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• Extra-articular manifestations in rheumatoid arthritis appear within the first 5 - years of disease. • After 15 years of disease, one of five patients have experienced extra-articular manifestations. • Extra-articular manifestations are associated with seropositivity as well as disease-modifying treatment. • Extra-articular manifestations are a clinical reality also in the era of modern treatment strategies. • The patients with Extra-articular manifestations had a 3-fold increase in mortality. Rheumatoid arthritis [RA) is a chronic inflammatory disease, with potential for extra-articular manifestations (ExRA). The incidence and predisposing factors for ExRA and the mortality were evaluated in an early RA inception cohort. Patients (n = 1468; 69 % females, mean age (SD) 57.3(16.3) years) were consecutively included at the date of diagnosis, between 1 January 1996 and 31 December 2016, and assessed prospectively. In December 2016 development of ExRA was evaluated by a patient questionnaire and a review of medical records. Cumulative incidence and incidence rates were compared between 5-year periods and between patients included before and after 1 January 2001. Cox proportional hazard regression models were used to identify predictors for ExRA, and models with ExRA as time-dependent variables to estimate the mortality. After a mean (SD) follow-up of 9.3(4.9) years, 238 cases (23.3 %) had ExRA and 151 (14.7 %) had ExRA without rheumatoid nodules. Most ExRA developed within 5 years from diagnosis. Rheumatoid nodules (10.5 %) and keratoconjunctivitis sicca (7.1 %) were the most frequent manifestations, followed by pulmonary fibrosis (6.1 %). The ExRA incidence among more recently diagnosed patients was similar as to the incidence among patients diagnosed before 2001. Seropositivity, smoking and early biological treatment were associated with development of ExRA. After 15 years 20 % had experienced ExRA. ExRA was associated with increased mortality, HR 3.029 (95 % CI 2.177–4.213). Early development of ExRA is frequent, particularly rheumatoid nodules. Predisposing factors were age, RF positivity, smoking and early biological treatment. The patients with ExRA had a 3-fold increase in mortality. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a chronic autoimmune condition frequently found in rheumatological patients that sometimes raises diagnosis and management problems. The pathogenesis of the disease is complex and involves the activation of many cells and intracellular signaling pathways, ultimately leading to the activation of the innate and acquired immune system and producing extensive tissue damage. Along with joint involvement, RA can have numerous extra-articular manifestations (EAMs), among which lung damage, especially interstitial lung disease (ILD), negatively influences the evolution and survival of these patients. Although there are more and more RA-ILD cases, the pathogenesis is incompletely understood. In terms of genetic predisposition, external environmental factors act and subsequently determine the activation of immune system cells such as macrophages, neutrophils, B and T lymphocytes, fibroblasts, and dendritic cells. These, in turn, show the ability to secrete molecules with a proinflammatory role (cytokines, chemokines, growth factors) that will produce important visceral injuries, including pulmonary changes. Currently, there is new evidence that supports the initiation of the systemic immune response at the level of pulmonary mucosa where the citrullination process occurs, whereby the autoantibodies subsequently migrate from the lung to the synovial membrane. The aim of this paper is to provide current data regarding the pathogenesis of RA-associated ILD, starting from environmental triggers and reaching the cellular, humoral, and molecular changes involved in the onset of the disease. [ABSTRACT FROM AUTHOR]

Objectives Autoantibody-negative RA differs from autoantibody-positive RA in several clinical aspects, possibly underpinned by pathogenetic differences. At present, the role of adaptive immune responses in autoantibody-negative RA remains unclear. Here, we investigated the synovial and serum immunophenotype indicative of B lymphocyte involvement across the spectrum of autoantibody-positive and -negative chronic arthritides. Methods Ultrasound-guided synovial biopsies were retrieved from 131 patients: 43 autoantibody-positive RA, 35 autoantibody-negative RA, 25 polyarticular PsA and 28 oligoarticular PsA. Samples were analysed for the degree of histological inflammation, B lymphocyte infiltration and the distribution of different pathotypes (lympho-myeloid, myeloid, pauci-immune). Serum levels of the B cell chemoattractant CXCL13 were compared among groups. Results Synovitis scores and CD68+ sublining macrophage infiltration were comparable irrespective of clinical diagnosis and disease subtype. In contrast, the degree of B lymphocyte infiltration and the frequency of lympho-myeloid synovitis in autoantibody-negative RA were lower than those of autoantibody-positive RA (mean [ s. d.] 1.8 [1] vs 2.4 [0.6], P  = 0.03, and 38.2% vs 62.9%, P  = 0.07, respectively), and similar to polyarticular PsA. Oligoarticular PsA had the lowest B cell scores. Serum CXCL13 was associated with lympho-myeloid synovitis and followed a similar gradient, with the highest levels in autoantibody-positive RA, intermediate and comparable levels in autoantibody-negative RA and polyarticular PsA, and low levels in oligoarticular PsA. Conclusions The synovial and serum immunophenotype indicative of B lymphocyte involvement in autoantibody-negative RA differs from that of autoantibody-positive RA and more closely resembles that observed in polyarticular PsA. The pathobiological stratification of chronic inflammatory arthritides beyond clinical diagnosis may fuel personalized treatment strategies. [ABSTRACT FROM AUTHOR]

Patients with rheumatoid arthritis (RA) have altered levels of exhaled nitric oxide (NO) compared with healthy controls. Here, we investigated whether the clinical features of and immunological factors in RA pathogenesis could be linked to the NO lung dynamics in early disease. A total of 44 patients with early RA and anti-citrullinated peptide antibodies (ACPAs), specified as cyclic citrullinated peptide 2 (CCP2), were included. Their exhaled NO levels were measured, and the alveolar concentration, the airway compartment diffusing capacity and the airway wall concentration of NO were estimated using the Högman–Meriläinen algorithm. The disease activity was measured using the Disease Activity Score for 28 joints. Serum samples were analysed for anti-CCP2, rheumatoid factor, free secretory component, secretory component containing ACPAs, antibodies against Porphyromonas gingivalis (Rgp) and total levels of IgA, IgA1 and IgA2. Significant negative correlations were found between the airway wall concentration of NO and the number of swollen joints (Rho −0.48, p = 0.004), between the airway wall concentration of NO and IgA rheumatoid factor (Rho −0.41, p = 0.017), between the alveolar concentration and free secretory component (Rho −0.35, p = 0.023) and between the alveolar concentration and C-reactive protein (Rho −0.36, p = 0.016), but none were found for anti-CCP2, IgM rheumatoid factor or the anti-Rgp levels. In conclusion, altered NO levels, particularly its production in the airway walls, may have a role in the pathogenesis of ACPA-positive RA. [ABSTRACT FROM AUTHOR]

Objective Obesity conveys a risk for RA development, while paradoxically, associating with less radiographic progression after RA diagnosis. Using MRI we can study this surprising association in detail from MRI-detected synovitis and osteitis to MRI-detected erosive progression, which precedes radiographic progression. Previous research suggested obesity associates with less osteitis and synovitis. We therefore aimed to (i) validate the previously suggested association between BMI and MRI-detected osteitis/synovitis; (ii) study whether this is specific for ACPA-positive or ACPA-negative RA or also present in other arthritides; (iii) study whether MRI-detected osteitis associates with MRI-detected erosive progression; and (iv) study whether obesity associates with MRI-detected erosive progression. Methods We studied 1029 early arthritis patients (454 RA, 575 other arthritides), consecutively included in Leiden Early Arthritis Clinic. At baseline patients underwent hand-and-foot MRI that were RAMRIS-scored, and 149 RA patients underwent follow-up MRIs. We studied associations between baseline BMI and MRI-detected osteitis/synovitis (using linear regression), and erosive progression (using Poisson mixed models). Results In RA, higher BMI associated with less osteitis at disease onset (β = 0.94; 95% CI: 0.93, 0.96) but not with synovitis. Higher BMI associated with less osteitis in ACPA-positive RA (β = 0.95; 95% CI: 0.93, 0.97), ACPA-negative RA (β = 0.97; 95% CI: 0.95, 0.99) and other arthritides (β = 0.98; 95% CI: 0.96, 0.99). Over 2 years, overweight and obesity associated with less MRI-detected erosive progression (P  = 0.02 and 0.03, respectively). Osteitis also associated with erosive progression over 2 years (P  < 0.001). Conclusions High BMI relates to less osteitis at disease onset, which is not confined to RA. Within RA, high BMI and less osteitis associated with less MRI-detected erosive progression. This suggests that the protective effect of obesity on radiographic progression is exerted via a path of less osteitis and subsequently fewer MRI-detected erosions. [ABSTRACT FROM AUTHOR]

