levels of carboxyterminal propeptide of collagen I (P = 0.0037), regardless of capillaroscopic alterations. Patients with a capillaroscopic scleroderma pattern (n = 55, 77.5%) had a higher prevalence of puffy fingers (P = 0.0001) and increased serum levels of soluble E-selectin (P = 0.0003) regardless of marker autoantibodies. CONCLUSION: These results suggest that the autoantibody and microvascular patterns in early SSc may each be related to different clinical-preclinical features and circulating activation markers at presentation. Longitudinal studies are warranted to investigate whether these subsets undergo a different disease course over time.

Background Systemic sclerosis (SSc) is a chronic autoimmune disease characterized by vascular changes and progressive fibrosis of skin and various internal organs. A variety of circulating autoantibodies have been detected in SSc. Some of them are well-established tools strongly associated with SSc and useful for diagnosis and management of the disease (e.g. ANA, anti-centromere, anti-topoisomerase I, anti-RNA polymerase III – RNAP III). Other autoantibodies are less frequent and non-specific for SSc even if potentially useful to better assess clinical subsets and prognosis (e.g. anti-Th/To, anti-Ku, anti-PM/Scl). Objectives To investigate the prevalence of anti-Th/To, anti-Ku, anti-RNAP III and anti-PM/Scl antibodies as well as to study the associations between these antibodies and clinical features in Italian SSc patients. Methods Sera from 176 consecutive patients with SSc collected at two Italian sites were tested for anti-Th/To (anti-Rpp25 and anti-Rpp38), anti-Ku, anti-RNAP III, anti-PM/Scl, and anti-centromere antibodies (anti-CENP-A and B) using QUANTA Flash chemiluminescence immunoassays (CIAs) on the BIO-FLASH instrument (Inova Diagnostics, San Diego, USA). Additionally, ANA screening was performed using QUANTA Flash CTD Screen Plus which includes the most relevant nuclear/cytoplasmic antigens. The Italian cohort included: women 162 (92%), limited cutaneous (lc-SSc) subtype 137 (78%), patients with present or past history of digital ulcers 56 (32%), calcinosis 25 (14%), lung fibrosis 69 (39%), heart involvement 22 (12.5%), pulmonary arterial hypertension 11 (6%) and esophageal involvement 84 (48%). Results 152 (86.4%) patients were CTD Screen Plus positive, 8 (4.5%) anti-Th/To positive, 8 (4.5%) anti-Ku positive, 20 (11.4%) anti-RNAP III positive, 0 (0%) anti-PM/Scl positive, 70 (39.8%) anti-CENP-A positive, and 87 (49.3%) anti-CENP-B positive. Anti-CENP-A, anti-CENP-B and anti-Ku antibodies were significantly associated with the lc-SSc subtype ( p p p =0.0469, respectively). None of patients positive for anti-Ku antibodies displayed digital ulcers ( p =0.0075). Anti-Th/To (Rpp25) antibodies were associated with the presence of calcinosis ( p =0.0399) while anti-RNAP III antibodies with its absence ( p =0.0353). All patients positive for anti-Th/To were positive for the Rpp25 component while only one of them was also positive for the Rpp38 component. Conclusions The present study points out the low prevalence of anti-RNAP III and highlights a lower prevalence of anti-Th/To in Italian SSc patients. Several known associations between autoantibodies as SSc subsets have been confirmed while others could not be verified in the study. This supports the potential utiliy of autoantibodies to stratify SSc into clinical subsets. Large international multi-center studies are required to define and validate rare autoantibodies such as anti-Th/To, anti-Ku and anti-PM/Scl antibodies. Disclosure of Interest None declared

Background The -455 G>A polymorphism in the proximal promoter region of β-fibrinogen gene is known to increase fibrinogen plasma levels and to promote arterial and venous thrombosis in the general population (1). We previously showed that systemic sclerosis (SSc) patients with pitting scars and/or ulcers have a higher frequency of the β-fibrinogen -455 A allele and higher plasma levels of fibrinogen (2). Objectives We aimed to investigate fibrinogen plasma levels in a prospective cohort of SSc patients and to search for associations with features of vascular involvement at follow-up (FU). Methods SSc patients meeting the 2013 ACR/EULAR criteria for disease classification with measurement of fibrinogen plasma levels and at least one FU visit with complete clinical evaluation were enrolled. The comparison of fibrinogen levels in different patient groups was performed using the Mann-Whitney U test; differences between patients with fibrinogen levels higher or equal/lower than the median value were analyzed by Kaplan-Maier curves applying the log-rank test. Results One-hundred and thirty-three SSc patients, aged 51±13 years, were investigated (91% females; median disease duration 8 years, range 1–49). Seventy-six % of patients had a limited SSc subset (lcSSc); 24% had a diffuse subset (dcSSc). All patients were ANA positive (40% had anti-centromere antibodies, 35% anti-Scl70+; 3% anti-RNApolymerase III; 2.3% anti-PmScl; 1.5% anti-U1RNP). All patients were under low-dose aspirin and oral vasodilators. Forty-one % of patients had already pitting scars and/or ulcers at baseline; 13% developed new digital ischemic injury during observation (median FU 3 years; range 0.7–22). Baseline fibrinogen levels were higher in patients who developed digital ischemic lesions during FU (385 mg/dl, range 180–688 versus 344 mg/dl, range 81–765; p 2 =4.3; p Conclusions Consistently with our previous findings, here we first show that higher fibrinogen levels at baseline in SSc patients are associated with a faster development of new digital ischemic lesions over time. Of note, all our patients were taking aspirin, which, however, does not affect platelet-fibrinogen interaction. In addition, all of them were taking an oral vasodilator, that is recommended for the treatment of peripheral vasculopathy in SSc (3). Randomized controlled clinical trials in larger SSc cohorts to explore the potential benefit of gpIIb/IIIa inhibitors± low-dose aspirin in these patients are needed. References van9t Hooft FM et al.Arterioscler Thromb Vasc Biol 1999;19:3063–70 Vettori S et al. Clin Exp Rheumatol 2010;28:923–4 Kowal-Bielecka O et al. Ann Rheum Dis 2009; 68:620–628 Disclosure of Interest None declared

