Christina Katsiari

Breg Cells Are Numerically Decreased and Functionally Impaired in Patients With Systemic Sclerosis.

Breg cells, a regulatory cell subset that produces interleukin‐10 (IL‐10), play a significant role in suppressing autoimmune responses and preventing autoimmunity. This study was undertaken to examine the number and function of Breg cells in patients with systemic sclerosis (SSc), a disease with many autoantibodies.

Early systemic sclerosis—opportunities for treatment

Systemic sclerosis (SSc) is characterized by microvasculopathy (Raynaud’s phenomenon and fibrointimal proliferation), presence of autoantibodies and collagen deposition in skin (scleroderma) and internal organs. Microvasculopathy, detected by nailfold capillaroscopy, and disease-specific autoantibodies (anti-topoisomerase I, anti-centromere, anti-RNA polymerase III antibodies) usually appear earlier, even years before scleroderma. At that stage of the disease, immune activation with T cells and B cells promote fibrosis. Diagnosis of SSc has been relied on scleroderma, and by this time, internal organs may have developed fibrosis, a lethal feature with no available treatment. The new EULAR/ACR 2013 criteria for the classification of SSc will help identify SSc patients before fibrosis of internal organs. The early diagnosis of SSc, before the development of fibrosis in internal organs, will allow the introduction of immunosuppressive medications in these patients in a controlled setting (randomized trials). It is anticipated that this approach will change the hitherto grim prognosis of SSc for the better.

Tuberculous pyomyositis: a re-emerging entity of many faces.

Tuberculosis (TB) has become a global concern due to its increasing incidence, particularly in immunocompromised patients, closely following the migratory patterns of populations. TB pyomyositis is a rare extrapulmonary manifestation of TB. Its clinical presentation varies and requires a high degree of suspicion for early diagnosis. We present three patients diagnosed with TB pyomyositis: a 46-year-old man with dermatomyositis (DM) and hepatitis B who presented with fever, muscle weakness, and an abscess at the right proximal arm; a 71-year-old immunocompetent male, with a past medical history of tuberculous lymphadenopathy in childhood, who presented with a 2-month history of fever and pain at the right thigh, and a 44-year-old woman with systemic lupus erythematosus (SLE) on prednisone and methotrexate who presented with skin eruption at her thighs mimicking lupus panniculitis. In all three patients, Mycobacterium tuberculosis was identified as the causative agent. The lack of specific signs, the false negative tuberculin skin test in some cases, and the unfamiliarity of many clinicians with this entity can cause diagnostic delays. Prompt diagnosis requires a high index of suspicion especially in immunocompromised patients with fever.

Curcumin for the Management of Periodontitis and Early ACPA-Positive Rheumatoid Arthritis: Killing Two Birds with One Stone

We propose curcumin as a preventive measure to avoid/manage periodontitis (PD), and as a natural immunosuppressant for rheumatoid arthritis (RA). PD, mainly caused by Porphyromonas gingivalis forming biofilm and leading to tooth decay, is a major public health issue and a risk factor for the development of RA in humans. P. gingivalis is able to trigger experimental autoimmune arthritis in animal models and in humans can induce citrullinated peptides, which not only are a source of anti-citrullinated antibodies (ACPAs), but also participate in autoreactive responses and disease development. Curcumin appears to have efficient anti-bacterial activity against P. gingivalis infection and biofilm formation. In addition to antibacterial, anti-oxidant, and anti-inflammatory action, curcumin exerts unique immunosuppressant properties via the inhibition of Th17 pro-inflammatory responses and promotion of regulatory T cells, thus suppressing autoimmunity. We introduce curcumin as a natural product for the management of both PD and RA-related autoreactivity, possibly also as a preventive measure in early RA or individuals at high risk to develop RA.