Purpose of Review: Host-microbiome interactions have been implicated in the pathophysiology of rheumatoid arthritis (RA), but the data linking specific microbes to RA is largely associative. Here, we review recent studies that have interrogated specific mechanistic links between microbes and host in the setting of RA. Recent Findings: Several candidate bacterial species and antigens that may trigger the conversion of an anti-bacterial to an autoimmune response have been recently identified. Additional studies have identified microbial metabolic pathways that are altered in RA. Some of these microbial species and metabolic pathways have been validated in mouse models to induce RA-like immune responses, providing initial evidence of specific mechanisms by which the microbiota contributes to the development of RA. Summary: Several microbial species, antigens, and metabolites have been identified as potential contributors to RA pathophysiology. Further interrogation and validation of these pathways may identify novel biomarkers of or therapeutic avenues for RA. [ABSTRACT FROM AUTHOR]

Background: SARS-CoV-2 vaccination in rheumatoid arthritis (RA) patients treated with B cell-depleting drugs induced limited seroconversion but robust cellular response. We aimed to document specific T and B cell immunity in response to vaccine booster doses and breakthrough infection (BTI). Methods: We included 76 RA patients treated with rituximab who received up to four SARS-CoV-2 vaccine doses or three doses plus BTI, in addition to vaccinated healthy donors (HD) and control patients treated with tumor necrosis factor inhibitor (TNFi). We quantified anti-SARS-CoV-2 receptor-binding domain (RBD) Spike IgG, anti-nucleocapsid (NC) IgG, 92 circulating inflammatory proteins, Spike-binding B cells, and Spike-specific T cells along with comprehensive high-dimensional phenotyping and functional assays. Findings: The time since the last rituximab infusion, persistent inflammation, and age were associated with the anti-SARS-CoV-2 RBD IgG seroconversion. The vaccine-elicited serological response was accompanied by an incomplete induction of peripheral Spike-specific memory B cells but occurred independently of T cell responses. Vaccine- and BTI-elicited cellular immunity was similar between RA and HD ex vivo in terms of frequency or phenotype of Spike-specific cytotoxic T cells and in vitro in terms of the functionality and differentiation profile of Spike-specific T cells. Interpretation: SARS-CoV-2 vaccination in RA can induce persistent effector T-cell responses that are reactivated by BTI. Paused rituximab medication allowed serological responses after a booster dose (D4), especially in RA with lower inflammation, enabling efficient humoral and cellular immunity after BTI, and contributed overall to the development of potential durable immunity. [ABSTRACT FROM AUTHOR]

Objective Sialylation of the crystallizable fragment (Fc) of ACPAs, which is catalysed by β-galactoside α-2,6-sialyltransferase 1 (ST6GAL1) could attenuate inflammation of RA. In this study, we screened the transcription factor of ST6GAL1 and elucidated the mechanism of transcriptionally upregulating sialylation of ACPAs in B cells to explore its role in the progression of RA. Methods Transcription factors interacting with the P2 promoter of ST6GAL1 were screened by DNA pull-down and liquid chromatography with tandem mass spectrometry (LC-MS/MS), and verified by chromatin immunoprecipitation (ChIP), dual luciferase reporter assay and electrophoretic mobility shift assay (EMSA). The function of the CCCTC-binding factor (CTCF) on the expression of ST6GAL1 and the inflammatory effect of ACPAs were verified by knocking down and overexpressing CTCF in B cells. The CIA model was constructed from B cell–specific CTCF knockout mice to explore the effect of CTCF on arthritis progression. Results We observed that the levels of ST6GAL1 and ACPAs sialylation decreased in the serum of RA patients and were negatively correlated with DAS28 scores. Subsequently, CTCF was screened and verified as the transcription factor interacting with the P2 promoter of ST6GAL1, which enhances the sialylation of ACPAs, thus weakening the inflammatory activity of ACPAs. Furthermore, the above results were also verified in the CIA model constructed from B cell–specific CTCF knockout mice. Conclusion CCCTC-binding factor is the specific transcription factor of β-galactoside α-2,6-sialyltransferase 1 in B cells that upregulates the sialylation of ACPAs in RA and attenuates the disease progression. [ABSTRACT FROM AUTHOR]

Objectives To determine whether an expanded antigen-specific ACPA profile predicts changes in disease activity in patients with RA initiating biologics. Methods The study included participants from a prospective, non-randomized, observational RA cohort. For this sub-study, treatment groups of interest included biologic-naïve initiating anti-TNF, biologic-exposed initiating non-TNF, and biologic-naïve initiating abatacept. ACPAs to 25 citrullinated peptides were measured using banked enrolment serum. Principal component analysis (PCA) was performed and associations of resulting principal component (PC) scores (in quartiles) and anti-CCP3 antibody (≤15, 16–250 or >250 U/ml) with EULAR (good/moderate/none) treatment response at 6 months were examined using adjusted ordinal regression models. Results Participants (n  = 1092) had a mean age of 57 (13) years and 79% were women. At 6 months, 68.5% achieved a moderate/good EULAR response. There were three PCs that cumulatively explained 70% of variation in ACPA values. In models including the three components and anti-CCP3 antibody category, only PC1 and PC2 were associated with treatment response. The highest quartile for PC1 (odds ratio [OR] 1.76; 95% CI: 1.22, 2.53) and for PC2 (OR 1.74; 95% CI: 1.23, 2.46) were associated with treatment response after multivariable adjustment. There was no evidence of interaction between PCs and treatment group in EULAR responses (P -value for interaction >0.1). Conclusion An expanded ACPA profile appears to be more strongly associated with biologic treatment response in RA than commercially available anti-CCP3 antibody levels. However, further enhancements to PCA will be needed to effectively prioritize between different biologics available for the treatment of RA. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a chronic inflammatory rheumatic disease with an autoimmune component that preferentially affects the small joints of the hands and feet. Extra-articular manifestations may be associated with it. These are rather long-term complications compared to the inaugural forms of the disease.

Copyright of Revista Latinoamericana de Hipertension is the property of Revista Latinoamericana de Hipertension and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Revista Latinoamericana de Hipertension is the property of Revista Latinoamericana de Hipertension and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Revista Latinoamericana de Hipertension is the property of Revista Latinoamericana de Hipertension and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Objectives The variable region of most ACPA IgG molecules in the serum of RA patients carries N -glycan (N -glycanV). To analyse the pathogenicity of N -glycanV of ACPAs, we analysed the pathogenicity of a monoclonal ACPA, CCP-Ab1, with or without N -glycanV, which had been isolated from a patient with RA. Methods CCP-Ab1 with no N -glycosylation site in the variable region (CCP-Ab1 N-rev) was generated, and antigen binding, the effect on in vitro differentiation of osteoclasts from bone marrow mononuclear cells of autoimmune arthritis–prone SKG mice (the cell size of TRAP+ cells and bone resorption capacity) and the in vivo effect on the onset or exacerbation of autoimmune arthritis in SKG mice were evaluated in comparison with glycosylated CCP-Ab1. Results Amino acid residues in citrullinated peptide (cfc1), which are essential for binding to CCP-Ab1 N-rev and original CCP-Ab1, were almost identical. The size of TRAP+ cells was significantly larger and osteoclast bone resorption capacity was enhanced in the presence of CCP-Ab1, but not with CCP-Ab1 N-rev. This enhancing activity required the sialic acid of the N -glycan and Fc region of CCP-Ab1. CCP-Ab1, but not CCP-Ab1 N-rev, induced the exacerbation of experimental arthritis in the SKG mouse model. Conclusions These data showed that N -glycanV was required for promoting osteoclast differentiation and bone resorption activity in both in vitro and in vivo assays. The present study demonstrated the important role of N -glycanV in the exacerbation of experimental arthritis by ACPAs. [ABSTRACT FROM AUTHOR]