Background Circumstantial evidence from preclinical studies indicates a key role of serotonin (5-HT) signaling via the 5-HT-2B receptor in the development of fibrosis. Terguride is an orally available 5-HT-2 receptor inhibitor that has shown beneficial effects in different animal models of systemic sclerosis (SSc). Objectives To evaluate the efficacy of Terguride for the treatment of fibrosis in patients with SSc in an investigator initiated phase 2 proof of concept study. Methods Main inclusion criteria were fulfillment of ACR classification criteria and diffuse cutaneous SSc (dcSSc). Patients with end-stage organ involvement and treatment with potentially disease modifying agents including immunosuppressives were excluded. Patients were treated with Terguride at up to 3 mg/d p.o. or standard of care (post hoc control) for three months. Primary efficacy endpoints were changes of pre-defined skin biopsy biomarkers over the three months treatment period. Secondary efficacy endpoints included change of mRSS and lung function parameters. Serious adverse events (SAEs) and AEs were coded using MedDRA. The study was externally monitored. Results Twelve patients were recruited into the Terguride group and 6 patients into the control group. The primary endpoints, skin biopsy biomarkers, showed a consistent and statistically significant down-regulation compared to the control group (Figure) for dermal thickness, myofibroblast counts and mRNA levels of col1a1, col1a2 as well as for the Lafyatis 4-gene biomarker set (COMP, THSP-1, SIGLEC-1, IFI-44). This was accompanied by a reduction in mRSS of - 32.3% versus baseline in the Terguride group versus stable values in the control group (p Conclusions Terguride was well tolerated in patients with dcSSc. Strong and consistent effects on fibrosis related skin biopsy biomarkers could be observed in this open-label controlled phase 2 proof of concept study, which was further supported by a significant improvement of the mRSS over the control group. These data justify further investigation in an upcoming randomized placebo controlled phase 3 study. Disclosure of Interest O. Distler Grant/research support from: Bayer, Sanofi, Ergonex, Boehringer Ingelheim, Actelion, Pfizer, Consultant for: 4 D Science, Actelion, Active Biotec, Bayer, BiogenIdec, BMS, Boehringer Ingelheim, EpiPharm, Ergonex, espeRare foundation, Genentech/Roche, GSK, Inventiva, Lilly, medac, MedImmune, Pharmacyclics, Pfizer, Serodapharm, Sinoxa, B. Maurer: None declared, S. Vettori: None declared, S. Blumhardt: None declared, D. Frey: None declared, A. Distler: None declared, C. Beyer: None declared, J. H. Distler Shareholder of: 4 D Science, Consultant for: Actelion, BMS, Celgene, Bayer Pharma, Boehringer Ingelheim, JB Therapeutics, Sanofi-Aventis, Novartis, UCB, GSK, Array Biopharma and Active Biotech

Background In SSc, the management of DU is a problem in clinical practice, therefore their prevention is highly warranted. Sildenafil, iloprost, calcium channel blockers (CCB), ACE inhibitors (ACEi) and bosentan, alone or in combination, are used for DU prevention without unanimous agreement. Objectives DeSScipher (“to decipher the optimal management of SSc”) is an EC-funded FP7 research project that consists of five observational trials (OT) addressing different aspects of SSc. The aim of OT1 was to evaluate the efficacy of vasoactive and vasodilating drugs or a combination therapy for the management of DU in SSc. Methods Longitudinal data were collected for up to 24 months by 28 DeSScipher centres. In SSc patients, the efficacy of sildenafil, bosentan, sildenafil+bosentan, iloprost or CCB /ACEi for the prevention of new DU was evaluated. Clinical features, ulcer type and other drugs were also recorded. Time without new DU, proportion of patients without new DU, risk of developing new DU and the mean number of new DU per patient were evaluated at 6 months follow-up. Results In OT1, 1394 patients were enrolled. The history of DU was a significant risk factor for developing new DU (OR=3.146; (95%CI:1.19–8.31), p=0.021), therefore analysis was focused on secondary prevention (prevention of new DU in patients with DU history). 473/1394 (33.9%) had a history of DU and 268/473 (56.7%) had follow up data (58.2% limited and 41.8% diffuse SSc subset). 47/268 (17.5%) were on Bosentan, 33/268 (12.3%) on Sildenafil, 40/268 (14.9%) on Iloprost, 31/268 (11.6%) on Sildenafil and Bosentan combination and 117/268 (43.7%) on CCB/ACEi alone. In patients with history of DU in the last 24 weeks, the treatment with CCB/ACEi alone was associated with 7 fold increased risk of developing new DU compared to all other treatment arms (OR =7.313; 95%CI:1.248–42.85, p=0.027). Patients on concomitant immunosuppressive treatment had a trend towards faster DU recurrence. Conclusions History of DU in the past 24 weeks should prompt a more aggressive preventive strategy other than therapy with CCB/ACEi alone due to its lower efficacy to prevent DU recurrence compared to the other treatment options. Acknowledgement The DeSScipher project was funded by the European Community9s Framework Programme 7 (FP7-HEALTH-2012.2.4.4–2 - Observational trials in rare diseases) under grant agreement N° 305495. We acknowledge the contribution of the following EUSTAR centers: Ancona (member N°34), Assiut (168), Bad Bramstedt (187), Belgrade (55), Bucharest (100), Cluj-Napoca (16), Erlangen (106), Frankfurt (124), Hamburg (64), Iasi (162), Istanbul (21), Istanbul (133), Koln (44), Monserrato (142), Moscow (78), Moscow (190), Padua (31), Rome (94), Salford/Manchester (80), Tubingen (56), Wuppertal (192). Disclosure of Interest None declared

Background In a previous study it has been shown shown that, in patients with systemic sclerosis (SSc), the cumulative number of micro-haemorrhages (MH) and micro-thrombosis (MT), computed in eight finger nailfold videocapillaroscopy (NVC), and called NEMO score, was strongly predictive of disease activity (DA) measured by means of European Scleroderma Study Group (ESSG) score. Even the number of giant capillaries (GC) was correlated to a lesser extent with the ESSG score, and then a modified (m) NEMO score was proposed in which the combined presence of a MH/MT and GC was considered [1]. Objectives The present study was aimed at validating NEMO score in different cohorts of patients. Methods NEMO score and the total number of GC were assessed by eight finger NVC in 122 patients with SSc referring to the Rheumatic Disease Unit of the “Istituto G.Pini” of Milan, where the preliminary study had been carried out (internal validation). The same procedure was performed in 99 patients collected in a different centre, i.e., the Rheumatic Dept. of the II University of Naples (external validation). In all of the patients DA, measured by ESSG score, was assessed at the same time of NVC performance. Results Demographic and clinical characteristics of the patients were summarised in Table 1. Table 2 shows the main statistical results obtained by the analysis of the two cohorts. The mNEMO score does not add anything in terms of sensitivity and specificity to the performance of the simple NEMO score in predicting DA. Conclusions This validation study confirms that the presence of a certain number of MH plus that of MT in eight finger NVC may represent a simple screening method to predict the presence of DA in patients with SSc. References Sambataro D, et al. Arthritis Res & Ther 2014;16:462. Disclosure of Interest None declared

Background Patients with UCTD (1) have been reported to present lung involvement most frequently characterized by nonspecific interstitial pneumonia (NSIP) at High Resolution Computerized Assial Tomography (HR-CAT), FVC and DLCO values lower than predicted (2). Objectives To investigate UCTD patients for the presence of lung involvement and search for demographic, serological, capillaroscopic and organ involvement features associated with it. Methods Eighty-six patients consecutively admitted to the Rheumatology Unit of the Second University of Naples and satisfying criteria for the classification of UCTD (1) were enrolled into the study. Patients presenting with any major CTD marker autoantibody other than anti-Ro (SSA) and/or scleroderma capillaroscopic specific pattern were excluded. Each patient underwent a careful history, a complete physical examination, evaluation of serum Antinuclear (ANA) and anti Extraible Nuclear Antigens antibody (anti-ENA), videocapillaroscopy, echo-Doppler-cardiography with evaluation of Left ventricular E/A ratio. Those who consented also underwent HR-CAT and esophagotonometry. Interstitial lung disease (ILD) was defined by HR-CAT or, if unavailable, by FVC Results The table lists the features detected in the patients subdivided according to the presence of lung involvement. DLCO An isolated reduction of DLCO resulted the be the most frequent finding and might depend on pulmonary vascular disease. Anti-SSA positivity and Capillaroscopic alterations were detected more frequently in patients with lung involvement (either isolated reduction of DLCO or ILD). Conclusions We detected a prevalence of ILD in UCTD lower than previously reported (2) (80%). Nevertheless, we found an isolated reduction of DLCO in about 35% of the UCTD patients. Lung involvement resulted to be significantly associated with capillaroscopic alterations and anti-SSA positivity. References Mosca M, Tani C, Vagnani S, Carli L, Bombardieri S.The diagnosis and classification of undifferentiated connective tissue diseases. J Autoimmun. 2014;48–49:50–2 Kinder BW, Collard HR, Koth L, Daikh DI, Wolters PJ, Elicker B, Jones KD, King TE Jr. Am J Respir Crit Care Med. Oct 1;176(7):691–7 Disclosure of Interest None declared