Multiparametric autoantibody profiling of patients with systemic sclerosis in Greece

Background: Systemic sclerosis (SSc) is an autoimmune rheumatic disease characterized by a wide range of disease-specific and disease-related autoantibodies (autoAbs). Profile assays have been developed and are currently in use to meet the demand for better characterization of all autoAbs found in SSc patients. Aim: To assess the clinical relevance of SSc-related autoantibodies in 158 patients with SSc, all from Central Greece, taking advantage of a multiparametric SSc autoantibody line immunoassay. Material and methods: 158 consecutive patients with SSc (137 females, mean age 53.2 ± 10 years; 63 patients with dcSSc and 95 with lcSSc) from central Greece were included in the study. Eighteen patients with morphea were also included. Serum samples were analyzed by a profile SSc nucleoli line assay (Euroimmun) to detect Abs against 13 autoantigens: Scl-70, Centromere (A, B), RNA polymerase III (subunits 11 & 155), fibrillarin, NOR90, Th/To, PM/Scl 100, PM/Scl75, Ku, PDGFR and Ro52. Antinuclear autoAbs (ANAs) were detected by indirect immunofluorescence. Results: ANAs were detected in 97.5% of SSc patients. Reactivities to specific autoantigens were as follows: Topo I, 40.5%; CENP, 32.9%; Ro52, 21.5%; RP11, 8.9%; RP155, 13.3%; NOR 90, 4.4%; Ku 3.8%; PM-Scl75, 3.2%; PM-Scl100, 1.3%; Th/To, 1.3%; Fibrillarin, 1.3%; PDGFR 0%; Ro52 21.5%. Twenty-one of SSc did not have any of the main autoAbs, namely anti-Topo I, anti-CENP, anti-RNA pol III Abs. Conclusions: Multiparametric autoAb test provides positive SSc-associated autoAb reactivities in SSc patients negative for the three main autoAbs and this may prove of significance in early disease diagnosis.

AB0163 No Association between The Percentage of Memory or Transitional B Regs and Disease-Related Autoantibodies in Systemic Sclerosis

Background Recently, we found that Breg cells are numerically decreased and functionally impaired in patients with systemic sclerosis (SSc)1 A subsequent study confirmed these data and reported a negative correlation between Breg levels and the titres of anti-topo I and anti-centromere antibodies (ab) in patients with SSc2. Objectives To assess the relation between memory or transitional Bregs with SSc-specific anti-topo I, anti-centromere (CEN) and anti-RNA pol I abs in our cohort of SSc patients. Methods Fifty patients with Ssc were tested for disease-specific abs (Topo I, CENP, RNA pol I) using line blot analysis (EUROIMMUN), and for memory [CD19(pos)CD27(pos)CD24(high)] and transitional [CD19(pos)CD24(high)CD38(high)] Bregs using flow cytometry. Results Anti-CEN, anti-Scl-70 and RNA pol I abs were present in 12 (24%), 23 (46%) and 7 (14%) patients with SSc, respectively. The % of memory and transitional B regs was comparable between anti-CEN ab-positive and -negative patients (1.56±1.40 vs 1.74±1.01 and 0.78 ±0.73 vs 0.65±0.68 respectively, p>0.05), and also beween anti-RNA ab-positive and -negative patients with SSc (1.96±1.26 vs 1.65±1.09 and 0.54±0.53 vs 0.7±0.72 respectively). While the % of memory Bregs did not differ between anti-topo I-positive and -negative patients (1.96±1.26 vs 1.65±1.09 and 0.54 ±0.53 vs 0.7±0.72 respectively), the % of transitional Bregs tended to be decreased in patients with anti-topo I-positive compared to anti-topo I ab-negative patients (0.49±0.46 vs 0.84±0.61 p=0.066). Levels of B regs (memory or transitional) did not negatively correlate with autoantibody titres by line blot analysis. Conclusions We failed to find an association between percentages of memory and transitional B regs with SSc-specific autoantibodies. References Mavropoulos A et al Arthritis Rheumatol 2016;68:494–504 Matsushita T et al Rheumatology 2016;55:263–7 Acknowledgement Financial support by ELKE, University of Thessaly Disclosure of Interest None declared

AB0614 A Comphehensive Analysis of 12 Disease-Specific Autoantibodies in Greek Patients with Systemic Sclerosis

