Umeå University's logo

Objectives: To investigate the occurrence and dynamics of secretory component-containing antibodies towards citrullinated proteins (SC ACPA) in plasma from pre-symptomatic individuals subsequently developing rheumatoid arthritis (RA).

Methods: We studied 319 individuals who had donated plasma prior to RA onset (median predating time 4.7 years), whereof 181 also donated samples after diagnosis. One hundred individuals were randomly selected from the same biobank cohorts to serve as controls. SC ACPA, total secretory IgA (TSIgA) and IgG ACPA were analysed in plasma by enzyme-linked immunoassays.

Results: Circulating SC ACPA levels in pre-symptomatic individuals and RA patients were significantly increased compared with controls [median (interquartile range) 108 (108), 179 (248) and 12.5 (537) AU/ml, respectively; P < 0.001], and SC ACPA levels in RA patients were significantly increased compared with pre-symptomatic individuals (P < 0.001). SC ACPA increased, in terms of both levels and proportion of positive samples, closer to symptom onset and diagnosis. TSIgA was not elevated compared with controls either during the pre-dating time or after diagnosis. The earliest detected SC ACPA positive sample was 9 years before symptom onset, as compared with 11 years for IgG ACPA. Only two pre-dating samples were positive for SC ACPA and negative for IgG ACPA.

Conclusions: Circulating SC ACPA responses arise and magnify during the asymptomatic phase of disease development in a subgroup of RA patients. This suggests mucosal involvement prior to both symptom onset and subsequent arthritis. As mirrored in the circulation, however, SC ACPA does not seem to precede the IgG ACPA response.

Clonal hematopoiesis (CH) becomes more prevalent with aging and may influence inflammatory diseases by altering immune function. While CH of indeterminate potential (CHIP) promotes inflammation in nonmalignant conditions, its relationship with rheumatoid arthritis (RA) remains unknown. We analyzed CHIP mutations in RA using two population-level cohorts and patients with newly diagnosed RA. CHIP was associated with prevalent RA in 10,089 FINRISK study participants with whole-exome sequencing (OR, 2.06; P = 0.029) and in the FinnGen cohort (n = 520,210; OR, 1.49; P < 0.001) using single-nucleotide polymorphism array–based CHIP annotation. In FinnGen, CHIP was also associated with inferior overall survival in participants with RA (P = 0.013). In newly diagnosed RA (n = 573), DNMT3A-mutated seropositive patients had increased inflammatory markers and disease activity compared with patients without CHIP. In contrast, TET2 mutations were enriched in seronegative RA (P = 0.009). Our findings provide further evidence for the context-dependent association between CHIP and inflammation, with potential therapeutic implications.

Objectives: To determine anticitrullinated protein antibody (ACPA) responses to novel peptides predicting the clinical outcomes of treatment-naïve early rheumatoid arthritis (RA) in the presymptomatic stage.

Methods: We analysed monoclonal ACPAs derived from RA patients, including a characterised protective ACPA (clone E4), along with plasma samples collected from 520 presymptomatic individuals, of whom 244 were also sampled at diagnosis of RA, and 530 population controls in Sweden. The validation cohort (The Nordic Rheumatic Diseases Strategy Trials and Registries, NORD-STAR) consisted of 690 treatment-naïve early RA patients. Responses to citrullinated or native alpha-enolase (ENO1) or peptidylarginine deiminase 4 (PAD4) peptides were analysed by bead-based multiplex flow immunoassay. Clinical outcomes included C-reactive protein (CRP) and the 28-joint disease activity score (DAS28) with its components: tender joint count (TJC), swollen joint count (SJC), and erythrocyte sedimentation rate (ESR).

Results: Monoclonal ACPAs displayed distinct binding patterns to ENO1 and PAD4 peptides. A time-dependent increase of ACPA response to citrullinated peptides was observed in the presymptomatic stage towards onset. In the presymptomatic (0.2-5 years before onset) and early RA stage, ACPA responses to several ENO1 and PAD4 peptides were associated with less severe RA, assessed as lower levels of CRP and DAS28 and its components. In early RA, the association was more pronounced in rheumatoid factor (RF)-negative patients based on lower SJC. In presymptomatic individuals, ACPA responses widely predicted lower disease activity in early RA and were more pronounced in 5 selected peptides.

Conclusions: Antibody responses to certain citrullinated epitopes are associated with lower disease activity in treatment-naïve early RA and appear years before symptom onset of RA.

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.