Difficult-to-treat Rheumatoid Arthritis (RA-D2T) is a condition in which patients do not achieve the treatment target despite multiple advanced therapies, more others features. Aims: to estimate the frequency of RA-D2T in a cohort comprehensively evaluated (clinical, serology, imaging), and to analyze the associated characteristics. In a second part, the frequency of RA-D2T after 1 year of follow-up, analyzing the predictive variables at baseline and therapeutic behavior. Cross-sectional and prospective study, consecutive RA were included, then those who completed the one-year follow-up were evaluated. RA-D2T frequency was estimated (DAS28-CDAI-SDAI-Ultrasonography (US)-HAQ) at baseline and 1 year. The variables associated and those baseline predictive characteristics of D2T at 1 year, and their independent association by logistic regression were analyzed. The treatment approach was described. Two hundred seventy-six patients completed the evaluation, frequency of RA-D2T (all scores): 27.5%. Anemia, RF high titers and higher HAQ score were independent associated. At year, 125 competed follow-up. RA-D2T (all scores): 33%, D2T-US and D2T-HAQ were 14 and 18.4% (p 0.001). Predictive baseline characteristics D2T (all score): ACPA + (OR: 13.7) and X-ray erosion (OR: 2.9). D2T-US: X-ray erosion (OR: 19.7). Conventional DMARDs, corticosteroids and TNF-blockers were the drugs most used by D2T patients, Jaki were the most used in the switch. We showed different frequencies of RA-D2T according to different objective parameters (scores, images) and their association with patient characteristics. In turn, predictive variables (erosions—ACPA) for RA-D2T at 1 year were analyzed. It was shown that the Jaki were the most used drug in these patients. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a chronic autoimmune condition frequently found in rheumatological patients that sometimes raises diagnosis and management problems. The pathogenesis of the disease is complex and involves the activation of many cells and intracellular signaling pathways, ultimately leading to the activation of the innate and acquired immune system and producing extensive tissue damage. Along with joint involvement, RA can have numerous extra-articular manifestations (EAMs), among which lung damage, especially interstitial lung disease (ILD), negatively influences the evolution and survival of these patients. Although there are more and more RA-ILD cases, the pathogenesis is incompletely understood. In terms of genetic predisposition, external environmental factors act and subsequently determine the activation of immune system cells such as macrophages, neutrophils, B and T lymphocytes, fibroblasts, and dendritic cells. These, in turn, show the ability to secrete molecules with a proinflammatory role (cytokines, chemokines, growth factors) that will produce important visceral injuries, including pulmonary changes. Currently, there is new evidence that supports the initiation of the systemic immune response at the level of pulmonary mucosa where the citrullination process occurs, whereby the autoantibodies subsequently migrate from the lung to the synovial membrane. The aim of this paper is to provide current data regarding the pathogenesis of RA-associated ILD, starting from environmental triggers and reaching the cellular, humoral, and molecular changes involved in the onset of the disease.

Patients with rheumatoid arthritis (RA) have altered levels of exhaled nitric oxide (NO) compared with healthy controls. Here, we investigated whether the clinical features of and immunological factors in RA pathogenesis could be linked to the NO lung dynamics in early disease. A total of 44 patients with early RA and anti-citrullinated peptide antibodies (ACPAs), specified as cyclic citrullinated peptide 2 (CCP2), were included. Their exhaled NO levels were measured, and the alveolar concentration, the airway compartment diffusing capacity and the airway wall concentration of NO were estimated using the Högman–Meriläinen algorithm. The disease activity was measured using the Disease Activity Score for 28 joints. Serum samples were analysed for anti-CCP2, rheumatoid factor, free secretory component, secretory component containing ACPAs, antibodies against Porphyromonas gingivalis (Rgp) and total levels of IgA, IgA1 and IgA2. Significant negative correlations were found between the airway wall concentration of NO and the number of swollen joints (Rho −0.48, p = 0.004), between the airway wall concentration of NO and IgA rheumatoid factor (Rho −0.41, p = 0.017), between the alveolar concentration and free secretory component (Rho −0.35, p = 0.023) and between the alveolar concentration and C-reactive protein (Rho −0.36, p = 0.016), but none were found for anti-CCP2, IgM rheumatoid factor or the anti-Rgp levels. In conclusion, altered NO levels, particularly its production in the airway walls, may have a role in the pathogenesis of ACPA-positive RA.

Abstract Introduction Clinical trial findings may not be generalizable to routine practice. This study evaluated sarilumab effectiveness in patients with rheumatoid arthritis (RA) and tested the real-world applicability of a response prediction rule, derived from trial data using machine learning (based on C-reactive protein [CRP] > 12.3 mg/l and seropositivity [anticyclic citrullinated peptide antibodies, ACPA +]). Methods Sarilumab initiators from the ACR-RISE Registry, with ≥ 1 prescription on/after its FDA approval (2017–2020), were divided into three cohorts based on progressively restrictive criteria: Cohort A (had active disease), Cohort B (met eligibility criteria of a phase 3 trial in RA patients with inadequate response/intolerance to tumor necrosis factor inhibitors [TNFi]), and Cohort C (characteristics matched to the phase 3 trial baseline). Mean changes in Clinical Disease Activity Index (CDAI) and Routine Assessment of Patient Index Data 3 (RAPID3) were evaluated at 6 and 12 months. In a separate cohort, predictive rule was tested based on CRP levels and seropositive status (ACPA and/or rheumatoid factor); patients were categorized into rule-positive (seropositive with CRP > 12.3 mg/l) and rule-negative groups to compare the odds of achieving CDAI low disease activity (LDA)/remission and minimal clinically important difference (MCID) over 24 weeks. Results Among sarilumab initiators (N = 2949), treatment effectiveness was noted across cohorts, with greater improvement noted for Cohort C at 6 and 12 months. Among the predictive rule cohort (N = 205), rule-positive (vs. rule-negative) patients were more likely to reach LDA (odds ratio: 1.5 [0.7, 3.2]) and MCID (1.1 [0.5, 2.4]). Sensitivity analyses (CRP > 5 mg/l) showed better response to sarilumab in rule-positive patients. Conclusions In real-world setting, sarilumab demonstrated treatment effectiveness, with greater improvements in the most selective population, mirroring phase 3 TNFi-refractory and rule-positive RA patients. Seropositivity appeared a stronger driver for treatment response than CRP, although optimization of the rule in routine practice requires further data.

Introduction: Clinical trial findings may not be generalizable to routine practice. This study evaluated sarilumab effectiveness in patients with rheumatoid arthritis (RA) and tested the real-world applicability of a response prediction rule, derived from trial data using machine learning (based on C-reactive protein [CRP] > 12.3 mg/l and seropositivity [anticyclic citrullinated peptide antibodies, ACPA +]). Methods: Sarilumab initiators from the ACR-RISE Registry, with ≥ 1 prescription on/after its FDA approval (2017–2020), were divided into three cohorts based on progressively restrictive criteria: Cohort A (had active disease), Cohort B (met eligibility criteria of a phase 3 trial in RA patients with inadequate response/intolerance to tumor necrosis factor inhibitors [TNFi]), and Cohort C (characteristics matched to the phase 3 trial baseline). Mean changes in Clinical Disease Activity Index (CDAI) and Routine Assessment of Patient Index Data 3 (RAPID3) were evaluated at 6 and 12 months. In a separate cohort, predictive rule was tested based on CRP levels and seropositive status (ACPA and/or rheumatoid factor); patients were categorized into rule-positive (seropositive with CRP > 12.3 mg/l) and rule-negative groups to compare the odds of achieving CDAI low disease activity (LDA)/remission and minimal clinically important difference (MCID) over 24 weeks. Results: Among sarilumab initiators (N = 2949), treatment effectiveness was noted across cohorts, with greater improvement noted for Cohort C at 6 and 12 months. Among the predictive rule cohort (N = 205), rule-positive (vs. rule-negative) patients were more likely to reach LDA (odds ratio: 1.5 [0.7, 3.2]) and MCID (1.1 [0.5, 2.4]). Sensitivity analyses (CRP > 5 mg/l) showed better response to sarilumab in rule-positive patients. Conclusions: In real-world setting, sarilumab demonstrated treatment effectiveness, with greater improvements in the most selective population, mirroring phase 3 TNFi-refractory and rule-positive RA patients. Seropositivity appeared a stronger driver for treatment response than CRP, although optimization of the rule in routine practice requires further data. Plain Language Summary: Rheumatoid arthritis (RA) is a condition that may cause joint damage, if untreated. Sarilumab is an advanced medication, approved for treating moderate-to-severe RA in patients not responding to initial standard medicines. Clinical trials have shown that sarilumab improves RA symptoms; however, some people do not respond. This is a common problem in RA treatment. Physicians measure proteins in people's blood (called biomarkers; e.g., anticyclic citrullinated peptide antibodies [ACPA], C-reactive protein [CRP], and rheumatoid factor [RF]) to predict a medicine's response. A previous study showed that people with positive blood tests for ACPA and CRP (> 12.3 mg/l) responded well to sarilumab; this study was based on machine learning (a branch of science using computers) and identified factors that could be linked to treatment benefits. The present study analyzed routine data of 2949 people from the ACR-RISE Registry and showed an improvement in RA symptoms after 6 and 12 months of sarilumab, with a greater improvement noted in patients previously treated with other medicines. Biomarkers were tested in 205 people to check whether they could predict treatment response in day-to-day life. People were called rule-positive if they tested positive for RF and/or ACPA with CRP > 12.3 mg/l, and otherwise rule-negative. After 24 weeks of treatment, rule-positive people had a greater chance of disease improvement than rule-negative people. These results showed the benefits of sarilumab in RA in routine care and suggested the usefulness of machine learning in identifying biomarkers that physicians can use to make treatment decisions. [ABSTRACT FROM AUTHOR]

