Huaqun Zhu

Regulatory T cells in rheumatoid arthritis showed increased plasticity toward Th17 but retained suppressive function in peripheral blood

Objective Regulatory T cells (Tregs) with the plasticity of producing proinflammatory cytokine IL-17 have been demonstrated under normal and pathogenic conditions. However, it remains unclear whether IL-17-producing Tregs lose their suppressive functions because of their plasticity toward Th17 in autoimmunity. The aim of this study was to investigate IL-17-producing Tregs from patients with rheumatoid arthritis (RA), and characterise their regulatory capacity and clinical significance. Methods Foxp3 and IL-17 coexpression were evaluated in CD4 T lymphocytes from RA patients. An in vitro T cell polarisation assay was performed to investigate the role of proinflammatory cytokines in IL-17-producing Treg polarisation. The suppressive function of IL-17-producing Tregs in RA was assessed by an in vitro suppression assay. The relationship between this Treg subset and clinical features in RA patients was analysed using Spearmans rank correlation test. Results A higher frequency of IL-17-producing Tregs was present in the peripheral blood of RA patients compared with healthy subjects. These cells from peripheral blood showed phenotypic characteristics of Th17 and Treg cells, and suppressed T cell proliferation in vitro. Tregs in RA synovial fluid lost suppressive function. The Th17 plasticity of Tregs could be induced by IL-6 and IL-23. An increased ratio of this Treg subset was associated with decreased levels of inflammatory markers, including the erythrocyte sedimentation rate and C-reactive protein level, in patients with RA. Conclusions Increased levels of IL-17-producing Tregs were identified in RA patients. This Treg subset with Th17 plasticity in peripheral blood retained suppressive functions and was associated with milder inflammatory conditions, suggesting that this Treg population works as a negative regulator in RA, but in RA synovial site it may be pathogenic.

Different expression patterns and clinical significance of mAxl and sAxl in systemic lupus erythematosus

Axl is one of the TAM family members that downregulates activated immune responses to maintain immune homeostasis. We analyzed the expression and clinical relevance of Axl on the surface of CD14+ monocytes/macrophages (mAxl, membrane Axl) and in the plasma (sAxl, soluble Axl) from patients with systemic lupus erythematosus (SLE). Compared to healthy subjects, the concentrations of sAxl were significantly elevated in plasma from SLE patients, while the mAxl expression on CD14+ monocytes/macrophages from SLE patients was significantly downregulated. A series of severe disease clinical manifestations and laboratory features such as presence of autoantibodies, 24-hour proteinuria excretion or SLEDAI ≥10 were associated with decreased mAxl expression on monocytes/macrophages but elevated sAxl levels in plasma. The plasma level of Gas6, the main ligand of Axl, was slightly decreased in SLE patients, and was negatively correlated with anti-dsDNA antibodies and C-reactive protein. SLE patients with SLEDAI ≥10 showed significantly lower Gas6 levels. Our study suggests that abnormal mAxl and sAxl expression may be involved in the imbalance of immune regulation in SLE.

Pathogenic conversion of regulatory B10 cells into osteoclast-priming cells in rheumatoid arthritis

Regulatory B10xa0cells were functionally impaired in rheumatoid arthritis (RA), yet the mechanisms were unclear. B cells are recently recognized as important participants in osteoclastogenesis by producing RANKL. In this study, we investigated whether regulatory B10xa0cells could convert into RANKL-producing cells, thus impairing their immunosuppressive functions in RA and exacerbating the disease progression. Our results showed that human regulatory B10xa0cells could ectopically express RANKL. Under RA circumstance, RANKL-producing B10xa0cells expanded dramatically, partially induced by TNF-α. The frequencies of these cells were positively correlated with RA patient disease activities and tender joint counts, but negatively correlated with the frequencies of regulatory B10xa0cells. Strikingly, RANKL-producing B10xa0cells from RA patients, but not healthy individuals significantly promoted osteoclast differentiation and bone erosion in a paracrine and cell-cell contact-dependent manner. Moreover, these pathogenic RANKL-producing B10xa0cells declined while regulatory IL-10-producing B10xa0cells increased in RA patients with disease remission after therapy. Collectively, these results showed that in RA, regulatory B10xa0cells demonstrated the potential of converting into RANKL-producing cells, thus exacerbating osteoclast formation, bone destruction and disease progression. Modulating the status of B10xa0cells might provide novel therapeutic strategies for RA.

