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Objectives JDM is an inflammatory myopathy characterized by prominent vasculopathy. AECAs are frequently detected in inflammatory and autoimmune diseases. We sought to determine whether AECAs correlate with clinical features of JDM, and thus serve as biomarkers to guide therapy or predict outcome. Methods Plasma samples from 63 patients with JDM, 49 patients with polyarticular JIA and 40 juvenile healthy controls were used to detect anti-heat shock cognate 71 kDa protein (HSC70) autoantibodies, a newly identified AECA, in ELISA assays. Clinical features were compared between JDM patients with and without anti-HSC70 autoantibodies. Results Anti-HSC70 autoantibodies were detected in 35% of patients with JDM, in 0% of patients with JIA (P Conclusion Anti-HCS70 autoantibodies are detected frequently in children with JDM and are novel myositis-associated autoantibodies correlating with disease severity.

Purpose of review One of the most important advances in medical research over the past 20 years has been the emergence of technologies to assess complex biological processes on a global scale. Although a great deal of attention has been given to genome-scale genetics and genomics technologies, the utility of studying the proteome in a comprehensive way is sometimes under-appreciated. In this review, we discuss recent advances in proteomics as applied to dermatomyositis/polymyositis as well as findings from other inflammatory diseases that may enlighten our understanding of dermatomyositis/polymyositis. Recent findings Proteomic approaches have been used to investigate basic mechanisms contributing to lung and skin disease in dermatomyositis/polymyositis as well as to the muscle disease itself. In addition, proteomic approaches have been used to identify autoantibodies targeting the endothelium in juvenile dermatomyositis. Studies from other inflammatory diseases have shown the promise of using proteomics to characterize the composition of immune complexes and the protein cargoes of exosomes. Summary There are many relevant scientific and clinical questions in dermatomyositis/polymyositis that can be addressed using proteomics approaches. Careful attention to both methodology and analytic approaches are required to obtain useful and reproducible data.

Background The response to treatment for juvenile idiopathic arthritis (JIA) can be staged using clinical features. However, objective laboratory biomarkers of remission are still lacking. In this study, we used machine learning to predict JIA activity from transcriptomes from peripheral blood mononuclear cells (PBMCs). We included samples from children with Native American ancestry to determine whether the model maintained validity in an ethnically heterogeneous population. Methods Our dataset consisted of 50 samples, 23 from children in remission and 27 from children with an active disease on therapy. Nine of these samples were from children with mixed European/Native American ancestry. We used 4 different machine learning methods to create predictive models in 2 populations: the whole dataset and then the samples from children with exclusively European ancestry. Results In both populations, models were able to predict JIA status well, with training accuracies > 74% and testing accuracies > 78%. Performance was better in the whole dataset model. We note a high degree of overlap between genes identified in both populations. Using ingenuity pathway analysis, genes from the whole dataset associated with cell-to-cell signaling and interactions, cell morphology, organismal injury and abnormalities, and protein synthesis. Conclusions This study demonstrates it is feasible to use machine learning in conjunction with RNA sequencing of PBMCs to predict JIA stage. Thus, developing objective biomarkers from easy to obtain clinical samples remains an achievable goal.

BACKGROUND: There is no effective way to predict cystic fibrosis (CF) pulmonary exacerbations (CFPE) before they become symptomatic or to assess satisfactory treatment responses. METHODS: RNA sequencing of peripheral blood neutrophils from CF patients before and after therapy for CFPE was used to create transcriptome profiles. Transcripts with an average transcripts per million (TPM) level>1.0 and a false discovery rate (FDR)1.5 fold change, FDR-adjusted P< 0.05). The model was able to successfully separate CF flare samples from those taken from the same patients in convalescence with an accuracy of 0.75 in both the training and testing cohorts. Six differently expressed genes were confirmed by real time PCR using both isolated neutrophils and whole blood from an independent cohort of CF patients before and after therapy, even though levels of myeloid related protein MRP8/14 dimers in plasma of CF patients were essentially unchanged by therapy. CONCLUSIONS: Our findings demonstrate the potential of machine learning approaches for classifying disease states and thus developing sensitive biomarkers that can be used to monitor pulmonary disease activity in CF.

Peripheral blood mononuclear cells (PBMCs) play an important role in the inflammation that accompanies intracranial aneurysm (IA) pathophysiology. We hypothesized that PBMCs have different transcriptional profiles in patients harboring IAs as compared to IA-free controls, which could be the basis for potential blood-based biomarkers for the disease. To test this, we isolated PBMC RNA from whole blood of 52 subjects (24 with IA, 28 without) and performed next-generation RNA sequencing to obtain their transcriptomes. In a randomly assigned discovery cohort of n = 39 patients, we performed differential expression analysis to define an IA-associated signature of 54 genes (q &lt, 0.05 and an absolute fold-change ≥ 1.3). In the withheld validation dataset, these genes could delineate patients with IAs from controls, as the majority of them still had the same direction of expression difference. Bioinformatics analyses by gene ontology enrichment analysis and Ingenuity Pathway Analysis (IPA) demonstrated enrichment of structural regulation processes, intracellular signaling function, regulation of ion transport, and cell adhesion. IPA analysis showed that these processes were likely coordinated through NF-kB, cytokine signaling, growth factors, and TNF activity. Correlation analysis with aneurysm size and risk assessment metrics showed that 4/54 genes were associated with rupture risk. These findings highlight the potential to develop predictive biomarkers from PBMCs to identify patients harboring IAs.

Systemic lupus erythematosus (SLE) is an autoimmune disease that is a challenge to diagnose and treat. There is an urgent need for biomarkers to help define organ involvement, and more effective therapies. A unique population of T cells, the CD3+CD4−CD8− (DNeg) cells, is significantly increased in lupus patients. Twenty-seven cases (53%) of pediatric SLE patients had elevated DNeg cells in their peripheral blood, which correlated with kidney function (R2 = 0.54). Significant infiltration of DNeg cells was observed in both adult and pediatric lupus kidneys by immunofluorescence. For the first time, this study provides direct evidence that DNeg cells facilitate kidney injury in preclinical 8-week-old MRL/lpr lupus mice. In lupus mice, the increase in DNeg cells tracked with worsening disease and correlated with kidney function (R2 = 0.85). Our results show that DNeg cells per se can cause kidney dysfunction, increase in number with increase in disease pathology, and could serve as a potential biomarker.

