Prior studies have demonstrated abnormalities in the composition of the gastrointestinal microbiota in pediatric and adult patients with spondyloarthritis (SpA). In particular, diminished fecal ...abundance of Faecalibacterium prausnitzii and abnormalities in both directions in the abundance of the Bacteroides genus have been identified.
We obtained fecal specimens from 30 children with treatment-naïve enthesitis-related arthritis (ERA) and 19 healthy controls, as well as specimens from 11 adult patients with longstanding SpA and 10 adult healthy controls. All of the samples underwent sequencing of the 16S ribosomal DNA. A subset of the pediatric fecal samples was subjected to shotgun metagenomics sequencing.
ERA patients had decreased abundance of the anti-inflammatory F. prausnitzii A2-165 strain (41 ± 28% versus 54 ± 20% of all sequences matching F. prausnitzii, p = 0.084) and an increased abundance of the control F. prausnitzii L2/6 strain (28 ± 28% versus 15 ± 15%, p = 0.038). Similar trends were observed in adults with longstanding SpA (n = 11) and controls (n = 10). In contrast, the fecal abundance of Bacteroides fragilis was increased in ERA subjects (2.0 ± 4.0% versus 0.45 ± 0.7% of all sequences, p = 0.045), yet was diminished in adult subjects (0.2 ± % versus 1.0 ± % of all sequences, p = 0.106). Shotgun metagenomics sequencing of the fecal DNA in the pediatric subjects revealed diminished coverage of the butanoate pathway (abundance normalized to controls of 1 ± 0.48 versus 0.72 ± 0.33 in ERA, p = 0.037).
The anti-inflammatory F. prausnitzii A2-165 strain appears to be depleted in both pediatric and adult SpA. In contrast, B. fragilis may be depleted in adult disease yet abundant in pediatric SpA, suggesting developmental effects on the immune system.
Objective
The nuclear oncoprotein DEK is an autoantigen associated with juvenile idiopathic arthritis (JIA), especially the oligoarticular subtype. DEK is a secreted chemotactic factor. Abundant ...levels of DEK and DEK autoantibodies are found in inflamed synovium in JIA. We undertook this study to further characterize the nature of DEK autoantibodies in screening serum samples from 2 different cohorts that consisted mostly of patients with JIA.
Methods
DEK autoantibody levels were analyzed in sera from 33 JIA patients, 13 patients with other inflammatory conditions, and 11 healthy controls, as well as in 89 serum samples from JIA patients receiving anti–tumor necrosis factor (anti‐TNF) therapy. Recombinant His‐tagged full‐length DEK protein (1–375 amino acids aa) and the 187–375‐aa and 1–350‐aa His‐tagged DEK fragments made in a baculovirus system were used for enzyme‐linked immunosorbent assay (ELISA) and immunoblotting. The C‐terminal 25‐aa fragment of DEK was expressed in a glutathione S‐transferase–tagged vector. ELISA results were calculated as area under the curve by the trapezoidal rule.
Results
DEK autoantibody levels were significantly higher in patients with polyarticular JIA than in those with oligoarticular JIA, and were higher in patients with polyarticular JIA who had more active disease after cessation of anti‐TNF therapy. Immunoblotting against the C‐terminal 25‐aa fragment of DEK confirmed that this section of the DEK molecule is the most immunogenic domain.
Conclusion
DEK autoantibody levels are higher in patients with polyarticular JIA than in those with oligoarticular JIA, and higher in patients who have disease flares after cessation of anti‐TNF therapy. The C‐terminal 25‐aa fragment is the most immunogenic portion of DEK. These findings are significant with respect to the nature of DEK autoantibodies, their contribution to JIA pathogenesis, and their implications for JIA management.
Abstract
Objectives
AECAs are detected in multiple forms of vasculitis or vasculopathy, including JDM. High levels of tropomyosin alpha-4 chain (TPM4) gene expression in cutaneous lesions and TPM4 ...protein expression in some endothelial cells (ECs) have been proven. Furthermore, the presence of autoantibodies to tropomyosin proteins have been discovered in DM. We therefore investigated whether anti-TPM4 autoantibodies are an AECA in JDM and are correlated with clinical features of JDM.