Soluble TAM receptor tyrosine kinases in rheumatoid arthritis: correlation with disease activity and bone destruction

The TAM receptor tyrosine kinases (TAM RTK) are a subfamily of receptor tyrosine kinases, the role of which in autoimmune diseases such as systemic lupus erythematosus has been well explored, while their functions in rheumatoid arthritis (RA) remain largely unknown. In this study, we investigated the role of soluble TAM receptor tyrosine kinases (sAxl/sMer/sTyro3) in patients with RA. A total of 306 RA patients, 100 osteoarthritis (OA) patients and 120 healthy controls (HCs) were enrolled into this study. The serum concentrations of sAxl/sMer/sTyro3 were measured by enzyme‐linked immunosorbent assay (ELISA), then the associations between sAxl/sMer/sTyro3 levels and clinical features of RA patients were analysed. We also investigated whether sTyro3 could promote osteoclast differentiation in vitro in RA patients. The results showed that compared with healthy controls (HCs), sTyro3 levels in the serum of RA patients were elevated remarkably and sMer levels were decreased significantly, whereas there was no difference between HCs and RA patients on sAxl levels. The sTyro3 levels were correlated weakly but positively with white blood cells (WBC), immunoglobulin (Ig)M, rheumatoid factor (RF), swollen joint counts, tender joint counts, total sharp scores and joint erosion scores. Conversely, there were no significant correlations between sMer levels and the above indices. Moreover, RA patients with high disease activity also showed higher sTyro3 levels. In‐vitro osteoclast differentiation assay showed further that tartrate‐resistant acid phosphatase (TRAP)+ osteoclasts were increased significantly in the presence of sTyro3. Collectively, our study indicated that serum sTyro3 levels were elevated in RA patients and correlated positively with disease activity and bone destruction, which may serve as an important participant in RA pathogenesis.

Impaired CD27+IgD+ B Cells With Altered Gene Signature in Rheumatoid Arthritis

Natural antibodies, particularly natural IgM, are proved to play indispensable roles in the immune defenses against common infections. More recently, the protective roles of these natural IgM were also recognized in autoimmune diseases. They are mainly produced by B-1 and innate-like B cells (ILBs). Human CD19+CD27+IgD+ B cells, also termed as un-switched memory B cells, were proposed to be a kind of ILBs. However, functional features and characteristics of these cells in rheumatoid arthritis (RA) remained poorly understood. In this study, we found that human CD27+IgD+ B cells could produce natural antibody-like IgM. Under RA circumstance, the frequencies of these cells were significantly decreased. Moreover, the IgM-producing capacities of these cells were also dampened. Interestingly, the BCR repertoire of these cells was altered in RA, demonstrating decreased diversity with preferential usage alteration from VH3-23D to VH1-8. Single cell sequencing further revealed the proinflammatory biased features of these cells in RA. These CD27+IgD+ B cells were negatively correlated with RA patient disease activities and clinical manifestations. After effective therapy with disease remission in RA, these cells could be recovered. Taken together, these results have revealed that CD27+IgD+ B cells were impaired in RA with dysfunctional features, which might contribute to the disease perpetuation.