Background: Dysregulated complement activation, increased protein citrullination, and production of autoantibodies against citrullinated proteins are hallmarks of rheumatoid arthritis (RA). Citrullination is induced by immune cell-derived peptidyl-Arg deiminases (PADs), which are overactivated in the inflamed synovium. We characterized the effect of PAD2- and PAD4-induced citrullination on the ability of the plasma-derived serpin C1-inhibitor (C1-INH) to inhibit complement and contact system activation. Methods: Citrullination of the C1-INH was confirmed by ELISA and Western blotting using a biotinylated phenylglyoxal probe. C1-INH-mediated inhibition of complement activation was analyzed by C1-esterase activity assay. Downstream inhibition of complement was studied by C4b deposition on heat-aggregated IgGs by ELISA, using pooled normal human serum as a complement source. Inhibition of the contact system was investigated by chromogenic activity assays for factor XIIa, plasma kallikrein, and factor XIa. In addition, autoantibody reactivity to native and citrullinated C1-INH was measured by ELISA in 101 RA patient samples. Results: C1-INH was efficiently citrullinated by PAD2 and PAD4. Citrullinated C1- INH was not able to bind the serine protease C1s and inhibit its activity. Citrullination of the C1-INH abrogated its ability to dissociate the C1-complex and thus inhibit complement activation. Consequently, citrullinated C1-INH had a decreased capacity to inhibit C4b deposition via the classical and lectin pathways. The inhibitory effect of C1-INH on the contact system components factor XIIa, plasma kallikrein, and factor XIa was also strongly reduced by citrullination. In RA patient samples, autoantibody binding to PAD2- and PAD4- citrullinated C1-INH was detected. Significantly more binding was observed in anti-citrullinated protein antibody (ACPA)-positive than in ACPA-negative samples. Conclusion: Citrullination of the C1-INH by recombinant human PAD2 and PAD4 enzymes impaired its ability to inhibit the complement and contact systems in vitro. Citrullination seems to render C1-INH more immunogenic, and citrullinated C1-INH might thus be an additional target of the autoantibody response observed in RA patients. [ABSTRACT FROM AUTHOR]

Objectives RA is a chronic inflammatory disease in which possible interstitial lung disease (ILD) is an extra-articular manifestation that carries significant morbidity and mortality. RF and ACPA are included in the RA classification criteria but prognostic and diagnostic biomarkers for disease endotyping and RA-ILD are lacking. Anti-protein arginine deiminase antibodies (anti-PAD) are a novel class of autoantibodies identified in RA. This study aimed to assess clinical features, ACPA and anti-PAD antibodies in RA patients with articular involvement and ILD. Methods We retrospectively collected joint erosions, space narrowing, clinical features and lung involvement of a cohort of 71 patients fulfilling the 2010 ACR/EULAR RA classification criteria. Serum samples from these patients were tested for ACPA IgG (QUANTA Flash CCP3), and anti-PAD3 and anti-PAD4 IgG, measured with novel assays based on a particle-based multi-analyte technology (PMAT). Results Anti-PAD4 antibodies were significantly associated with radiographic injury (P  = 0.027) and erosions (P  = 0.02). Similarly, ACPA levels were associated with erosive disease (P  = 0.014). Anti-PAD3/4 double-positive patients displayed more joint erosions than patients with anti-PAD4 antibodies only or negative for both (P  = 0.014 and P  = 0.037, respectively). RA-ILD (15.5%, 11/71 patients) was associated with older age (P  < 0.001), shorter disease duration (P  = 0.045) and less erosive disease (P  = 0.0063). ACPA were elevated in RA-ILD, while anti-PAD4 were negatively associated (P  = 0.043). Conclusion Anti-PAD4 and anti-PAD3 antibodies identify RA patients with higher radiographic injury and bone erosions. In our cohort, ILD is associated with lower radiographic and erosive damage, as well as low levels of anti-PAD4 antibodies. [ABSTRACT FROM AUTHOR]

Abstract Synthetic and natural cannabinoid derivatives are highly investigated as drug candidates due to their antinociceptive, antiepileptic and anticancer potential. Arachidonoylcyclopropylamide (ACPA) is a synthetic cannabinoid with antiproliferative and apoptotic effects on non-small cell lung cancer and pancreatic and endometrial carcinoma. Thus, ACPA has a great potential for being used as an anticancer drug for epithelial cancers. Therefore, determining the levels of ACPA in biological fluids, cells, tissues and pharmaceutical dosage forms is crucial in monitoring the effects of various pharmacological, physiological and pathological stimuli on biological systems. However, the challenge in the quantification of ACPA is its short half-life and lack of UV signal. Therefore, we developed a liquid chromatography-tandem mass spectrometric (LC–MS/MS) method for sensitive and selective quantification of ACPA in cell culture medium and intracellular matrix. Multiple reaction monitoring in the positive ionization mode was used for detection with 344 → 203 m/z transitions. The separation of ACPA was performed on C18 column (50 × 3.0 mm, 2.1 μm) with the mobile phase run in the gradient mode with 0.1% formic acid (FA) in water and 0.1% FA in acetonitrile at a flow rate of 0.3 ml/min. The assay was linear in the concentration range of 1.8–1000 ng/mL (r = 0.999). The validation studies revealed that the method was linear, sensitive, accurate, precise, selective, repeatable, robust and rugged. Finally, the developed method was applied to quantify ACPA in cell culture medium and intracellular matrix.

ObjectivesPeriodontitis and underlying bacteria have been linked to the development of rheumatoid arthritis (RA). One suggested pathogen is Aggregatibacter actinomycetemcomitans (A.a.), which expresses leukotoxin A (LtxA) that can citrullinate human proteins, providing a possible trigger for the production of anti-citrullinated protein antibodies (ACPA). In this study, we seek to determine the presence of antibodies toward LtxA in patients at risk of developing RA.MethodsTwo prospective observational patient cohorts (one Swedish and one British) with symptomatic at-risk patients were studied. Anti-LtxA antibodies were analyzed by a cell-based neutralization assay in baseline serum and compared to 100 Swedish blood donors that served as controls.ResultsSerum anti-LtxA levels or positivity did not differ between patients and blood donors. In the British cohort, anti-LtxA was more prevalent among ACPA-positive arthralgia patients compared with ACPA-negative arthralgia cases (24% vs. 13%, p < 0.0001). In the Swedish at-risk cohort, anti-LtxA positive patients were at increased risk of progression to arthritis (hazard ratio (HR) 2.10, 95% CI 1.04–4.20), but this was not confirmed in the UK at-risk cohort (HR 0.99, CI 0.60–1.65).ConclusionSerum anti-LtxA is not elevated before RA diagnosis, and associations with disease progression and ACPA levels differ between populations. Other features of the oral microbiome should be explored in upcoming periodontitis-related RA research.

