A. Murgo

Differences in serum and synovial CD4+ T cells and cytokine profiles to stratify patients with inflammatory osteoarthritis and rheumatoid arthritis

BackgroundThe aim was to investigate CD4+T-cell subsets, immune cells and their cytokine profiles in blood and synovial compartments in rheumatoid arthritis (RA) and inflammatory osteoarthritis (OA) to define specific immune signatures.MethodsPeripheral blood, synovial fluid (SF) and synovial membranes (SM) of RA and OA patients were analyzed. CD4+T-cell subset frequencies were determined by flow cytometry, and cytokine concentrations in serum and SF were measured by ELISA.ResultsIn peripheral blood, OA patients had altered frequencies of regulatory T-cell subsets, and higher frequencies of Th17 and of Th1/17 cells than RA patients. In the synovial compartment of OA patients, conventional Th17 cells were largely excluded, while Th1/17 cells were enriched and more frequent than in RA patients. Conversely, in the synovial compartment of RA patients, regulatory T cells and Tfh cells were enriched and more frequent then in OA patients. IL-17 and Blys were increased both in serum and SF of RA patients, and correlated with autoantibodies and disease activity. Notably, Blys levels were already significantly elevated in RA patients with low disease activity score in 28 joints (DAS28) and without autoantibody positivity.ConclusionsAlthough patients with inflammatory OA have immune activation in the synovial compartment, they display different T-cell subset frequencies and cytokine profiles. Soluble mediators such as Blys might help to discriminate mild clinical forms of RA from inflammatory OA particularly at the onset of the disease.

N-TproBNP as biomarker in systemic sclerosis

Systemic sclerosis (SSc) is a connective tissue disorder characterized by tissue fibrosis affecting the skin and internal organs, fibroproliferative vasculopathy, and autoimmune activation. SSc still heralds a poor prognosis with significant morbidity and mortality. Early detection of organ involvement is critical as currently available treatments are most effective when started early. Many candidate biomarkers have been investigated in the past two decades. However, despite the enormous efforts, no accurate tool to predict the pattern of organ involvement and to assess disease activity has been yet identified. The N-terminal fragment of probrain natriuretic peptide (N-TproBNP) is a neurohormone released by ventricular myocytes in response to pressure overload. N-TproBNP is highly relevant for diagnosis, prognosis, and prediction of pulmonary arterial hypertension in SSc. Moreover, several studies support its potential benefit for cardiac assessment of scleroderma patients. Conversely, the role of N-TproBNP as surrogate marker of pulmonary fibrosis and skin involvement is much less clear. We provide an extensive review of the studies that have previously investigated the role of N-TproBNP as candidate biomarker in scleroderma manifestations, presenting also the findings of a recent study we conducted in a cohort of 87 SSc patients.

Baseline ultrasound examination as possible predictor of relapse in patients affected by juvenile idiopathic arthritis (JIA)

Objectives To define the correlation between joint ultrasonography and clinical examination in patients with juvenile idiopathic arthritis (JIA) and to assess whether synovitis detected by ultrasonography in clinically inactive patients predicts arthritis flares. Methods 88 consecutive patients with JIA—46 (52%) with persistent oligoarthritis, 15 (17%) with extended oligoarthritis, 15 (17%) with rheumatoid factor-negative polyarthritis and 12 (14%) with other forms of JIA, all clinically inactive for a minimum of 3 months—underwent ultrasound (US) assessment of 44 joints. Joints were scanned at study entry for synovial hyperplasia, joint effusion and power Doppler (PD) signal. Patients were followed clinically for 4 years. Results US was abnormal in 20/88 (22.7%) patients and in 38/3872 (0.98%) joints. Extended oligoarthritis and rheumatoid factor-negative polyarthritis were more frequent in US-positive than in US-negative patients (35.0% vs 11.8% and 30.0% vs 13.2%, respectively; P=0.005). During 4 years of follow-up, 41/88 (46.6%) patients displayed a flare; 26/68 (38.2%) were US-negative and 15/20 (75%) were US-positive at baseline. Abnormality on US examination, after correction for therapy modification, significantly increased the risk of flare (OR=3.8, 95% CI 1.2 to 11.5). The combination of grey scale and PD abnormalities displayed a much higher predictive value of relapse (65%, 13/20) than grey scale alone (33%, 6/18). Conclusions US abnormalities are a strong predictor of relapse at individual patient level. Irrespective of treatment, the risk of flare in US-positive versus US-negative patients was almost four times higher. In case of US abnormalities, patients should be carefully followed regardless of both the International League of Associations for Rheumatology and Wallace categories.

