Biomarkers For Rheumatoid Arthritis Research Articles

Cytokine expression and cytokine-based T cell profiling in South Indian rheumatoid arthritis

Rheumatoid arthritis (RA), a chronic inflammatory disease affects up to 1% of the general population. Early diagnosis and treatment are limited by the absence of specific and reliable diagnostic/prognostic biomarkers. This study was carried out in 48 Tamil South Indian RA patients and 49 healthy controls (HC) to identify any cytokine signature(s) that could potentially serve as biomarkers. Expression profiles of Th1, Th2, Th17 and Tregs cell type-specifying cytokines and transcription factors were analyzed using real time quantitative PCR (qPCR) assay. To explore if such expression profiles mirror their steady state plasma levels, a bead-based multiplex fluorescent assay was carried out. We found that the expression of transcription factors T-bet (for Th1), GATA-3 (for Th2) and FoxP3 (for Tregs) were significantly lower in patients than in healthy controls (P<0.0001) similar to lowering of IFNγ (P=0.004) and IL-10 (P=0.04). The transcript levels of IL-12p40 and TNF-α were higher among patients as compared to HC (P<0.0001 and P=0.02, respectively). Circulating levels of assessed cytokines were in general higher in RA patients as compared to controls. These alterations in the expression of transcription factors and cytokines highlight the underlying dysregulation of T cell subsets in RA that reflects a predominantly inflammatory phenotype. Despite dissecting these cellular and molecular processes, no specific signature that could be of diagnostic and/or prognostic value was identified. Additional longitudinal follow-up studies, especially on newly diagnosed treatment-naïve patients are warranted to uncover clinically useful biomarkers of RA.

Therapeutic efficacy of infused molecular hydrogen in saline on rheumatoid arthritis: a randomized, double-blind, placebo-controlled pilot study.

The aim of this study was to demonstrate the safety and efficacy of H2-saline infusion for treatment of rheumatoid arthritis (RA). We conducted a randomized, double-blind, placebo-controlled investigation of the infusion of 1 ppm H2-dissolved saline (H2-saline) in 24 RA patients. Patients were randomized 1:1 to receive 500 ml of either H2-saline or placebo-saline, which was drop infused intravenously (DIV) daily for 5 days. The disease activity score in 28 joints (DAS28) was measured at baseline, immediately post infusion, and after 4 weeks. Therapeutic effects of H2-saline on joint inflammation were estimated by measuring serum biomarkers for RA, tumor necrosis factor-α (TNFα), interleukin-6 (IL-6), matrix metalloproteinase-3 (MMP-3), and urinary 8-hydroxydeoxyguanosine (8-OHdG). In the H2-infused group, average DAS28 decreased from 5.18 ± 1.16 to 4.02 ± 1.25 immediately post infusion and reached 3.74 ± 1.22 after 4 weeks. No significant decrease in DAS28 was observed in the placebo group throughout the study. IL-6 levels in the H2 group significantly decreased in 4 weeks by 37.3 ± 62.0% compared to baseline, whereas it increased by 33.6 ± 34.4% in the placebo group. TNFα levels did not change remarkably in the H2 or placebo groups in 4 weeks post-infusion compared to baseline. The relative ratio of 8-OHdG in the H2 group also significantly decreased by 4.7%. After 4 weeks, MMP3 was significantly reduced by 19.2% ± 24.6% in the H2 group, and increased by 16.9% ± 50.2% in the placebo group. Drop infusion of H2 safely and effectively reduced RA disease activity.

Global metabolite profiling of synovial fluid for the specific diagnosis of rheumatoid arthritis from other inflammatory arthritis.