Background The pathology of juvenile dermatomyositis (JDM) is characterized by prominent vessel wall and perivascular inflammation. This feature of the disease has remained unexplained and under-investigated. We have hypothesized that plasma exosomes, which play an important role in inter-cellular communication, may play a role in the vascular injury associated with JDM. Objective To characterize the circulating exosomes of children with JDM and determine whether the small RNA cargoes within those exosomes are capable of altering transcriptional programs within endothelial cells. Design/methods We purified exosomes from plasma samples of children with active, untreated JDM (n = 6) and healthy controls (n = 9). We characterized the small RNA cargoes in JDM and control exosomes by RNA sequencing using the Illumina HiSeq 2500 platform. We then incubated isolated exosomes from healthy controls and children with JDM with cultured human aortic endothelial cells (HAEC) for 24 h. Fluorescence microscopy was used to confirm that both control and JDM exosomes were taken up by HAEC. RNA was then purified from HAEC that had been incubated with either control or JDM exosomes and sequenced on the Illumina platform. Differential expression of mRNAs from HAEC incubated with control or JDM exosomes was ascertained using standard computational methods. Finally, we assessed the degree to which differential gene expression in HAEC could be attributed to the different small RNA cargoes in JDM vs control exosomes using conventional and novel analytic methods. Results We identified 10 small RNA molecules that showed differential abundance when we compared JDM and healthy control exosomes. Fluorescence microscopy of labeled exosomes confirmed that both JDM and control exosomes were taken up by HAEC. Differential gene expression analysis revealed 59 genes that showed differential expression between HAEC incubated with JDM exosomes vs HAEC incubated with exosomes from controls. Statistical analysis of gene expression data demonstrated that multiple miRNAs exerted transcriptional control on multiple genes with HAEC. Conclusions Plasma exosomes from children with active, untreated JDM are taken up by HAEC and are associated with alterations in gene expression in those cells. These findings provide new insight into potential mechanisms leading to the targeting of vascular tissue by the immune system in JDM. Electronic supplementary material The online version of this article (10.1186/s12969-019-0347-0) contains supplementary material, which is available to authorized users.

Background The rupture of an intracranial aneurysm (IA) causes devastating subarachnoid hemorrhages, yet most IAs remain undiscovered until they rupture. Recently, we found an IA RNA expression signature of circulating neutrophils, and used transcriptome data to build predictive models for unruptured IAs. In this study, we evaluate the feasibility of using whole blood transcriptomes to predict the presence of unruptured IAs. Methods We subjected RNA from peripheral whole blood of 67 patients (34 with unruptured IA, 33 without IA) to next-generation RNA sequencing. Model genes were identified using the least absolute shrinkage and selection operator (LASSO) in a random training cohort (n = 47). These genes were used to train a Gaussian Support Vector Machine (gSVM) model to distinguish patients with IA. The model was applied to an independent testing cohort (n = 20) to evaluate performance by receiver operating characteristic (ROC) curve. Gene ontology and pathway analyses investigated the underlying biology of the model genes. Results We identified 18 genes that could distinguish IA patients in a training cohort with 85% accuracy. This SVM model also had 85% accuracy in the testing cohort, with an area under the ROC curve of 0.91. Bioinformatics reflected activation and recruitment of leukocytes, activation of macrophages, and inflammatory response, suggesting that the biomarker captures important processes in IA pathogenesis. Conclusions Circulating whole blood transcriptomes can detect the presence of unruptured IAs. Pending additional testing in larger cohorts, this could serve as a foundation to develop a simple blood-based test to facilitate screening and early detection of IAs.

Background Bennett et al identified the presence of interferon gene expression signatures in peripheral blood of children with systemic lupus erythematosus (pSLE). Here, we aim to identify how B cells contribute to those signatures. We hypothesize that disordered transcription in pSLE will be prominent in B cells and attributed to disease-specific epigenetic alterations. Thus we will not only identify important disease mechanisms in SLE, we will shed light on the genetics of SLE. Our previously published work demonstrated that most of the genetic risk for SLE located within non-coding regions of the genome appears to also contain higher than baseline epigenetic modifications of DNA and transcription factor binding sites that regulate and coordinate transcription. Methods Our specific aim is to assess regions of open chromatin in untreated pSLE and compare findings with healthy children. We propose to use assays of transposase-accessible chromatin with sequencing (ATACseq) to broadly survey open regions of chromatin and clarify the functional epigenome. In this pilot study, we propose to determine feasibility of performing this assay and developing methods for data analysis using 5 pediatric lupus patients and 5 healthy children. Anticipated results and conclusions Our long-term goal is to gain a mechanistic understanding of the aberrant transcriptional signatures in untreated SLE. The data generated by this pilot study will provide the basis for a rigorous power analysis, and firmly establish the working relationship between the Buffalo and Cohen Children’s Medical Center groups, both of which will be essential for a competitive application to NIH. Our preliminary findings from this pilot study will also allow us to begin the exciting process of linking the genetics and epigenetics of pSLE to the well-established transcriptional aberration. Acknowledgements This study is funded, in part, by an Arthritis Foundation-Childhood Arthritis and Rheumatology Research Alliance Small Pilot Grant.

Objective: Systemic lupus erythematosus (SLE) is a systemic autoimmune disease with varied morbidity and mortality. We assessed clinical presentations, autoantibody specificities and therapeutic interventions in Native American (NA) patients with SLE. Methods: Patients with SLE meeting 1997 American College of Rheumatology classification criteria (n=3148) were enrolled between 1992 and 2010 in the multiethnic, cross-sectional Lupus Family Registry and Repository. Clinical, demographic and therapeutic information were extracted from medical records using a standardised form and formalised training. Autoantibodies were assessed by indirect immunofluorescence (antinuclear antibodies (ANA) and antidouble-stranded DNA), precipitin (ENA) and ELISA (IgG and IgM anticardiolipins). Results: NA patients met SLE classification at a younger age (29.89±12.3 years) than European Americans (EA; 32.02±12.87, P=0.0157) and a similar age to African-Americans (AAs) and Hispanics (HIS). More NA patients had concurrent rheumatic diseases or symptoms, such as Raynaud's phenomenon, interstitial lung disease, Sjögren's syndrome and systemic sclerosis. Compared with EAs, NAs were more likely to have high-titre ANA (≥1:3240; P

Objective Although generally classified within the group of inflammatory myopathies, JDM displays many pathological features of vasculitis. Previous work has shown that AECA are abundant in other forms of vasculitis. We therefore investigated whether such antibodies might also be detected in JDM. Methods We screened plasma from children with JDM for the presence of AECA by western blotting and 2D gel electrophoresis (2DE) using proteins extracted from human aortic endothelial cells as the substrate. We performed mass spectrometry to identify candidate antigens from 2DE gels and used ELISA to confirm the presence of specific antibodies. Results We identified 22 candidate target autoantigens for AECA probed with JDM plasma. Interestingly, 17 of these 22 target antigens were proteins associated with antigen processing and protein trafficking. ELISA confirmed the presence of antibodies to heat shock cognate 71 kDa protein in JDM plasma, particularly in children with active, untreated disease. Conclusion Children with JDM express antibodies to autoantigens in endothelial cells. The clinical and pathological significance of such autoantibodies require further investigation.

