M. Iwata

Intercellular adhesion molecule-1 expression on human corneal epithelial outgrowth from limbal explant in culture

Aim: To investigate the relation between intercellular adhesion molecule (ICAM)-1 expression and cellular dynamics occurring concomitantly with epithelial cell movement. Methods: Outgrowing epithelial sheets of human corneal epithelial (HCE) cells from cultured limbal explants were examined by immunoperoxidase staining with anti-ICAM-1 monoclonal antibody. An adhesion assay was performed using the epithelial sheets of HCE cells and an Epstein-Barr virus (EVB) infected B cell lymphoma cell line (EVB+BJAB) expressing CD11a/CD18, a counter-receptor of ICAM-1. Also, the effect of calphostin C, a specific protein kinase C (PKC) inhibitor, on ICAM-1 expression on the outgrowing epithelial sheets of HCE cells was examined. Results: Strong positive staining for ICAM-1 was found predominantly on HCE cells in the marginal segment of the epithelial sheet, particularly on the cells at the leading edge. EBV+BJAB cells adhering to the HCE cells corresponded well to the area of ICAM-1 staining. Treatment of outgrowing epithelial sheets with calphostin C markedly decreased the ICAM-1 expression on the HCE cells. Conclusion: ICAM-1 is actively expressed on HCE cells in the marginal segment of the outgrowing epithelial sheets where there is active movement mediated through a PKC dependent mechanism, suggesting the role of ICAM-1 in epithelial cell motility such as the spreading and migration of cells.

Differential effects of protein Tyrosine kinase inhibitors on interferon-γ-induction of major histocompatibility complex class II and intercellular adhesion molecule-1 expression in human corneal epithelial cells

PURPOSE Interferon (IFN)-gamma induces major histocompatibility complex (MHC) class II and intercellular adhesion molecule-1 (ICAM-1) expression on human corneal epithelial (HCE) cells. So far, it has not been clarified whether both inductions by IFN-gamma use the same signal transduction pathway. Therefore, in the present study, we tried to determine the significance of the protein tyrosine kinase (PTK)-dependent signaling pathway in the induction of both MHC class II and ICAM-1 expression by IFN-gamma in cultured HCE cells. METHODS Cultured HCE cells were treated with human recombinant IFN-gamma. The induction of protein tyrosine phosphorylation of proteins including PTKs, janus kinase (JAK)1, and JAK2, was examined by Western blotting and immunoprecipitation. The effects of treatment of HCE cells with specific PTK inhibitors on IFN-gamma-induction of MHC class II and ICAM-1 expression were examined by flow cytometry. RESULTS IFN 1 (Interferon) induced tyrosine phosphorylation of multiple substrates, particularly that of 75,000; 90,000; 130,000; and 160,000 molecular weight proteins including JAK1 and JAK2 in cultured HCE cells. The PTK inhibitors, herbimycin A and genistein, inhibited tyrosine phosphorylation of those proteins. Also, these PTK inhibitors prevented IFN-gamma-induction of MHC class II synthesis and surface expression. However, neither herbimycin A nor genistein had any effect on IFN-gamma-induction of ICAM-1 expression. CONCLUSIONS Tyrosine phosphorylation of proteins including JAK1 and JAK2 is essential for IFN-gamma-induction of MHC class II expression, but not critical for that of ICAM-1 expression in cultured HCE cells. In addition, it is suggested that the IFN-gamma-induction of MHC class II requires PTK activities not only in the primary JAK-signal transducers and activators of transcription (STAT) pathway but also in the subsequent pathway mediated by IFN-gamma-induced intermediate proteins.

Efficacy and safety of minodronic acid hydrate in patients with steroid-induced osteoporosis

Minodronic acid hydrate, an oral bisphosphonate, has a greater inhibitory effect on bone resorption than do other approved drugs; however, this has been studied only in patients with primary osteoporosis. Here, we administered minodronic acid hydrate to patients with steroid‐induced osteoporosis who have been treated with steroids for rheumatoid arthritis or other collagen diseases, and the efficacy and safety of minodronic acid hydrate were prospectively investigated.