Currently, reliable biomarkers that can be used to distinguish rheumatoid arthritis (RA) from other inflammatory diseases are unavailable. To find possible distinctive metabolic patterns and biomarker candidates for RA, we performed global metabolite profiling of synovial fluid samples. Synovial fluid samples from 38 patients with RA, ankylosing spondylitis, Behçet's disease, and gout were analyzed by gas chromatography/time-of-flight mass spectrometry (GC/TOF MS). Orthogonal partial least-squares discriminant and hierarchical clustering analyses were performed for the discrimination of RA and non-RA groups. Variable importance for projection values were determined, and the Wilcoxon-Mann-Whitney test and the breakdown and one-way analysis of variance were conducted to identify potential biomarkers for RA. A total of 105 metabolites were identified from synovial fluid samples. The score plot of orthogonal partial least squares discriminant analysis showed significant discrimination between the RA and non-RA groups. The 20 metabolites, including citrulline, succinate, glutamine, octadecanol, isopalmitic acid, and glycerol, were identified as potential biomarkers for RA. These metabolites were found to be associated with the urea and TCA cycles as well as fatty acid and amino acid metabolism. The metabolomic analysis results demonstrated that global metabolite profiling by GC/TOF MS might be a useful tool for the effective diagnosis and further understanding of RA.

AB0351 The Novel Adipokine Chemerin is A Biomarker of Cardiovascular Risk in Patients with Rheumatoid Arthritis

BackgroundWhereas chemerin contributes to the pathophysiology of rheumatoid arthritis (RA) (1), its role in the enhanced cardiovascular disease risk in RA is unknown. In non-RA subjects, dependent on physiological context,...

AB0142 Mir-223 Expression Profile as A Biomarker in Rheumatoid Arthritis

BackgroundRheumatoid arthritis (RA) is associated with altered miRNA expression profile in peripheral blood, synovial fluid and synovial membrane. There are several reports on the role of miR-223 in the disease...

SAT0220 Measurement of Serum Leucine-Rich Alpha-2 Glycoprotein, A Novel Disease Activity Biomarker in Rheumatoid Arthritis, for the Detection of Biologic-Associated Tuberculosis

BackgroundTuberculosis is considered to be a severe complication during biologic therapies in patients with rheumatoid arthritis (RA). Importantly, conventional inflammatory biomarkers such as C-reactive protein (CRP) and erythrocyte sedimentation rate...

AB0262 Seroconversion and Fluctuation of Current and Novel Biomarkers during Disease Course of RA

BackgroundIn rheumatoid arthritis (RA), rheumatoid factor (RF) and antibodies against citrullinated proteins (ACPA) are established diagnostic markers. However, up to one third of the RA patients is not detected using...

OP0177 A High Frequency of N-Glycans in the Acpa-Igg Variable Domain Modulates Reactivity to Citrullinated Antigens

BackgroundAntibodies against citrullinated proteins antigens (ACPA) are the most relevant prognostic and diagnostic biomarker for rheumatoid arthritis (RA). ACPA positive patients are characterized by progressive disease, a high rate of...

THU0226 Evaluation of Serum 14-3-3Eta Levels in A Japanese RA Cohort Treated with Tocilizumab

BackgroundSerum 14-3-3η is a novel protein biomarker for rheumatoid arthritis (RA) and is available for clinical use in the United States, Europe, and Canada. It precedes the onset of RA...

Secreted Semaphorin 5A Activates Immune Effector Cells and Is a Biomarker for Rheumatoid Arthritis

To investigate the role of the multifunctional protein semaphorin 5A (Sema5A) in modulating cellular immune responses and as a biomarker in rheumatoid arthritis (RA). A soluble form of recombinant Sema5A was used to assess its effect on the functions of primary T cells and natural killer (NK) cells isolated from the peripheral blood of healthy donors. Cell proliferation and expression of transcription factors were determined by flow cytometry. Cytokine secretion was analyzed using Luminex technology. Serum samples obtained from 145 patients with RA and control serum samples obtained from healthy individuals or patients with non-RA rheumatic diseases were analyzed for the presence of secreted Sema5A, using enzyme-linked immunosorbent assays and immunoblotting. Soluble Sema5A strongly increased T cell and NK cell proliferation and induced the secretion of proinflammatory Th1/Th17 cytokines. Accordingly, Sema5A stimulation caused significant up-regulation of T-bet and retinoic acid receptor-related orphan nuclear receptor γt levels in T cells. In addition, significantly elevated levels of secreted Sema5A were detected in the serum of patients with RA compared with control serum. Sema5A levels were highest in patients with RA who were positive for the RA biomarker anti-cyclic citrullinated peptide (P < 0.001 versus patients with systemic lupus erythematosus and patients with Sjögren's syndrome) and correlated with the levels of rheumatoid factor. Soluble Sema5A is a potent activator of T cells and NK cells in vitro, and high serum levels of Sema5A are associated with RA. Taken together, the results indicate that Sema5A contributes to the pathogenesis of RA through antigen-independent T cell and NK cell activation. Hence, Sema5A is a promising complementary biomarker for the diagnosis of RA.

