Insulin signaling regulates cellular metabolism in an epigenetic manner, but its role in the immune cell homeostasis remains unknown. High plasma insulin obstructs efficient insulin signaling and rewires metabolic activity in autoimmunity. In this study, we explored the functional consequences of insulin signaling for the metabolism and phenotype of effector CD4+ T cells in blood and synovial tissue of patients with rheumatoid arthritis (RA). Transcriptome profiling of CD4+ cells in RA blood and synovia revealed high metabolic activity and effector function of the survivin/BIRC5hiPD1hi T peripheral helper cell population. Low insulin signaling and deficient histone acetylation in RA T cells amplified proinflammatory IFNγ and TNF expression. Co-deposition of survivin with acetylated histone H3K27 on regulatory chromatin controlled the transcription of histone acetylation complex subunits and insulin-dependent genes. Insulin stimulation and histone deacetylase inhibition induced an increase in histone acetylation. In CD4+ cell cultures and in aggressive PD1hiTph cells in RA synovial tissue, exposure to insulin synergized with inhibition of histone deacetylation to upregulate IL7 production suppressing IFNγ and PD1. This activated IL7R-signaling mediators STAT5A/B, BCL2, and promoted acquisition of CD27+CD45RO+ central memory phenotype in the PD1hiTph cells. Likewise, the CD4+ cells in hyperinsulinemic T2D patients showed enrichment of IL7R+T cell cluster. In RA patients, antagonizing folate transport and JAK/STAT signaling activated insulin signaling and histone acetylation-dependent metabolism of CD4+ cells. Concomitant with CTLA4-dependent signaling, this enabled the adoption of an incipient IL7R+ T cell phenotype. This study demonstrates that insulin binds together metabolic activity and histone acetylation in CD4+ cells. Sufficient insulin signaling promotes IL7R+ memory phenotype accrual in aggressive PD1hiTph cells. Hence, achieving insulin sensitivity via histone acetylation disarms effector CD4+ T cell function and presents an attractive interventional goal to restore immune cell homeostasis in RA.

Background In the American College of Rheumatology/European Alliance of Rheumatology Associations 2010 classification criteria for rheumatoid arthritis (RA), rheumatoid factor (RF) and anti-citrullinated protein/peptide antibodies (ACPA) are assigned the same weight, although ACPA have proven more specific than RF. We aimed to refine the serological weight factors for RA classification.Materials and methods Diagnostic samples from 398 patients with RA and 448 diseased controls (DC) were evaluated with RF and ACPA assays from five different manufacturers. A grid search was conducted across all possible integer score pairs for RF and ACPA to identify the weighting combination that optimised the specificity of RA classification. This combination was applied in a refined scoring system (RSS) for three confirmatory cohorts comprising, respectively, 67, 157 and 132 patients with RA and 441, 153 and 193 DC.Findings In the investigative cohort, the optimal combination of weighting scores was characterised by higher scores for ACPA, higher antibody levels and double RF/ACPA positivity.Applying the optimal combination of weighting scores decreased misclassification compared with the 2010 RA classification criteria, minimally affecting sensitivity. For seropositive patients, RSS significantly lowered misclassification from 38.3–55.8% to 15.0–26.9%.The improved specificity of RSS (compared with the 2010 criteria) was validated in the confirmatory cohorts. Considering only seropositive patients, the RSS significantly decreased RA misclassification (8.6–36.8% vs 20.0–78.9%), minimally affecting the sensitivity (86.7–97.3% vs 86.7–96.0%).Interpretation Serological weight factors for RA classification can be improved by taking into account the antibody type, the antibody levels and single or combined positivity. The application of an RSS reduced the number of RA misclassifications.