AB1033 Comorbidities and metabolism indexes in patients with hua comparing with normal uric acid: a data from 33319 patients between 2014 and 2015

Objectives To investigate and compare the differences in Comorbidities and metabolism indexes between peoples with high and normal uric acid. Methods All the outpatients were enrolled in this cross–sectional study from September 2014 to September 2015 in peking university people’s hospital. The patients were divided into high uric acid (HUA) and normal uric acid according to the international definition of HUA. The distribution characteristics of UA level and correlations between UA level and prevalence rate of comorbidities and metabolism indexes, including glutamic-pyruvic transaminase, glutamic-oxalacetic transaminase, serum total cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), blood glucose, creatinine (CRE), carbonyldiamide, estimated glomerular filtration rate (eGFR) were analysed. Results (1 Among 33u2009319 outpatients, with an average age of 56.4, 44.4% were men with a higher uric acid level than women (368±113.1u2009umol/ vs 300±95.5u2009umol/L, p<0.001). In all patients, 25.6% had HUA and the sex ratio was about 1:1, while 50–59 age were most common in men and women with 60–69 age were the most.2 Both the levels of uric acid in males and females had a significant relationship with the metabolism indexes above-mentioned. However, the levels of uric acid in females had a positive correlation with age and blood glucose, the levels of uric acid in males were negative correlated with age and bloodglucose. And the uric acid levels in females displayed a higher relationship with carbonyldiamide and eGFR.3 The prevalences of common chronic metabolic disorders, were significantly increased in HUA patients than those with normal uric acid, such as hypertension, hypertensive renal disease, hyperlipoidemia, diabetes mellitus (DM), diabetic nephropathy (DN), chronic renal failure. And male patients with HUA had higher prevalence of gout than females (10.76% VS 2.58%, p<0.001). In patients with normal uric acid, the prevalence of DM and DN in males was higher than females. In contrast, the prevalence of DM and DN in males was lower than females in the patients with HUA (25.7% VS 57.1%, p<0.05;6.52% VS 41.76%, p<0.001). Conclusions The prevalence of HUA in females was showing a increasing trend and the uric acid level in females had more influence in blood glucose and renal function than males. Disclosure of Interest None declared

387 A cross-sectional study on application of glucocorticoid in systemic lupus erythematous patients in china

Background and aims To explore the status of glucocorticoid application in patients with systemic lupus erythematosus (SLE) in China. Methods The SLE patients who meet the 1997 classification criteria of American College of rheumatology were enrolled. Epidemiological survey was used. The usage of glucocorticoid and related adverse reactions were recorded and analysed. Results The 400 cases with SLE were enrolled. In these patients, the male to female ratio was 1:19. The average age was 37.37±13.96 years old, the average duration was 6.7±5.8 years. Among them, 310 patients were in glucocorticoid maintenance stage. 61% of patients received the medium dosage (30–60u2009mg/d) as the initial treatment dosage of glucocorticoid. However, patients receiving different initial dosage had no discrepancy on glucocorticoid in the maintenance therapy. In the maintenance stage, 51% of patients received 2.5–5u2009mg/d prednisone, while the dosage of 5–10u2009mg/d could maintain for a longer time. Patients with internal organs involvement had a higher tendency to receive 60–100u2009mg/d or pulse-dose therapy in the initial treatment, nevertheless there had no difference on the dosage of glucocorticoid in the maintenance stage. Among the 400 patients, 62 patients had glucocorticoid withdrawal, including 17 patients due to disease remission (17/400, 4.25%), 44 patients due to self-withdrawal (44/400, 11%) and one patient due to adverse reaction (1/400, 0.25%). Conclusions In China, the medium dosage of glucocorticoid is the most common choice in the initial treatment of SLE patients, and the dosage of 2.5–5u2009mg/d was most common in maintenance stage. Currently, the proportion of glucocorticoid withdrawal remains low after SLE patients achieving the remission.

Elevated serum interleukin‐35 is associated with disease activity in patients with systemic lupus erythematosus