Objectives To investigate the expression of citrullinated and carbamylated proteins in extracellular microvesicles (EMVs) from RA patients. Methods We enrolled 24 RA naïve for biological therapy and 20 healthy donors (HD), matched for age and sex. For each patient, laboratory and clinical data were recorded and clinical indexes were measured (Clinical Disease Activity Index, Simplified Disease Activity Index, DAS28). EMVs in RA patients and HD were purified from plasma and measured by nanoparticle tracking analysis (NanoSight). Further, EMVs were incubated with anti-citrullinated/carbamylated proteins antibodies and processed by flow cytometry and western blot to evaluate the expression of citrullinated/carbamylated antigens. Results NanoSight revealed a significant increase of EMVs in RA compared with HD. Moreover, cytofluorimetric analysis showed a significative higher expression of citrullinated antigens on EMVs' surface in RA than donors, while no substantial difference was found in the expression of carbamylated antigens. These data were confirmed by western blot which identified vimentin, glycolytic enzyme alpha-enolase 1 and collagen type II as the main citrullinated and carbamylated proteins carried by EMVs. Finally, a relevant correlation between the expression of citrullinated antigens and disease activity was found. Conclusions The results of this study suggest an involvement of EMVs in the pathogenesis of RA by inducing autoimmunity. [ABSTRACT FROM AUTHOR]

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Background: Why the adaptive immune system turns against citrullinated antigens in rheumatoid arthritis (RA) and whether anti-citrullinated protein antibodies (ACPAs) contribute to pathogenesis are questions that have triggered intense research, but still are not fully answered. Neutrophils may be crucial in this context, both as sources of citrullinated antigens and also as targets of ACPAs. To better understand how ACPAs and neutrophils contribute to RA, we studied the reactivity of a broad spectrum of RA patient-derived ACPA clones to activated or resting neutrophils, and we also compared neutrophil binding using polyclonal ACPAs from different patients. Methods: Neutrophils were activated by Ca2+ ionophore, PMA, nigericin, zymosan or IL-8, and ACPA binding was studied using flow cytometry and confocal microscopy. The roles of PAD2 and PAD4 were studied using PAD-deficient mice or the PAD4 inhibitor BMS-P5. Results: ACPAs broadly targeted NET-like structures, but did not bind to intact cells or influence NETosis. We observed high clonal diversity in ACPA binding to neutrophil-derived antigens. PAD2 was dispensable, but most ACPA clones required PAD4 for neutrophil binding. Using ACPA preparations from different patients, we observed high patient-to-patient variability in targeting neutrophil-derived antigens and similarly in another cellular effect of ACPAs, the stimulation of osteoclast differentiation. Conclusions: Neutrophils can be important sources of citrullinated antigens under conditions that lead to PAD4 activation, NETosis and the extrusion of intracellular material. A substantial clonal diversity in targeting neutrophils and a high variability among individuals in neutrophil binding and osteoclast stimulation suggest that ACPAs may influence RA-related symptoms with high patient-to-patient variability. [ABSTRACT FROM AUTHOR]

Citrullination is catalyzed by the peptidyl arginine deiminase 4 (PAD4) enzyme, encoded by the PADI4 gene. Increased PAD4 activity promotes the onset and progression of rheumatoid arthritis (RA). This study aimed to evaluate the association of PADI4 haplotypes with RA risk, mRNA expression, and the PAD4 activity in patients with RA from Mexico. Methodology: 100 RA patients and 100 control subjects (CS) were included. Genotyping was performed by PCR-RFLP method, PADI4 mRNA expression was quantified by real-time PCR, the contribution of PADI4 alleles (PADI4_89 G>A, PADI4_90 T>C, and PADI4_92 G>C) to mRNA expression by the ASTQ method, and PAD4 activity by HPLC. Also, the anti-CCP and anti-PADI4 antibodies were quantified by ELISA. Results: The three PADI4 polymorphisms were associated with RA susceptibility (OR = 1.72, p = 0.005; OR = 1.62; p = 0.014; OR = 1.69; p = 0.009; respectively). The 89G, 90T, and 92G alleles have a higher relative contribution to PADI4 mRNA expression from RA patients than 89A, 90C, and 92C alleles in RA patients. Moreover, the GTG/GTG haplotype was associated with RA susceptibility (OR = 2.86; p = 0.024). The GTG haplotype was associated with higher PADI4 mRNA expression (p = 0.04) and higher PAD4 enzymatic activity (p = 0.007) in RA patients. Conclusions: The evaluated polymorphisms contribute to PADI4 mRNA expression and the enzymatic activity of PAD4 in leukocytes. Therefore, the GTG haplotype is a genetic risk factor for RA in western Mexico, and is associated with increased PADI4 mRNA expression and higher PAD4 activity in these patients.

Monica Leu Agelii,1 Ingiäld Hafström,2 Björn Svensson,3 Sofia Ajeganova,4,5 Kristina Forslind,3,5 Maria Andersson,3,5 Inger Gjertsson1 1Department of Rheumatology and Inflammation Research, Institute of Medicine, Sahlgrenska Academy, Gothenburg University, Gothenburg, Sweden; 2Department of Medicine Huddinge, Division of Gastroenterology and Rheumatology, Karolinska Institutet, Karolinska University Hospital, Stockholm, Sweden; 3Department of Clinical Sciences Lund, Section of Rheumatology, Lund University, Faculty of Medicine, Lund, Sweden; 4Department of Clinical Sciences, Rheumatology Division, Universitair Ziekenhuis Brussel, Vrije Universiteit Brussel, Brussels, Belgium; 5Spenshult Research and Development Center, Halmstad, SwedenCorrespondence: Inger Gjertsson, Department of Rheumatology and Inflammation Research, Institute of Medicine, Sahlgrenska Academy, Gothenburg University, Box 480, 40530 Gothenburg, Sweden, Tel +46- 031-342 4692, Email inger.gjertsson@rheuma.gu.seObjective: Correct diagnosis of early rheumatoid arthritis (RA) is essential for optimal treatment choices. No pathognomonic test is available, and diagnosis is based on classification criteria, which can result in misdiagnosis. Here, we examined the differences between actual and misdiagnosed RA cases in a long-term cohort of patients included based on the ACR-1987 classification criteria.Methods: Patients in the BARFOT (Better Anti-Rheumatic PharmacOTherapy) cohort (n=2543) with at least four follow-up visits during the initial 5 years from enrolment were assessed, and a change in diagnosis was reported by the treating rheumatologist. The groups were analysed with respect to the individual classification criteria, antibodies to citrullinated proteins (ACPA), disease activity (DAS28) and radiographic changes from inclusion up to 2 years.Results: Forty-five patients (1.8%) were misdiagnosed (RA-change group). When compared to those in the RA-change group, the patients who kept their diagnosis (RA-keep) were more often RF positive (64% vs 21%, p< 0.001) or ACPA positive (59% vs 8%, p< 0.001). They were also more likely to fulfil more than four ACR-1987 criteria (64% vs 33%, p< 0.001) and to have radiographic changes at inclusion (RA-keep 27% vs RA-change 12%, p=0.04). The groups had a similar evolution of DAS28 and its components as well as of radiological joint destruction.Conclusion: Diagnosis of RA according to the ACR-1987 criteria had a high precision in this long-term cohort. A diagnosis of RA should be re-evaluated in patients who do not fulfil more than four ACR-1987 criteria especially in patients negative for RF.Keywords: arthritis, precision, ACR-1987 classification criteria, rheumatoid factor, ACPA

Abstract Background Studies on the association between coffee, a modifiable lifestyle factor, and rheumatoid arthritis (RA), a chronic autoimmune disease primarily affecting the joints, have been conflicting. The aim of the present study was to study the association between coffee consumption and risk of RA in the context of different lifestyle factors. Methods We included 2184 cases (72% women, mean age 55 years) newly diagnosed with RA during 2005–2018 in Sweden and 4201 controls matched on age, sex, and residential area. Data on coffee consumption was collected through a food frequency questionnaire and categorized into < 2 (reference), 2–< 4, 4–< 6, and ≥ 6 cups/day. We calculated odds ratios (OR) with 95% confidence intervals (CI) for coffee consumption and risk of RA, in a crude model (taking matching factors into account), and then adjusted first for smoking and further for BMI, educational level, alcohol consumption, and physical activity. We also stratified analyses on sex, smoking, rheumatoid factor, and anti-CCP2 status. Results In the crude model, high coffee consumption was associated with increased risk of RA (OR = 1.50, 95% CI 1.20–1.88 for ≥ 6 cups/day compared to < 2 cups). After adjusting for smoking, the OR decreased and was no longer statistically significant (OR = 1.16, 95% CI 0.92–1.46) and decreased further in the full model (OR = 1.14 95% CI 0.89–1.45). This pattern held true in all strata. Conclusion The findings from this large, population-based case-control study did not support a significant association between coffee consumption and risk of RA as a whole nor within different subgroups.