Patients with systemic sclerosis (SSc) who express autoantibodies to centromeric proteins (CENPs) are at risk of developing pulmonary vascular disease and pulmonary arterial hypertension without fibrosis. Currently no biomarkers are available to predict these complications. We previously characterized the fine specificity of anti-CENP-A antibodies in SSc by screening a phage display library (expressing random 12-mer peptides), and identified phage clones whose peptides were differentially recognized by patients’ autoantibodies. Here, we examined if subgroups of SSc patients with different anti-CENP-A antibody subspecificities also differ clinically, and if serum reactivity to phage-displayed peptides can predict pulmonary vascular disease. Clinical data and serum samples were collected from 84 anti-CENP-A-positive SSc patients. Indirect ELISAs were used to test serum reactivity. Pulmonary vascular disease was defined as high systolic pulmonary arterial pressure (sPAP) and low diffusing lung capacity for carbon monoxide (DLCO; percent of predicted values). Sera were screened for reactivity to peptides expressed by phage clones pc4.2 and pc14.1, confirming our earlier observation of differential specificities. Linear regression showed that the levels of antibodies specific for the 2 phage clones were associated with clinical features of pulmonary vascular disease, but in opposite ways: anti-pc4.2 antibodies were positively associated with sPAP and inversely associated with DLCO, whereas anti-pc14.1 antibodies were inversely associated with sPAP and positively associated with DLCO. Anti-pc4.2 and anti-pc14.1 antibody levels predicted sPAP independently of DLCO. These associations were confirmed by logistic regression using antibodies as predictors and dichotomized sPAP (cutoff, 45 mm Hg) as outcome. The ratio of the 2 antibody levels was a useful marker in predicting high sPAP. This study demonstrates that some SSc clinical features associate with subspecificities of anti-CENP-A antibodies. Moreover, it shows that a simple, inexpensive phage-based assay can predict which SSc patients have high sPAP and low DLCO, hence who are at greater risk of developing pulmonary arterial hypertension. The ability to identify these at-risk patients can contribute to clinical efficiency and effectiveness. Further research into the peptides expressed by the phage clones may reveal the molecular mechanisms that put some anti-CENP-A-positive patients at greater risk than others for pulmonary vascular disease.

Background Systemic sclerosis (SSc) can greatly impact the patients9 quality of life due to the multisystem manifestations. The health assessment questionnaire (HAQ) is one of the most commonly used measures of disability in musculoskeletal disorders and was extended to form the scleroderma HAQ (SHAQ), a more disease-specific disability scale that incorporates the HAQ and the 5 SHAQ-visual analogue scales (VAS) into one score. Objectives This study aims to identify predictors of disability in SSc by means of the SHAQ. Methods Patients from the DeSScipher cohort were included in the analysis if they had at least one complete SHAQ recorded, fulfilled either the 1980 ACR or the 2013 ACR/EULAR criteria for SSc and were above 18 years of age. Multiple linear regression analysis was used to assess the combined effect of factors possibly associated with disability. Variables included in the model were defined a priori. Results 813 patients had one complete SHAQ recorded between June 2013 and January 2016 (34.1% of all patients followed in the DeSScipher cohort). Of these, 12% fulfilled only the 2013 ACR/EULAR criteria, 2% fulfilled only the 1980 ACR criteria and 86% fulfilled both, the median age was 58 years (IQR 49–67) and 15% were male. 26% of the patients were in the diffuse SSc subset, 56% in the limited and 18% had SSc sine sclerosis. The patients had a median SHAQ score of 0.79 (IQR 0.28–1.31) and a median HAQ score of 0.88 (IQR 0.25–1.5). The medians of the five VASs included in the SHAQ were (in order of the VAS magnitude): overall disease severity (30, IQR 10–51), Raynaud9s phenomenon (21, IQR 3–50), pulmonary symptoms (10, IQR 1–40), gastrointestinal symptoms (6, IQR 1–31) and digital ulcers (2, IQR 1–30). 60% of the patients were in the lowest SHAQ category (score of 0 to 1) which is regarded as “mild to moderate difficulty”, 34% in the category regarded as “moderate to severe disability” (score of 1 to 2) and 6% in the category regarded as “severe to very severe disability” (score of 2 to 3). In multiple linear regression, the main factors associated with high SHAQ scores were dyspnoea (NYHA-stage 4 (β=0.67), 3 (β=0.59) and 2 (β=0.18; all vs. stage 1), fibromyalgia (β=0.42), muscle weakness (β=0.25), current digital ulcers (β=0.20) and gastrointestinal symptoms (oesophageal symptoms, β=0.18; stomach symptoms, β=0.14; intestinal symptoms, β=0.14; figure). Conclusions Patients perceive dyspnoea, pain, digital ulcers, muscle weakness and gastrointestinal symptoms as the main factors driving their level of disability. Acknowledgement The DeSScipher project was funded by the European Community9s Framework Programme 7 (FP7-HEALTH-2012.2.4.4–2-Observational trials in rare diseases) under grant agreement N° 305495. We acknowledge the contribution of the following EUSTAR centres: Moscow(78), Padova(31), Istanbul(133), Wuppertal(192), Monserrato(142), Roma(94), Cologne(44), Cluj-Napoca(16), Ancona(34), Iasi(162), Frankfurt(124). Disclosure of Interest None declared

Background A reduced basal lower esophageal sphincter (LES) pressure (LES Objectives We aimed to investigate potential associations of a reduced basal LES pressure at first observation with the development of new gastrointestinal symptoms and with self-administered questionnaires devoted to assess the impact of gastrointestinal involvement on quality of life during follow-up (FU) in SSc patients. Methods SSc patients meeting the 2013 ACR/EULAR criteria for disease classification with measurement of basal LES pressure at the first observation and at least one FU visit with complete clinical evaluation were enrolled. The gastroesophageal reflux disease questionnaire (GerdQ) (2, 3) was administered to explore the impact of esophageal symptoms on patient9s disease perception and to estimate the risk for GERD. Associations between reduced basal LES pressure and new onset gastrointestinal symptoms at FU were analyzed by Fisher9s exact test, while the GerdQ score in SSc patients with or without a reduced basal LES pressure was analyzed by Mann-Whitney U test. Results Fifty-nine SSc patients, aged 47±13 years, were investigated (75% females; median disease duration 7 years, range 0.5–43). Eighty % of patients had a limited SSc subset (lcSSc); 20% had a diffuse subset (dcSSc). All patients were ANA positive (36% had anti-centromere antibodies, 39% anti-Scl70+; 5% anti-RNApolymerase III; 1.7% anti-PmScl; 1.7% anti-U1RNP). All patients were under proton pomp inhibitors ± prokinetics during FU. Forty-two % of patients had already esophageal symptoms at baseline; 32% developed new esophageal symptoms and 40% new gastric and/or intestinal symptoms during observation (median FU 4 years; range 0.5–15.5). Baseline basal LES pressure was reduced in 63% of patients. Out of these, 49% had also esophageal symptoms. Median GerdQ score at the end of FU was significantly higher in patients with baseline reduced LES pressure (7, range 3–14 versus 6, range 2–11; p Conclusions Our data suggest that evaluation at baseline of esophageal involvement in SSc patients by esophageal manometry can predict the development of more severe symptoms over time, as evaluated by the association with a higher score at GerdQ also in initially asymptomatic patients. Larger studies need to be carried out to confirm this association. References Sjogren RW. Arthritis Rheum 1994; 37: 1265–82 Jonasson C et al. Aliment Pharmacol Ther 2013; 37: 564–72 Chunlertrith K et al. Clin Exp Rheumatol 2014; 32 (6 Suppl 86): S98–102 Disclosure of Interest None declared