Background Patients with systemic sclerosis (SSc) are characterized by ANAs against various autoantigens, none of which is 100% sensitive for the disease. The need for profiling testing based on a combination of the most important SSc-related autoantigens has led to the development of commercial SSc-profile assays. Objectives To determine the diagnostic and clinical relevance of systemic sclerosis (SSc)-related autoantibodies tested by a line SSc-profile assay in a well defined cohort of Caucasian SSc patients, all from Central Greece. Methods 98 consecutive patients with SSc (80 female, 18 male, mean age 57,9±14,9 y) including 60 patients with diffuse cutaneous SSc (dcSSc) and 38 with limited cutaneous SSc (lcSSc) were analyzed by a profile SSc line assay (Euroimmun) for autoantibodies against 12 autoantigens: Scl-70, Centromere (A, B), RNA polymerase III (subunits 11 & 155), fibrillarin, NOR90, Th/To, PM/Scl 100, PM/Scl75, Ku and PDGFR. Results ANA by indirect immunofluorescence was present in 98 (98%) patients with SSc. Overall, 81 (82.6%) patients had abs against at least one of the 12 autoantigens. Anti-Scl-70 abs were present in 49 (49.4%) SSc patients, anti-centromere A in 22 (22.5%), anti-centromere B in 22 (22.5%, all anti-centromere A positive) anti-RNA polymerase III RP155) in 18 (18.3%) anti-RNA polymerase III RP11 in 14 (14.3%), anti-fibrillarin in 3 (3.1%), anti-Ku in 5 (5.1%), anti-NOR90 in 7 (7.1%), anti-PM/Scl100 in 4 (4.1%), anti-PM/Scl75 in 7 (7.1%), anti-Th/To in 4 (4.1%) and anti-PGDFR in 1 (1%) SSc patients. Overall, the most frequent anti-Scl-70, anti-centromere or anti-RNA polymerase III abs were present in 81 (81%) SSc patients while 6 (6.1%) lacked those antibodies and were positive for at least one of the other 9. Anti-Scl-70 were associated with diffuse SSc and ACA with limited SSc (p<0.001 for both). Positivity of Scl-70 was also correlated with the presence of lung fibrosis (p=0.004) and tended to be associated with the presence of digital ulcers (p=0.073). Anti-centromere abs were less prevalent in SSc patients with lung fibrosis (p=0.004). Anti-PM/Scl75 abs tended to be associated with the presence of digital ulcers (p=0.055). Anti-NOR90 abs were more frequent in male SSc patients (p=0.019). Conclusions Anti-Scl70, anti-centromere and anti-RNA polymerase are by far the most dominant autoantibodies in SSc but other disease-related autoantibodies can also be present in SSc patients lacking those three autoantibodies. The presence of anti-Scl-70 abs identifies patients with lung fibrosis and along with anti-PM/Scl75 tend to be associated with the presence of digital ulcers. Disclosure of Interest C. Liaskos: None declared, E. Marou: None declared, T. Simopoulou: None declared, M. Barmakoudi: None declared, T. Scheper Employee of: Euroimmun AG, W. Meyer Employee of: Euroimmun AG, C. Katsiari: None declared, D. Bogdanos: None declared, L. Sakkas: None declared

AB0613 Anti-C1q Autoantibodies Are Frequently Detected in Patients with Systemic Sclerosis and Lung Fibrosis

Background Anti-C1q autoantibodies are associated with systemic lupus erythematosus (SLE) but their presence in other rheumatic diseases has not been investigated in great detail. Objectives We aim to assess the clinical significance of anti-C1q autoantibodies in systemic sclerosis (SSc). Methods Seventy five patients with SSc (median age 59 years; females, 61; diffuse cutaneous SSc 35, limited cutaneous SSc, 40) were studied. Thirty eight patients with SLE, 25 patients with Sjögrens syndrome (SjS), 19 with rheumatoid arthritis (RA), and 31 healthy individuals (NC) were also included. IgG anti-C1q antibodies were tested by ELISA. Anti-nuclear antibodies (ANA), and SSc-specific autoantibodies were also tested. Results A receiver operator characteristic curve (ROC) confirmed that the cut off at 20 RU/mL for the anti-C1q test (specificity, 0.94, 95% CI, 0.79 to 0.99, LR: 5.0) was appropriate. Anti-C1q antibodies were present in 32.0% of SSc patients compared to 16.0% SS, 10.5% RA patients (p<0.05, for all) and in 6.5% NC (p<0.01). The frequent presence of anti-C1q antibodies in SLE was confirmed (13/38, 34.2%). Amongst the 24 anti-C1q (+) SSc patients, 15 had high/moderate levels (>40 RU/mL). There was no association between anti-C1q antibodies and anti-topoisomerase I, anti-centromere, or anti-RNA polymerase III antibodies. Anti-C1q antibodies were associated with male sex (p=0.04) and lung fibrosis (LF) (p=0.008), and co-occurrence of LF and skin ulcers (p=0.006). Ulcers were more prevalent in anti-C1q(+) than anti-C1q(−) patients (64% vs 44%), but the difference did not reach statistical significance (p=0.1). Conclusions Anti-C1q antibodies were frequently detected in SSc, were associated with PF and co-occurrence of PF and skin ulcers and may contribute to the autoimmune process in SSc. Acknowledgement Fianancial support by ELKE, University of Thessaly Disclosure of Interest None declared