Objectives: The impact of body mass index (BMI) on cardiovascular risk in rheumatoid arthritis (RA) is unclear. RA characteristics may influence the association between BMI and risk. Disease activity, which predicts cardiovascular risk, is associated with obesity only among anticitrullinated antibody (ACPA)-positive patients. Biologics alter body composition and mitigate cardiovascular risk in RA. We explored the association of BMI with cardiovascular risk and whether this varied across ACPA status and biologic use.

Methods: We evaluated 3982 patients from an international observational cohort. Outcomes included (a) first major adverse cardiovascular event (MACE) encompassing myocardial infarction, stroke or cardiovascular death; and (b) all events comprising MACE, angina, revascularisation, transient ischaemic attack, peripheral arterial disease and heart failure. Multivariable Cox models stratified by centre risk evaluated the impact of BMI, ACPA, biologics and their two- and three-way interactions on outcomes.

Results: We recorded 192 MACE and 319 total events. No main effects of BMI, ACPA or biologics were observed. A three-way interaction between them on MACE (p-interaction<0.001) and all events (p-interaction=0.028) was noted. Among ACPA negative patients, BMI was inversely associated with MACE (HR 0.38 (95% CI 0.25 to 0.57)) and all events (HR 0.67 (0.49 to 0.92)) in biologic users but not non-users (p-for-interaction <0.001 and 0.012). Among ACPA-positive patients, BMI was associated with MACE (HR 1.04 [1.01–1.07]) and all events (HR 1.03 (1.00 to 1.06)) independently of biologic use.

Conclusions: BMI is inversely associated with cardiovascular risk only among ACPA-negative biologic users. In contrast, BMI is associated with cardiovascular risk in ACPA-positive patients independently of biologic use.

Objective: Patients with rheumatoid arthritis (RA) experience higher cardiovascular risk. Methotrexate may decrease this risk, although it is unclear whether males and females similarly benefit. We explored the influence of sex on the effect of methotrexate use on cardiovascular risk in RA.

Methods: An observational cohort of 4362 patients, 3223 (73.9%) females, without cardiovascular disease were included from an international cardiovascular consortium for RA. Outcomes were (a) major adverse cardiovascular events (MACE) including cardiovascular death, myocardial infarction, or stroke and (b) any ischemic cardiovascular events (iCVE) including MACE, angina, revascularization, transient ischemic attack, and peripheral arterial disease. The effects of sex, prevalent methotrexate use at enrollment visit and their interaction on MACE and iCVE were assessed with multivariable Cox regression models, reporting adjusted hazard ratios (HRs) and 95% confidence intervals (CIs).

Results: There were 237 first MACE and 358 first iCVE. The sex by methotrexate interaction was significant for MACE (p = 0.005) and iCVE (p = 0.006), suggesting the effect of methotrexate use on cardiovascular risk differed among males and females. In males, methotrexate use associated with lower risk of MACE (HR 0.32, [95% CI 0.12–0.83]) and iCVE (HR 0.43 [95% CI 0.21–0.85]). In females, methotrexate use was not associated with MACE (p = 0.267) or iCVE (p = 0.407). In sensitivity analyses, models with inverse probability of treatment weighting and models additionally adjusting for inflammation yielded similar results.

Conclusion: Methotrexate use associated with cardiovascular benefit in males but not females with RA and the effect was independent of inflammation.

Objective: Platelet activation is thought to participate in polymyalgia rheumatica (PMR) pathogenesis. Upon platelet activation, mitochondria are expelled into the extracellular space. However, whether extracellular mitochondria are present in patients with PMR and whether they can induce platelet activation is not known.

Methods: To investigate this, we measured markers of platelet activation (thrombospondin-1 [TSP-1]), mitochondrial-derived N-formyl methionine peptide (fMET), and autoantibodies directed toward specific mitochondrial antigen mitofusin-1 (MFN1) by enzyme-linked immunosorbent assay in plasma of healthy controls (HCs, n = 30) and patients with PMR without giant cell arteritis (GCA) (n = 45) and patients with PMR with GCA (n = 9) before and after treatment with glucocorticoid therapy. Ultrapure mitochondria were opsonized with plasma from patients with PMR without GCA (n = 45) or HCs (n = 10) and were subsequently incubated with HC platelets. Platelet activation was assessed by P-selectin levels using flow cytometry.