OBJECTIVES:Idiopathic inflammatory myopathies (IIM) can affect multiple organs, including the heart, potentially leading to arrhythmia, heart failure, and thereby a poor prognosis. We hypothesised that cardiac and skeletal muscle involvement in patients with IIM share pathological mechanisms, and that severe skeletal muscle involvement may be associated with cardiac involvement. The aim of this study was to identify disease-related parameters that indicate cardiac involvement in newly diagnosed patients with IIM. METHODS:In this prospective study, 34 newly diagnosed patients with IIM and 9 age- and gender-matched healthy controls underwent cardiac magnetic resonance imaging, blood analyses for skeletal muscle markers, and assessments of IIM-specific disease features. RESULTS:Cardiac involvement was detected by cardiac magnetic resonance imaging in 47% of patients with newly diagnosed IIM, presenting as ongoing myocarditis/perimyocarditis (44%), ongoing pericarditis (25%) or previous myocarditis (31%). IIM patients with cardiac involvement had significantly more prevalent myositis (p=0.018) and higher levels of serum markers of muscle inflammation (myoglobin, p=0.039; alanine aminotransferase, p=0.045 and aspartate aminotransferase p=0.005) compared to IIM without cardiac involvement. IIM with ongoing myocarditis/peri-myocarditis displayed significantly elevated cardiac troponin I (cTnI) levels than IIM with ongoing pericarditis (p=0.015) or previous myocarditis (p=0.015). Additionally, cTnI levels were strongly correlated to myositis (as clinical manifestation, p=0.011), creatin kinase (p=0.001), myoglobin (p=0.001), lactate dehydrogenase (p=0.008) and aspartate aminotransferase (p=0.0.001). CONCLUSIONS:Cardiac involvement as detected by cardiac magnetic resonance imaging is common at time of diagnosis in patients with IIM and is closely linked to the severity of skeletal muscle involvement.

ObjectiveAnti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitides (AAV) are chronic, relapsing inflammatory diseases, yet reliable biomarkers for detecting relapse and organ involvement remain limited. This study aimed to identify plasma protein biomarkers that distinguish active disease from remission and to explore markers associated with lung and kidney involvement.MethodsPlasma samples from 113 patients with granulomatosis with polyangiitis or microscopic polyangiitis (68 active disease and 45 remission) were profiled using a proximity extension assay targeting 181 inflammation- and cardiovascular-related proteins. Clinical data, including CRP and Birmingham vasculitis activity score (BVAS), were collected at sampling. Differential protein expression was assessed using ANOVA, with top candidates validated in independent sample cohorts (plasma samples, n = 74; serum samples, n = 34). Correlations with BVAS and CRP and discriminatory performance (AUC) were evaluated. Associations with chest- and kidney-specific BVAS were also examined.ResultsA total of 57 proteins were differentially expressed between active disease and remission. Seven proteins (ST2, OPN, IL-2RA, CCL23, IL-6, Flt3L, and SCF) were validated in independent cohorts and showed strong associations with disease activity. Several demonstrated high discriminatory ability (AUC ≥ 0.80) between active disease and remission. Multiple proteins correlated with organ-specific BVAS scores: seven for chest involvement and 16 for kidney involvement, after adjustment for kidney function-related protein variation.ConclusionThis study identifies a robust panel of plasma proteins that differentiate active AAV from remission and correlate with global and organ-specific disease activity. These biomarkers may enhance non-invasive disease monitoring and support earlier recognition of relapse and organ involvement in AAV.

Abstract Background Handgrip strength (HGS) in rheumatoid arthritis (RA) is commonly attributed to joint pathology, but may also reflect extra-articular manifestations, including atrophy of motor-related brain regions. We investigated HGS as a marker of peripheral joint status, systemic immune regulation, and central motor integrity. Methods Maximal HGS was assessed using a dynamometer. Joint pathology was evaluated using radiographic and clinical measures, and upper limb disability using questionnaire. Brain volumes were quantified using MRI and MAPER software. Transcriptome sequencing was performed on circulating CD4⁺ and CD14⁺ cells. In a six-month, single-arm pilot trial, a subgroup of patients performed daily hand exercises. Associations and longitudinal changes were analysed using linear mixed-effects models accounting for repeated measurements across hands and timepoints, with variable selection performed using LASSO regression. Results A total of 59 women with established RA were included in the cross-sectional analysis (median age 64 years [range 23–76], DAS28 2.46 [1.1–5.8], disease duration 11 years [0–45]). Lower HGS was associated with greater disability. HGS was also independently associated with premotor and supplementary motor cortex (PMA/SMA) volume after adjustment for age, hand dominance, and joint pathology. Among joint pathology measures, tender joint count showed a significant negative association with HGS. Transcriptome analyses of CD4⁺ and CD14⁺ cells indicated that lower HGS was associated with reduced immune responsiveness and altered cytokine signalling pathways. In a six-month pilot hand exercise trial (n = 12; median age 54 years [28–68], DAS28 2.87 [1.2–3.6], disease duration 14 years [1–40]), HGS increased at 3 months, with a non-significant trend at 6 months. Baseline PMA/SMA volume showed a non-significant trend towards predicting HGS improvement. Longitudinal analyses revealed region-specific brain changes, with a decrease in PMA/SMA volume and an increase in insular volume over time. Conclusions Handgrip weakness in RA may reflect both joint pathology and motor cortex atrophy in the PMA/SMA. Hand exercise improved HGS and induced certain structural changes in the brain, though effects on motor regions remain uncertain and warrant further study. Trial registration Clinical trial registration: ClinicalTrials.gov, NCT04378621. Registration date: May 5, 2020.