Acute and chronic effects of two different intravenous iloprost regimens in systemic sclerosis: a pragmatic non-randomized trial

Objectives I.v. iloprost (ILO) may be used in the treatment of refractory RP and digital ulcers. We aim to evaluate the acute and chronic effects of two different ILO regimens by power Doppler US (PDUS) and nailfold videocapillaroscopy. Methods In this 3-month single-centre pragmatic non-randomized trial, 96 SSc patients were included and stratified according to ILO treatment as: no ILO (group A), ILO once monthly (group B) and ILO for five consecutive days (group C). Resistivity index (RI), finger pulp blood flow and periungual vascularization by PDUS, and sum of capillaries apex width in 1 mm by nailfold videocapillaroscopy were evaluated. Results were adjusted for the average outdoor temperature at the place of residence. Results An acute ILO effect was observed for only finger pulp blood flow in groups B and C (P < 0.001 and P < 0.005, respectively). An acute effect was observed for RI and periungual vascularization only in group B. A progressive increase was observed for other parameters without statistical difference. ILO effects were not observed any longer before the following infusion. Some parameters (finger pulp blood flow in group B and RI in group C) showed a statistically higher increase the lower the outdoor temperature was. Conclusion ILO had an acute effect as assessed by PDUS, especially in group B. By contrast, an ILO chronic effect was not detectable before the following infusion in both treatment groups. More studies are needed to define how often ILO should be administered.

Fibrosis biomarkers in isolated Raynaud's phenomenon: too little, too soon?

Raynauds phenomenon (RP) can be the first manifestation or may be present before the development of an overt systemic sclerosis (SSc).1 Enhanced liver fibrosis (ELF) test, an algorithm combining tissue inhibitors of matrix metalloproteinases (TIMP-1), hyaluronic acid (HA) and aminoterminal propeptide of type III procollagen (PIIINP), has been related to several measures of fibrosis in SSc patients.2–⇓4 We evaluated whether these biomarkers could discriminate between primary and secondary RP. Consecutive adult patients with RP at the first rheumatologic evaluation were recruited from February 2011 to May 2012 with ethics committee approval. Exclusion criteria were history of any fibrosing disorder, organ transplantation, hepatocellular carcinoma or treatment with interferon. One patient was excluded for interferon treatment. Fifteen limited cutaneous SSc and 15 diffuse …

Evidence of macro- and micro-angiopathy in scleroderma: An integrated approach combining 22-MHz power Doppler ultrasonography and video-capillaroscopy

OBJECTIVE We aimed to study in SSc patients macrovascular involvement by using power Doppler ultrasound (PDUS) and microvascular one by PDUS and nailfold video-capillaroscopy (NVC) and to examine the association between history of digital ulcers (HDU) and imaging (PDUS and NVC) parameters. METHODS NVC and PDUS were systematically performed in 106 consecutive SSc patients at the 3rd and 4th finger of the dominant hand after exclusion of ulnar artery occlusion (UAO). 22 MHz PDUS measurements included nailbed and fingertip qualitatively graded, and resistivity index (RI) of ulnar and radial proper digital arteries. Capillary number/mm was calculated by NVC on the same digits examined by PDUS. RESULTS Vascularization at fingertip and nailbed showed a good correlation between them and to capillary number. RI, representative of macrovascular involvement, did not correlate to microvascular involvement as assessed by PDUS and NVC. RI and capillary number at NVC showed significant correlation to HDU while fingertip and nailbed vascularization as assessed by PDUS did not. As such, PDUS and NVC provide different and potentially complementary information on SSc-related peripheral macro- and micro-vascular involvement. CONCLUSION Macro- and micro-vascular involvement in SSc patients are different processes and are not present at the same time in every patient. Thus, both these aspects should be carefully evaluated in SSc patients.