Results: Plasma levels of anti-MFN1 IgG were elevated in patients with PMR with and without GCA before glucocorticoid therapy when compared with HCs (P < 0.01 for both groups). Levels of anti-MFN1 IgG significantly reduced after treatment with glucocorticoids in both groups (P < 0.01). Levels of fMET were also significantly higher in patients with PMR with and without GCA before glucocorticoid therapy in comparison with HCs (P < 0.001 and P < 0.01, respectively). However, the levels of fMET only dropped significantly after therapy in patients with PMR without GCA (P < 0.001). Plasma levels of TSP-1 were elevated in patients with PMR with and without GCA before glucocorticoid therapy when compared to HC (P < 0.001 for both groups). After glucocorticoid therapy, plasma levels of TSP-1 decreased significantly only in patients with PMR without GCA (P = 0.023). Mitochondria opsonized with plasma from patients with PMR without GCA induced higher platelet activation regardless of treatment status as compared with plasma from HCs (P < 0.0001 and P < 0.01 for pretreatment and posttreatment).

Conclusion: Our results indicate increased platelet activation and the presence of mitochondrial antigens and antibodies in the circulation of patients with PMR. Blocking mitochondrial-mediated platelet activation may reduce inflammation in patients with PMR, with potential therapeutic implications.

Objective: Systemic inflammatory autoimmune diseases (SIADs) such as systemic lupus erythematosus (SLE), primary Sj & ouml;gren disease (pSS), and idiopathic inflammatory myopathies (myositis) are complex conditions characterized by shared circulating autoantibodies and clinical manifestations, including skin rashes, among others. This study was aimed at elucidating the genetics underlying these common features.

Methods: We performed targeted DNA sequencing of coding and regulatory regions from approximately 1,900 immune-related genes in a large cohort of 2,292 well-characterized Scandinavian patients with SIADs with SLE, pSS, and myositis as well as 1,252 controls. A gene-based functionally weighted genetic score for aggregate testing of all genetic variants, including rare variants, was complemented by in silico functional analyses and in vitro reporter experiments.

Results: Case-control association analysis detected known and potentially novel genetic loci in agreement with previous genetic and transcriptomics findings linked to the SIAD autoimmune background. Intriguingly, case-case comparisons between patient subgroups with and without specific autoantibodies revealed that the subgroups defined by antinuclear antibodies and anti-double-stranded DNA antibodies have unique genetic profiles reflecting their heterogeneity. When focusing on clinical features, we overall showed that dual-specificity phosphatase 1 (DUSP1) protective genetic variants lead to increased gene expression and potentially to anti-inflammatory effects on the SIAD-associated skin phenotype. This is consistent with recent genetic findings on eczema and with the previously reported down-regulation of the MAPK signaling-related gene DUSP1 in other skin disorders.

Conclusion: Together, this suggests common molecular mechanisms potentially underlying overlapping clinical manifestations shared among different disorders and informs clinical heterogeneity, which could be translated to improve disease diagnostic and treatment, also in more generalized disease frameworks.

The complement system plays a central role in the pathogenesis of Systemic Lupus Erythematosus (SLE), but most studies have focused on the classical pathway. Ficolin-3 is the main initiator of the lectin pathway of complement in humans, but its role in systemic autoimmune disease has not been conclusively determined. Here, we combined biochemical and genetic approaches to assess the contribution of ficolin-3 to SLE risk and disease manifestations. Ficolin-3 activity was measured by a functional assay in serum or plasma samples from Swedish SLE patients (n = 786) and controls matched for age and sex (n = 566). Genetic variants in an extended 300 kb genomic region spanning the FCN3 locus were analyzed for their association with ficolin-3 activity and SLE manifestations in a Swedish multicenter cohort (n = 985). Patients with ficolin-3 activity in the highest tertile showed a strong enrichment in an SLE cluster defined by anti-Sm/DNA/nucleosome antibodies (OR 3.0, p < 0.001) and had increased rates of hematological disease (OR 1.4, p = 0.078) and lymphopenia (OR = 1.6, p = 0.039). Genetic variants associated with low ficolin-3 activity mapped to an extended haplotype in high linkage disequilibrium upstream of the FCN3 gene. Patients carrying the lead genetic variant associated with low ficolin-3 activity had a lower frequency of hematological disease (OR 0.67, p = 0.018) and lymphopenia (OR 0.63, p = 0.031) and fewer autoantibodies (p = 0.0019). Loss-of-function variants in the FCN3 gene were not associated with SLE, but four (0.5 %) SLE patients developed acquired ficolin-3 deficiency where ficolin-3 activity in serum was depleted following diagnosis of SLE. Taken together, our results provide genetic and biochemical evidence that implicate the lectin pathway in hematological SLE manifestations. We also identify lectin pathway activation through ficolin-3 as a factor that contributes to the autoantibody response in SLE.