Abstract Chronic systemic inflammation has been implicated in age-related neurodegeneration, but whether rheumatoid arthritis (RA) is associated with accelerated brain aging remains unclear. We combined structural magnetic resonance imaging (MRI), circulating neurodegeneration biomarkers, and peripheral monocyte transcriptomics to investigate brain aging in RA across two independent cohorts. A brain-age prediction model trained in healthy controls from the IXI imaging dataset was applied to RA patient cohorts from Gothenburg (n = 71) and Glasgow (n = 50). RA was associated with significantly elevated corrected brain-age gap relative to healthy controls (+6.5 years, 95% CI 4.2–8.8 years, p = 1.2 × 10 −7 ), with substantially stronger effects in patients ≥60 years. Older RA patients demonstrated a significant ventricular enlargement together with reduced frontal and parietal lobe volumes. Serum brain-derived tau and glial fibrillary acidic protein levels were elevated in RA. The increased brain-age gap was associated with altered myeloid transcriptional signatures. These findings demonstrate that RA is associated with age-related neurostructural alterations consistent with accelerated brain aging.

Abstract Objective Cardiac involvement (CI) is a major determinant of poor prognosis in idiopathic inflammatory myopathies (IIM). Soluble suppression of tumorigenicity 2 (sST2) is a biomarker linked to cardiac inflammation and remodelling, but its role has scarcely been explored in IIM. We aimed to investigate the levels of sST2 and its longitudinal profile in patients with IIM and CI. Methods Serum sST2 levels were measured longitudinally over the course of two years in 34 patients with newly diagnosed IIM and in 109 patients with established IIM from a cross-sectional cohort, as well as in age and gender matched healthy controls (HC) and in patients with rheumatoid arthritis (RA). CI was assessed using cardiac magnetic resonance (CMR) in newly diagnosed IIM patients. Associations between sST2 and clinical parameters were analyzed. Results Patients with newly diagnosed IIM exhibited significantly higher sST2 levels at diagnosis compared to HC (P < 0.0001) and patients with established IIM (P < 0.0001). sST2 levels were highest at diagnosis, decreased significantly by 6 months, and remained lower during follow-up. sST2 levels were positively associated with cardiac troponin I and N-terminal pro-brain natriuretic peptide (P = 0.0001 and P = 0.0001, respectively). Importantly, IIM patients with active CMR-verified CI exhibited significantly higher sST2 levels than those without ongoing CI. In contrast, patients with established IIM had sST2 levels comparable to HC and RA patients. Conclusion sST2 is elevated in early IIM and closely associated with cardiac biomarkers and CMR-verified cardiac inflammation. Its dynamic decline with treatment suggests potential utility as a biomarker for early detection and monitoring of CI in IIM.