Dear Editor, Systemic lupus erythematosus (SLE) is a systemic autoimmune disease with a wide variety of presenting features. Many studies have suggested cytokines are involved in the pathogenesis of SLE. Interleukin (IL)35, a heterodimeric cytokine composed of the subunits EBI3 and p35, was recently identified as a new member of the IL-12 cytokine family. The role of IL-35 remains controversial in inflammatory diseases. Some studies have shown its anti-inflammatory properties given its effect on Treg, regulator B cells and associated cytokines in inflammatory bowel disease, collagen-induced arthritis or other autoimmune diseases. In contrast, IL-35 enhanced the inflammatory response in Lyme arthritis and IL-35 gene therapy exacerbates experimental rheumatoid arthritis in mice. The expression and significance of IL-35 in patients with SLE is still obscure. Several studies have shown that the level of IL-35 was decreased and the percentage of CD4 EBI3 T cells were also significantly decreased in patients with active SLE. Another study showed that plasma IL-35 and soluble gp130 levels were significantly higher in patients with severe SLE. We conducted this study to investigate the IL-35 level in patients with SLE and their potential association with clinical manifestations and disease activity. We enrolled 145 patients with SLE (133 females and 12 males, mean age 35.6 13.9 years) and 69 ageand sex-matched healthy volunteers without diseases.

THU0273 Decreased Interleukin-35 Is Associated with Higher Risk of Pregnancy Morbidity in Patients with Antiphospholipid Syndrome

Background Antiphospholipid syndrome (APS) is an autoimmune disease characterized mainly by recurrent pregnancy failures and/or thrombosis 1. Interleukin (IL)-35, a new member of IL-12 family, was demonstrated to play a role in the immunopathogenesis of several autoimmune diseases 2,3. The expression of IL-35 was observed to be abnormal in idiopathic recurrent pregnancy loss 4. Objectives To determine the expression of IL-35 and assess its association with clinical /laboratory features in patients with APS. Methods Serum IL-35 was detected by the enzyme-linked immunosorbent assay (ELISA). 126 patients with APS, 38 patients with primary pregnancy morbidity, and 69 healthy controls were enrolled in the study. Clinical and laboratory features of APS patients were recorded. The Mann-Whitney U test was used to evaluate differences of non-normal distribution data and the Students t test was used for normal distribution data. The differences in the frequency of clinical /laboratory characteristics were assessed by the Chi-square (χ2) test. The relevances between IL-35 and clinical/laboratory features were further analyzed by the Spearman test (measurement data) or Chi-square (χ2) test (qualitative data). Results Serum IL-35 was significantly decreased in APS patients (median 14.73, IQR 0–28.09 pg/ml) and patients with primary pregnancy morbidity (16.00, 10.56–25.00 pg/ml), compared to healthy controls (28.87, 20.16–47.47 pg/ml; all p<0.001). Female APS patients with decreased IL-35 had higher risk of pregnancy morbidity than those with normal IL-35, with an odds ratio (OR) of 2.565 (95% confidence interval (CI) 1.138–5.785, p=0.022), especially events of fetus death ≥ the 10th week (OR=6.545, 95% CI 6.03–21.101; p<0.001). In contrast, the frequency of thrombosis was significantly lower in APS patients with decreased IL-35 and the OR value was 0.382 (0.185- 0.789; p=0.009).Figure 1 Conclusions Serum IL-35 was decreased in APS patients and was associated with pregnancy morbidity and thrombosis, raising the hypothesis of its complicated pathogenic roles in APS. References Miyakis S, Lockshin MD, Atsumi T, et al. International consensus statement on an update of the classification criteria for definite antiphospholipid syndrome (APS). J Thromb Haemost 2006;4:295–306. Niedbala W, Wei XQ, Cai B, et al. IL-35 is a novel cytokine with therapeutic effects against collagen-induced arthritis through the expansion of regulatory T cells and suppression of Th17 cells. Eur J Immunol 2007;37:3021–9. Collison LW, Workman CJ, Kuo TT, et al. The inhibitory cytokine IL-35 contributes to regulatory T-cell function. Nature 2007;450:566–9. Ozkan ZS, Deveci D, Simsek M, et al. What is the impact of SOCS3, IL-35 and IL17 in immune pathogenesis of recurrent pregnancy loss? J Matern Fetal Neonatal Med 2015;28:324–8. Acknowledgement This study was funded by the 973 program of China (No. 2012CB517702) and National Natural Science Foundation of China (No. 81471536). We thank all co-authors for their contribution to this work. We also thank all patients and healthy donors whom took part in this study. Disclosure of Interest None declared