Identification of Potential Serum Biomarkers for Rheumatoid Arthritis by High-Resolution Quantitative Proteomic Analysis

The aim of this study was to find serum biomarkers of rheumatoid arthritis (RA) by high-resolution proteomic analysis. Low-abundance proteins from pooling serum sample of early RA patients and healthy controls were enriched using ProteoMiner™ enrichment kits. The enriched proteins were separated on SDS-PAGE, digested by trypsin, labeled with tandem mass tag (TMT) reagents, and desalted by C18 stage tip column. Then, the labeled peptides were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) with nano-LC combined with Orbitrap Q Exactive mass spectrometer, and experiments were carried out three times using different specimens, and differentially expressed proteins were screened by intensity ratios of identified peptides. Enzyme-linked immunosorbent assays (ELISAs) were performed to confirm differentially expressed proteins. Twenty-six proteins were found differentially expressed in RA serum by high-resolution proteomic analysis. Among them, levels of thrombospondin-1, ficolin-2, isoform 10 of fibronectin, and apolipoprotein E were higher in RA patients than in healthy controls (RA/healthy control (HC) ≥ 1.5, p<0.05), while levels of angiotensinogen, paraoxonase/arylesterase 1, isoform E of proteoglycan 4, and plasminogen were significantly lower (RA/HC ≤ 0.67, p<0.05). Further study by ELISA showed a higher level of ficolin-2 in the serum of RA patients compared to healthy controls; the level of ficolin-2 was found to be positively correlated with swollen joint counts (SJCs), disease activity score (DAS28), rheumatoid factor, and IgM in RA patients and with DAS28 and IgM in early RA patients through statistical analysis. The results of this study suggest that ficolin-2, as a newly screened biomarker by high-resolution quantitative proteomic analysis, offers the potentiality to become a diagnostic or disease evaluation tool in RA.

Effects of Lactobacillus casei supplementation on disease activity and inflammatory cytokines in rheumatoid arthritis patients: a randomized double‐blind clinical trial

The present study aimed at investigating the effects of Lactobacillus casei 01 supplementation on symptoms and inflammatory biomarkers of rheumatoid arthritis (RA) in women. In this randomized double-blind clinical trial, female patients with established RA for more than 1 year, 20-80 years of age and body mass index (BMI) lower than 40, who followed stable medication for 3 months prior to the supplementation, were randomly allocated to receive either one capsule containing 10(8) colony forming units (CFU) of L. casei 01, or a placebo for 8 weeks; allocation was stratified by BMI and menopausal status. Disease activity score-28 (DAS28) was calculated, European League Against Rheumatism (EULAR) response was evaluated and the cytokines, interleukin (IL)-1β, IL-6, IL-10, IL-12 and tumor necrosis factor (TNF)-α were measured. Thirty patients were recruited in each group; 22 and 24 patients were analyzed in the probiotic and placebo groups, respectively. L. casei 01 supplementation decreased serum high-sensitivity C-reactive protein (hs-CRP) levels, tender and swollen joint counts, global health (GH) score and DAS28 (P < 0.05). More patients in the L. casei 01 group had moderate response to the treatment, based on the EULAR criteria, at the end of the study (P < 0.01). At the end of the study, a significant difference was observed between the two groups for IL-10, IL-12 and TNF-α changes through the study course (P < 0.05), in favor of the probiotic group. No adverse effects were reported for the intervention. Probiotic supplementation may be an appropriate adjunct therapy for RA patients and help alleviate symptoms and improve inflammatory cytokines.