THU0337 Raynaud Phenomenon Is Associated with Myocardial Fibrosis in Primary Sjögren Syndrome, Assessed by A Cardiac Magnetic Resonance Approach: A Prospective Pilot Study at A Single Center

Background Primary Sjögren syndrome (pSS) shares many clinical, inflammatory, and immunological features with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). Both SLE and RA are characterized by a high risk of cardiac involvement. However, there are limited data on the risk of overt cardiac involvement in pSS. Objectives We sought to use a cardiac magnetic resonance imaging (CMR) approach to assess cardiac involvement and determine its association with disease characteristics in pSS patients without cardiac symptoms. Methods Consecutive pSS patients, according to ACR classification criteria (2012), without a history or clinical findings of hypertension, cardiovascular disease, diabetes, or dyslipidemia underwent contrast CMR. Late gadolinium enhancement (LGE) was used for the assessment of myocardial fibrosis. Using a black-blood T2-weighted image (T2-WI), myocardial inflammation could be assessed. Sjögren syndrome disease activity index (ESSDAI) was determined. Eighty percent patients had documentation of a minor salivary gland biopsy. Salivary gland biopsy data were classified by focus score (FS). We investigated the patients in terms of prevalence of CMR abnormalities and explored possible associations between CMR abnormalities and pSS disease characteristics. Results Thirty-seven female pSS patients were enrolled (mean age: 55.5 ± 7.0 years). On an average, cardiovascular risk was low for the group, with patients demonstrating no ECG abnormalities, and the patients had generally low traditional cardiovascular risk factors, with a mean Framingham 10-year hard cardiovascular risk score of 4 ± 2%. The mean ESSDAI was 3.3 ± 2.1. Thirteen patients (35%) demonstrated myocardial abnormalities. Myocardial edema was seen in 5 patients (13%) on T2-WI. LGE was found in 11 patients (29%), 3 of whom demonstrated edema on T2-WI. The main finding observed in 7 among 11 LGE-positive patients (63%) was a linear LGE pattern without coronary distribution. A patchy nodular LGE pattern was observed in 4 among 11 patients (37%). The patients with CMR abnormalities showed no significant difference of ESSDAI, compared with those with no CMR abnormalities. Antibodies to La/SSB antigens were significantly higher in LGE-positive than LGE-negative patients (p=0.003). Raynaud phenomenon was significantly associated with LGE-positive and T2-WI-positive patients (p=0.001 and p=0.04, respectively). Other pSS characteristics such as disease duration, commodities, and cardiovascular risk factors were not significantly associated with myocardial abnormalities. The greatest relative difference between LGE-positive and -negative patients was observed in FS >3, with an adjusted odds ratio of 3.0. After adjusting for confounding by age, pSS duration, and anti-SSB antigen, the association of LGE with Reynaud phenomenon remained significant (p=0.02). Conclusions Subclinical myocardial involvement, as detected by CMR, was frequent in pSS patients without cardiac symptoms. Our results suggest that Raynaud phenomenon has a role in promoting cardiac involvements in patients with pSS. Disclosure of Interest None declared

FRI0472 Detection of Left Ventricular Morphology and Myocardial Abnormalities Using Contrast Cardiac Magnetic Resonance Imaging at 3.0 Tesla in Systemic Sclerosis Without Cardiac Manifestations

