D.-H. Zheng

Serum Matrix Metalloproteinase-3 as a Noninvasive Biomarker of Histological Synovitis for Diagnosis of Rheumatoid Arthritis

Objective. To explore the correlation between matrix metalloproteinase- (MMP-) 3 and histological synovitis in rheumatoid arthritis (RA). Methods. Serum MMP-3 of 62 patients with active RA was detected by ELISA. Serial synovial tissue sections from all RA patients, 13 osteoarthritis, and 10 orthopedic arthropathies patients were stained with hematoxylin and eosin and immunohistochemically for MMP-3, CD3, CD20, CD38, CD68, and CD15. Results. The percentage of lining MMP3+ cells was significantly higher in RA patients especially with high grade synovitis and it was significantly correlated with Krenns synovitis score (r = 0.574, P < 0.001) and sublining inflammatory cells. Multivariate stepwise linear regression analysis revealed that the association of the percentage of lining MMP3+ cells with activation of synovial stroma, sublining CD68+ macrophages, and CD15+ neutrophils was stronger than other histological indicators. The percentage of lining MMP3+ cells was significantly correlated with serum MMP-3 in RA (r = 0.656, P < 0.001). Serum MMP-3 was higher in RA patients with high grade synovitis than that of low grade synovitis and significantly correlated with synovitis score and activation of synovial stroma subscore (all P < 0.05). Conclusion. Serum MMP-3 may be an alternative noninvasive biomarker of histological synovitis and RA diagnosis.

Synovial Infiltration with CD79a-positive B Cells, But Not Other B Cell Lineage Markers, Correlates with Joint Destruction in Rheumatoid Arthritis

Objective. The efficacy of B cell depletion in the treatment of patients with rheumatoid arthritis (RA) has revitalized interest in the pathogenic role(s) of B cells in RA. We evaluated the distribution of synovial B lineage cells and their correlation with histologic disease activity and joint destruction in RA. Methods. Synovial tissue samples were obtained by closed-needle biopsy from 69 Chinese patients with active RA, from 14 patients with osteoarthritis (OA), and from 15 with orthopedic arthropathies (OrthA) as disease controls. Serial tissue sections were stained immunohistochemically for CD79a (pro-B cell to plasma cell), CD20 (B cells), CD38 (plasma cells), CD21 (follicular dendritic cells), CD68 (macrophages), CD3 (T cells), and CD34 (endothelial cells). Densities of positive-staining cells were determined and correlated with histologic disease activity (Krenn 3-component synovitis score) and radiographic joint destruction (Sharp score). Results. Mean sublining CD79a-positive cell density was significantly higher in RA than in OA (p <0.001) or OrthA (p = 0.003). Receiver operating characteristic curve analysis showed that CD79a-positive cell density differentiated RA well from OA [area under the curve (AUC) = 0.79] or OrthA (AUC = 0.75). Spearman’s rank order correlation showed significant correlations between sublining CD79a-positive cell density and the synovitis score (r = 0.714, p < 0.001), total Sharp score (r = 0.490, p < 0.001), and the erosion subscore (r = 0.545, p < 0.001), as well as the joint space narrowing subscore (r = 0.468, p = 0.001) in RA. Conclusion. Synovial CD79a-positive B cells may be a helpful biomarker for histologic disease activity in RA and may be involved in the pathogenesis of joint destruction in RA.

Elevated Serum IgG4 Defines Specific Clinical Phenotype of Rheumatoid Arthritis

Objectives. To explore the correlation of serum IgG4 (sIgG4) with clinical manifestations or therapeutic response in rheumatoid arthritis (RA). Methods. Consecutive 136 RA patients were recruited and followed up at regular interval. SIgG4 was detected by immunonephelometry. Serial synovial tissue sections from 46 RA patients were stained immunohistochemically for IgG4. Results. Forty-six percent of 136 RA patients had elevated sIgG4. Patients with elevated sIgG4 had higher sIgG4/sIgG ratio, C-reactive protein, erythrocyte sedimentation rate, rheumatoid factor, and anticyclic citrullinated peptide antibodies than those with normal sIgG4 (all P < 0.05). Among 45 patients who received methotrexate and leflunomide therapy, 50% (9/18) of patients with elevated sIgG4 and 85% (23/27) of patients with normal sIgG4 reached therapeutic target (disease activity score of 28 joints < 3.2) at 6-month visit (χ 2 = 6.508, P = 0.011). IgG4-positive plasma cell count correlated positively with sIgG4, total synovitis score, and CD3-, CD20-, and CD38-positive cell counts (all P < 0.05). Conclusions. Our results showed that elevated sIgG4 in RA is common and disproportional to total IgG and RA with elevated sIgG4 may be a specific clinical phenotype with higher disease activity, higher level of autoantibodies, and poor response to methotrexate and leflunomide therapy.