OBJECTIVE: To investigate health-related quality of life (HRQOL) in patients affected by early systemic sclerosis (eSSc) and to compare it with that of patients with undifferentiated connective tissue disease (UCTD). METHODS: At baseline, 31 eSSc and 35 UCTD patients underwent clinical evaluation, laboratory investigations, nailfold videocapillaroscopy, echocardiography, and lung function tests. All patients and 40 controls, matched for sex and age completed the Short Form-36 (SF-36) questionnaire and the Health Assessment Questionnaire Disability Index (HAQ-DI). RESULTS: SF-36 scores were significantly lower in eSSc and UCTD patients than in healthy controls as regards the following domains: physical component score (PCS), mental component score (MCS), physical functioning, role-physical, bodily pain, general health and mental health. PCS was negatively correlated to the HAQ-DI (rho -0.59; p=0.0004) and ESR >20 mm/h (rho -0.58; p=0.0006) in eSSc patients. No statistically significant correlation was found between PCS, MCS and HAQ-DI in UCTD patients. Age, sex, disease duration, history of arthritis, low levels of either C3 or C4, a low DLCO (carbon monoxide lung diffusion) and inversion of the E/A ratio were not correlated to PCS and MCS in either eSSc or UCTD patients. CONCLUSION: Many eSSc or UCTD patients perceive they have an impaired quality of life in both physical and mental domains. This condition has to be taken into account by the clinicians involved in the care of these patients.

We investigated early systemic sclerosis (SSc) (that is, Raynaud's phenomenon with SSc marker autoantibodies and/or typical capillaroscopic findings and no manifestations other than puffy fingers or arthritis) versus undifferentiated connective tissue disease (UCTD) to identify predictors of short-term disease evolution. Thirty-nine early SSc and 37 UCTD patients were investigated. At baseline, all patients underwent clinical evaluation, B-mode echocardiography, lung function tests and esophageal manometry to detect preclinical alterations of internal organs, and were re-assessed every year. Twenty-one early SSc and 24 UCTD patients, and 25 controls were also investigated for serum endothelial, T-cell and fibroblast activation markers. At baseline, 48.7% of early SSc and 37.8% of UCTD patients had at least one preclinical functional alteration (P > 0.05). Ninety-two percent of early SSc patients developed manifestations consistent with definite SSc (that is, skin sclerosis, digital ulcers/scars, two or more teleangectasias, clinically visible nailfold capillaries, cutaneous calcinosis, X-ray bibasilar lung fibrosis, X-ray esophageal dysmotility, ECG signs of myocardial fibrosis and laboratory signs of renal crisis) within five years versus 17.1% of UCTD patients (X 2 = 12.26; P = 0.0005). Avascular areas (HR = 4.39 95% CI 1.18 to 16.3; P = 0.02), increased levels of soluble IL-2 receptor alpha (HR = 4.39; 95% CI 1.03 to 18.6; P = 0.03), and of procollagen III aminopropeptide predicted disease evolution (HR = 4.55; 95% CI 1.18 to 17; P = 0.04). Most early SSc but only a few UCTD patients progress to definite SSc within a short-term follow-up. Measurement of circulating markers of T-cell and fibroblast activation might serve to identify early SSc patients who are more likely to develop features of definite SSc.

OBJECTIVE: To investigate survival in Italian systemic sclerosis (SSc) patients from a tertiary center, reporting death causes. MATERIALS AND METHODS: We analyzed the charts of 251 SSc patients prospectively enrolled in our Rheumatology Unit from 2000 to 2008. Baseline characteristics were recorded. In 2008 the vital status and the causes of death were assessed. Overall and subgroup survival were analyzed by the Kaplan-Meier method and the log-rank test. RESULTS: In 2008, 82% of patients were alive, 8% were known to have died and 10% were lost to follow-up. Overall 5- and 8-year survival were 94.8% and 77.1%, respectively. Patients with an age greater than the median value of the cohort (χ²=4.4; p=0.036), diffuse cutaneous SSc (χ²=3.9; p=0.048), digital ulcers (χ²=6; p=0.015), articular (χ²=5.3; p=0.021), lung (χ²=5.6; p=0.018) and heart involvement (χ²=9.3; p=0.002) had a poorer survival than patients without these features. The majority of SSc-related deaths (60%) were secondary to interstitial lung disease and heart involvement (both 33.3%); 50% of non-SSc-related deaths were due to cancer. CONCLUSIONS: Our study reports an improvement in survival of Italian SSc patients during the last decade with respect to the previous ones. Moreover, a reduction in deaths from renal involvement and an increase in deaths from interstitial lung disease were recorded in Italian SSc patients. Our data are consistent with those from recent survival studies carried out on SSc patients from other geographic areas

The European League Against Rheumatism Scleroderma Trials and Research Group (EUSTAR) has established an online database with clinical data of currently more than 8200 patients with systemic sclerosis (SSc). In addition to clinical research, EUSTAR fosters biomolecular studies to develop novel biomarkers and therapies for SSc. High-quality biospecimens are the basis for successful biomolecular studies. The EUSTAR biobanking group has therefore developed recommendations to standardise the collection, storage and distribution of SSc biospecimens at EUSTAR centres. These recommendations consider the scientific challenges associated with biomolecular research in SSc and the organisational requirements of EUSTAR. They were approved by the EUSTAR executive committee as well as the EUSTAR board. Once they become effective, these recommendations will be the basis for international EUSTAR studies with large numbers of SSc biospecimens. These recommendations might also be followed by other SSc consortia to enable exchange of biosamples between different SSc initiatives and might serve as a template for biobanking initiatives in other rheumatic diseases.