Background Unruptured intracranial aneurysms (IAs) are typically asymptomatic and undetected except for incidental discovery on imaging. Blood-based diagnostic biomarkers could lead to improvements in IA management. This exploratory study examined circulating neutrophils to determine whether they carry RNA expression signatures of IAs. Methods Blood samples were collected from patients receiving cerebral angiography. Eleven samples were collected from patients with IAs and 11 from patients without IAs as controls. Samples from the two groups were paired based on demographics and comorbidities. RNA was extracted from isolated neutrophils and subjected to next-generation RNA sequencing to obtain differential expressions for identification of an IA-associated signature. Bioinformatics analyses, including gene set enrichment analysis and Ingenuity Pathway Analysis, were used to investigate the biological function of all differentially expressed transcripts. Results Transcriptome profiling identified 258 differentially expressed transcripts in patients with and without IAs. Expression differences were consistent with peripheral neutrophil activation. An IA-associated RNA expression signature was identified in 82 transcripts (p

Objective To determine whether gene expression profiles identified in peripheral whole blood samples could be used to determine therapeutic outcome in a cohort of children with newly diagnosed polyarticular juvenile idiopathic arthritis (JIA). Methods Whole blood samples from the Trial of Early Aggressive Therapy (TREAT) in JIA patients were analyzed on Illumina microarrays, and differential gene expression was compared to expression in healthy controls. Microarray results were validated by real-time quantitative polymerase chain reaction in an independent cohort of samples. Pathway analysis software was used to characterize gene expression profiles. Support vector machines were used to develop predictive models for different patient classes. Results Differential gene expression profiles for rheumatoid factor (RF)–positive and RF-negative patients were remarkably similar. Pathway analysis revealed a broad range of affected pathways, consistent with current mechanistic theories. Modeling showed that the prognosis at 6 months was strongly linked to gene expression at presentation, irrespective of treatment. Conclusion Gene expression is linked to therapeutic outcome, and gene expression in the peripheral blood may be a suitable target for a prognostic test.

Juvenile Idiopathic Arthritis (JIA) is one of the most common chronic disease conditions affecting children in the USA. As with many rheumatic diseases, there is growing interest in using genomic technologies to develop biomarkers for either diagnosis or to guide treatment (“personalized medicine”). Here, we explore the use of gene expression patterns in peripheral blood mononuclear cells (PBMC) as a first step approach to developing such biomarkers. Although PBMC carry many theoretical advantages for translational research, we have found that sample heterogeneity makes RNASeq on PBMC unsuitable as a first-step method for screening biomarker candidates in JIA. RNASeq studies of homogeneous cell populations are more likely to be useful and informative.

Author(s): Fotis, Lampros; Shaikh, Nur; Baszis, Kevin; French, Anthony; Tarr, Phillip; Grevich, Sriharsha; Lee, Peggy; Ringold, Sarah; Leroux, Brian; Leahey, Hannah; Yuasa, Megan; Foster, Jessica; Sokolove, Jeremy; Lahey, Lauren; Robinson, William; Newsom, Joshua; Stevens, Anne; Karasawa, Rie; Tamaki, Mayumi; Tanaka, Megumi; Sato, Toshiko; Yudoh, Kazuo; Jarvis, James N; Moncrieffe, Halima; Bennett, Mark F; Tsoras, Monica; Luyrink, Lorie; Xu, Huan; Prahalad, Sampath; Morris, Paula; Dare, Jason; Nigrovic, Peter A; Rosenkranz, Margalit; Becker, Mara; O’Neil, Kathleen M; Griffin, Thomas; Lovell, Daniel J; Grom, Alexei A; Medvedovic, Mario; Thompson, Susan D; Zhu, Lisha; Jiang, Kaiyu; Wong, Laiping; Buck, Michael J; Chen, Yanmin; Moncrieffe, Halima; Brungs, Laura; Liu, Tao; Wang, Ting; Jarvis, James N; Alsaeid, Khaled; Alfailakawi, Jasim; Alenezi, Hamid; Alsaeed, Hazim; Beukelman, Tim; Natter, Marc; Ilowite, Norm; Mieszkalski, Kelly; Burrell, Grendel; Best, Brian; Bristow, Helen; Carr, Shannon; Dennos, Anne; Kaufmann, Rachel; Kimura, Yukiko; Schanberg, Laura; Blier, Peter R; Boneparth, Alexis; Wenderfer, Scott E; Moorthy, L Nandini; Radhakrishna, Suhas M; Sagcal-Gironella, Anna Carmela P; von Scheven, Emily; Gedik, Kader Cetin; Siddique, Salma; Aguiar, Cassyanne L; Erkan, Doruk; Cohen, Ezra; Lee, Yvonne; Dossett, Michelle; Mehta, Darshan; Davis, Roger; Gilbert, Mileka; Goilav, Beatrice; Meidan, Esra

The Trial of Early Aggressive Therapy in Juvenile Idiopathic Arthritis (TREAT trial) was accompanied by a once-in-a-generation sample collection for translational research. In this paper, we report the results of whole blood gene expression analyses and genomic data-mining designed to cast light on the immunopathogenesis of polyarticular juvenile idiopathic arthritis (JIA). TREAT samples and samples from an independent cohort were analyzed on Affymetrix microarrays and compared to healthy controls. Data from the independent cohort were used to validate the TREAT data. Pathways analysis was used to characterize gene expression profiles. Furthermore, we correlated differential gene expression with new information about functional regulatory elements within the genome to develop models of aberrant gene expression in JIA. There was a strong concordance in gene expression between TREAT samples and the independent cohort. In addition, rheumatoid factor (RF)-positive and RF-negative patients showed only small differences on whole blood expression profiles. Analysis of the combined samples showed 158 genes represented by 176 probes that showed differential expression between TREAT subjects at baseline and healthy controls. None of the differentially expressed genes were encoded within linkage disequilibrium blocks containing single nucleotide polymorphisms known to be associated with risk for JIA. Functional analysis of these genes showed functional associations with multiple processes associated with innate and adaptive immunity, and appeared to reflect overall suppression of STAT1–3/interferon response factor-mediated pathways. Despite their limitations, whole blood expression profiles clearly distinguish children with polyarticular JIA from healthy controls. Whole blood expression profiles identify several immunologic pathways of biologic relevance that will need to be pursued in homogeneous cell populations in order to clarify mechanisms of pathogenesis. ClinicalTrials.gov registry #NCT00443430 , originally registered 2 March 2007 and last updated 30 May 2013.

This study sought to examine the differential effect of youth age on the association between parental uncertainty and both parent distress and youth depressive symptoms within the context of juvenile rheumatic disease.: The sample comprised 51 youth aged 9 to 17 years diagnosed with a juvenile rheumatic disease and their parents. Youth completed a measure of depressive symptoms (i.e., Children's Depression Inventory); parents completed measures of illness uncertainty (i.e., Parental Perceptions of Uncertainty Scale) and parental distress (i.e., Brief Symptom Inventory).: Parental uncertainty demonstrated a significant main effect on both parent distress and youth depressive symptoms. However, these main effects were qualified by significant parental uncertainty × youth age interactions. Parental uncertainty was significantly related to parent distress for both younger and older youth but was only significantly related to youth depressive symptoms in the sample of older youth.: Parental uncertainty due to illness seems to be more predictive of both parent and youth distress with increasing youth age. These findings indicate that the impact of parental illness uncertainty on adjustment outcomes is pronounced as children transition into developmental periods of increasing autonomy and independence.