Elevated Serum Glucose-6-Phosphate Isomerase Correlates with Histological Disease Activity and Clinical Improvement After Initiation of Therapy in Patients with Rheumatoid Arthritis

Objective. To determine serum glucose-6-phosphate isomerase (GPI) concentrations in patients with rheumatoid arthritis (RA), and to test whether they correlate with objective measures of disease activity. Methods. Sera from 116 patients with RA, 69 patients with non-RA rheumatic diseases, and 101 healthy controls were analyzed. Levels of soluble serum GPI were measured by ELISA. Histological disease activity was determined with the synovitis score in synovial needle biopsies from 58 of the 116 patients with RA. Thirty-one of the 58 synovium samples were stained for CD68, CD3, CD20, CD38, CD79a, and CD34 by immunohistochemistry. Demographic data were collected, as well as serological and clinical variables that indicate RA disease activity, for Spearman correlation analysis. Results. Serum GPI level correlated positively with the synovitis score (r = 0.278, p = 0.034). Significantly higher soluble GPI levels were detected in the RA sera compared with sera from healthy controls and the non-RA disease controls (2.25 ± 2.82 vs 0.03 ± 0.05 and 0.19 ± 0.57 μg/ml, respectively; p < 0.0001). The rate of serum GPI positivity was significantly higher in the RA patients than in the non-RA disease controls (64.7% vs 10.1%; p < 0.0001). Spearman analysis showed no significant correlation between serum GPI level and Disease Activity Score in 28 joints at baseline. After initiation of antirheumatic treatments, GPI levels decreased significantly (2.81 ± 3.12 vs 1.44 ± 2.09 μg/ml; p = 0.016), paralleling improvement of the disease activity indices. Conclusion. Elevated serum GPI may be involved in the synovitis of RA and may prove useful as a serum marker for disease activity of RA.

Short-course tocilizumab increases risk of hepatitis B virus reactivation in patients with rheumatoid arthritis: a prospective clinical observation

To investigate the impact of short‐course tocilizumab (TCZ) on hepatitis B virus (HBV) reactivation in rheumatoid arthritis (RA) patients.

Joint damage is amplified in rheumatoid arthritis patients with positive thyroid autoantibodies