FRI0677 Role of nailfold videocapillaroscopy and 22-MHZ doppler ultrasound in the assessment of systemic sclerosis-related digital vasculopathy

Background Microvascular damage plays a critical role in the initiation and perpetuation of systemic sclerosis (SSc). A comprehensive approach should investigate both superficial and deep layers of peripheral microcirculation. In addition to nailfold videocapillaroscopy (NVC), a well-established technique to evaluate outer skin layer vessels, power Doppler ultrasound (PDUS) has been recently used to study microcirculation in the inner levels [1]. Objectives To study the severity of microvascular involvement in patients with SSc by using both NVC to measure capillary density (outer layer at the nailfold area) and PDUS to detect perfusion (deeper layers at the nailfold and pulp area). Methods 100 SSc consecutive patients fulfilling the 2013 EULAR classification criteria were enrolled. PDUS was performed at the 3rd and 4th finger of the dominant hand after exclusion of ulnar artery occlusion (UAO). In case of UAO non-dominant hand was examined. Ultrasound investigation was performed with Esaote MyLab 70 XVG by means of linear array transducer (10–22 MHz). Power Doppler settings were standardized (Doppler frequency 14.3 MHz, Gain 55%, PRF 750 Hz). PDUS measurements included sagittal scan of nailbed and fingertip qualitatively graded from 1 (no signal) to 4 (marked hyperemia) [2], and resistivity index (RI) of ulnar and radial proper digital arteries. Capillary density (number/mm) was calculated by NVC with magnification 200X performed on two images of the same digits examined by PDUS. Results 100 SSc patients, 87 (87%) women, 86 (86%) limited cutaneous SSc, median age 62.2 years old, median disease duration 8 years were evaluated. 7 (7%) patients had UAO. Concordance between fingertip and nailbed perfusion as assessed by PDUS is reported in Table 1.Table 1 Nailbed PDUS Sum Grade 1 Grade 2 Grade 3 Grade 4 Fingertip PDUS Grade 1 15 19 3 1 38 Grade 2 13 13 6 6 38 Grade 3 3 5 10 10 28 Grade 4 2 8 15 71 96 Sum 33 45 34 88 200 Concordance between fingertip and nailbed perfusion as assessed by PDUS is equal to 0.7398. The lower 97.5% confidence interval limit is 0.6433. Association between capillary density, and fingertip and nailbed perfusion as assessed by PDUS is shown in Table 2.Table 2 Capillary density p-value of the difference between the mean of the category, with respect to reference (grade 1) Fingertip PDUS  Grade 1 2.895  Grade 2 3.763 0.038  Grade 3 3.500 0.181  Grade 4 3.844 0.007 Nailbed PDUS  Grade 1 3.212  Grade 2 3.433 0.597  Grade 3 3.294 0.854  Grade 4 3.949 0.049 Conclusions To our knowledge, this is the first study to correlate NVC and PDUS finding in SSc patients. Fingertip and nailbed PDUS grade concordance was found to be satisfactory. The mean capillary density tends to be greater with respect to grade 1. This is particularly evident comparing grade 4 and grade 1. As such, these two imaging techniques provide different and potentially complementary information on SSc-related peripheral microvascular involvement. There is potential clinical utility in these observations that has yet to be unlocked fully. References Lescoat A et al. Arthritis Care Res. 2016;Epub ahead of print. Newman JS et al. Radiology. 1996,198:582–584. Disclosure of Interest None declared

OP0224 Th17 Cells and TFH Cells and their Cytokine Products Are Enriched in the Synovium of Rheumatoid Arthritis Patients and Correlate with Disease Activity