OBJECTIVE To examine the longitudinal relationships between parent and child distress in a sample of children with juvenile rheumatic diseases (JRDs). DESIGN Cross-lagged panel correlation analysis tested the temporal precedence of parent distress versus child distress over a 1-year period. PARTICIPANTS Thirty-seven children (ages 9-17 years; 22 girls) diagnosed with JRD and their parents completed self-report measures on 2 occasions (assessment interval M = 12 months). PRIMARY OUTCOME MEASURES Child Depression Inventory and Brief Symptom Inventory. RESULTS Significant cross-sectional parent-child distress associations were observed at both time points. Moreover, Time 1 parent distress predicted child distress at Time 2 after child-reported functional ability was controlled; Time 1 child distress was unrelated to Time 2 parent distress. Cross-lagged panel correlations demonstrated the temporal precedence of parent distress relative to child distress in the parent-child distress relationship. CONCLUSIONS These preliminary findings underscore the importance of parent distress in parent-child transactional adjustment, and suggest a predominant role for parent distress in children's adjustment to JRDs.

Objective To compare the criteria for Wegener's granulomatosis (WG) of the American College of Rheumatology (ACR) with those of the European League Against Rheumatism/Pediatric Rheumatology European Society (EULAR/PRES) in a cohort of children with WG and other antineutrophil cytoplasmic antibody (ANCA)–associated vasculitides (AAVs), and to describe the interval to diagnosis, presenting features, and initial treatment for WG. Methods Eligible patients had been diagnosed by site rheumatologists (termed the “MD diagnosis”) since 2004. This diagnosis was used as a reference standard for sensitivity and specificity testing of the 2 WG classification criteria. Descriptive analyses were confined to ACR-classified WG patients. Results MD diagnoses of 117 patients (82 of whom were female) were WG (n = 76), microscopic polyangiitis (n = 17), ANCA-positive pauci-immune glomerulonephritis (n = 5), Churg-Strauss syndrome (n = 2), and unclassified vasculitis (n = 17). The sensitivities of the ACR and EULAR/PRES classification criteria for WG among the spectrum of AAVs were 68.4% and 73.6%, respectively, and the specificities were 68.3% and 73.2%, respectively. Two more children were identified as having WG by the EULAR/PRES criteria than by the ACR criteria. For the 65 ACR-classified WG patients, the median age at diagnosis was 14.2 years (range 4–17 years), and the median interval from symptom onset to diagnosis was 2.7 months (range 0–49 months). The most frequent presenting features by organ system were constitutional (89.2%), pulmonary (80.0%), ear, nose, and throat (80.0%), and renal (75.4%). Fifty-four patients (83.1%) commenced treatment with the combination of corticosteroids and cyclophosphamide, with widely varying regimens; the remainder received methotrexate alone (n = 1), corticosteroids alone (n = 4), or a combination (n = 6). Conclusion The EULAR/PRES criteria minimally improved diagnostic sensitivity and specificity for WG among a narrow spectrum of children with AAVs. Diagnostic delays may result from poor characterization of childhood WG. Initial therapy varied considerably among participating centers.

Objective: Examine age-related patterns of association between parent-reported illness intrusiveness andparent distress in parents of youth with juvenile rheumatic diseases (JRDs). Study Design: Cross-sectional multiple regression analysis tested child age as a moderator in the illness intrusiveness–distressrelationship. Participants: Fifty-two parents of children ages 9–17 diagnosed with JRD. Main OutcomeMeasures: The Illness Intrusiveness Scale—Parent Version and the Brief Symptom Inventory. Results:Parent-reported illness intrusiveness demonstrated a signi cant main effect on distress for all parents inthe sample. This was quali ed by an Illness Intrusiveness Child Age interaction. Illness intrusivenesswas found to be signi cantly related to distress among parents of older youth, but was only marginallyrelated to distress for parents of younger children. Conclusions: Results are consistent with family lifecycle development models of adjustment to childhood chronic illness. The clinical implications of the ndings are also discussed.Keywords: illness intrusiveness, parent adjustment, juvenile rheumatic disease

Objectives Serum cytokines play an important role in the pathogenesis of psoriatic arthritis (PsA) by initiating and perpetuating various cellular and humoral autoimmune processes. The aim of this study was to describe a broad spectrum of T- and B-cell cytokines, growth factors and chemokines in patients with PsA and healthy individuals. Methods A novel protein array system, denoted as multiplex cytokine assay was utilized to measure simultaneously the levels of 23 circulating cytokines of patients with PsA and healthy individuals. Additionally, correlational clustering and discriminant function analysis (DFA), two multivariate, supervised analysis methods were employed to identify a subset of biomarkers in order to describe potential functional inter-relationships among these pathological cytokines and identify biomarkers with prognostic and diagnostic utility. Results Univariate analysis demonstrated that serum levels of a complex set of immune and inflammatory modulating cytokines are significantly up-regulated in patients with PsA relative to unaffected controls including interleukin (IL)-10, IL-13, interferen (IFN)-alpha, epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), fibroblast growth factor [CCL3 macrophage inflammatory protein (MIP)-1alpha], CCL4 (MIP-1beta) and CCL11 (Eotaxin), while granulocyte-colony stimulating factor was significantly reduced in PsA patients. Correlational clustering was able to discriminate among, and hence subclassify, patients with varying levels of disease activity, which may prove useful in guiding therapy in these apparently phenotypically distinct disease subsets. DFA identified EGF, IFN-alpha, VEGF, CCL3 (MIP-1alpha) and IL-12p40 as analytes with the strongest discriminatory power among various PsA patients and controls. Conclusions Our findings suggest that these factors modulate PsA pathology and the articular involvement in a synergistic manner. Identifying factors could be used in the development of clinical diagnostic tests, which are valuable to guide evidence-based diagnosis and disease management of PsA.

Background We have previously shown that childhood-onset rheumatic diseases show aberrant patterns of gene expression that reflect pathology-associated co-expression networks. In this study, we used novel computational approaches to examine how disease-associated networks are altered in one of the most common rheumatic diseases of childhood, juvenile idiopathic arthritis (JIA). Methods Using whole blood gene expression profiles derived from children in a pediatric rheumatology clinical trial, we used a network approach to understanding the impact of therapy and the underlying biology of response/non-response to therapy. Results We demonstrate that therapy for JIA is associated with extensive re-ordering of gene expression networks, even in children who respond inadequately to therapy. Furthermore, we observe distinct differences in the evolution of specific network properties when we compare children who have been treated successfully with those who have inadequate treatment response. Conclusions Despite the inherent noisiness of whole blood gene expression data, our findings demonstrate how therapeutic response might be mapped and understood in pathologically informative cells in a broad range of human inflammatory diseases.