Background Autoimmune thyroid disease (AITD), which is characterized by an increased presence of thyroid autoantibodies (TAbs), such as antibodies against thyroid peroxidase (TPOAbs) and antibodies against thyroglobulin (TgAbs), has been reported to be associated with rheumatoid arthritis (RA) because AITD and RA both involve autoimmunity. However, few data are available on the incidence of TAbs in Chinese RA patients, and studies on the association between TAbs and joint damage as well as synovitis in RA patients remain sparse. Here, we aimed to evaluate the incidence of TAbs in a consecutive Chinese RA cohort and to investigate whether the elevated presence of TAbs is associated with joint damage and synovitis in RA patients. Methods A total of 125 hospitalized RA patients were consecutively recruited. Clinical data and available synovial tissues were collected at baseline, and TAbs and thyroid function were detected by chemiluminescent immunoassay. Patients who tested positive for TPOAbs or TgAbs were classified as the TAbs-positive group, and patients who tested positive for neither TPOAbs nor TgAbs were recruited as the TAbs-negative group. Disease activity was assessed using DAS28-ESR (the disease activity score in 28 joints and including the erythrocyte sedimentation rate). X-ray assessment of the hand/wrist was performed according to the Sharp/van der Heijde-modified Sharp score (mTSS), and patients with an mTSS score >10 were defined as having radiographic joint damage (RJD). Serial tissue sections were stained immunohistochemically for CD3, CD15, CD20, CD34, CD38, and CD68, and synovitis were assessed according to Krenn’s synovitis score. Results A total of 44 (35%) patients were positive for either TPOAbs or TgAbs. Importantly, there was a significantly greater percentage of patients with RJD in the TAbs-positive group versus the TAbs-negative group (68% vs. 42%, p = 0.005). Compared with the TAbs-negative group, significantly more CD38-positive plasma cells infiltrated the TAbs-positive synovium, and a higher percentage of patients with high-grade synovitis were observed in the TAbs-positive group (5/8, 63% vs. 5/14, 36%). Moreover, RF positivity and disease activity indicators, including TJC28, DAS28-ESR, and CDAI, were significantly higher in the TAbs-positive group (all p < 0.05). Adjusted logistic regression analysis revealed that positive TAbs (OR 2.999, 95% CI [1.301–6.913]; p = 0.010) and disease duration (OR 1.013, 95% CI [1.006–1.019]; p < 0.001) were independently associated with RJD, and an odds ratio of 2.845 (95% CI [1.062–7.622]) was found for RJD in women with positive TAbs (n = 37) compared with those without TAbs (n = 59) (p = 0.038). Conclusion Our data showed that joint destruction was amplified in RA patients with an elevated presence of TAbs, which supports the importance and necessity of TAbs and thyroid function screening and monitoring in RA patient management in clinical practice.

Deleterious role of hepatitis B virus infection in therapeutic response among patients with rheumatoid arthritis in a clinical practice setting: a case-control study

BackgroundPrevious studies have revealed that hepatitis B virus (HBV) infection may be associated with rheumatoid arthritis (RA), while there are no further clinical studies regarding the role of HBV infection in RA progression during disease-modifying anti-rheumatic drug (DMARD) therapy. Here, we aimed to explore the influence of HBV infection on radiographic and clinical outcomes among patients with RA in a clinical practice setting.MethodsThirty-two consecutive patients with RA (Disease Activity Score 28-joint assessment based on C-reactive protein (DAS28-CRP) ≥2.6) with chronic HBV infection (CHB) were retrospectively recruited as the CHB group and 128 age-matched, sex-matched, and disease activity-matched contemporary patients with RA without CHB were included in the non-CHB group. Clinical data were collected at baseline and visits at month 1, 3, 6, and 12. The therapeutic target was defined as DAS28-CRP <2.6 in all patients or <3.2 in patients with long disease duration (>24 months). The primary outcome was the percentage of patients with one-year radiographic progression (a change in modified total Sharp score ≥0.5).ResultsCompared with the non-CHB group, a significantly higher percentage of patients with one-year radiographic progression was observed in the CHB group (53% vs. 17%, p < 0.001), with smaller proportions of patients achieving therapeutic target at month 6 and month 12 (53% vs. 82% and 53% vs. 75%, both p < 0.05), remission at month 6 (DAS28-CRP <2.6, 50% vs. 72%, p = 0.039), and American College of Rheumatology (ACR)20/50 responses and good or moderate European League Against Rheumatism (EULAR) responses mainly at month 6 and 12 (all p < 0.05). Multivariate logistic regression analysis revealed that CHB status was significantly associated with one-year radiographic progression and failure to achieve therapeutic target within 6 months. HBV reactivation occurred in 34% of patients with CHB during one-year follow up, with two patients suffering hepatitis flare.ConclusionsHBV infection may play a deleterious role in radiographic and clinical outcomes in patients with RA, and HBV reactivation should be paid close attention during immunosuppressive therapy.