Background Rheumatoid arthritis (RA) and osteoarthritis (OA) patients are affected by joint degradation caused by autoimmune pathology and cartilage loss, respectively. The balance between effector and regulatory T cell subsets that secrete respectively IL-17 (Th17 cells), IL-21 and BAFF (follicular helper T cells, TFH) or IL-10 (Tr1 cells and Tregs) are thought to be crucial in RA pathology, while little is known on the role of these T cell subsets in OA. Objectives To monitor the frequency and function of T cell subsets in RA and OA patients in peripheral blood or affected tissue (synovial fluid and synovial membrane) and their cytokine products in serum and synovial fluid, and to correlate these parameters with disease activity. Methods Blood samples from 24 clinically well-characterized RA patients, 11 OA and 25 healthy donors (HD) were included. Additionally, synovial fluid as well as synovial membranes were analysed when available. Composition of different T cell subsets according to surface marker expression and intracellular cytokine production were assessed by flow cytometry. Cytokines in serum and secreted by T cell subsets were correlated with disease activity assessed as das28 index or the presence of autoantibodies. Synovial B cell IgG production was measured in the absence and presence of helper and regulatory T cell subsets by ELISA. Results Th17 cells were decreased in the peripheral blood of RA patients, while OA patients had surprisingly an altered ratio of CD25+Tregs and Tr1 cells in circulation. In synovial tissues Th17 cells, TFH and regulatory T cell subsets were enriched among CD4+ T cells in RA as compared to OA patients, while frequencies of Th1 cells were similar. RA patients had higher serum levels of IL-17, IL-10 and BAFF, while serum IL-21 was elevated in both RA and OA patients. Importantly, serum levels of IL-17, IL-10 and BAFF were enhanced only in patients with active RA or with detectable autoantibodies. Moreover, BAFF and IL-17 levels in synovial fluid were also higher in RA as compared to OA patients, while IL-10 concentrations were similar. B cells from synovial fluid spontaneously released high amounts of IgG in the absence of CD4+ helper T cells, and regulatory T cell subsets were unable to suppress B cell IgG production. Conclusions Th17 cells and TFH cells accumulate in synovial tissues of RA patients, and IL-17 and BAFF both in serum and synovial fluid correlate with disease activity, suggesting a role for TFH cells and/or Th17 cells in disease progression and autoantibody production. Moreover, although regulatory T cell subsets are efficiently recruited to the synovium they seem to be functionally impaired, since they failed to suppress B cell antibody production. Interestingly, also OA patients show a peculiar modulation of T cell subsets in peripheral blood, but in contrast to RA patients only the anti-inflammatory cytokine IL-10 is expressed in the synovium. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.5392

FRI0403 How early starts increased collagen synthesis in systemic sclerosis

Background One of the hallmarks of overt Systemic Sclerosis (SSc) is the interstitial and perivascular fibrosis resulting in functional impairment of affected organs. During the fibrotic process, the increased collagen synthesis has been correlated to the increase of some biomarkers of collagen metabolism, but it is not know how early these biomarkers can be detected during the disease course. Objectives To evaluate the ongoing fibrotic process based on collagen biomarkers in a cohort of patients in different phases of the disease: from isolated Raynaud’s phenomenon (RP) to overt SSc. For this purpose tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), hyaluronic acid (HA), aminoterminal propeptide of type III procollagen (PIIINP) and Enhanced Liver Fibrosis (ELF) test were chosen. Methods 140 consecutive adult subjects referring in approximately 6 months time were enrolled. They were classified as: 1) primary RP (pRP) [1]; 2) RP suspected secondary to SSc; 3) very early SSc [2]; 4) limited cutaneous (lc)-SSc; and 5) diffuse cutaneous (dc)-SSc. Patients with any other fibrosing disorder or treated with interferon were excluded. TIMP-1, HA, PIIINP and ELF score (derived from an algorithm of the above mentioned biomarkers) were blindly determined by an independent commercial service (iQur, UK). Statistical analysis was performed by regression modelling and ROC curve analysis adjusting for age. Results The study population consisted in 50 pRP (47 women, median age 44.5 yrs), 43 RP suspected secondary to SSc (43 women, median age 43 yrs), 16 very early SSc (16 women, median age 57.5 yrs),15 lc-SSc (14 women, median age 67 yrs), and 15 dc-SSc (13 women, median age 65 yrs). 1 patient was excluded for a recent treatment with interferon. The ELF test and HA were significantly higher in patients with very early SSc compared to subjects with RP suspected secondary to SSc (ELF test: median 8,46 vs 7,84 ng/ml, p= 0.0286; HA: median 30,62 vs 13,6 ng/ml, p= 0.03). PIIINP was significantly higher in patients with dc-SSc compared to very early SSc (median 7,23 vs 5,42 ng/ml; p= 0.006). These differences were not confirmed when these results were weighted for age. When patients with overt SSc were studied an increase of these biomarkers was observed as a trend, but only PIIINP reached a statistical significance. This last result was also confirmed once being weighed for age. Conclusions In our study, the selected biomarkers were not able to distinguish among subjects between pRP, suspected secondary RP, and patients with very early SSc. Nevertheless, these fibrosis biomarkers showed an increasing trend in different subgroups and with age, with no relation to RP duration. These data support the hypothesis that the immune perturbations and vascular injury precede the development of fibrosis, which, in turn, further exacerbates vascular and immune damage. References LeRoy EC, Medsger TA, Jr. Clin Exp Rheumatol. 1992;10:485-8. Avouac J, Fransen J, Walker UA, Riccieri V, Smith V, Muller C, et al. Ann Rheum Dis. 2011;70:476-81. Disclosure of Interest None Declared