Neonatal-onset multisystem inflammatory disease is characterized by fever, urticarial rash, aseptic meningitis, deforming arthropathy, hearing loss, and mental retardation. Many patients have mutations in the cold-induced autoinflammatory syndrome 1 (CIAS1) gene, encoding cryopyrin, a protein that regulates inflammation.We selected 18 patients with neonatal-onset multisystem inflammatory disease (12 with identifiable CIAS1 mutations) to receive anakinra, an interleukin-1-receptor antagonist (1 to 2 mg per kilogram of body weight per day subcutaneously). In 11 patients, anakinra was withdrawn at three months until a flare occurred. The primary end points included changes in scores in a daily diary of symptoms, serum levels of amyloid A and C-reactive protein, and the erythrocyte sedimentation rate from baseline to month 3 and from month 3 until a disease flare.All 18 patients had a rapid response to anakinra, with disappearance of rash. Diary scores improved (P0.001) and serum amyloid A (from a median of 174 mg to 8 mg per liter), C-reactive protein (from a median of 5.29 mg to 0.34 mg per deciliter), and the erythrocyte sedimentation rate decreased at month 3 (all P0.001), and remained low at month 6. Magnetic resonance imaging showed improvement in cochlear and leptomeningeal lesions as compared with baseline. Withdrawal of anakinra uniformly resulted in relapse within days; retreatment led to rapid improvement. There were no drug-related serious adverse events.Daily injections of anakinra markedly improved clinical and laboratory manifestations in patients with neonatal-onset multisystem inflammatory disease, with or without CIAS1 mutations. (ClinicalTrials.gov number, NCT00069329 [ClinicalTrials.gov].).

PURPOSE OF THIS REVIEW Gene expression profiling is emerging as a promising methodology in pediatric rheumatology research. There is considerable interest in using this technology as the basis for diagnostic assays. This review will summarize the new knowledge. RECENT FINDINGS Most gene expression studies of children have been exploratory in nature. However, preliminary gene expression studies in juvenile dermatomyositis, systemic lupus erythematosus, and chronic forms of arthritis demonstrate both the promise and limits of this technology. It seems likely that gene expression profiling will significantly enhance our understanding of the immunopathology of childhood-onset rheumatic diseases; however, considerable impediments must be overcome before these assays move into the clinical arena. SUMMARY Gene expression profiling carries considerable potential to provide novel insights into the rheumatic diseases of childhood. Future developments will determine whether these technologies provide new clinical diagnostic or prognostic tools.

Several recommendations in the American Academy of Pediatrics policy statement “The Pediatrician's Role in Community Pediatrics” underscore the essential role of research as an agent of change to promote the health and well-being of children. This article provides (1) a framework for thinking about research in community pediatrics, (2) special considerations important in conducting community-level research, (3) an example of community-level research that has significantly decreased mortality in children (prevention of sudden infant death syndrome), (4) an example of a current issue illustrating the importance of community pediatrics research (promotion of school readiness), and (5) a discussion of future directions for research. Many of the leading health problems facing the United States as outlined in Healthy People 2010 are problems that affect children or have their roots in childhood and are likely to be addressed by community-level research and interventions. It seems clear that pediatricians should be learning to participate in, advocate for, and conduct more community pediatrics research.

Objective. To examine complaints for which children were referred to a pediatric rheumatology service and to determine whether there are specific complaints that are more likely to indicate the presence of chronic arthritis or chronic, systemic inflammatory disease.Methods. A retrospective chart review of 414 children referred to the pediatric rheumatology service at the Children’s Hospital of Oklahoma from April 1998 to July 2001.Results. Musculoskeletal pain was the most common complaint for which children were referred (n = 226). Of these, 111 had musculoskeletal pain as an isolated complaint. One of these children had a chronic inflammatory disease. Another 115 children had pain as 1 of several reasons for seeking a rheumatology consultation, including positive results on laboratory tests (antinuclear antibody, erythrocyte sedimentation rate, and rheumatoid factor). Nineteen of these children had a chronic inflammatory disease, including 12 with juvenile rheumatoid arthritis (JRA). Thus, musculoskeletal pain as a presenting complaint had a strong negative predictive value for the presence of either JRA (0.95) or any other chronic inflammatory disease that might be characterized by arthritis. Children who were referred, in part, because of positive antinuclear antibody and/or rheumatoid factor tests were no more likely to have a chronic inflammatory disease than children who did not include such results as a reason for referral. Joint swelling, in contrast, was the most likely complaint to be associated with a diagnosis of JRA.Conclusions. Musculoskeletal pain was the most common reason for referral to our pediatric rheumatology clinic. However, isolated musculoskeletal pain, in the absence of other signs or symptoms, is almost never a presenting complaint of children with chronic forms of arthritis. Children with arthritis more commonly present with complaints of joint swelling and/or gait disturbance. Neither ANA nor rheumatoid factor evaluations were useful in evaluating children with musculoskeletal complaints.

You have probably heard this dozens of times: “This might be an autoimmune disease. Order an ANA and a rheumatoid factor.” It is a common occurrence on adult medicine wards, and all too common in pediatrics as well. It probably stems from the fact that, until recently, very few medical schools had a pediatric rheumatologist; a third of the medical schools in the United States still do not have one. 1 Thus, pediatricians have had to undertake their evaluations of children with suspected rheumatic disease or musculoskeletal complaints using models derived from adult medicine. One of the most important advances in pediatric rheumatology over the past 15 years, and the one that has received the least attention, I think, has been the growing body of research that has shown just how inappropriate those models are for the evaluation of children with musculoskeletal complaints and/or suspected rheumatic disease. In this short article, I will summarize the most important findings from this research, inviting the reader to access the primary resources to better understand the significant differences between adult and pediatric rheumatology. Perhaps the most surprising finding has been the documentation that isolated musculoskeletal pain is almost never the complaint with which children with arthritis present to their primary care physicians. At the same time, evaluation of musculoskeletal pain is the single most common reason that primary care physicians refer children to pediatric rheumatologists. 2 In a retrospective study of 414 children evaluated at a university-based pediatric rheumatology clinic, McGhee and colleagues 3 reported that more that 75% of children referred for evaluation reported musculoskeletal pain as an isolated complaint or part of a complex complaint (eg, rash and pain). However, not a single child among 76 diagnosed with juvenile rheumatoid arthritis (JRA) presented with a chief complaint of isolated musculoskeletal pain. Indeed, only 16% of the children with arthritis included pain as one of their reasons for consulting their primary care physician. Rather, children with JRA invariably presented with joint swelling and/or gait disturbance as their primary complaint. The gait disturbance, when present, was typically better with activity and worse with rest, exactly the opposite of what one finds in mechanical musculoskeletal pain syndromes. The rarity with which children with chronic forms of arthritis verbalize pain probably accounts for the large number of children with arthritis (more than 60% in one

Background Juvenile dermatomyositis (JDM) and psoriasis are inflammatory disorders that share interferon- α induced responses and dysregulation of cytokines, including tumor necrosis factor alpha. Although 13% of patients with JDM have a family history of psoriasis there is little information concerning children with JDM and psoriasis. Observations We identified 3 children with both JDM and psoriasis. In 2 cases, psoriatic lesions occurred after the child's JDM symptoms had diminished following effective immunosuppressive therapy (high-dose intermittent intravenous methylprednisolone, methotrexate, and low dose oral corticosteroids). Patient 2, initially diagnosed as having psoriasis, was treated with prednisone and methotrexate but then developed classic JDM, which worsened following use of tumor necrosis factor alpha inhibitor and reduction of prednisone and methotrexate dosage. For each child, their history of JDM complicated the choice of therapy for psoriasis. Conclusions Two therapies commonly used to treat psoriasis —phototherapy and tumor necrosis factor-alpha antagonists —must be used with caution in patients with both JDM and psoriasis owing to their potential to exacerbate clinical manifestations of JDM. We discuss the implications affecting treatment of children with these dual diagnoses and consider the pathophysiology linking these 2 conditions.