Synthetic and natural cannabinoid derivatives are highly investigated as drug candidates due to their antinociceptive, antiepileptic and anticancer potential. Arachidonoylcyclopropylamide (ACPA) is a synthetic cannabinoid with antiproliferative and apoptotic effects on non-small cell lung cancer and pancreatic and endometrial carcinoma. Thus, ACPA has a great potential for being used as an anticancer drug for epithelial cancers. Therefore, determining the levels of ACPA in biological fluids, cells, tissues and pharmaceutical dosage forms is crucial in monitoring the effects of various pharmacological, physiological and pathological stimuli on biological systems. However, the challenge in the quantification of ACPA is its short half-life and lack of UV signal. Therefore, we developed a liquid chromatography-tandem mass spectrometric (LC–MS/MS) method for sensitive and selective quantification of ACPA in cell culture medium and intracellular matrix. Multiple reaction monitoring in the positive ionization mode was used for detection with 344 → 203 m/z transitions. The separation of ACPA was performed on C18 column (50 × 3.0 mm, 2.1 μm) with the mobile phase run in the gradient mode with 0.1% formic acid (FA) in water and 0.1% FA in acetonitrile at a flow rate of 0.3 ml/min. The assay was linear in the concentration range of 1.8–1000 ng/mL (r = 0.999). The validation studies revealed that the method was linear, sensitive, accurate, precise, selective, repeatable, robust and rugged. Finally, the developed method was applied to quantify ACPA in cell culture medium and intracellular matrix. [ABSTRACT FROM AUTHOR]

Objective Mucosal initiated immune responses may be involved in the pathophysiology of RA. The most abundant immunoglobulin at mucosal surfaces is IgA, of which two subclasses exist: IgA1 and IgA2. IgA2 is mainly present at mucosal sites and has been ascribed pro-inflammatory properties. As IgA subclasses might provide insights into mucosal involvement and pro-inflammatory mechanisms, we investigated IgA responses in sera of RA patients. Methods In two cohorts of RA patients, the EAC and IMPROVED, total IgA1 and IgA2 were measured by ELISA. Furthermore, IgA subclass levels of RF and anti-citrullinated protein antibodies (anti-CCP2) were determined. The association of these IgA subclass levels with CRP and smoking was investigated. Results Total IgA1 and IgA2 were increased in RA patients compared with healthy donors in both cohorts. This increase was more pronounced in seropositive RA vs seronegative RA. For RF and anti-CCP2, both IgA1 and IgA2 could be detected. No strong associations were found between IgA subclasses (total, RF and anti-CCP2) and CRP. In smoking RA patients, a trend towards a selective increase in total IgA2 and RF IgA1 and IgA2 was observed. Conclusion RA patients have raised IgA1 and IgA2 levels. No shift towards IgA2 was observed, indicating that the increase in total IgA is not due to translocation of mucosal IgA into the bloodstream. However, mucosal inflammation might play a role, given the association between smoking and total IgA2 levels. Despite its pro-inflammatory properties, IgA2 does not associate strongly with pro-inflammatory markers in RA patients. [ABSTRACT FROM AUTHOR]

Post-translational modifications (PTMs) influence cellular processes and consequently, their dysregulation is related to the etiologies of numerous diseases. It is widely known that a variety of autoimmune responses in human diseases depend on PTMs of self-proteins. In this review we summarize the latest findings about the role of PTMs in the generation of autoimmunity and, specifically, we address the most relevant PTMs in rheumatic diseases that occur in synovial tissue. Citrullination, homocitrullination (carbamylation) and acetylation are responsible for the generation of Anti-Modified Protein/Peptide Antibodies (AMPAs family), autoantibodies which have been implicated in the etiopathogenesis, diagnosis and prognosis of rheumatoid arthritis (RA). Synthetic peptides provide complete control over the exact epitopes presented as well as the specific positions in their sequence where post-translationally modified amino acids are located and are key to advancing the detection of serological RA biomarkers that could be useful to stratify RA patients in order to pursue a personalized rheumatology. In this review we specifically address the latest findings regarding synthetic peptides post-translationally modified for the specific detection of autoantibodies in RA patients. [ABSTRACT FROM AUTHOR]

OBJECTIVE: Cancer ranks first among the causes of morbidity and mortality all over the world, and it is expected to continue to be the main cause of death in the coming years. Therefore, new molecular targets and therapeutic strategies are urgently needed. In many cases, some reports show increased levels of endocannabinoids and their receptors in cancer, a condition often associated with tumour aggressiveness. Recent studies have suggested that cannabinoid-1/2 receptors contribute to tumour growth in a variety of cancers, including pancreatic, colon, prostate, and breast cancer. Understanding how cannabinoids can regulate key cellular processes involved in tumorigenesis, such as: cell proliferation and cell death, is crucial to improving existing and new therapeutic approaches for the cancer patients. The present study was aimed to characterize the in-vitro effect of L-759633 (a selective CB2 receptor agonist), ACPA (a selective CB1 receptor agonist) and ACEA (a selective CB1 receptor agonist) on the cell proliferation, clonogenicity, and apoptosis in pancreatic (PANC1) and breast (MDA-MB-231) cancer cells. METHODS: The viability and/or proliferation of cells were detected by MTS assay. A clonogenic survival assay was used to detect the ability of a single cell to grow into a colony. Apoptosis was determined with Annexin V staining (Annexin V-FITC/PI test) and by analyzing the expression of Bcl-2-associated X protein (Bax) and B-cell lymphoma 2 (Bcl-2). RESULTS: We found that selective CB1/2 agonists suppressed cell proliferation, clonogenicity and induced proapoptotic function in human PANC1 pancreatic and MDA-MB-231 breast cancer cells. Based on our fi ndings, these agonists led to the inhibition of both cell viability and clonogenic growth in a dose dependent manner. CB1/2 agonists were observed to induce intrinsic apoptotic pathway by upregulating Bax, while downregulating Bcl-2 expression levels. CONCLUSION: Our data suggests that CB1/2 agonists have the therapeutic potential through the inhibition of survival of human PANC1 pancreatic and MDA-MB-231 breast cancer cells and also might be linked with further cellular mechanisms for the prevention (Fig. 5, Ref. 49). [ABSTRACT FROM AUTHOR]

Oral microbiome changes take place at the initiation of rheumatoid arthritis (RA); however, questions remain regarding the oral microbiome at pre-RA stages in individuals with clinically suspect arthralgia (CSA). Two cross-sectional cohorts were selected including 84 Tatarstan women (15 early-RA as compared to individuals with CSA ranging from CSA = 0 [n = 22], CSA = 1 [n = 19], CSA = 2 [n = 11], and CSA ≥ 3 [n = 17]) and 42 women with established RA (median: 5 years from diagnosis [IQ: 2–11]). Amplicon sequence variants (ASVs) obtained from oral samples (16S rRNA) were analyzed for alpha and beta diversity along with the abundance at the genus level. A decrease in oral Porphyromonas sp. is observed in ACPA-positive individuals, and this predominates in early-RA patients as compared to non-RA individuals irrespective of their CSA score. In the RA-established cohort, Porphyromonas sp. and Aggregatibacter sp. reductions were associated with elevated ACPA levels. In contrast, no associations were reported when considering individual, genetic and clinical RA-associated factors. Oral microbiome changes related to the genera implicated in post-translational citrullination (Porphyromonas sp. and Aggregatibacter sp.) characterized RA patients with elevated ACPA levels, which supports that the role of ACPA in controlling the oral microbiome needs further evaluation. [ABSTRACT FROM AUTHOR]

Autoantibodies represent a hallmark of rheumatoid arthritis (RA), with the rheumatoid factor (RF) and antibodies against citrullinated proteins (ACPA) being the most acknowledged ones. RA patients who are positive for RF and/or ACPA ("seropositive") in general display a different etiology and disease course compared to so-called "seronegative" patients. Still, the seronegative patient population is very heterogeneous and not well characterized. Due to the identification of new autoantibodies and advancements in the diagnosis of rheumatic diseases in the last years, the group of seronegative patients is constantly shrinking. Aside from antibodies towards various post-translational modifications, recent studies describe autoantibodies targeting some native proteins, further broadening the spectrum of recognized antigens. Next to the detection of new autoantibody groups, much research has been done to answer the question if and how autoantibodies contribute to the pathogenesis of RA. Since autoantibodies can be detected years prior to RA onset, it is a matter of debate whether their presence alone is sufficient to trigger the disease. Nevertheless, there is gathering evidence of direct autoantibody effector functions, such as stimulation of osteoclastogenesis and synovial fibroblast migration in in vitro experiments. In addition, autoantibody positive patients display a worse clinical course and stronger radiographic progression. In this review, we discuss current findings regarding different autoantibody types, the underlying disease-driving mechanisms, the role of Fab and Fc glycosylation and clinical implications. [ABSTRACT FROM AUTHOR]

Abstract Background A specific feature of the autoimmune response in rheumatoid arthritis (RA) is the presence of anti‐citrullinated protein antibodies (ACPAs) in patient sera. These antibodies can appear several years before disease onset and are involved in the development of RA. Objective We performed proteomic analysis by mass spectroscopy to identify novel citrullinated antigens and autoantibodies in RA patients. Methods Polypeptides isolated from the sera of RA patients were identified by Orbitrap high‐precision mass spectrometry and then citrulline‐containing proteins were selected. The levels of ACPAs against these newly identified citrullinated autoantigens in sera of 100 RA patients and 50 healthy controls were determined by enzyme‐linked immunosorbent assays. Results A total of 135 proteins were identified in RA patients and the protein profile included 11 citrulline‐containing antigens. Three of the 11 citrullinated proteins had been reported in previous studies. ACPAs against the novel citrullinated epitopes from these proteins were increased in sera from the RA patients compared with those from healthy controls. Autoantibodies against one of the citrullinated antigens, lipopolysaccharide‐binding protein (LBP), was significantly increased in RA patients and associated with disease activities. The titer of anti‐citrullinated LBP antibodies (anti‐cLBP) was closely related to the infection incidence in RA patients. Conclusion Serum protein analysis by high‐precision proteomic technology is a feasible method to identify novel citrullinated epitopes in RA patients. Anti‐cLBP antibodies are associated with disease severity and infection in RA patients.