Methods
The expression of TPM4 protein in cultured normal human dermal microvascular ECs was investigated by Western blotting. Plasma samples from 63 children with JDM, 50 children with polyarticular JIA (pJIA) and 40 healthy children (HC) were tested for the presence of anti-TPM4 autoantibodies using an ELISA. Clinical features were compared between JDM patients with and without anti-TPM4 autoantibodies.
Results
Autoantibodies to TPM4 were detected in the plasma of 30% of JDM, 2% of pJIA (P < 0.0001) and 0% of HC (P < 0.0001). In JDM, anti-TPM4 autoantibodies were associated with the presence of cutaneous ulcers (53%; P = 0.02), shawl sign rash (47%; P = 0.03), mucous membrane lesions (84%; P = 0.04) and subcutaneous edema (42%; P < 0.05). Anti-TPM4 autoantibodies significantly correlated with the use of intravenous steroids and IVIG therapy in JDM (both P = 0.01). The total number of medications received was higher in patients with anti-TPM4 autoantibodies (P = 0.02).
Conclusion
Anti-TPM4 autoantibodies are detected frequently in children with JDM and are novel myositis-associated autoantibodies. Their presence correlates with vasculopathic and other cutaneous manifestations of JDM that may be indicative of more refractory disease.
Objective
Autoantibodies recognizing 3‐hydroxy‐3‐methylglutaryl‐coenzyme A reductase (HMGCR) are associated with statin exposure, the HLA allele DRB1*11:01, and necrotizing muscle biopsies in adult ...myositis patients. The aim of this study was to characterize the features of juvenile anti‐HMGCR‐positive myositis patients.
Methods
The sera of 440 juvenile myositis patients were screened for anti‐HMGCR autoantibodies. Demographic and clinical features, responses to therapy, and HLA alleles were assessed. The features of anti‐HMGCR‐positive patients were compared to those of previously described adult patients with this autoantibody and to children with other myositis‐specific autoantibodies (MSAs).
Results
Five of 440 patients (1.1%) were anti‐HMGCR‐positive; none had taken statin medications. Three patients had rashes characteristic of juvenile dermatomyositis and 2 patients had immune‐mediated necrotizing myopathies. The median highest creatine kinase (CK) level of anti‐HMGCR‐positive subjects was 17,000 IU/liter. All patients had severe proximal muscle weakness, distal weakness, muscle atrophy, joint contractures, and arthralgias, which were all more prevalent in HMGCR‐positive subjects compared to MSA‐negative patients or those with other MSAs. Anti‐HMGCR‐positive patients had only partial responses to multiple immunosuppressive medications, and their disease often took a chronic course. The DRB1*07:01 allele was present in all 5 patients, compared to 26.25% of healthy controls (corrected P = 0.01); none of the 5 juvenile patients had DRB1*11:01.
Conclusion
Compared to children with other MSAs, muscle disease appears to be more severe in those with anti‐HMGCR autoantibodies. Like adults, children with anti‐HMGCR autoantibodies have severe weakness and high CK levels. In contrast to adults, in anti‐HMGCR‐positive children, there is a strong association with HLA–DRB1*07:01.
Objective
Novel autoantibody specificities including anti‐CCAR1 were recently discovered in adult patients with anti‐transcriptional intermediary factor (TIF1)–positive dermatomyositis (DM) and were ...associated with attenuated cancer emergence. The aims of the present study were to examine whether these autoantibodies occur in patients with juvenile‐onset DM (JDM) and to determine their associated features.
Methods
Sera from 150 patients with anti‐TIF1γ autoantibody‐positive JDM in a cross‐sectional cohort and 90 juvenile healthy controls were assayed for anti‐CCAR1, anti‐C1Z1, anti‐IMMT, anti‐TBL1XR1, and anti‐Sp4 autoantibodies. Demographics, myositis autoantibodies, clinical features, medications, outcomes, and HLA‐DRB1 and HLA‐DQA1 alleles were compared between those with and without these autoantibodies.