Staphylococcus aureus is a leading cause of skin and soft tissue infections (SSTIs), which can escalate into systemic disease. While innate immune responses play a critical role in bacterial clearance, the bacterial components themselves can exacerbate inflammation. Here, we demonstrate that S. aureus lipoproteins (Lpp) and polymeric peptidoglycan (PG) synergistically induce skin abscesses in mice, in a process that requires both the lipid moiety of Lpp and the intact polymeric structure of PG. This synergy is mediated by Toll-like receptor 2 (TLR2) and nucleotide-binding oligomerization domain-containing protein 2 (NOD2) and depends on infiltrating neutrophils and monocytes. Co-administration of Lpl1 and PG results in a 5-fold to 10-fold increase in macrophage inflammatory protein-2 (MIP-2) levels in the skin compared to either ligand alone, indicating a clear synergistic effect. Furthermore, we show that local alteration in coagulation and fibrinolysis contributes to the inflammatory response, as fibrinogen depletion significantly reduced lesion size. To extend these findings to a clinically relevant model, we employed an S. aureus double mutant that lacked both lipidation (Δlgt) and peptidoglycan O-acetyltransferase (ΔoatA). This strain exhibited markedly attenuated virulence in a murine skin infection model. Importantly, this attenuation was fully reversed by neutrophil depletion, indicating that neutrophils are essential mediators of the host responses to these bacterial structures. Our findings reveal a cooperative mechanism through which S. aureus cell wall components drive skin lesion development, and we identify potential therapeutic targets for reducing the severity of SSTIs. IMPORTANCE:Staphylococcus aureus is a bacterium that often causes skin infections, including painful abscesses. We discovered that two components of S. aureus, lipoproteins on its surface and peptidoglycan in its cell wall, collaborate to drive the formation of skin abscesses. This combination triggers potent immune responses by activating the receptor Toll-like receptor 2 (TLR2) and nucleotide-binding oligomerization domain-containing protein 2 (NOD2) on host cells. As a consequence, high numbers of neutrophils and monocytes swarm the infection site. The resulting immune overreaction, together with activation of the coagulation system, produces intense inflammation. We confirmed the importance of these bacterial components using mutant S. aureus strains in a skin infection model. These mutants generated much smaller abscesses in our experiments. Our findings highlight a cooperative mechanism that exacerbates staphylococcal infections. Targeting this synergy could be a valuable strategy to reduce disease severity.

OBJECTIVE:The antineutrophil cytoplasmic antibody (ANCA)-associated vasculitides are heterogeneous disorders. The aim of this study was to identify and characterize subgroups of patients based on sex, ANCA, age at diagnosis, and organ involvement. METHODS:In total, 1,167 patients with granulomatosis with polyangiitis (GPA) or microscopic polyangiitis (MPA) were retrospectively recruited to the study. Data including cumulative involvement of 10 different organ systems, end-stage kidney disease (ESKD), sex, proteinase (PR) 3-ANCA, myeloperoxidase (MPO)-ANCA, age at diagnosis, disease duration, and relapse were obtained from medical records. Clinical variables were analyzed for associations with sex, age at diagnosis, and relapse using logistic regression analysis. Thirteen clinical variables were included in hierarchical cluster analyses using the Ward method. RESULTS:In patients with GPA, PR3-ANCA, renal and pulmonary involvement, and ESKD were significantly associated with male sex, whereas MPO-ANCA was associated with female sex. Patients with GPA who were younger than 32 years of age at diagnosis were significantly more often females and had more ear-nose-throat involvement than patients older than 32 years. In patients with MPA, female patients were significantly younger at diagnosis than male patients. Relapse was significantly associated with young age at diagnosis and pulmonary involvement in GPA and with musculoskeletal involvement in MPA. Hierarchical cluster analyses identified five and seven patient clusters among individuals with GPA and MPA, respectively. PR3-/MPO-ANCA defined the largest clusters, whereas heart, gastrointestinal, and central nervous system involvement were hallmarks for three clusters for both patients with GPA and MPA. CONCLUSION:Sex, age at diagnosis, and specific organ involvements define clinically relevant subgroups among patients with ANCA-associated vasculitides.