Patients with Coexistence of Circulating Hepatitis B Surface Antigen and Its Antibody May Have a Strong Predisposition to Virus Reactivation During Immunosuppressive Therapy: A Hypothesis

Hepatitis B virus (HBV) reactivation is a well-recognized complication in patients who undergo immunosuppressive drug therapy. Although the recommendation of antiviral prophylaxis made by the American Gastroenterological Association in 2015 focuses on the risk stratification of different immunosuppressive drugs, risk factors for HBV reactivation are also worth identifying in clinical practice. Recent studies have shown that the uncommon serological pattern of coexistent circulating HBV surface antigen (HBsAg) and its antibody (anti-HBs) was associated with double mutations (A1762T/G1764A) in the basal core promoter (BCP) region of the HBV genome, which is critical for HBV replication. Here, we depicted rheumatoid arthritis (RA) patients with coexistent HBsAg and anti-HBs in our medical center, who developed HBV reactivation during immunosuppressive drug therapy. DNA sequencing analysis of the HBV genome revealed triple mutations (A1762T, G1764A, and T1753V) in the BCP region, which could further enhance the ability of HBV replication. Hence, a novel hypothesis is advanced for the first time that patients with coexistent HBsAg and anti-HBs may have a strong predisposition to HBV reactivation due to specific BCP mutations. This hypothesis would, if correct, justify the concurrent detection of HBsAg and anti-HBs in HBV screening in patients with rheumatic diseases and quickly recognize patients with high risk of HBV reactivation. Further controlled studies are needed to confirm this hypothesis.

AB0813 High Prevalence of Urolithiasis Due To Excess Urinary Uric Acid Concentration and Calcium Excretion in South Chinese Gout Patients: Table 1.

Background Urolithiasis greatly impact urate lowering therapy in Chinese gout patients. Benzbromarone is contraindicated and Chinese is high risk of allopurinol hypersensitivity syndrome. Febuxostat is too expensive for many patients. Few study about urolithiasis in Chinese gout patients are presented. Objectives To investigate the prevalence and risk factors of urolithiasis in south Chinese gout patients. Methods Patients with primary gout were recruited in the present cross-section study. Urolithiasis was defined as positive stone history and/or ultrasonography. All inpatients with estimated glomerular filtration rate (eGFR) over 30 ml/min/1.73 m2 underwent 24 hours urinary chemistry assay including uric acid (UA), UA concentration, clearance of UA, Cr, calcium, phosphorus, potassium, sodium and chloride. Logistic regression were performed to examine association of urolithiasis with independent variables including age, gender, duration of gout, sUA, previous urate lowering therapy, serum creatinine, eGFR, body mass index, urine pH and 24 hours urinary chemistry assay. Results (1) Two hundred and nighty-four patients were recruited. Mean age of male (N=255) and female (N=39) were 54.4±16.4 and 67.2±15.3, respectively. Average serum UA was 9.0±2.5mg/dl and 25.9% of patients presented tophi. Allopurinol, benzbromarone, or a combination of these two drugs had been prescribed in 97 patients (33.0%), 18 patients (6.1%) and 36 patients (12.2%), respectively. The rest 143 patients (48.6%) had not received urate lowering therapy before. (2) One hundred and eleven gout patients (37.8%) complicated with urolithiasis and urolithiasis was observed in 69 of them (62.1%) on ultrasonography. Compared with non urolithiasis group, urolithiasis group had significant higher serum creatinine (1.6±0.6 VS 1.3±0.3 mg/dl], lower eGFR (56.3±18.0 VS 64.1±18.4 ml/min/1.73m2) and higher alcohol overuse (38.7% VS 26.2%) (P<0.05). (3) Among 102 patients performed 24 hours urinary chemical assay, 45 patients complicated with urolithiasis. Urolithiasis group had significantly higher 24 hours urine UA concentration than non-urolithiasis group (30.4±13.1 VS 23.9±11.9 mg/dl, P=0.01). There were no significant difference of other variables between two groups. Prevalence of urolithiasis increased significantly across increasing quartiles of urinary UA concentration (P=0.012, Table 1). (4) Logistic regression suggested that risk factors of urolithiasis were urinary UA concentration grouped by quartile (P=0.05) and urinary calcium excretion [OR 1.26 (1.01, 1.57), P=0.04]. Urinary UA concentration over 24.4mg/dl was associated with increasing risk of urolithiasis (Table 1).Table 1. Prevalence of urolithiasis among quartiles of urinary UA concentration Quartiles of urinary UA concentration (mg/dl) I (<17.6) II (17.6–24.3) III (24.4–34.9) IV (≥35.0) Median (mg/dl) 12.8 20.8 30.0 42.5 Prevalence* 13.3% (6) 22.2% (10) 33.3% (15) 31.1% (14) Adjusted OR# 1 2.47 (0.66, 9.24) 5.5 (1.50, 20.72) 4.58 (1.24, 16.92) P# – 0.18 0.01 0.02 *Test for a linear trend P=0.012. #By logistic regression. Conclusions Prevalence of urolithiasis in Chinese gout patients were high and predicted by excess urinary UA concentration and urinary calcium excretion. Acknowledgement The present study was supported by Guangdong Natural Science Foundation, China (Grant no. 2014A030310086) to Qian-Hua Li. Disclosure of Interest None declared