Results
Any one of the anti‐TIF1γ‐associated autoantibodies was present in 44 patients (29%) overall, including 25 (17%) with anti‐Sp4, 22 (15%) with anti‐TBL1XR1, 14 (9%) with anti‐CCAR1, 2 (1%) with anti‐C1Z1, and 2 (1%) with anti‐IMMT autoantibodies. These anti‐TIF1γ‐associated autoantibodies frequently co‐occurred. Patients with any of the anti‐TIF1γ‐associated autoantibodies had less frequent falling (34% 15 vs. 53% 56, P = 0.032) and lower peak muscle enzymes. None of the patients had cancer. Among White patients, HLA‐DRB1*03 was protective against an anti‐TIF1γ‐associated autoantibody (odds ratio 0.20, 95% confidence interval 0.07–0.52).
Conclusion
Autoantibodies associated with anti‐TIF1γ were found in isolation and in combination among a subset of patients with JDM. Patients with these autoantibodies had less severe muscle disease and were not enriched for HLA‐DRB1*03. Additional autoantibodies among patients with positive anti‐TIF1γ with JDM likely contribute to the heterogeneity of the anti‐TIF1γ serologic subgroup.
Objective
To determine the relationship between serum levels of S100A8/A9 and S100A12 and the maintenance of clinically inactive disease during anti–tumor necrosis factor (anti‐TNF) therapy and the ...occurrence of disease flare following withdrawal of anti‐TNF therapy in patients with polyarticular forms of juvenile idiopathic arthritis (JIA).
Methods
In this prospective, multicenter study, 137 patients with polyarticular‐course JIA whose disease was clinically inactive while receiving anti‐TNF therapy were enrolled. Patients were observed for an initial 6‐month phase during which anti‐TNF treatment was continued. For those patients who maintained clinically inactive disease over the 6 months, anti‐TNF was withdrawn and they were followed up for 8 months to assess for the occurrence of flare. Serum S100 levels were measured at baseline and at the time of anti‐TNF withdrawal. Spearman's rank correlation test, Mann‐Whitney U test, Kruskal‐Wallis test, receiver operating characteristic (ROC) curve, and Kaplan‐Meier survival analyses were used to assess the relationship between serum S100 levels and maintenance of clinically inactive disease and occurrence of disease flare after anti‐TNF withdrawal.
Results
Over the 6‐month initial phase with anti‐TNF therapy, the disease state reverted from clinically inactive to clinically active in 24 (18%) of the 130 evaluable patients with polyarticular‐course JIA; following anti‐TNF withdrawal, 39 (37%) of the 106 evaluable patients experienced a flare. Serum levels of S100A8/A9 and S100A12 were elevated in up to 45% of patients. Results of the ROC analysis revealed that serum S100 levels did not predict maintenance of clinically inactive disease during anti‐TNF therapy nor did they predict disease flare after treatment withdrawal. Elevated levels of S100A8/A9 were not predictive of the occurrence of a disease flare within 30 days, 60 days, 90 days, or 8 months following anti‐TNF withdrawal, and elevated S100A12 levels had a modest predictive ability for determining the risk of flare within 30, 60, and 90 days after treatment withdrawal. Serum S100A12 levels at the time of anti‐TNF withdrawal were inversely correlated with the time to disease flare (r = −0.36).
Conclusion
Serum S100 levels did not predict maintenance of clinically inactive disease or occurrence of disease flare in patients with polyarticular‐course JIA, and S100A12 levels were only moderately, and inversely, correlated with the time to disease flare.
Four-and-a-half LIM domains 1 (FHL1) is a muscle-specific protein. Autoantibodies against FHL1 were recently discovered in adults with idiopathic inflammatory myopathies (IIMs) and were found to be ...associated with clinical features and outcomes indicative of increased disease severity. Anti-FHL1 autoantibodies have not been described in children. Here, the prevalence and clinical features associated with anti-FHL1 autoantibodies were examined in a large North American cohort of juvenile patients with IIM.
Sera from 338 juvenile IIM patients and 91 juvenile healthy controls were screened for anti-FHL1 autoantibodies by ELISA. Clinical characteristics and HLA alleles of those with and without anti-FHL1 autoantibodies were compared among those with juvenile IIM.