Abstract Background Musculoskeletal (MSK) symptoms are among the commonest reasons for primary care assessments; however, few individuals will be diagnosed with an inflammatory arthritis (IA) within the following year. The purpose of this study was to investigate, in individuals with new MSK symptoms, the association between patient factors and risk of progression to IA, in order to optimise primary care referrals to rheumatology. Methods Individuals ≥16 years old with new non-specific MSK symptoms and no clinical synovitis were recruited by primary care across the UK from July 2007 until May 2019. Those testing positive for the anti-CCP2 assay (anti-CCP+) were invited to Leeds for follow-up. Subjects with a negative result (anti-CCP−) were sent a 1-year questionnaire, and general practitioners were contacted to confirm whether the individual had been diagnosed with an IA by a rheumatologist. Predictors for progression were assessed using multivariable regression analysis. Results Six thousand seven hundred eighty individuals were recruited: 3% were anti-CCP+, of whom 45% progressed to IA, predominantly rheumatoid arthritis. Anti-CCP+ participants with high antibody levels had an odds ratio (OR) for progression to IA of 9.42 [P < 0.001, 95% CI (3.13–28.30)], hand pain, OR 2.74 [P = 0.043, 95% CI (1.03–7.27)] and foot pain, OR 4.10 [P = 0.003, 95% CI (1.59–10.54)]. In low-level anti-CCP+ individuals, absence of pain in hands or feet had a negative predictive value of 96% for progression to IA. One-year follow-up data were available for 5640 anti-CCP− individuals, of whom 53 were diagnosed with IA (0.93%). Pain in hands, OR 2.51 [P = 0.018, 95% CI (1.17–5.39)] or knees, OR 3.03 [P = 0.003, 95% CI (1.47–6.25)] were associated with development of IA within 12 months. Conclusions This is the largest prospective primary care study of individuals at risk of IA, and the first one to prospectively investigate the outcome of MSK symptoms in a large anti-CCP− cohort. High anti-CCP levels and pain in hands/feet indicated an increased likelihood of progression to IA. In patients with low anti-CCP level and no pain in the hands/feet, progression is unlikely. In anti-CCP− patients, those with hand or knee pain were at increased risk of progression. This study demonstrates that routinely available tests and joint symptoms provide useful discrimination that may be used to prioritise referrals to rheumatology and avoid a delayed diagnosis. Trial registration NCT, NCT02012764 . Registered 25 January 2007.

• Optical biosensors were critically reviewed, including ECL, SERS-based, and SPR-based biosensors. • Lateral flow immunoassays (LFIA) as point-of-care (POC) rapid tests were discussed. • Amperometric and label-free electrochemical biosensors developed for anti-CCP were analyzed. • The clinical significance of these biosensors and their advantages and disadvantages were demonstrated. • Various signal amplification systems with their peptide sequences and modifiers were revealed. Biosensors are unique analytical tools for the detection of biomarkers. Of these, autoantibodies against citrullinated proteins (ACPA) are useful for the differential diagnosis of rheumatoid arthritis (RA). The autoantibodies may be detected by immunoassay technology using synthetic cyclic citrullinated peptides (CCP), ie, anti-CCP. Recently, several biosensors have been developed for anti-CCP using CCP and mutated citrullinated vimentin (MCV) as recognition elements. In this review we highlight all currently available ACPA biosensor technology including those based on fluorescence, chemiluminescence, electrochemiluminescence (ECL), surface-enhanced Raman scattering (SERS)-based, surface plasmon resonance (SPR), lateral flow immunoassays (LFIA), and electrochemical. We explore various peptides as recognition elements, electrode modifiers and signal amplification systems thus providing new opportunities for next-generation biosensor design in RA. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis is an autoimmune disease that affects joints, leading to swelling, inflammation, and dysfunction in the joints. Recently, research efforts have been focused on finding novel curative approaches for rheumatoid arthritis, as current therapies are associated with adverse effects. Here, we examined the effectiveness of dabigatran, the antithrombotic agent, in treating complete Freund's adjuvant (CFA)-induced arthritis in rats. Subcutaneous injection of a single 0.3 mL dosage of CFA into the rat's hind leg planter surface resulted in articular surface deformities, reduced cartilage thickness, loss of intercellular matrix, and inflammatory cell infiltration. There were also increased levels of the Anti-cyclic citrullinated peptide antibody (ACPA), oxidative stress, and tissue Receptor activator of nuclear factor–kappa B ligand (RANKL). Proteins of the kallikrein-kinin system (KKS) were also elevated. The inhibitory effects of dabigatran on thrombin led to a subsequent inhibition of KKS and reduced Toll-like receptor 4 (TLR4) expression. These effects also decreased RANKL levels and showed anti-inflammatory and antioxidant effects. Therefore, dabigatran could be a novel therapeutic strategy for arthritis. [ABSTRACT FROM AUTHOR]

The number of cannabis users is increasing in the world. However, the mechanisms involved in the psychiatric effects and addiction formation remain unclear. Medical treatments against cannabis addiction have not yet been established. Δ9-Tetrahydrocannabinol (THC), the main active substance in cannabis, binds and affects cannabinoid type 1 receptors (CB1R) in the brain. The mice were intraperitoneally (i.p.) administered arachidonylcyclopropylamide (ACPA), a CB1R-selective agonist, and then two behavioral experiments on anxiety and addiction were performed. Administration of ACPA caused anxiolytic-like behavior in the elevated plus maze test. In addition, ACPA increased place preference in a conditioned place preference (CPP) test. The basolateral amygdala (BLA), which is the focus of this study, is involved in anxiety-like behavior and reward and is reported to express high levels of CB1R. We aimed to reveal the role of CB1R in BLA for ACPA-induced behavior. AM251, a CB1R selective antagonist, was administered intra-BLA before i.p. administration of ACPA. Intra-BLA administration of AM251 inhibited ACPA-induced anxiolytic-like behavior and place preference. These results suggest that CB1R in the BLA contributes to behavior disorders caused by the acute or chronic use of cannabis. [ABSTRACT FROM AUTHOR]

Observations from numerous clinical, epidemiological and serological studies link periodontitis with severity and progression of rheumatoid arthritis. The strong association is observed despite totally different aetiology of these two diseases, periodontitis being driven by dysbiotic microbial flora on the tooth surface below the gum line, while rheumatoid arthritis being the autoimmune disease powered by anti-citrullinated protein antibodies (ACPAs). Here we discuss genetic and environmental risk factors underlying development of both diseases with special emphasis on bacteria implicated in pathogenicity of periodontitis. Individual periodontal pathogens and their virulence factors are argued as potentially contributing to putative causative link between periodontal infection and initiation of a chain of events leading to breakdown of immunotolerance and development of ACPAs. In this respect peptidylarginine deiminase, an enzyme unique among prokaryotes for Porphyromonas gingivalis, is elaborated as a potential mechanistic link between this major periodontal pathogen and initiation of rheumatoid arthritis development. [ABSTRACT FROM AUTHOR]