Septic arthritis, the most severe joint disease, is frequently caused by Staphylococcus aureus (S. aureus). A substantial proportion of patients with septic arthritis experience poor joint outcomes, often necessitating joint replacement surgery. Here, we show that monocyte depletion confers full protection against bone erosion in a septic arthritis mouse model. In the infected synovium, Ly6Chi monocytes exhibited increased expression of osteoclastogenesis-related molecules, including CCR2, c-Fms, and RANK. S. aureus lipoproteins induced elevated levels of RANKL, MCSF, and CCL2 in joints, with synovial fibroblasts identified as the major RANKL producer. Anti-RANKL treatment prevented bone destruction in both local and hematogenous septic arthritis murine models. Importantly, combining anti-RANKL treatment with antibiotics provided robust protection against joint damage. Our results indicate that the infiltration and transformation of monocytes into bone-destructive, osteoclast-like cells are key mechanisms in septic arthritis. Combining anti-RANKL and antibiotic therapy represents a promising therapy against this devastating disease.

Methotrexate (MTX), the most common first-line treatment in rheumatoid arthritis, is often insufficient, with no model capable of predicting response. The RA classification criteria, including autoantibodies and inflammation, were applied to 257 patients with newly diagnosed inflammatory arthritis in the cohort study, estimating MTX response. A total of 172 patients received MTX as the first anti-rheumatic drug and response was recorded at 1 year follow-up. A multivariable logistic regression used variables distinct between MTX-responders and non-responders to build the predictive model of response. Overall, 53.5% of MTX treated patients responded. Non-responders were frequently autoantibody positive, and responders were older, had lower RA classification scores, frequent corticosteroid use, and high insulin levels at baseline. Inflammation parameters were comparable between the groups. In the multiple regression analysis, the RA classification score and age at the first visit were strong predictors of MTX response (AUC 0.697, p < 0.0001). Including blood levels of insulin and IFNg improved AUC to 0.782 (p < 0.0001), offering early discrimination between responders and non-responders with high accuracy. Cellular experiments showed that insulin could be used to estimate MTX response by demonstrating that insulin changed the transcription of MTX target genes in the folate metabolism after exposing CD4+ cells ex vivo, which could facilitate MTX response in immune cells.

Septic arthritis is a severe and rapidly destructive joint infection, primarily caused by Staphylococcus aureus. The interplay between bacterial virulence factors and host immune responses determines disease progression and clinical outcomes. This review discusses the key bacterial factors that contribute to septic arthritis, including S. aureus cell wall components, surface proteins, and secreted toxins. In parallel, host-related factors, such as aging, immune responses, and genetic predispositions, are examined in conjunction with the impact of S. aureus infection on bone integrity and osteoimmunological mechanisms. Finally, this review highlights emerging therapeutic approaches, including targeted anti-virulence strategies, immune modulation, and anti-osteoclastogenic interventions, in mitigating joint damage. Understanding the multifaceted interactions between S. aureus and the host immune system is crucial for advancing treatment strategies and reducing morbidity associated with septic arthritis.

Background Insulin has epigenetic effect influencing gene expression. High peripheral insulin concentrations promote insulin resistance in autoimmunity. Oncoprotein survivin/ BIRC5 modulates glucose metabolism through chromatin binding and propagates IFNg effects in CD4+ cells. In this study, we explored how insulin influences chromatin binding and metabolic activity in autoimmune CD4+ cells of patients with rheumatoid arthritis (RA). Methods We profiled the metabolic activity of CD4+ cell clusters using single-cell transcriptome analysis in blood, synovial fluid and synovial tissue of RA patients. Through chromatin immunoprecipitation and sequencing, we identified the genes controlled by deposition of survivin and acetylated lysine 27 on histone H3 (H3K27ac) in CD4+ cells. Treating CD4+ cells with insulin and histone deacetylase inhibitors (HDACi), we identified changes in H3K27ac, linked those to transcription of the H3K27-survivin-controlled genes and the pathogenic phenotype of CD4+ cells using flow cytometry. Finally, we explored if anti-diabetic and anti-rheumatic drugs affect the metabolic profile and memory phenotype of the metabolic active CD4+ cells. Results Transcription of survivin/ BIRC5 and histone acetylation enzymes strongly correlate with active metabolism in blood CD4+ cells of RA patients. In RA synovial tissue, these BIRC5hi active T cell clusters are inflammatory, exhausted, and memory-like. Genome co-deposition of H3K27ac-survivin pinpointed the insulin-dependent genes in metabolic active CD4+ cells. These genes favored histone acetylation by suppressing methylating enzymes EZH2 and KMT2A , and T cell development by activating CD27, CD3G, and SCIMP . Inhibition of histone deacetylation reverted these transcriptional effects and supported cellular sensitivity to insulin. Insulin stimulation increased H3K27ac and together with HDACi, suppressed PDCD1 and IFNg transcription and production in CD4+CD27+CD45RO+ memory T cells. Immune modulation impacted metabolic activity and synergized with the effect of histone acetylation on insulin responsiveness in RA patients. Conclusions RA synovia is enriched with the metabolic active BIRC5 hiCD4+ T cell clusters. The metabolic activity of these cells is histone acetylation-dependent and mediates insulin effects through the H3K27ac-survivin epigenetic mechanism. Increasing plasma insulin levels when combined with insulin sensitivity, can be protective in RA dearmoring effector T cell function. Hence, increasing the insulin sensitivity by enabling histone acetylation presents a reasonable interventional goal to restore immune cell homeostasis in RA. ### Competing Interest Statement The authors have declared no competing interest.