Anti-FHL1 autoantibodies were present in 10.9% of juvenile IIM patients and 1.1% of controls. The frequency of anti-FHL1 autoantibodies among clinical and serologic subgroups did not differ. A higher percentage of Asian patients had anti-FHL1 autoantibodies (11% vs 0.7%; P = 0.002). Myositis-associated autoantibodies (MAAs) odds ratio (OR) 2.09 (CI 1.03, 4.32), anti-Ro52 autoantibodies specifically OR 4.17 (CI 1.83, 9.37) and V-sign rash OR 2.59 (CI 1.22, 5.40) were associated with anti-FHL1 autoantibodies. There were no differences in other features or markers of disease severity. No HLA associations with anti-FHL1 autoantibodies in Caucasian myositis patients were identified.
Anti-FHL1 autoantibodies are present in ∼11% of juvenile IIM patients and commonly co-occur with MAAs, including anti-Ro52 autoantibodies. In contrast to adult IIM, anti-FHL1 autoantibodies in juvenile myositis are associated with V-sign rash but not with other distinctive clinical features or worse outcomes.
Objective
To determine the frequency, time to flare, and predictors of disease flare upon withdrawal of anti–tumor necrosis factor (anti‐TNF) therapy in children with polyarticular forms of juvenile ...idiopathic arthritis (JIA) who demonstrated ≥6 months of continuous clinically inactive disease.
Methods
In 16 centers 137 patients with clinically inactive JIA who were receiving anti‐TNF therapy (42% of whom were also receiving methotrexate MTX) were prospectively followed up. If the disease remained clinically inactive for the initial 6 months of the study, anti‐TNF was stopped and patients were assessed for flare at 1, 2, 3, 4, 6, and 8 months. Life‐table analysis, t‐tests, chi‐square test, and Cox regression analysis were used to identify independent variables that could significantly predict flare by 8 months or time to flare.
Results
Of 137 patients, 106 (77%) maintained clinically inactive disease while receiving anti‐TNF therapy for the initial 6 months and were included in the phase of the study in which anti‐TNF therapy was stopped. Stopping anti‐TNF resulted in disease flare in 39 (37%) of 106 patients by 8 months. The mean/median ± SEM time to flare was 212/250 ± 9.77 days. Patients with shorter disease duration at enrollment, older age at onset and diagnosis, shorter disease duration prior to experiencing clinically inactive disease, and shorter time from onset of clinically inactive disease to enrollment were found to have significantly lower hazard ratios for likelihood of flare by 8 months (P < 0.05).
Conclusion
Over one‐third of patients with polyarticular JIA with sustained clinically inactive disease will experience a flare by 8 months after discontinuation of anti‐TNF therapy. Several predictors of lower likelihood of flare were identified.
Biologics treatment with antitumour necrosis factor alpha (TNFα) is efficacious in patients with juvenile idiopathic arthritis (JIA). Despite displaying clinical inactivity during treatment, many ...patients will flare on cessation of therapy. The inability to definitively discriminate patients who will relapse or continue to remain in remission after therapy withdrawal is currently a major unmet medical need. CD4 T cells have been implicated in active disease, yet how they contribute to disease persistence despite treatment is unknown.
We interrogated the circulatory reservoir of CD4
immune subsets at the single-cell resolution with mass cytometry (cytometry by time of flight) of patients with JIA (n=20) who displayed continuous clinical inactivity for at least 6 months with anti-TNFα and were subsequently withdrawn from therapy for 8 months, and scored as relapse or remission. These patients were examined prior to therapy withdrawal for putative subsets that could discriminate relapse from remission. We verified on a separate JIA cohort (n=16) the dysregulation of these circulatory subsets 8 months into therapy withdrawal. The immunological transcriptomic signature of CD4 memory in relapse/remission patients was examined with NanoString.
An inflammatory memory subset of CD3
CD4
CD45RA
TNFα
T cells deficient in immune checkpoints (PD1
CD152
) was present in relapse patients prior to therapy withdrawal. Transcriptomic profiling reveals divergence between relapse and remission patients in disease-centric pathways involving (1) T-cell receptor activation, (2) apoptosis, (3) TNFα, (4) nuclear factor-kappa B and (5) mitogen-activated protein kinase signalling.
A unique discriminatory immunomic and transcriptomic signature is associated with relapse patients and may explain how relapse occurs.
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