Background Systemic sclerosis (SSc) has an increased prevalence of cardiac involvement despite often being clinically silent. When clinically evident, cardiac involvement decreased the 70% 5-year mortality of SSc. Cardiac magnetic resonance imaging (CMR) is useful in SSc beause it focuses on late gadolinium enhancement (LGE) abnormalities and ventricular morphology and function. Objectives We aimed to assess the prevalence of subclinical myocardial involvement by left ventricular (LV) function and structure on CMR. We evaluated the relation between myocardial abnormalities and LV geometry. Methods This study compared consecutive female SSc patients without cardiac symptoms and healthy female controls with no history or clinical findings of systemic and pulmonary hypertension by echocardiography, coronary artery disease, valvular heart disease, atrial fibrillation, diabetes mellitus, and dyslipidemia. All underwent non-contrast or contrast CMR on a 3.0-T scanner. LV function was measured using ejection fraction (EF), end-systolic volume (ESV), end-diastolic volume (EDV), stroke volume (SV), and cardiac output (CO). LV hypertrophy was measured by absolute LV mass (LVM) and LVM index (LVMI) determined by LVM/body surface area. LGE was obtained to assess myocardial fibrosis. Myocardial inflammation was assessed by black- blood T2WI. Serum BNP concentrations were measured simultaneously. Results There were 44 SSc patients with a mean age of 57.1±8.7 years; 20 had diffuse type and 24 had limited type. There were 20 healthy controls with a mean age of 56.9±3.1 years. There were no significant differences in terms of age, gender, and cardiovascular risk factors. Compared with the control, SSc patients had a significantly higher EDV with tendency towards a high LVMI. There was no difference in EF. LGE (+) was detected in 25 of 44 (57%) SSc patients; LGE was in a linear pattern without coronary distribution in 13 (52%) SSc patients. T2WI was observed in 11 of 44 (25%) SSc patients. There were no differences in LGE and T2WI between the diffuse and limited type. The BNP level of the SSc group was significantly higher than that of the control group (P=0.04). The mean BNP level of SSc patients with LGE was significantly higher than that of SSc patients without LGE (P<0.0001). BNP level in SSc patients was significantly correlated with LVMI (P<0.0001) but not correlated with EF. Eccentric hypertrophy was observed in 52% of LGE (+) patients. LGE (+) was correlated with (+) anti Scl-70 antibody (P=0.004). After adjustment for age, disease duration, anti Scl-70 antibody, and BNP, SSc with LGE did not have a modified association with LVMI. Conclusions SSc patients without cardiac symptoms have a high prevalence of cardiac abnormalities. Our data suggest that SSc-specific autoimmunity against Scl-70 mediates these changes. SSc patients with LGE had cardiac abnormalities associated with LVMI and serum BNP, leading to cardiac remodeling and possible development of cardiac involvement, even with a normal EF. Disclosure of Interest None declared

AB0321 IL-6 Blockade Reduces Circulating N-Terminal Pro-Brain Natriuretic Peptide Levels in Patients with Active Rheumatoid Arthritis

Background Patients with rheumatoid arthritis (RA) have a 1.5–2.0 fold higher risk of developing congestive heart failure than the general population. Small increases in N-terminal pro-brain natriuretic peptide (NT-proBNP) levels predict left ventricular (LV) dysfunction, and the LV myocardium is the primary site of NT-proBNP production. Data relating to the effects of interleukin (IL)-6 blocking agents on circulating NT-proBNP levels in patients with active RA are lacking but may be informative. To our knowledge, there are no published reports regarding the effect of tocilizumab (TCZ) treatment on NT-proBNP levels. Objectives To test the hypothesis anti-IL-6 therapy might reduce circulating NT-proBNP levels. Methods RA patients with active disease without a clinical diagnosis of cardiovascular disease (CVD) and with an inadequate clinical response to DMARDs were enrolled. The patients received TCZ once a month after 24 weeks. Serum NT-pro BNP levels were measured on the Cobas 6000 modular analyzer simultaneously on stored baseline and 24-week samples, and NT-pro-BNP levels ≥100 pg/mL were considered elevated. We explored the associations between NT-pro BNP and the RA disease activity score for 28 joints: erythrocyte sedimentation rate (DAS28-ESR) and Simple Disease Activity Index (SDAI) scores. The anti-citrullinated protein antibody (ACPA) titre was divided into high and low levels using a cut-off of 30 units/mL. Correlations between the biomarkers and changes in circulating NT-proBNP levels were evaluated using the Spearman rank test, and multivariable linear regression analyses of the correlates were performed. Results Sixty RA patients (mean age, 60.4±10.4 years; 75% female) were enrolled. The DAS28-ESR and SDAI at baseline were 4.57±1.35 and 22.5±12.7, respectively. The 24-week DAS28-ESR and SDAI scores were significantly lower than those at baseline (p=0.04, p=0.03, respectively). The NT-proBNP levels at baseline were approximately 31% higher than normal levels, and the median (interquartile range) levels significantly decreased from baseline (131.78 [52.81–230.24] pg/mL) to 24 weeks (57.13 [29.50–128.67] pg/mL, p=0.004) following TCZ treatment. The change in NT-proBNP levels was significantly correlated with the change in the SDAI score and swollen joints count (SJC) (r =0.455, p=0.003, r =0.395, p=0.004, respectively). The baseline NT-proBNP levels in the high ACPA group tended to be higher than in the low ACPA group (p=0.07). After adjustment for age, gender, ESR, and RA duration, the association between the change in NT-proBNP levels and the change in SJC remained significant (p=0.023). Conclusions The NT-proBNP level was higher than normal in patients with active RA without CVD; this may indicate subclinical left ventricular dysfunction. Furthermore, our results indicate the NT-proBNP levels decreased by approximately 38% with TCZ treatment, which was related to a reduction in disease activity. Therefore, TCZ treatment may directly influence the anti-inflammatory effect of IL-6 on the myocardium. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.2189