Objectives To investigate whether anti-CCP2-positive at-risk individuals with musculoskeletal (MSK) symptoms but without clinical synovitis (CCP2+ at-risk) develop US subclinical synovitis before inflammatory arthritis and if US subclinical synovitis can be predicted. Methods First, US scans of CCP2+ at-risk individuals who developed inflammatory arthritis ('progressors') were reviewed for subclinical synovitis prior to inflammatory arthritis development. Patients in whom the pre-progression US scan was negative but the scan was conducted >6 months before progression were excluded. Subsequently, regression analyses were performed to identify predictors of US synovitis in CCP2+ at-risk individuals without baseline US abnormalities who had one or more longitudinal US scan and a complete dataset. Results US subclinical synovitis was detected in one or more scan in 75 of 97 progressors (77.3%) {median time to inflammatory arthritis development from first evidence of US synovitis 26.5 weeks [interquartile range (IQR) 7–60]}, in whom one or more scan was available, excluding those with a negative scan >6 months from inflammatory arthritis development (n  = 38). In 220 CCP2+ at-risk individuals with normal baseline US scans, who had one or more longitudinal US scan and a complete dataset, US synovitis was detected in 69/220 (31.4%) [median time to first developing US synovitis 56.4 weeks (IQR 33.0–112.0)]. In the multivariable analysis, only anti-CCP3 antibodies were predictive for the development of US synovitis [odds ratio 4.75 (95% CI 1.97, 11.46); P  < 0.01]. Conclusions In anti-CCP2+ at-risk individuals, a stage of subclinical synovitis usually precedes the development of inflammatory arthritis. Anti-CCP2+/CCP3+ individuals without clinical or US subclinical synovitis may represent the optimal window of opportunity for intervention to prevent joint disease. [ABSTRACT FROM AUTHOR]

This study aimed to analyze the factors that influence anti-citrullinated protein antibody (ACPA) titers in a seropositive early arthritis (EA) population under non-protocolized treatment with disease-modifying anti-rheumatic drugs (DMARDs). A total of 130 ACPA-positive patients from the PEARL (Princesa Early Arthritis Longitudinal) study were studied along a 5-year follow-up. Sociodemographic, clinical, and therapeutic variables, along with serum samples, were collected at five visits by protocol. Anti-cyclic citrullinated peptide 2 (CCP2) ACPA titers were measured by ELISA. The effect of different variables on anti-CCP2 titers was estimated using longitudinal multivariate analysis models, nested by visit and patient. Data from 471 visits in 130 patients were analyzed. A significant decrease in anti-CCP2 titers was observed at all time-points, compared to baseline, following the decline of disease activity. In the multivariate analysis, active or ever smoking was significantly associated with the highest anti-CCP2 titers while reduction in disease activity was associated with titer decline. After adjusting for these variables, both conventional synthetic (cs) and biologic (b) DMARDs accounted for the decline in anti-CCP2 titers as independent factors. Conclusion: In patients with EA, an early and sustained reduction in ACPA titers can be detected associated with the decline in disease activity, irrespective of the treatment used. [ABSTRACT FROM AUTHOR]

Background: The presence of autoantibodies against citrullinated proteins (ACPA) significantly increases the risk of developing rheumatoid arthritis (RA). Dysregulation of lymphocyte subpopulations was previously described in RA., Objectives: To propose the predictive model for progression to clinical arthritis based on peripheral lymphocyte subsets and ACPA in individuals who are at risk of RA., Methods: Our study included 207 at-risk individuals defined by the presence of arthralgias and either additional ACPA positivity or meeting the EULAR definition for clinically suspect arthralgia. For the construction of predictive models, 153 individuals with symptom duration ≥12 months who have not yet progressed to arthritis were included. The lymphocyte subsets were evaluated using flow cytometry and anti-CCP using ELISA., Results: Out of all individuals with arthralgia, 41 progressed to arthritis. A logistic regression model with baseline peripheral blood lymphocyte subpopulations and ACPA as predictors was constructed. The resulting predictive model showed that high anti-CCP IgG, higher percentage of CD4+ T cells, and lower percentage of T and NK cells increased the probability of arthritis development. Moreover, the proposed classification decision tree showed, that individuals having both high anti-CCP IgG and low NK cells have the highest risk of developing arthritis., Conclusions: We propose a predictive model based on baseline levels of lymphocyte subpopulations and ACPA to identify individuals with arthralgia with the highest risk of progression to clinical arthritis. The final model includes T cells and NK cells, which are involved in the pathogenesis of RA. This preliminary model requires further validation in larger at-risk cohorts., (© The Author(s) 2024. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Abstract Background There is a need for biomarker to identify patients “at risk” for rheumatoid arthritis (risk-RA) and to better predict the therapeutic response and in this study we tested the hypothesis that novel native and citrullinated heterogeneous nuclear ribonucleoprotein (hnRNP)-DL autoantibodies could be possible biomarkers. Methods Using protein macroarray and ELISA, epitope recognition against hnRNP-DL was analysed in sera from different developed RA disease and diagnosed SLE patients. Toll-like receptor (TLR) 7/9 and myeloid differentiation primary response gene 88 (MyD88)-dependency were studied in sera from murine disease models. HnRNP-DL expression in cultivated cells and synovial tissue was analysed by indirect immunofluorescence, immunoblot and immunohistochemistry. Results HnRNP-DL was highly expressed in stress granules, citrullinated in the rheumatoid joint and targeted by autoantibodies either as native or citrullinated proteins in patient subsets with different developed RA disease. Structural citrullination dependent epitopes (SCEs) of hnRNP-DL were detected in 58% of the SLE patients although 98% of these sera were α-CCP-2-negative. To obtain a specific citrullinated signal value, we subtracted the native antibody value from the citrullinated signal. The citrullinated/native index of autoantibodies against hnRNP-DL (CNDL-Index) was identified as a new value for an “individual window of treatment success” in early RA and for the detection of RF IgM/α-CCP-2 seronegative RA patients (24–46%). Negative CNDL-index was found in SLE patients, risk-RA and early RA cohorts such as EIRA where the majority of these patients are DAS28-responders to methotrexate (MTX) treatment (87%). High positive CNDL-values were associated with more severe RA, shared epitope and parenchymal changes in the lung. Specifically, native α-hnRNP-DL is TLR7/9-dependent, associated with pain and ROC analysis revealed an association to initial MTX or etanercept treatment response, especially in seronegative RA patients. Conclusion CNDL-index defines people at risk to develop RA and the “window of treatment success” thereby closing the sensitivity gap in RA.

Abstract Background Tea is a popular beverage around the world and has properties that can affect the immune system. The association between tea consumption and the risk of rheumatoid arthritis (RA), a chronic autoimmune disease primarily affecting the joints, is not well studied and results are conflicting. Methods We collected data on tea consumption for 2237 incident RA cases diagnosed 2005–2018 and 4661 controls matched on age, sex, and residential area. Tea consumption was classified into no (0 cups/day), irregular (

Observations from numerous clinical, epidemiological and serological studies link periodontitis with severity and progression of rheumatoid arthritis. The strong association is observed despite totally different aetiology of these two diseases, periodontitis being driven by dysbiotic microbial flora on the tooth surface below the gum line, while rheumatoid arthritis being the autoimmune disease powered by anti-citrullinated protein antibodies (ACPAs). Here we discuss genetic and environmental risk factors underlying development of both diseases with special emphasis on bacteria implicated in pathogenicity of periodontitis. Individual periodontal pathogens and their virulence factors are argued as potentially contributing to putative causative link between periodontal infection and initiation of a chain of events leading to breakdown of immunotolerance and development of ACPAs. In this respect peptidylarginine deiminase, an enzyme unique among prokaryotes for Porphyromonas gingivalis, is elaborated as a potential mechanistic link between this major periodontal pathogen and initiation of rheumatoid arthritis development.

The risk to develop ACPA positive rheumatoid arthritis (RA), the most destructive type of autoimmune arthritis, is carried by HLA-DRB1 alleles containing a 5 amino acid motif: the shared epitope (SE). RA is preceded by the emergence of disease specific anti citrullinated protein antibodies (ACPA). SE positive HLA-DRB1 alleles are associated with ACPA and ACPA positive RA, not with ACPA negative RA, suggesting that ACPA contribute to the pathogenesis of RA. Understanding how HLA-DRB1 genotypes influence ACPA could lead to a curative or preventive treatment of RA. The "Shared epitope binds citrullinated peptides " hypothesis suggests that RA associated HLA-DR alleles present citrullinated peptides to T cells that help ACPA producing B cells. The "Hapten carrier model" suggests that PAD4 is the target of the T cells which help ACPA specific B cells through a hapten carrier mechanism in which PAD4 is the carrier and citrullinated peptides are the haptens. Direct binding assay of citrullinated peptides to purified HLA-DR molecules does not support the "shared epitope binds citrullinated peptides" hypothesis. The Odds Ratios to develop ACPA positive RA associated with each of 12 common HLA-DRB1 genotypes match the probability that the two HLA-DR molecules they encode can bind at least one peptide from PAD4, not from citrullinated fibrinogen. Thus, PAD4 tolerization might stop the carrier effect and switch off production of ACPA. [ABSTRACT FROM AUTHOR]