OBJECTIVE To assess internal organ involvement in early SSc at presentation. METHODS One hundred and fifteen patients admitted to a tertiary centre because of RP, who did not present any routinely detectable scleroderma-related internal organ involvement, were investigated for ANA and videocapillaroscopy, and underwent history and physical examination to detect symptoms/signs suggestive of SSc. Patients were then subdivided into three groups: (i) early SSc, constituted by patients without clinical manifestations other than RP, but with scleroderma marker autoantibodies and/or typical capillaroscopic abnormalities; (ii) probable SSc, constituted by patients with the same autoantibody and/or capillaroscopic status as early SSc patients, but with any of the following manifestations: digital ulcers/scars, puffy fingers, arthritis, telangiectasia, dysphagia/heartburn, shortness of breath; (iii) UCTD, constituted by patients with a specific (i.e. disease antibody marker) ANA and capillaroscopic findings plus any disease manifestation. All patients were investigated by lung functional study and B-mode echo-Doppler-cardiography. Patients who consented underwent oesophageal manometry. RESULTS An inverted mitral E : A ratio (i.e. early scleroderma cardiac involvement) and/or a diffusing lung capacity for CO

OBJECTIVE: To investigate the prevalence of clinical and subclinical atherosclerosis in systemic sclerosis (SSc) patients and its associated features. METHODS: Fifty unselected SSc patients and 41 controls, matched for sex and age, were investigated for previous cardiovascular events, cardiovascular risk factors, and ultrasonographic features of subclinical atherosclerosis in the carotid arteries, that is intima-media thickness (IMT) > 0.9 mm or plaques. SSc patients were also investigated for disease features and previous treatment. Finally, blood samples were randomly selected from 27 patients and 18 controls to evaluate concentrations of amino-terminal propeptide of type III procollagen (PIIINP), transforming growth factor (TGF)-β, hepatocyte growth factor (HGF), soluble interleukin-2 receptor (sIL-2R), IL-13, E-selectin, intercellular adhesion molecule (ICAM)-1, plasminogen activator inhibitor (PAI)-1, tissue plasminogen activator (t-PA), D-dimer, and prothrombin fragments (F1+2). RESULTS: Previous cardiovascular events were recorded in three SSc patients and no controls (p > 0.05). Mean IMT (0.613 ± 0.240 vs. 0.654 ± 0.173 mm) did not differ between patients and controls (p > 0.05), but subclinical atherosclerosis was detected in 14/50 SSc patients and 4/41 controls (p = 0.036). At multiple logistic regression analysis, mean IMT was correlated with older age [p = 0.006; odds ratio (OR) 1.276, 95% confidence interval (CI) 1.043-1.516] and a higher cumulative corticosteroid intake (p = 0.017; OR 1.155, 95% CI 1.027-1.300). No correlation was found with any soluble marker of disease activity and of coagulation/fibrinolysis system activation. CONCLUSION: Our study confirms an increased prevalence of subclinical atherosclerosis in SSc patients and demonstrates a hitherto unknown association with corticosteroid cumulative dosage.

OBJECTIVES Oligoclonal T-cell infiltrates have been detected in the skin of patients with early dcSSc. Peripheral T cells from patients with early dcSSc co-cultured with autologous fibroblasts have been found to expand the same T-cell clonotypes found in the affected skin. Here, we characterize oligoclonally expanded T lymphocytes and investigate functional changes occurring in early-dcSSc co-cultured T lymphocytes and fibroblasts. METHODS Peripheral T lymphocytes from five patients with early (

The occurrence of non-Hodgkin’s lymphoma in systemic sclerosis is an uncommon event. In our scleroderma cohort, a case of primary gastric B-cell lymphoma was diagnosed in 2007. The patient was a 45-year-old woman suffering from late systemic sclerosis sine scleroderma in whom non-Hodgkin’s lymphoma presented with progressive weight loss, later with gastrointestinal symptoms. Subsequently, we retrospectively analyzed the charts of 251 systemic sclerosis patients consecutively admitted to our Unit from 2000 to 2008 to search for other non-Hodgkin’s lymphoma cases (prevalence, 0.49%). Then we performed a Pubmed search for “systemic sclerosis & non-Hodgkin’s lymphoma,” limited to the English language. Twenty detailed cases of such an association were found, pointing out the following: non-Hodgkin’s lymphoma seems to be associated to old age, female sex, diffuse cutaneous subset and early disease; B-cell lymphoma subtypes are the majority; the interval between systemic sclerosis and lymphoma onset is usually short; systemic sclerosis could present as a paraneoplastic syndrome in some cases. We concluded that, although rare, the association of systemic sclerosis and non-Hodgkin’s lymphoma may not be coincidental and the clinician should be aware of the risk for lymphoproliferative disorders in scleroderma patients. This is the first description of non-Hodgkin’s lymphoma in systemic sclerosis sine scleroderma. The insidious onset of gastric lymphomas, mimicking the most common features of gastrointestinal involvement in systemic sclerosis is underlined.

BACKGROUND: Hypocomplementemia has been detected in about 15% of unselected series of SSc patients. It constitutes one of the 10 parameters needed to evaluate the European Scleroderma Study Group (EScSG) activity index. A few studies have been so far devoted to investigate the clinical manifestations correlated with this finding. OBJECTIVE: To investigate SSc patients for hypocomplementemia and point out clinical manifestations associated with it. METHODS: 302 patients with SSc consecutively admitted to the Rheumatology Unit of the Second University of Naples were enrolled in the study. SSc patients were all investigated for sex, age, disease duration, clinical and serological subset, disease activity, organ/system severity and functional status. Patients were divided into 2 groups: normo-complementemic and hypocomplementemic (low C3 and/or C4) as measured by nephelometry. RESULTS: 252 of the 302 patients had normal complementemia; 50 (16,5%) had hypocomplementemia. Significant associations were found between hypocomplementemia and EScSG activity index (p0.5)( p=0.04); and the severity of general manifestations (p

Hudson et al. argue that the conclusions of our study on hypocomplementemia in systemic sclerosis cannot be extended to other series. First of all, we would like to underline that our study is the 3rd one at least pointing out a definite prevalence of hypocomplementemia in Systemic Sclerosis (Della Rossa et al. (1); Hudson et al. (2); Cuomo et al. (3). Therefore, hypocomplementemia can be considered a defined feature occurring in about 15-20% of SSc patients. Secondly, differences in the autoantibody profile between Italian...

Objective:To investigate the possible implication of SDF1-3′ polymorphism in systemic sclerosis (SSc) susceptibility or clinical phenotype, or both.Methods:150 patients with SSc and 150 controls were enrolled. Skin involvement, autoantibodies, interstitial lung disease, pulmonary arterial hypertension (PAH), scleroderma renal crisis, past and/or current skin ulcers were assessed. Genotyping was performed by PCR-RFLP.Results:Genotype distribution and allele frequency were similar in SSc and controls. SDF1-3′A allele and SDF1-3′GA/AA genotype frequencies were significantly higher in SSc-PAH than in SSc-non-PAH (33.3% vs 18.3%, p = 0.01) and in SSc with skin ulcers than in SSc without ulcers (27.3% vs 16.9%, p = 0.03). The SDF1-3′A allele influenced the predisposition to SSc-related PAH (OR = 2.52, 95% CI 1.11 to 5.69, p = 0.02) and skin ulcers (OR = 2.31, 95% CI 1.18 to 4.52, p = 0.01). After adjustment for age and gender, the SDF1-3′A allele remained a susceptibility factor for the SSc-related vascular manifestations (PAH: OR = 2.37, 95% CI 1.04 to 5.42, p = 0.04; ulcers: OR = 2.33, 95% CI 1.78 to 4.62, p = 0.01).Conclusion:The SDF1-3′A allele is significantly associated with microvascular involvement in SSc.