FRI0437 How do we Treat Patients with Focus Score ≥1, but not Consistent with the New American College of Rheumatology Classification Criteria for SjÖGren's Syndrome? Evaluation from Study in Japanese Patients

Background One of the criteria for Sjögrens syndrome (SS) is focal lymphocytic infiltration in minor salivary gland biopsy (MSGB). Few studies have revealed that the proportion of patients with focus score ≥1 (FS≥1) in SS and even in the dry mouth (DM) group. Objectives To diagnose SS using the 2012 American College of Rheumatology (ACR) classification criteria for SS, to assess MSGB, and to investigate pathological features, such as FS≥1. Methods Patients underwent MSGB from September 2009 to November 2013 at one institution. The indications for MSGB included the presence of DM symptoms and serological features. Regardless of connective tissue disease (CTD) before FS evaluation, patients who met the ACR criteria were divided into: Group I, primary SS; Group II, secondary SS; Group III, secondary DM; or Group IV, primary DM (Table). We investigated the correlation between FS≥1 and the diagnosis, and the correlation of the predictor with FS≥1. Statistical analyses were performed using the Fishers exact or Mann-Whitney tests, Bonferroni multiple comparisons, and multivariate analysis. These analyses were conducted for patients with rheumatoid arthritis (RA) and those with a CTD other than RA. Results MSGBs were performed on 192 patients (93% female); those with lymphoma, IgG4-related disease, and graft-versus-host disease (7 patients) were excluded. There were 185 patients with SS and DM, including primary SS (82), secondary SS (38), DM with CTDs (22), and DM without CTDs (43). The CTDs included RA (29), systemic lupus erythematosus (15), mixed CTD (3), myositis (6), and systemic sclerosis (8). The classification of the 185 patients by diagnosis and FS≥1 is shown in the Table. More patients in Groups I and II had FS≥1 (P<0.001). Factors such as anti-Ro, anti-La, RF, and IgG correlated with the presence of FS≥1 by univariate analysis, but not by multivariate analysis. The numbers of FS≥1 were significantly different between Groups I and III, I and IV, II and III, and II and IV (all, P<0.05) but not between Groups I and II or Groups III and IV (both, P>0.08). Among the 29 RA patients, 24 had FS≥1, but SS was confirmed in 14 patients only. The numbers of RA patients with FS≥1 in both secondary SS and secondary DM group were not significantly different (P=0.134). However, among patients with CTDs other than RA, the number of patients with FS≥1 in the secondary SS group was significantly different from that in the secondary DM group (P=0.005). Even RA patients with FS≥1 were assigned to DM group. Conclusions Pathologically, FS≥1 are useful for diagnosing SS. A significant number of patients with FS≥1 was found in both the SS and DM groups. However, patients with noncharacteristic SS may also have RA. Consequently, this study suggests especially the necessity of further investigation and follow-up studies in RA patients who showed inconsistent results with the ACR criteria for DM symptoms. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.2684