Background Clinical evidence connects hyperinsulinemia with obesity, and development of type 2 diabetes (T2D). However, its role in autoimmune conditions was questioned. We investigated consequences of hyperinsulinemia for development of T2D and CD4 T cell function in rheumatoid arthritis (RA). Methods Incident T2D was prospectively studied in two independent RA cohorts and in gout patients matched to RA by age and gender, for 10 years. Effect of hyperinsulinemia and JAK-STAT signaling inhibition (JAKi) in CD4 T cells was studied by integrating transcriptional sequencing with direct effect of insulin, and JAKi on cell proliferation, DNA enrichment, and cytokine production. Results T2D was 3.2-2.5 times less prevalent in RA compared to gout, particularly in females. Hyperinsulinemia predicted the development of T2D, regardless of metabolic parameters and insulin resistance. Additionally, hyperinsulinemia correlated with the senescence-associated high serum levels of IL6, IL8, and VEGF. Hyperinsulinemia, along with ex-vivo exposure of CD4 cells to insulin, inhibited cell cycle progression and induced DNA enrichment through the suppression of the PI3K-Src kinases and cell cycle promoting genes. It also reduced IFNγ production. JAKi-treated CD4+ cells regained insulin sensitivity, which activated glucose metabolism and facilitated senescence. This insulin-dependent mechanism promoted the accumulation of naïve CD4 cells in JAKi-treated patients. Conclusions This study shows that insulin has important immunosuppressive ability controlling the adaptive immunity by suppressing IFNγ production and inducing senescence in the effector CD4 T cells. Inhibition of JAK-STAT signaling enhances insulin sensitivity and rejuvenates CD4 cell population in RA patients. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This work has been funded by grants from the Swedish Research Council (MB, 2017-03025 and 2017-00359), the Swedish Association against Rheumatism (MB, R-566961, R-751351 and R-860371; MD, R-968867; RP, R-969562, R-862061), the King Gustaf V:s 80-year Foundation (MB, FAI-2018-0519, FAI-2020-0653, FAI-2022-0882), the Regional agreement on medical training and clinical research between the Western Gotaland county council and the University of Gothenburg (MB, ALFGBG-717681, ALFGBG-965623; RP, ALFGBG-965012, ALFGBG-926621; MD, ALFGBG-888321), the University of Gothenburg. The authors declare that the funding sources have no role in study design; in the collection, analysis, and interpretation of data; in the writing of the report; and in the decision to submit the paper for publication. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The included swedish RA study is approved by the Swedish Ethical Review Authority and was registered at the Clinical Trials.gov with ID [NCT03449589][1]. The data collection of gout patients and their comorbidities was approved by the Ethical Review Board of Gothenburg, Sweden. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes This study includes publically available transcriptional data which is deposited at the Gene Expression Omnibus at the National Centre for Biotechnology Information, accession numbers GSE201669 and GSE138747. Other data from the present study are available upon request to the authors [1]: /lookup/external-ref?link_type=CLINTRIALGOV&access_num=NCT03449589&atom=%2Fmedrxiv%2Fearly%2F2024%2F03%2F10%2F2024.03.08.24303970.atom