OBJECTIVE: To investigate the relationship of the polymorphic enhancer HS1,2 central to the 3' enhancer complex regulatory region (IgH3'EC) of the immunoglobulin heavy chain genes with systemic sclerosis (SSc) disease and compare it with HLA-DR and DQ associations. METHODS: A total of 116 patients with SSc were classified as diffuse (dSSc) or limited (lSSc), and as carriers of antitopoisomerase I (anti-Scl70) or anticentromere (ACA) antibodies. Allele and genotype frequencies were assessed in the population as a whole and in the two major subsets, dSSc and lSSc. The concentration of peripheral blood immunoglobulin levels was also determined and analysed according to the genotypes. RESULTS: The analysis of genotypes for the four alleles of the HS1,2A enhancer showed an increased frequency of allele *2 in the SSc cohort highly significant versus controls (57% vs. 40%, p

Background Systemic sclerosis (SSc) is classically dichotomized in limited or diffuse cutaneous subsets associated with different prognosis. However, it appears that SSc is a highly heterogeneous disease, with probably more subtle clinical phenotypes. Objectives The primary objective of our study was to identify and characterize homogeneous phenotypes in the EUSTAR SSc population using a cluster analysis. The secondary objective is to assess the survival in the different clusters. Methods The participants were SSc patients of the EUSTAR prospective cohort. Inclusion criteria were: age ≥18 years, fulfilled American College of Rheumatology criteria for SSc, and had a calculable follow up. Hierarchical clustering based on the Ward method was performed using 25 clinically meaningful variables. The clinical relevance of the clusters was analysed by their characteristics and survival outcomes. Survival curves were plotted by the Kaplan-Meier method. Crud and adjusted (on the age at diagnosis and gender) Cox models were performed. Results A total of 6.927 patients were analyzed. Cluster analysis identified 5 clusters (Table 1). The first cluster C1 (n=1,963) was mainly characterized by a low Rodnan skin score contrasting with a relatively similar percentage of anti topoisomerase 1 and anti-centromere Ab (28% and 40%) and a high percentage of lung fibrosis (48%); the second cluster C2 (n=1,186) by a large majority of limited SSc (89%) and anti-centromere (79%) and the lowest percentage of lung fibrosis (22%); the third cluster C3 (n=1,249) and the fourth cluster C4 (n=856) by a high percentage of diffuse SSc/anti topoisomerase 1 (72%/50% and 92%/77% respectively) and the highest proportion of male patients. Digital ulcers, renal crisis/anti RNA polymerase III antibodies, joint involvement were more frequently found in C4 than in the other clusters. Patients in the cluster C5 (n=1,673) were mainly female patients, older than in the other clusters, had a rather low mean Rodnan skin score, a high percentage of anti-topoisomerase 1 antibodies (46%) and a high percentage of lung fibrosis. Concerning survival, C2 had the best survival and C4 the worst. When compared to C1, C2 to C5 clusters had a significantly different prognosis (C2: adjusted hazard ratio [95% CI]: 0.70 [0.51-0.96]; C3: 2.27 [1.76-2.94]; C4: 4.52 [3.57-5.72]; and C5: 1.94 [1.51-2.44]. Conclusions This cluster analysis on the largest ever worldwide database of SSc patients showed 5 different clusters characterized by different sex ratio, maximum Rodnan skin scores, autoantibodies status, joint and organ involvement as well as by a different prognosis. Autoantibodies status seemed to play an important role for association with organ involvement. Although systemic sclerosis is a heterogeneous disease, this study shows that homogeneous groups beyond the classical limited/diffuse dichotomy can be delineated in this disease. Disclosure of Interest None declared

Background Long noncoding RNAs (LncRNAs) are a novel class of noncoding transcripts with diverse regulatory functions, e.g. imprinting regulation, dosage compensation, cell cycle regulation, pluripotency and retrotransposon silencing. Dysregulation of lncRNAs is increasingly recognized to contribute to the disease pathogenesis, such as cancer, autoimmune disorders and neurodegeneration. Objectives To identify candidate lncRNAs in systemic sclerosis (SSc) and investigate their function, in particular in relation to the TGFβ pathway and myofibroblast phenotype development. Methods RNA Sequencing Ilumina HiSeq2000 was performed in healthy and SSc skin biopsies. Human skin fibroblasts were isolated from biopsies of SSc patients and healthy controls (HC), human pulmonary smooth muscle cells were purchased from Lonza. The cells were treated with 10 ng/ml TGFβ. TGFβR1 inhibitors (SD208 and SB431542) and siRNA against SMAD3 were used to investigate TGFβ driven gene expression. The lncRNA MIR503HG was silenced in skin fibroblasts using locked nucleic acid antisense oligonucleotides (LNA GapmeRs), followed by qPCR analyses and immunofluorescence staining. The expression of MIR503HG was measured in tissue samples of liver and lung fibrosis. Results RNA sequencing showed a significant upregulation of the lncRNA MIR503HG (H19X) in SSc versus HC skin biopsies. Importantly, the upregulation of MIR503HG was not limited to SSc skin, but present also in the tissues from liver and lung fibrosis, indicating a broader role of MIR503HG in fibrotic diseases. While there was no difference in the basal expression of MIR503HG between SSc and HC cultured dermal fibroblasts, MIR503HG was strongly and consistently induced by TGFβ. Induction of MIR503HG by TGFβ was not limited to skin fibroblasts, but evident also in other cell types relevant for SSc, e.g. pulmonary vascular smooth muscle cells. Time curve analysis revealed that the upregulation of MIR503HG by TGFβ was strongest after 6h reaching 12.8±0.7 induction; and dose curve analysis showed a steady increase of MIR503HG over physiologically relevant TGFβ concentrations. These effects were TGFβR1 dependent as shown by the inhibition experiments with the chemical inhibitors SD208 and SB431542. Moreover, the upregulation of MIR503HG by TGFβ was significantly impaired by silencing of SMAD3, further pointing to an important role of the canonical TGFβ pathway in MIR503HG expression. The knockdown of MIR503HG in skin fibroblasts led to a strong down regulation of COL1A1, fibronectin and αSMA mRNAs, indicating a potential involvement of MIR503HG in the development of a myofibroblast phenotype. Additionally, immunofluorescence staining showed reduced αSMA formation in MIR503HG silenced skin fibroblasts. Conclusions This is the first study reporting changes in long-non coding RNAs in SSc and across fibrotic organs. It opens new perspectives in the pathogenesis of fibrotic diseases by this novel class of regulatory, non-coding RNAs. Disclosure of Interest E. Pachera: None declared, S. Assassi: None declared, G. Salazar Cintora: None declared, M. Frank-Bertoncelj: None declared, V. Haunerdinger: None declared, R. Dobrota Grant/research support from: Pfizer, Actelion, M. Brock: None declared, S. Vettori Grant/research support from: Pfizer, Roche, Consultant for: Phadia-Thermofischer, Speakers bureau: Pfizer, Abbvie, C. Hellerbrand: None declared, C. Feghali-Bostwick: None declared, J. Distler: None declared, G. Kania: None declared, O. Distler Grant/research support from: 4D Science, Actelion, Active Biotec, Bayer-Schering, Biogen, Biovitrium, BMS, Boehringer Ingelheim Pharma, EpiPharm, Ergonex, GSK, Inventiva, Medac, Novartis, Pfizer, Pharmacyclics, Roche/Genentech, Sanofi/Genzyme, Serodapharm, Sinoxa and United BioSource Corporation, Consultant for: 4D Science, Actelion, Active Biotec, Bayer-Schering, Biogen, Biovitrium, BMS, Boehringer Ingelheim Pharma, EpiPharm, Ergonex, GSK, Inventiva, Medac, Novartis, Pfizer, Pharmacyclics, Roche/Genentech, Sanofi/Genzyme, Serodapharm, Sinoxa and United BioSource Corporation