FRI0011 IL-10-Producing Regulatory B Cells are Decreased in Systemic Sclerosis

Background Systemic sclerosis (SSc) is a characterized by extensive fibrosis and a plethora of autoantibodies, the latter being indicative of breakage of tolerance. Regulatory B cells (Bregs) producing interleukin (IL)-10 play a significant role in suppressing inflammatory immune responses and preventing autoimmunity Objectives To investigate the significance of IL-10-producing Bregs in SSc Methods The study groups consisted of 45 patients with SSc (12 with early SSc, 33 with established SSc [of whom 16 with SSc-associated lung fibrosis,SSc-LF]) and 10 healthy controls (HCs). As a disease controls for SSc-LF, 12 patients with rheumatoid arthritis-associated LF (RA-LF) were included. Peripheral blood monunuclear cells were isolated from patients and controls. Phenotypic analysis of immature/transitional Bregs (CD19+CD24highCD38high) and memory Bregs (Cd19+CD27+Cd24high) was carried out by flow cytometry using FACS Calibur. The function of bregs was evaluated by IL-10 expression after Bcell culture with toll-like receptor (TLR)9 stimulation, and flow cytometry analysis Results Memory Bregs were decreased in early SSc (1.85±0.38), established SSc (1.6±0.88), and SSc-LF (1.52±0.17) compared to HCs (6.3±0.49, p<0.001). There were more decreased in diffuse cutaneous SSc (dcSSc) than limited cutaneous SSc (lcSSc), but not significantly so. The lowest percentage of memory Bregs was in dcSSc-LF (1.36±0.16). Memory Bregs were also numerically decreased in RA-LF compared to HCs (1.58±0.26, p<0.001). Transitional Bregs were also numerically decreased in early SSc, and established SSc compared to HCs (p<0.02). Bregs IL-10 expression after Toll-like receptor (TLR)-9 stimulation (innate stimulus) was impaired in SSc, particularly in SSc-LF. Conclusions This is the first study to demonstrate that Bregs are reduced and functionally impaired in SSc, particularly SSc-LF. The impaired IL-10 production after innate stimulus may have clinical implications. The impairment of Bregs along with the reported increased expression of the stimulatory Bcell receptor CD19 in SSc support B cell autoaggression in SSc. These findings may offer a new therapeutic strategy for SSc, namely expanding Bregs ex-vivo and re-administering them to patients with SSc. Disclosure of Interest None declared

Treatment of ANCA-Negative Small Vessel Vasculitis

Cryoglobulinaemia refers to circulating cryoglobulins. Cryoglobulins are immunoglobulins, which precipitate in temperatures below 37oC and dissolve upon rewarming [2]. Cryoglobu‐ linaemia is classified in 3 types based on clonality and immunoglobulin class. In particular, type I consists of monoclonal IgM or IgG immunoglobulin, type II is a mixture of monoclo‐ nal IgM and polyclonal IgG, while type III is a mixture of polyclonal IgM and IgG. Type II and III are also called “mixed”, since both contain a mixture of IgM and IgG immunoglobu‐ lins [3]. The IgM component of type II cryoglobulins has rheumatoid factor activity (can bind to the Fc portion of IgG). All 3 types of cryoglobulinaemia may or may not result from an underlying disorder. In the absence of an identifiable cause cryoglobulinaemia is charac‐ terized as “essential”.