Rheumatic disease manifestations and autoimmune phenomena are common in adults infected with human immunodeficiency virus (HIV). On the other hand, rheumatic disease manifestations are rare in children congenitally infected with HIV. We studied the relationship between autoimmune phenomena and rheumatic diseases by examining HIV-infected children for the presence of rheumatoid factors (RFs), immune complexes, and complement activation fragments. RFs (principally IgA) were detected in 12 of the 24 HIV-infected patients (50%) and in none of the uninfected (HIV-exposed, seroreverted) controls (n = 22). Mean levels of complement activation fragments C3a and Bb were elevated in the HIV-infected children compared with the controls. A correlation was seen between the presence of IgA RF and alternative complement pathway activation as measured by plasma levels of Bb. None of the children had clinical or laboratory evidence of rheumatic disease.

Studies in model organisms and humans have begun to reveal the complexity of the transcriptome. In addition to serving as passive templates from which genes are translated, RNA molecules are active, functional elements of the cell whose products can detect, interact with, and modify other transcripts. Gene expression profiling is the method most commonly used thus far to enrich our understanding of the molecular basis of rheumatoid arthritis in adults and juvenile idiopathic arthritis in children. The feasibility of this approach for patient classification (for example, active versus inactive disease, disease subsets) and improving prognosis (for example, response to therapy) has been demonstrated over the past 7 years. Mechanistic understanding of disease-related differences in gene expression must be interpreted in the context of interactions with transcriptional regulatory molecules and epigenetic alterations of the genome. Ongoing work regarding such functional complexities in the human genome will likely bring both insight and surprise to our understanding of rheumatoid arthritis.

Objective. To investigate the association between CD5+ B cell expression and IgM rheumatoid factor (IgM-RF) in juvenile rheumatoid arthritis (JRA). Methods. CD5+ B cell levels analyzed by flow cytometry and IgM-RF expression determined by enzymelinked immunosorbent assay were compared in children with JRA, children with other collagen vascular diseases, and healthy controls. Results. Children with polyarticular JRA had expanded populations of CDS+ B cells, and expansion of CDS+ B cells and IgM-RF both correlated with disease activity. Conclusion. The results indicate that an expanded CD5+ B cell population leads to IgM-RF production in patients with polyarticular JRA, as well as patients with RA.

Juvenile idiopathic arthritis (JIA) is a term used to denote a family of diseases of unknown etiology characterized by chronic inflammation of synovial membranes (1). Distinct phenotypes are recognized clinically, with specific immunogenetic markers associated with each of the phenotypes (2,3). While the JIA subtypes have commonly been assumed to have an “autoimmune” origin, our growing understanding of biologic complexity makes any such simple, linear hypothesis of disease pathogenesis unlikely (4). We have hypothesized that the pathogenesis of a common JIA subtype, polyarticular disease, involves complex interactions between innate and adaptive immunity not readily subsumed under a simple “autoimmunity” model (5,6). In support of this notion, we have demonstrated the presence of a population of hyperreactive neutrophils in children with polyarticular-onset JIA (7). Given our growing knowledge of how neutrophils regulate adaptive immunity (8), it is plausible to hypothesize that these abnormal neutrophils have a significant effect on adaptive immune mechanisms and the disease course in polyarticular JIA. The disease process in polyarticular JIA as seen in the normal clinical setting is not static. That is, children can be categorized based on their disease activity and response to therapy (i.e., active disease, inactive disease, remission of disease while taking medication, remission of disease while not taking medication), as Wallace and colleagues have shown (9). We have recently demonstrated that these clinically derived criteria for disease state have objective biologic identities, based on gene transcription profiling in peripheral blood mononuclear cells (PBMCs) (10). Thus, we have hypothesized that a practical way of gaining insight into the potential role of neutrophils in JIA pathogenesis is to study their function in specific disease states in conjunction with PBMCs. In the current study, we used a systems biology approach (gene transcription profiling and in silico modeling) to determine whether and how the neutrophil function may be altered in polyarticular JIA at different stages of the disease.

The development of biomarkers to predict response to therapy in polyarticular juvenile idiopathic arthritis (JIA) is an important issue in pediatric rheumatology. A critical step in this process is determining whether there is biologic meaning to clinically derived terms such as "active disease" and "remission." The aim of this study was to use a systems biology approach to address this question.We performed gene transcriptional profiling on children who fulfilled the criteria for specific disease states as defined by the consensus criteria developed by Wallace and colleagues. The study group comprised children with active disease (n = 14), children with clinical remission on medication (CRM; n = 9), children with clinical remission off medication (CR; n = 6), and healthy control children (n = 13). Transcriptional profiles in peripheral blood mononuclear cells (PBMCs) were obtained using Affymetrix U133 Plus 2.0 arrays.Hierarchical cluster analysis and predictive modeling demonstrated that the clinically derived criteria represent biologically distinct states. Minimal differences were seen between children with active disease and those with disease in CRM. Thus, underlying immune/inflammatory abnormalities persist despite a response to therapy. The PBMC transcriptional profiles of children whose disease was in remission did not return to normal but revealed networks of proinflammatory and antiinflammatory genes, suggesting that remission is a state of homeostasis, not a return to a normal state.Gene transcriptional profiling of PBMCs revealed that clinically derived criteria for JIA disease states reflect underlying biology. We also demonstrated that neither CRM nor CR status results in resolution of the underlying inflammatory process, but that these conditions are more likely to be states of balanced homeostasis between proinflammatory and antiinflammatory mechanisms.

One of the least-appreciated advances in pediatric rheumatology over the past 25 years has been the delineation of the many ways in which children with rheumatic disease differ from adults with the same illnesses. Furthermore, we are now learning that paradigms that are useful in evaluating adults with musculoskeletal complaints have limited utility in children. Nowhere is that more true than in the use of commonly used laboratory tests, particularly antinuclear antibody (ANA) and rheumatoid factor (RF) assays. This short review will provide the practitioner with the evidence base that supports a more limited use of ANA and RF testing in children.

Objective To investigate cognitive appraisal–adjustment relationships in Native American (NA) and Caucasian parents of children diagnosed with juvenile rheumatic diseases. Methods NA (n ¼ 16) and Caucasian (n ¼ 24) parents completed measures of disease status, illness intrusiveness, and adjustment; the rheumatologist provided estimates of disease severity. Results Hierarchical regression analysis revealed a moderating effect for racial group membership on the illness intrusiveness–parent adjustment relationship. Specifically, parent-perceived illness intrusiveness was more closely related to poorer adjustment among NA parents relative to Caucasian parents. Post hoc tests indicated that illness intrusiveness was significantly associated with poorer adjustment in NA parents, but was unrelated to parent adjustment in the Caucasian sample. Conclusions Results highlight the importance of examining racial group differences in cognitive appraisal–adjustment outcome relationships. Results are discussed with respect to the need for incorporating cultural issues into pediatric chronic illness research and treatment.