Background Due to lack of adequate clinical research data, drug treatment of the rare disease systemic sclerosis (SSc) is commonly off-label. The international EC-funded research project DeSScipher (acronym for “to decipher the optimal management of systemic sclerosis”) was designed to increase the evidence-based treatment strategies for SSc patients and subsequently to improve their long-term quality-of-life. Objectives The primary objective is to compare the outcomes of different treatments with respect to efficacy and safety of currently used off-label drugs from the early to the advanced phases of the main SSc-associated organ dysfunctions in a routine rheumatology in- and outpatient setting. Methods Five prospective observational trials (OTs), carried out within the EULAR Scleroderma Trials and Research (EUSTAR) group, have been designed to analyze current treatment approaches of early functionally relevant manifestations such as digital ulcers (OT1) and hand arthritis (OT2) to the morbidity and mortality-driving manifestations such as interstitial lung disease (OT3), pulmonary hypertension (OT4) and severe heart disease (OT5). The study protocols are accessible at clinicaltrials.gov Identifiers NCT01836263, NCT01834157, NCT01858259, NCT01840748, NCT01829126. Results Between April 2013 and January 2015, 1781 SSc patients have been screened at 27 contributing EUSTAR centers. 1577 (89%) patients have been enrolled into at least one of the five OTs. In particular, 1179, 127, 981, 237 and 716 patients have been enrolled into OT1-5, respectively (3240 in total; a given patient could be enrolled into multiple OTs), which represents a baseline patient recruitment rate of 79% of the target number of 4098 patients (accordingly 226%, 79%, 59%, 25% and 91% of the required number of 522, 160, 1670, 960, 786 patients for OT1-5, respectively). The completion of 1-year (OT3, OT5) and 2-year follow-up visits (OT1, OT2, OT4) are pending. Conclusions DeSScipher is currently the largest prospective observational research project ever for SSc. While patient recruitment is still ongoing, preliminary results of the five OTs are expected in late 2015 and the final results depending on the completion of follow-up visits are expected between 2017 and 2018. Acknowledgements The DeSScipher project was funded by the European Community9s Framework Programme 7 (FP7-HEALTH-2012.2.4.4-2 - Observational trials in rare diseases) under grant agreement N° 305495. We acknowledge the contribution of the following EUSTAR centers: Wuppertal (member N°192), Lille (93), Bad Bramstedt (187), Moscow (78), Assiut (168), Moscow (190), Bucharest (100), Monserrato (142), Iasi (162), Cluj-Napoca (16), Frankfurt (124), Salford/Manchester (80), Tubingen (56), Ancona (34), Zagreb (51) and Roma (94). Disclosure of Interest None declared

Background Heart disease in patients with systemic sclerosis (SSc) can cause manifestations such as cardiac blocks (CBs), ventricular arrhythmias (VA), and congestive heart failure (CHF) which may require a pacemaker/defibrillator implant, are the cause of death or sudden death (SD) in about 20% of patients and can be referred to as severe heart disease (SHD). The European League against Rheumatism – Scleroderma Trial and Research (EUSTAR) database provides an unique opportunity to address this topic. The results of such analysis are instrumental for the DeSScipher project, an EU funded project devoted to decipher the optimal management of SSc . Objectives To investigate the prevalence of each SHD manifestation and their associations with demographic, serological, clinical and therapeutic aspects in a large series of adult SSc patients. Methods Seven EUSTAR-DeSScipher centers provided clinical charts at study entry including all the items of SHD. The prevalence of any and each SHD manifestation was calculated. The associations with demographic, serological, clinical features and treatment with vasodilators, i.e. calcium channel blockers (CCBs) and /or ACE inhibitors (ACEi), were investigated by univariate analysis and confirmed by multivariate logistic regression analysis. Results At July 6th 2012, 1119 SSc patients from the 7 EUSTAR centers had no missing data (995 females, 164 males; median age 53.6 years, range 14-86). Out of them, 211 patients (19%) had at least 1 SHD manifestation, 152 patients had CBs (14%), 71 had VA (6%), 132 had CHF (12%), and 27 received a pacemaker/defibrillator implant (2%). CBs were the only SHD manifestation in 100 patients, VA in 31, and CHF in 80. No association was found between SHD and clinical (diffuse or limited) and serological (ACA or anti-Scl-70) subset. No association was found between either CHF or pacemaker/defibrillator implant and any other disease feature. In multiple logistic regression analysis, CBs were associated with bibasilar lung fibrosis at chest X-Ray (OR=2.6; 95%CI=1-6.4; p=0.04), while VA were associated with increased CPK levels (OR=11; 95%CI=1-117; p=0.02) and, unexpectedly, current CCB use (OR 10; 95%CI 1.4-76.1; p=0.02). Conclusions This is the first study devoted to investigate SHD in a large series of carefully assessed SSc patients. SHD was detected in about 19% of the cases, CBs in 14%, VA in 6%, CHF in 12%. Pacemaker/defibrillator implant was needed in 2.4%. These data highlight the importance of an accurate heart assessment in SSc patients. Moreover, the associations between VA and CPK elevation and current CCB use should stimulate the clinician to avoid these drugs in patients with myositis/myocarditis. Acknowledgements The DeSScipher project was funded by the European Community9s Framework Programme 7 (FP7-HEALTH-2012.2.4.4-2 - Observational trials in rare diseases) under grant agreement N° 305495 Disclosure of Interest S. Vettori Grant/research support from: Roche, Consultant for: Phadia Thermofisher, Pfizer, Abbvie, G. Cuomo: None declared, V. Jaeger: None declared, M. Frerix: None declared, E. Siegert: None declared, V. Lorand: None declared, S. Jordan: None declared, G. Riemekasten: None declared, Y. Allanore: None declared, L. Czirjak: None declared, I. Tarner: None declared, O. Distler Grant/research support from: 4D Science, Actelion, Active Biotec, Bayer-Schering, Biogen, Biovitrium, BMS, Boehringer Ingelheim Pharma, EpiPharm, Ergonex, GSK, Inventiva, Medac, Novartis, Pfizer, Pharmacyclics, Roche/Genentech, Sanofi/Genzyme, Serodapharm, Sinoxa and United BioSource Corporation, Consultant for: 4D Science, Actelion, Active Biotec, Bayer-Schering, Biogen, Biovitrium, BMS, Boehringer Ingelheim Pharma, EpiPharm, Ergonex, GSK, Inventiva, Medac, Novartis, Pfizer, Pharmacyclics, Roche/Genentech, Sanofi/Genzyme, Serodapharm, Sinoxa and United BioSource Corporation, C. Denton: None declared, M. Matucci-Cerinic: None declared, F. Del Galdo: None declared, U. Walker: None declared, U. Mueller-Ladner: None declared, G. Valentini: None declared

