Gene expression profiling in neutrophils from children with polyarticular juvenile idiopathic arthritis.

OBJECTIVE: We have previously reported a defect in neutrophil activation in children with polyarticular juvenile idiopathic arthritis (JIA). The current study was undertaken to determine whether gene expression abnormalities persist in JIA in remission and to use systems biology analysis to elucidate pathologic pathways in polyarticular JIA. METHODS: We performed gene expression profiling on neutrophils from children with polyarticular JIA. Children were grouped according to disease status. We studied 14 children with active disease who were taking medication, 8 children with clinical remission of disease who were taking medication (CRM status), and 6 children with clinical remission of disease who were not taking medication (CR status). We also studied 13 healthy children whose age ranges overlapped those of the patients. RESULTS: Neutrophil abnormalities persisted in children with polyarticular JIA even after disease remission was achieved. Children with active disease and those with CRM status showed no differences in expression of specific genes, although they could be separated on cluster analysis. A comparison of children with CR status and healthy control children revealed networks of pro- and antiinflammatory genes that suggested that remission is a state of homeostasis and balance rather than a return to normal immune function. Furthermore, gene overexpression in patients with CR status supports the hypothesis that neutrophils play a role in regulating adaptive immunity in this disease. CONCLUSION: Neutrophil gene profiling in polyarticular JIA suggests important roles for neutrophils in disease pathogenesis. These findings suggest the presence of complex interactions between innate and adaptive immunity, that are not easily modeled in conventional, linear, reductionist systems.

The meaning of clinical remission in polyarticular juvenile idiopathic arthritis: gene expression profiling in peripheral blood mononuclear cells identifies distinct disease states.

OBJECTIVE: 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. METHODS: 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. RESULTS: 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. CONCLUSION: 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.

Evidence for chronic, peripheral activation of neutrophils in polyarticular juvenile rheumatoid arthritis.

Although strong epidemiologic evidence suggests an important role for adaptive immunity in the pathogenesis of polyarticular juvenile rheumatoid arthritis (JRA), there remain many aspects of the disease that suggest equally important contributions of the innate immune system. We used gene expression arrays and computer modeling to examine the function in neutrophils of 25 children with polyarticular JRA. Computer analysis identified 712 genes that were differentially expressed between patients and healthy controls. Computer-assisted analysis of the differentially expressed genes demonstrated functional connections linked to both interleukin (IL)-8- and interferon-gamma (IFN-gamma)-regulated processes. Of special note is that the gene expression fingerprint of children with active JRA remained essentially unchanged even after they had responded to therapy. This result differed markedly from our previously reported work, in which gene expression profiles in buffy coats of children with polyarticular JRA reverted to normal after disease control was achieved pharmacologically. These findings suggest that JRA neutrophils remain in an activated state even during disease quiescence. Computer modeling of array data further demonstrated disruption of gene regulatory networks in clusters of genes modulated by IFN-gamma and IL-8. These cytokines have previously been shown to independently regulate the frequency (IFN-gamma) and amplitude (IL-8) of the oscillations of key metabolites in neutrophils, including nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) and superoxide ion. Using real-time, high-speed, single-cell photoimaging, we observed that 6/6 JRA patients displayed a characteristic defect in 12% to 23% of the neutrophils tested. Reagents known to induce only frequency fluctuations of NAD(P)H and superoxide ion induced both frequency and amplitude fluctuations in JRA neutrophils. This is a novel finding that was observed in children with both active (n = 4) and inactive (n = 2) JRA. A subpopulation of polyarticular JRA neutrophils are in a chronic, activated state, a state that persists when the disease is well controlled pharmacologically. Furthermore, polyarticular JRA neutrophils exhibit an intrinsic defect in the regulation of metabolic oscillations and superoxide ion production. Our data are consistent with the hypothesis that neutrophils play an essential role in the pathogenesis of polyarticular JRA.

Clinical utility of antinuclear antibody tests in children.

BACKGROUND: Antinuclear antibody (ANA) tests are frequently used to screen children for chronic inflammatory diseases such as systemic lupus erythematosus (SLE). However, the diagnostic utility of this test is limited because of the large number of healthy children who have low-titer positive tests. We sought to determine the clinical utility of ANA tests in screening children for rheumatic disease and to determine whether there are specific signs or symptoms that enhance the clinical utility of ANA tests in children. METHODS: We undertook a retrospective analysis of 509 new patient referrals. Charts of patients referred because of results of ANA testing were selected for further analysis. Children with JRA, SLE, and other conditions were compared using demographic data, chief complaints at the time of presentation, and ANA titers. RESULTS: One hundred ten patients were referred because of an ANA test interpreted as positive. Ten patients were subsequently diagnosed with SLE. In addition, we identified one patient with mixed connective tissue disease, and an additional child with idiopathic Raynaud's phenomenon. Eighteen children of the children referred for a positive ANA test had juvenile rheumatoid arthritis (JRA). Another 80 children with positive ANA tests were identified, the majority of whom (n = 39, 49%) had musculoskeletal pain syndromes. Neither the presence nor the titer of ANA served to distinguish children with JRA from children with other musculoskeletal conditions. Children with JRA were readily identified on the basis of the history and physical examination. Children with SLE were therefore compared with children with positive ANA tests who did not have JRA, designated the "comparison group." Non-urticarial rash was more common in children with SLE than in children without chronic inflammatory disease (p = 0.007). Children with SLE were also older (mean +/- sd = 14.2 +/- 2.5 years) than the comparison group (11.0 +/- 3.6 years; p = 0.001). ANA titer was also a significant discriminator between children with SLE and children without chronic inflammatory disease. The median ANA titer in children with SLE was 1: 1,080 compared with 1:160 for other children (p < 0.0001). ANA titers of >/=1,080 had a positive predictive value for SLE of 1.0 while titers of

Identifying children with chronic arthritis based on chief complaints: absence of predictive value for musculoskeletal pain as an indicator of rheumatic disease in children.

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.