The term juvenile rheumatoid arthritis (JRA) is a erm used to describe a heterogeneous group of disorers that share the common characteristic of chronic nflammation and hyperplasia of synovial membranes. he term overlaps, but is not synonymous with, the erms juvenile chronic arthritis (used in the British iterature) and juvenile idiopathic arthritis (a term oming into more common use internationally). For all hree major subtypes of JRA (ie, systemic, polyarticlar, and pauciarticular), the histologic appearance of he inflamed synovium is indistinguishable from that een in adult rheumatoid arthritis, leading many invesigators to conclude that similar pathogenic mechaisms must be involved in these phenotypically disinct diseases. The pathogenesis of JRA is unknown. Current theries of disease pathogenesis have been informed by he following two observations: (1) the presence of D4 T-lymphocytes demonstrating a CD45RO “memory”) phenotype in inflamed synovium; and (2)

Rheumatoid arthritis and juvenile rheumatoid arthritis are histopathologically similar diseases characterized by chronic inflammation of the synovium. The pathogenesis of these diseases is unknown, but the emergence of gene expression profiling provides considerable promise that some of the complex, interconnected immunopathologic events underlying these diseases will soon be better understood. This review will summarize the potential use of gene expression profiling as a diagnostic or prognostic modality, and the potential benefits or limits of such uses. It will conclude with a short discussion of the potential for using gene expression profiling to identify novel targets of therapy in rheumatoid arthritis and related diseases.

Rheumatic diseases are prevalent in Native American adults at rates five to seven times higher than those seen in the Caucasian population. Little, however, has been published concerning rheumatic diseases in Native American children. The authors’ work in Oklahoma and with tribes on the northern Great Plains demonstrates high rates of childhood-onset rheumatic disease in this population. Familial disease is common, and large, multiplex kindreds showing strong founder effects are likely found within specific Native American tribes. However, a deeper understanding of the genetic and environmental triggers of rheumatic disease in Native Americans will require a respect and appreciation for the specific and unique cultural and social issues that impinge on research with Native Americans and other indigenous people.

SUMMARY Both innate and adaptive immune systems are thought to participate in the pathogenesis of rheumatoid arthritis in adults and children. The experiments reported here were undertaken to examine how immune complexes, potent stimulators of inflammation, may regulate cells of the adaptive immune system. Human T cells were prepared from peripheral blood by negative selection and incubated with bovine serum albumin (BSA)–anti-BSA immune complexes that were formed in the presence or absence of human C1q. C1q-bearing immune complexes, but not unopsonized complexes, elicited both TNF-α and IFN-γ secretion from human T cells. Secretion of both cytokines was time- and dose-dependent. Cross-linking C1q on the cell surface of T cells produced the same results. Cytokine secretion was not inhibited by blocking the C3b receptor (CR1, CD35) on T cells prior to incubation with immune complexes. Reverse transcriptase polymerase chain reaction (RT-PCR) of immune complex-stimulated cells revealed accumulation of both TNF-α and IFN-γ mRNA within 2 h post-stimulation. IL-2 was not detected in cell culture supernatants, but IL-2 receptor α chain (CD25) was detected in low density on a small proportion of T cells activated by C1q-bearing immune complexes. Secretion of both cytokines was inhibited partially, but not completely, by IL-10. These experiments show that immune complexes, potent inflammatory mediators, may activate T cells through a novel mechanism. These findings have implications for chronic inflammatory diseases in humans.

Background We have recently demonstrated the feasibility of developing prognostic biomarkers from whole blood gene expression profiles in children with newly diagnosed juvenile idiopathic arthritis (JIA). However, determining disease status at one year was more challenging. Objectives To determine whether prognostic biomarkers for disease status at 12 months can be developed from whole blood gene expression profiles in children with JIA after the initiation of therapy. Methods We studied whole blood expression profiles from children enrolled in the TREAT study, an NIH-funded clinical trial comparing methotrexate (MTX) with MTX + etanercept (Et) in children with newly-diagnosed JIA. We extracted RNA from whole blood (PAXgene) samples using conventional methods and removed globin genes using Ambion GlobinClear reagents. Labeling, hybridization reactions, and scanning were performed using conventional methods for Illumina WG-6_V3 whole genome microarrays. Gene expression profiles were then examined to determine those genes whose expression levels best predicted outcome (active vs inactive disease) at 12 months. Results Predictive models could be developed at 6 months from baseline data, as we have previously reported. Baseline expression profiles could not predict status at 12 months. Using 4 month data (the earliest point at which samples were collected from children on treatment), we were able to construct receiver operator curves (ROC) with strong predictive properties for disease status at 12 months (AUC =0.89 for RF-negative patients). Thus, longer term outcome is predictable only after children have initiated therapy. When examining the transcriptional changes that occur between baseline and 4 months, we found numerous genes showing significant differential expression (absolute fold change >1.2, p Conclusions These studies show the feasibility of developing whole blood prognostic biomarkers in children with JIA. Longer term prognoses can only be made after therapy has been initiated. A curious additional finding in this study was the appearance of different mechanisms of response in RF positive and RF negative patients after 4 months of therapy, a finding that may explain the relative refractoriness of RF positive patients to otherwise effective therapies. Acknowledgements This work was supported by R01-AR-060604, R01-AI-084200, and R01-AR-049762 from the National Insitutes of Health. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.4625