Background In Systemic sclerosis, anti-CENP Ab positive patients (pts) are at higher risk of developing pulmonary hypertension compared to pts expressing other ANA specificities. In a previous investigation (1), IgG against the immunodominant epitope of CENP-A spanning amino acid 1-17 (peptide Ap1-17), were purified from sera of patient (pt) 4 and pt14 and used to isolate specific phage clones expressing peptide (pc). Pcs specific for Pt4 IgG (pc4s) and pt14 IgG (pc14s) displayed differential reactivity with anti-Ap1-17 IgG from other SSc pts (1,2), indicating they can distinguish different subsets within anti-CENP-A Ab population. Objectives To assess the reactivity of sera from 84 anti-CENP-A Ab positive pts with pc4.1, pc4.2, pc14.1 and pc14.2, and to search for clinical correlates. Methods Clinical and serological data were collected as previously described (2). Systolic pulmonary arterial pressure (sPAP) was measured using Doppler echocardiography. The reactivity and specificity of serum with pc was assessed by indirect ELISA using mouse anti-M13 mAb to capture pc to the plates and xeno-Ab to the Fc portion of human IgG to detect pc-bound sera IgG. Multivariate forward stepwise linear (or logistic) regression analysis was used to define a predictive association between a clinical variable and the levels of anti-pcs Ab. Results Following a linear regression analysis, anti-pc4.2 and pc14.1 Ab were the only serological variables retained in the model able to predict sPAP and DLCO, albeit in a opposite way: an increased of anti-pc4.2 predicted a sPAP increase (p 45 mmHg) as outcome variable and anti-pc Ab as predictors, along with possible confounding factors namely gender, disease duration and age at diagnosis. Again, anti-pc4.2 and pc14.1 were retained in the model: positivity for sPAP was associated with high levels of anti-pc4.2 Ab (p=0.038), while negativity for sPAP was associate with high levels of anti-pc14.1 Ab (p=0.002). Conclusions Overall the data indicate that patients expressing high levels of Ab to the sub-specificities of CENP-A defined by pc4.1 are at higher risk of developing pulmonary hypertension, whereas those expressing high levels of Ab to the specificity defined by pc14.1 are more protected from this complication. References Favoino, E. et al., 2013. Autoantibodies recognizing the amino terminal 1-17 segment of CENP-A display unique specificities in systemic sclerosis. PLoS. One. 8: e61453. Perosa, F., et al., 2013. Clinical correlates of a subset of anti-CENP-A antibodies cross-reacting with FOXE3p53-62 in systemic sclerosis. Arthritis Res. Ther. 15: R72. Acknowledgements This work was supported by a grant from the “Italian Group against Systemic Sclerosis” (GILS) Disclosure of Interest None declared

Early systemic sclerosis (SSc) is a newly identified condition characterized by the association of Raynaud9s phenomenon (RP) with SSc marker autoantibodies and/or with a scleroderma pattern at nailfold videocapillaroscopy (NVC), without clinical manifestations of definite SSc. Patients with early SSc do not meet the old 1980 American College of Rheumatology (ACR) nor the new 2013ACR/European League Against Rheumatism (EULAR) classification criteria for the disease. Nevertheless, they show a scleroderma-type preclinical involvement of internal organs at second level examinations in up to 42% of cases and increased serum levels of markers of endothelial, T-cell, and fibroblast activation. A number of these circulating markers, like sICAM-1, CCL2, CXCL8, and IL-13 increase their serum levels in parallel with the onset and the extent of skin sclerosis. They have been found to be elevated in patients with early SSc, but reach the highest levels in definite SSc patients with a diffuse skin disease. Other circulating markers display a peculiar behavior, being associated with distinct early SSc subsets. In fact, the carboxyterminal telopeptide of type I collagen (ICTP) has been found to be higher in early SSc patients with marker autoantibodies and impaired DLCO, while sE-selectin has been found to be higher in early SSc patients with an NVC scleroderma pattern and puffy fingers. More interestingly, IL-33, an inflammatory cytokine belonging to the IL-1β superfamily enriched in epithelial and endothelial cells, has been found to be higher in patient with early SSc than in patients with a definite, classifiable disease. Taken together, these data suggest that the cellular pathways that are implicated in the complex vascular/immune pathogenesis of SSc are active in patients with an early pre-fibrotic stage of the disease, when an increased collagen synthesis can also be suspected. In addition, they suggest that serum levels of some adhesion molecules, chemokines and cytokines may be helpful to follow the evolution of early SSc into definite SSc, while IL-33 may be investigated in large patient series as a potential biomarker of the early subset. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.6230

Background We had reported that miRNA-193b (miR-193b) is significantly downregulated in SSc and that down regulation of miR-193b causes overproduction of urokinase type plasminogen activator (uPA). However, the effects of increased levels of uPA may be limited in SSc because of inactivation of the cellular receptor for uPA (uPAR) after cleavage by matrix metalloproteinase-12. Objective To investigate whether the effects of uPA in SSc are mediated by uPAR independent pathways. Methods: Expression of miR-193b in SSc skin fibroblasts and skin sections from SSc were analysed by Real-time PCR. Transfection with miR-193b precursor and inhibitor were used to identify targets of miR-193b. Basal expression and localisation of uPA were examined by Real-time PCR, Western blot and immunohistochemistry. Furthermore, human pulmonary artery smooth muscle cells (HPASMCs) were treated with uPA and the proliferative effects of uPA were determined by WST-1 assay and by analysis of proliferating cell nuclear antigen expression. FACS was performed to investigate the effect of uPA on apoptosis. For inhibition of the uPA-uPAR pathway, uPAR neutralising antibodies, siRNA against uPAR and low molecular weight uPA, which does not bind to uPAR, were used. Results MiR-193b was significantly down regulated in SSc fibroblasts (n = 12) as compared with healthy controls (HC) (n = 6) (31 ± 33%, p Conclusion Our results suggest that the down regulation of miR-193b in SSc induces expression of uPA, which leads to increased numbers of vascular smooth muscle cells via uPAR independent pathways and thereby contributes to the proliferative vasculopathy characteristic of SSc. This is the first link between miRNA dysregulation and vasculopathy in SSc.

Background Cardiac blocks (CBs) (atrioventricular blocks, left bundle branch block, left anterior fascicle block ± right bundle block) are known to reflect myocardial fibrosis in patients with Systemic Sclerosis (SSc). Their prevalence has been already assessed in EUSTAR cohort on 3656 patients (1). Nevertheless, only a few associated features were investigated. Objectives To assess the prevalence of CBs in a larger series and investigate associated clinical and therapeutic features. Methods Data from 10183 SSc patients (87% females, 13% males; 31% diffuse disease, 69% limited disease); aged from10-91 years-median age 56; with a disease duration ranging from (0.83-76 years - median 9 years) entered in the EUSTAR MEDS-online system, the largest online SSc database in the world, from 2003 to July 2012 were analyzed retrospectively. CBs as well as other epidemiological, clinical and therapeutic aspects were deduced from Minimal Essential Data Set charts. Results CBs were detected at enrollment in 968 patients (10%). In univariate analysis CBs were found to be associated with epidemiological [ male sex ( OR 1.6, 1.4-1.9); age Conclusions We confirm the already reported association of CCBs with anti-Scl-70 positivity. The detected association with CCB and ACE is likely to reflect confounding for indication. References Walker UA, Tyndall A, Czirjak L. Ann Rheum Dis 2007; 66:754-63 Disclosure of Interest None Declared