Background: Rheumatoid arthritis today has no cure. Several clinical trials [1] have demonstrated ability of immunosuppressive drugs to prevent development of arthritis whereas others did not. Accurate selection of individuals at risk to develop arthritis from arthralgia is the key to increase efficacy and choice of prevention treatment. This enables an intensive search for molecular characteristics of processes preceding arthritis. Objectives: To identify inflammatory subtypes with highest and lowest probability of arthritis development. Methods: The study includes the cohort of 1012 patients assessed for joint pain at the Rheumatology Clinic during the period of two calendar years. The records of the 1st visit were carefully revised to exclude 303 patients with diagnosis of gout, polymyalgia rheumatica, spondylarthritis, primary Sjögren’s syndrome, SLE, trauma and paraneoplastic skeletal complaints. The remaining 709 patients with arthritis (ARTH) and clinically suspect arthralgia (ALG) were included in the 5-year prospective follow-up for new cases of arthritis. In addition to traditional inflammation parameters of CRP, ESR, hemoglobin, WBC, and platelet counts, blood samples of the 1st visit were investigated for autoantibodies ACPA, RF, cytokines IFNγ and IL8, growth factors Flt3-ligand, insulin, and oncoprotein survivin. These variables combined with gender and age, were put in the unsupervised learning algorithm to identify groups with similar profiles. Clinical characteristics of the formed clusters were analyzed to identify subtypes with high and low probability for arthritis development. Results: The unsupervised clustering resulted in 6 groups with diverse inflammatory profiles as summarized in Table 1. The largest group 5 had almost equal proportion of Arthritis and ALG diagnosis at the 1st visit, two of 6 groups were dominated by ARTH, and three of 6 groups were dominated by ALG. Individuals in the ALG groups 1,2,3 were significantly younger compared to the ARTH groups 4,6 (p<0.0001) and had higher frequency of females (p=0.0012). The ARTH groups had high IL8 and were remarkably different in autoantibody profile, and levels of IFNγ and insulin. The ALG groups were varying in WBC count, survivin and Flt3-ligand levels and, also, in presence of autoantibodies. In total, 83% of the new arthritis cases developed within 2 years and were most prevalent in the ARTH group 4, which dominated by autoantibody presence compared to all other groups (OR 4.25[2.33-7.69], p<0.0001) including the ALG group 2 with autoantibodies (OR 4.25[2.33-7.69], p=0.00027). The group with highest prevalence of new arthritis cases was also recognized by a combination of highest levels of IFNγ, IL8 and insulin compared to all other groups, while traditional inflammatory parameters CRP, ESR, platelet and WBC counts were not different from other groups. The highest WBC count coexisted with high survivin found in ALG group 1. The ALG group 3 had the longest time to new arthritis and was recognized by a combination of high Flt3-ligand and survivin. Conclusion: Here, we demonstrated that the presence of autoantibodies shared with high levels of Insulin, IFNγ and IL8 characterize Arthritis and identify the ALG patients at risk of progressing to arthritis. High levels of survivin and Flt3-ligand are valuable for long-term prognosis in arthritis development. REFERENCES: [1] Van der Helm-van Mil, A.H.M. Joint Bone Spine 2023 Jul;90(4):105543. doi: 10.1016/j.jbspin.2023.105543. Acknowledgements: NIL. Disclosure of Interests: None declared.Group 1Group 2Group 3Group 4Group 5Group 6Patients, n30999019927021Age, y47,2±16.847,8±13.942,7±16.154,7±15.450,1±14.757,4±16.8Gender, F90%81%78%70%70%71%Diagnosisat 1st visitALG, 93%Arth, 7%ALG, 100%Arth, 0ALG,83%Arth, 17%ALG, 35%Arth, 65%ALG, 59%Arth, 41%ALG, 33%Arth, 67%Presence of autoantibodies017%0100%00New ARTH14%11%12%34,8%10%14%Dominating variableWBCsurvivinaAB 17%Flt3-ligandsurvivinaAB 100%,InsulinIFNγ, IL8NoneESR+CRPPlateletsIL8