Background Juvenile idiopathic arthritis (JIA) is the most common rheumatic disease of childhood. The aetiology of JIA remains largely unknown. The number of effective medications for the treatment of RA has rapidly expanded, and studies have demonstrated that aggressive treatment of early JIA results in improved clinical outcomes. Therefore biomarkers for early diagnosis and prediction of therapeutic outcome are highly desirable. Exosomes, endosome-derived membrane microvesicles, contain specific RNA transcripts that are thought to be involved in cell-cell communication. Of these RNA transcripts, microRNAs are the most abundant. These microRNAs have great potential as disease biomarkers. Objectives To characterize exosomal microRNA profiles in children with polyarticular JIA and determining whether such profiles might be disease state-specific. Methods Plasma samples were obtained from children with RF-negative, polyarticular JIA as well as healthy controls (HC; n=5). Children with JIA had either active, untreated disease (AD, n=5) or were in clinical remission on medication (CRM; n=5) as established by the Wallace criteria. Exosomes were isolated from plasma using ExoQuick Precipitation methods. Expression of CD63 (a marker for exosomes) in all samples was validated by western blot. MicroRNA samples were isolated from exosomes using the miRNeasy Micro kit. Small RNA libraries were prepared using standard methods. Sequencing was performed using the Illumina HiSeq 2500 platform. The pass filter reads were mapped to the genome (GRCH37/hg19) using Bowtie 2. Differentially expressed (DE) microRNA were determined using edgeR using a P-value of 0.05. Results Plasma exosomal RNAs demonstrated both disease and disease-state specificity. For example, when we compared AD to HC, there were 30 DE miRNAs (24 down-regulated, 6 up-regulated) and 15 small nucleolar RNA (24 down-regulated, 6 up-regulated) that demonstrated a 1.6 fold change or greater. We found 11 DE miRNAs (down-regulated) and 35 DE snoRNA (up-regulated) that showed a 2.0 fold of greater difference when we compared CRM vs HC. Ingenuity pathway analysis demonstrated that the DE miRNAs are associated with inflammatory disease, connective tissue disorders and skeletal and muscular disorders (e.g. miR-10, miR-22, miR-29, miR-146, miR-154, miR-320, miR-374 and miR-500). Four miRNAs (miR-146a, miR-146b, miR-24-2 and miR-320b-2) were common to the AD vs HC and the CRM vs HC comparisons. Further evidence that the plasma exosomal RNA repertoire is disease-state specific was seen in direct comparison of the AD vs CRM samples. There were 21 DE miRNAs (2 down-regulated,19 up-regulated) and 24 DE snoRNA in the AD vs CRM comparison. These miRNA included let-7, miR-21, miR-143, miR-148, miR-154, miR-379 and miR-432, which associated with hereditary, skeletal, muscular, and developmental disorders. Conclusions The repertoire of small RNA molecules in the plasma of children with JIA differs significantly from those seen in the plasma of healthy children. Furthermore, these RNAs show disease-state specificity, suggesting that they may be directly involved in therapeutic response. Further characterization of these exosomal miRNAs in JIA may provide greater insight into pathobiology and open new doors for the development of blood-based biomarkers for JIA. Acknowledgements This work was supported by R01-AR-060604 from the National Institutes of Health and an Innovative Research Grant from the Arthritis Foundation. Disclosure of Interest : None declared DOI 10.1136/annrheumdis-2014-eular.4607

The presence of CD4+ cells in the synovium of children with juvenile rheumatoid arthritis has led to the generally accepted hypothesis that aberrant activation or regulation of acquired immunity is central to the pathogenesis of this family of diseases; however, this hypothesis remains unproven, and, indeed, a specific role for T cells in the process of chronic synovitis in rheumatoid disease has yet to be identified for either adults or children. In contrast, processes associated with innate immunity are undeniably involved in the pathophysiology of chronic synovitis in both rheumatoid arthritis and juvenile rheumatoid arthritis. The presence of neutrophils in the synovial fluid, complement activation, and immune complex accumulation in the synovial fluid and serum all indicate an active inflammatory process. It is reasonable to hypothesize, therefore, that there are important clues to the cause of juvenile rheumatoid arthritis to be gleaned from a more careful study of inflammation or innate immunity. This review will focus on the roles of complement, immune complexes, and the vascular endothelium in the pathogenesis of juvenile rheumatoid arthritis. In so doing, it will reexamine the dogma of the central role of the T cell in juvenile rheumatoid arthritis disease pathogenesis and offer new paradigms with which to understand this and other rheumatic diseases of childhood.

SUMMARY Data published from In vitro studies have shown that IgM-rheumatoid factor (RF)-bearing immune complexes possess several biological features that may contribute to their pathogenicity. However, no studies have demonstrated that such complexes exist at sites of inflammation in children with rheumatoid disease. We used two methods of sequential column chromatography to purify immune complexes from synovial fluids of children with JRA. We demonstrate that high molecular weight complexes contain IgM-RF, have not bound C4 in vivo, but activate the classical pathway in vitro. In contrast, complexes which have bound C3 in vivo do not contain IgM- RF and are weak complement activators in vitro.

Juvenile rheumatoid arthritis (JRA) has a complex, poorly characterized pathophysiology. Modeling of transcriptosome behavior in pathologic specimens using microarrays allows molecular dissection of complex autoimmune diseases. However, conventional analyses rely on identifying statistically significant differences in gene expression distributions between patients and controls. Since the principal aspects of disease pathophysiology vary significantly among patients, these analyses are biased. Genes with highly variable expression, those most likely to regulate and affect pathologic processes, are excluded from selection, as their distribution among healthy and affected individuals may overlap significantly. Here we describe a novel method for analyzing microarray data that assesses statistically significant changes in gene behavior at the population level. This method was applied to expression profiles of peripheral blood leukocytes from a group of children with polyarticular JRA and healthy control subjects. Results from this method are compared with those from a conventional analysis of differential gene expression and shown to identify discrete subsets of functionally related genes relevant to disease pathophysiology. These results reveal the complex action of the innate and adaptive immune responses in patients and specifically underscore the role of IFN-γ in disease pathophysiology. Discriminant function analysis of data from a cohort of patients treated with conventional therapy identified additional subsets of functionally related genes; the results may predict treatment outcomes. While data from only 9 patients and 12 healthy controls was used, this preliminary investigation of the inflammatory genomics of JRA illustrates the significant potential of utilizing complementary sets of bioinformatics tools to maximize the clinical relevance of microarray data from patients with autoimmune disease, even in small cohorts.

Objectives To examine referral patterns from primary care physicians for children with pauciarticular juvenile rheumatoid arthritis (JRA) and to determine whether children with pauciarticular JRA referred to pediatric rheumatologists differ in clinical presentation from children referred to other specialists. Design A retrospective records review of 49 patients with pauciarticular JRA was performed. Records were reviewed to determine the specialty of the referring physician and whether the children referred had symptoms and signs compatible with a synovitis at the time primary care was sought. Setting Inner-city tertiary pediatric rheumatology referral center. Participants Children with pauciarticular JRA. Main Outcome Measures Identification of referral patterns of primary care physicians. Associated morbidity owing to JRA was ascertained at the time of referral. Results Most children with pauciarticular JRA (62%) were referred to orthopedic surgeons prior to referral for pediatric rheumatology care. No differences in clinical symptoms were seen between children referred to pediatric rheumatologists and those referred to orthopedic surgeons. Children referred initially to orthopedic surgeons were younger than those referred to pediatric rheumatologists. Conclusion A notable number of children with pauciarticular JRA are referred to orthopedic surgeons prior to the establishment of that diagnosis, even when such children present with unequivocal signs of synovitis. This may be owing to the misconception that arthritis is rare in preschool-aged children or to the difficulty of ascertaining the presence of synovitis in younger children.

Epidemiologic evidence supports the hypothesis that genetic factors are involved in disease risk and severity in juvenile rheumatoid arthritis (JRA). Because the prevalence of specific HLA alleles differs among ethnic groups, we have hypothesized differences in JRA disease expression and immunogenetics in children with JRA.

JRA SYNOVIAL FLUID IgG FRACTIONS INDUCE A BROAD SPECTRUM OF PROINFLAMATORY CYTOKINES FROM PERIPHERAL BLOOD MONONUCLEAR CELLS . • 2344