A127: Validating Popliteal Lymph Node Phenotype and Bin Expansion as Biomarkers of Rheumatoid Arthritis Knee Flare in Clinical Pilot Studies

Background/Purpose:Factors precipitating joint flare in rheumatoid arthritis (RA) patients are poorly understood. Contrast enhanced (CE) MRI studies in murine models have identified popliteal lymph node (PLN) phenotypes that correlate with arthritic flare in the adjacent knee defined by a sudden increase in synovial volume and focal erosions. Prior to flare, the PLN expands in size and CE due to increased lymph volume and accumulation of CD23+CD21hiCD1dhi B cells (Bin). Arthritic flare following PLN collapse is characterized by decreased CE and translocation of Bin to the paracortical sinuses with loss of efferent lymphatic drainage. To validate lymphatic biomarkers in RA, we performed pilot studies to demonstrate the feasibility of detecting expanding and collapsed PLN via CE‐MRI. We analyzed PLN from RA patients undergoing total knee replacement or amputation for ischemic limbs in patients without RA.Methods:RA patients experiencing knee flare had a 3T CE‐MRI prior to initiation of biologic therapy. PLN and synovium volumes were measured with Amira. PLN from RA patients and amputees were harvested, cells were stained and examined by flow cytometry.Results:MRI and 3D volume rendering of a representative flaring knee from early vs. long‐standing RA (Fig. 1) show evidence of expanding PLN (n = 3; 3.57, 1.23, 2.2 cm3) adjacent to highly vascular synovitis (203.40 cm3). In contrast, knee flare in longstanding RA (&gt;30 yrs) presents with collapsed PLN (n = 2; 0.24, 0.16 cm3) adjacent to pannus (159.88 cm3) that contains both bright (vascular) and dark (necrotic) regions on MRI. Flow cytometry confirmed large numbers of Bin‐like cells (CD23hiCD21hi) in RA vs. non‐RA PLN. Clinical endpoints measured by disease activity score (25% improvement) and C‐reactive protein levels (40% decrease) preliminarily show a correlation between effective therapy and CE‐MRI changes. imageConclusion:We present evidence that similar to murine inflammatory arthritis, knee flare in RA is modulated by altered dynamics of draining lymph nodes and lymphatic vessels. We found that the flare in early arthritis is characterized by extensive joint inflammation that appears to exceed the capacity of expanded lymphatics. In contrast, knee flare in long‐standing arthritis is associated with loss of lymphatic drainage, which in mice, is caused by loss of the lymphatic pulse and clogging of efferent lymphatics by Bin cells. Additional studies are warranted to demonstrate the reliability of these findings of knee flare in early vs. chronic RA. Figure 1. Representative images of MRI and corresponding 3D rendering from RA patients with knee flare shows inflamed synovium with bright PLN in the T2 fat‐suppressed MRI (A) and uniform inflammatory tissue and large PLN (B) in early RA. In contrast, note marked focal erosions in the proton density weighted MRI (C) and patchy synovium with collapsed PLN (D) in long‐standing RA.

Updating the OMERACT Filter: Implications for Imaging and Soluble Biomarkers

The Outcome Measures in Rheumatology (OMERACT) Filter provides a framework for the validation of outcome measures for use in rheumatology clinical research. However, imaging and biochemical measures may face additional validation challenges because of their technical nature. The Imaging and Soluble Biomarker Session at OMERACT 11 aimed to provide a guide for the iterative development of an imaging or biochemical measurement instrument so it can be used in therapeutic assessment. A hierarchical structure was proposed, reflecting 3 dimensions needed for validating an imaging or biochemical measurement instrument: outcome domain(s), study setting, and performance of the instrument. Movement along the axes in any dimension reflects increasing validation. For a given test instrument, the 3-axis structure assesses the extent to which the instrument is a validated measure for the chosen domain, whether it assesses a patient-centered or disease-centered variable, and whether its technical performance is adequate in the context of its application. Some currently used imaging and soluble biomarkers for rheumatoid arthritis, spondyloarthritis, and knee osteoarthritis were then evaluated using the original OMERACT Filter and the newly proposed structure. Breakout groups critically reviewed the extent to which the candidate biomarkers complied with the proposed stepwise approach, as a way of examining the utility of the proposed 3-dimensional structure. Although there was a broad acceptance of the value of the proposed structure in general, some areas for improvement were suggested including clarification of criteria for achieving a certain level of validation and how to deal with extension of the structure to areas beyond clinical trials. General support was obtained for a proposed tri-axis structure to assess validation of imaging and soluble biomarkers; nevertheless, additional work is required to better evaluate its place within the OMERACT Filter 2.0.