THU0025 Over-Expressed PGC-1β in CD14+ Monocytes of Rheumatoid Arthritis Patients and Its Down-Regulation Can Suppress Osteoclastogenesis and Bone Resorption through Inhibition of Cathepsin K and Trap

Background Peroxisome proliferator-activated receptor-gamma coactivator-1 (PGC-1) is a transcriptional coactivator that plays important roles in regulating energy metabolism and cytokine signaling pathways. Our previous study showed that down-regulating PGC-1β could inhibit rheumatoid arthritis fibroblast-like synoviocytes induced osteoclastogenesis by RAW264.7 (used as precursor of osteoclasts). Objectives To investigate the expression of PGC-1β in CD14+ monocytes from RA patients and its effect and underlying mechanism on osteoclastogenesis and bone resorption. Methods (1) Peripheral blood mononuclear cells (PBMCs) were collected from 3 patients with active RA and 3 healthy controls and CD14+ monocytes were selected by flow cytometry. PGC-1β expression was detected by immunofluorescence staining or flow cytometry. (2) PGC-1β in RAW264.7 was depleted by lentivirus short hairpin RNAs and then cultured with recombinant murine RANKL (50ng/ml) and M-CSF (25ng/ml). Tartrate-resistant acid phosphatase (TRAP) staining and western blot (WB) for detecting the expression of DC-STAMP, cathepsin K and TRAP were performed at day 7. Toluidine blue staining for examining resorption pits by reflected light microscopy and image analysis system was performed at day 14. (3) After PGC-1β knockdown in RAW264.7, cytoplasmic protein was extracted for WB determining the expression of JNK, p-JNK, p38, p-p38, p-ERK1/2, NFATc-1, NF-κB and IκB, while nuclear protein was extracted for WB determining the expression of PGC-1β, p- NF-κB, ERK1/2, c-jun and c-fos. Results (1) Immunofluorescence staining showed that the percentage of PGC-1β positive PBMCs in RA was significantly higher than that in healthy controls (94±3% vs. 57±7%, P=0.021, Figure 1A). Flow cytometric analyses showed that PGC-1β expression in CD14+ monocytes from RA patients was higher than that of healthy controls (Figure 1B) and the mean fluorescence intensity was significantly higher than that of healthy controls (274±23 vs. 167±15, P=0.034).(2) At day 7, the number of TRAP-positive multinucleated osteoclasts in sh-PGC-1β group was significantly lower than that in sh-GFP group (33.4 ± 3.4 vs. 192.5 ± 5.1 cells/per well, P=0.021, Figure 1C). At day 14, the pit area of bone resorption lacunae on the slices of sh-PGC-1β group was significantly reduced than that of sh-GFP group (13 ± 2 vs. 197 ± 15 μm2/per slice, P=0.012, Figure 1C). (3) WB assay showed that the protein expression of cathepsin K and TRAP,but not DC-STAMP, were obviously suppressed in sh-PGC-1β group (Figure 1D). Further analysis of signaling pathways showed the expression of NFATc1 and phosphorylate NF-κB p65 were also significantly decreased (Figure 1E). Conclusions Our results showed that over-expressed PGC-1β in CD14+ monocytes of RA patients and down-regulation of PGC-1β can suppress RANKL induced osteoclastogenesis and bone resorption through inhibition of cathepsin K and TRAP by NF-κB and NFATc1 pathway. Acknowledgement This work was supported by National Natural Science Foundation of China (81471597), Specialized Research Fund for the Doctoral Program of Higher Education (20130171110075) and Guangdong Natural Science Foundation (2014A030313074). Disclosure of Interest None declared