AB0998 Medium and high molecular weight hyaluronic acid injected in hip joint by US-guided technique in patients with primary or secondary hip osteoarthritis: A 2 year observational study

Background Guidelines for treatment of osteoarthritis (OA) of the hip of the American College of Rheumatology provide intra-articular therapy with steroids and hyaluronic acid (HA) in addition to standard therapy. The hip joint is difficult to approach and several studies in the literature show greater safety and accuracy when the US-guided procedure is used. The HA used are different although high molecular weight (MW) HA allows smaller number of infiltrations. However, there are no data on the best administration schedule (i.e. number or intervals of infiltrations) and on the clinical characteristics of the patients in order to get the highest efficacy. Objectives To evaluate the efficacy and safety of medium and high MWHA injection in hip joint by US-guided technique with a follow up of 2 years. To study if there are negative predictive factor for this therapy. Methods 95 hips of 91 patients (45 M; 46 F), mean age 57,6±15,35, 80 primary hip OA, 15 hip OA secondary to inflammatory rheumatic diseases were randomly assigned to treatment with Hylan G-F 20 [Synvisc®-Genzyme] (56 hips), or Sodium Hyaluronate >1.6KDa [Hyalubrix®-Fidia] (39 hips), intra-articularly in the hip. The treatment consisted of three injections: at the inclusion, after 1 and 2 months; then the treatment was repeated every 6 months. Efficacy was assessed by pain evaluation on a 100mm VAS scale and WOMAC questionnaire for hip osteoarthritis recorded at the inclusion, after 1, 6, 12 and 24 months. Data on age, BMI, radiologic degree of OA (Kellgren-Lawrence scale) and duration of symptoms were also recorded at the inclusion. Viscosupplementation was performed with an anterior sagittal approach, under ultrasound guidance. Patient who had to recur to prosthesis up to six months after the end of the study were excluded. Friedman test for repeated measures with Bonferroni correction was used to test efficacy of the treatment. Multivariate logistic regression analisys was used to find if there were predictive indices for the response variables. Results The treatment produced a significant improvement on VAS pain and on WOMAC total score (p<0.0001) rapidly after 1 month maintained over 2 years independently from radiological degree of OA, independently from the type of HA used and present both in primary and secondary OA group. The improvement was faster in the group treated with higher MW HA that was showed by a significant improvement on VAS pain (p=0.039) between T0 and T1 in the higher MW group vs. medium MW group. Multivariate logistic regression analysis on age, disease duration, BMI and radiological degree didn’t showed any significance. However there was a trend towards significance that shows that patients non responder to treatment were older than average (p=0.08) and with higher BMI (p=0.09). Conclusions With our scheme of treatment there was an improvement on pain function immediately one month after intraarticular HA treatment that was maintained over the 2 years. In our study high MW Synvisc seems to be more rapid in the effect than medium MW Hyalubrix. Advanced age and higher BMI seems to be negative predictive factors for the result of the therapy, while radiological scoring is not. Further studies with larger numbers of patients and control arms are needed to confirm our results. Disclosure of Interest None Declared