A5.16 miRNA expression profile in Bulgarian patients with rheumatoid arthritis compared to patients with osteoarthritis and healthy controls in regard to their use as biomarkers in the clinical practice

Background and ObjectivesThe pathogenesis of rheumatoid arthritis (RA) is still unknown. Recent data suggest that micro-ribonucleic acids (microRNAs) and their targets are involved in the inflammatory process and may perpetuate...

A1.45 Hyperglycosylation of ACPA-IGG variable domains modulates reactivity to citrullinated antigens

Background and ObjectivesAutoantibodies specific for citrullinated antigens are highly relevant diagnostic and prognostic biomarkers in rheumatoid arthritis and have been considered to be involved in disease pathogenesis. Previous studies have...

A1.9 Pilot work towards discovery of novel diagnostic biomarker for rheumatoid arthritis using combinations of chemokine receptor expression on peripheral blood cell-subsets

Background and ObjectivesRheumatoid Arthritis (RA) is an autoimmune inflammatory joint disease resulting in joint damage and functional loss. Management is centred on early intervention, proven to prevent disabling joint damage...

Assessment of ischemia-modified albumin levels in patients with rheumatoid arthritis.

Oxygen metabolism has an important role in the pathogenesis of rheumatoid arthritis (RA). Abundant amounts of ROS have been identified in the synovial fluid of RA patients. The accumulation of ROS in cells also serves as an important intracellular signaling of molecules that amplify the synovial inflammatory-proliferative response. Thus, the aim of this study was to assess the IMA levels and other oxidative stress and inflammatory biomarkers in RA subjects. IMA, AOPP, CRP, hemoglobin, Hct, MCV, RF, creatinine, urea levels were assessed in 16 RA subjects and 20 healthy controls. IMA levels were significantly higher in the RA group than in the control group (0.495 +/- 0.01 vs. 0.433 +/- 0.02 ABSU, p = 0.038). No significant differences were observed for the other markers studied. This study demonstrated that RA is related to oxidative stress and inflammation. We also showed for the first time an increase of IMA levels in RA subjects, suggesting that this pathology promotes an increase in the oxidative stress process.

Adipokines as potential biomarkers in rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic systemic inflammatory autoimmune disease characterized by severe joint injury. Recently, research has been focusing on the possible identification of predictor markers of disease onset and/or progression, of joint damage, and of therapeutic response. Recent findings have uncovered the role of white adipose tissue as a pleiotropic organ not only specialized in endocrine functions but also able to control multiple physiopathological processes, including inflammation. Adipokines are a family of soluble mediators secreted by white adipose tissue endowed with a wide spectrum of actions. This review will focus on the recent advances on the role of the adipokine network in the pathogenesis of RA. A particular attention will be devoted to the action of these proteins on RA effector cells, and on the possibility to use circulating levels of adipokines as potential biomarkers of disease activity and therapeutic response.

Cytokines as biomarkers in rheumatoid arthritis.

RA is a complex disease that develops as a series of events often referred to as disease continuum. RA would benefit from novel biomarker development for diagnosis where new biomarkers are still needed (even if progresses have been made with the inclusion of ACPA into the ACR/EULAR 2010 diagnostic criteria) and for prognostic notably in at risk of evolution patients with autoantibody-positive arthralgia. Risk biomarkers for rapid evolution or cardiovascular complications are also highly desirable. Monitoring biomarkers would be useful in predicting relapse. Finally, predictive biomarkers for therapy outcome would allow tailoring therapy to the individual. Increasing numbers of cytokines have been involved in RA pathology. Many have the potential as biomarkers in RA especially as their clinical utility is already established in other diseases and could be easily transferable to rheumatology. We will review the current knowledge's relation to cytokine used as biomarker in RA. However, given the complexity and heterogeneous nature of RA, it is unlikely that a single cytokine may provide sufficient discrimination; therefore multiple biomarker signatures may represent more realistic approach for the future of personalised medicine in RA.