FcγRIIIa-Syk Co-signal Modulates CD4+ T-cell Response and Up-regulates Toll-like Receptor (TLR) Expression.

CD4(+) T-cells in systemic lupus erythematosus (SLE) patients show altered T-cell receptor signaling, which utilizes Fc-receptor γ-chain FcRγ-Syk. A role for FcγRIIIa activation from immune complex (IC) ligation and sublytic terminal complement complex (C5b-9) in CD4(+) T-cell responses is not investigated. In this study, we show that the ICs present in SLE patients by ligating to FcγRIIIa on CD4(+) T-cells phosphorylate Syk and provide a co-stimulatory signal to CD4(+) T-cells in the absence of CD28 signal. This led to the development of pathogenic IL-17A(+) and IFN-γ(high) CD4(+) T-cells in vitro. Cytokines IL-1ß, IL-6, TGF-ß1, and IL-23 were the only requirement for the development of both populations. SLE patients CD4(+) T-cells that expressed CD25, CD69, and CD98 bound to ICs showed pSyk and produced IFN-γ and IL-17A. This FcγRIIIa-mediated co-signal differentially up-regulated the expression of IFN pathway genes compared with CD28 co-signal. FcγRIIIa-pSyk up-regulated several toll-like receptor genes as well as the HMGB1 and MyD88 gene transcripts. ICs co-localized with these toll-like receptor pathway proteins. These results suggest a role for the FcγRIIIa-pSyk signal in modulating adaptive immune responses.

Induced expression of FcγRIIIa (CD16a) on CD4+ T cells triggers generation of IFN-γhigh subset.

Whether or not CD4(+) T-cells express low affinity receptor FcγRIIIa (CD16a) in disease pathology has not been examined in great detail. In this study, we show that a subset of activated CD4(+) T-cells in humans express FcγRIIIa. The ligation of FcγRIIIa by immune complexes (ICs) in human CD4(+) T-cells produced co-stimulatory signal like CD28 that triggered IFN-γ production. The induced expression of FcγRIIIa on CD4(+) helper T-cells is an important finding since these receptors via ITAM contribute to intracellular signaling. The induced expression of FcγRIIIa on CD4(+) T helper cells and their ability to co-stimulate T-cell activation are important and novel findings that may reveal new pathways to regulate adaptive immune responses during inflammation and in autoimmunity.

Laboratory Studies in Autoimmune Diseases.

Autoimmune diseases (AID) are a great percentage of the patients needing to be seen by Rheumatologists. Their initial work-up by their primary care physician (PCP) can be helpful in determining who needs to be seen in consultation. If interpreted properly, initial laboratory studies can help the PCP know when to initiate a consultation. The following laboratory studies as outlined will provide the PCP with the necessary information to determine if a further AID work-up is indicated.

Laboratory Evaluation in Pediatric Autoimmune Diseases.

Pediatric autoimmune diseases are chronic lifelong disorders associated with potential disability and increased morbidity and mortality if not properly recognized and treated. On the basis of largely expert opinion in addition to observational studies, children with suspected autoimmune disease should undergo general laboratory and autoantibody screening. (1)(2)(4)(6)(7)(11) There can be many causes of positive antinuclear antibody test results, including, but not limited to, autoimmune disease. On the basis of expert opinion and observational studies, a thorough history and physical examination as well as laboratory evaluation is recommended (often in consultation with a pediatric rheumatologist) to elucidate the cause for a positive test result. (4) (6)(11)

Measurement and evaluation of isotypes of anti-citrullinated fibrinogen and anti-citrullinated alpha-enolase antibodies in juvenile idiopathic arthritis.

OBJECTIVES: Our objective was to evaluate sera from juvenile idiopathic arthritis (JIA) patients to investigate the presence of isotypes (IgA, IgG, IgM) of anti-citrullinated fibrinogen and anti-α-enolase antibodies and their association with rheumatoid factor (RF) and anti-cyclic citrullinated peptide (anti-CCP) antibody isotypes. METHODS: Sera were obtained from 89 JIA patients and were measured for isotypes (IgA, IgM) of anti-citrullinated and native fibrinogen and anti-α-enolase antibodies by enzyme-linked immunosorbent assay. Results were compared to anti-CCP antibody isotypes and RF isotypes, in addition to previously measured IgG anti-citrullinated fibrinogen and α-enolase antibodies. RESULTS: IgA anti-citrullinated fibrinogen antibodies were positive in 20 JIA patients and IgM in 11 JIA patients. Two IgM RF-positive polyarthritis patients were positive for all 3 isotypes of anti-citrullinated fibrinogen antibodies. IgA anti-citrullinated α-enolase antibodies were positive in 7 JIA patients and IgM in 9 JIA patients. IgA and IgG anti-citrullinated fibrinogen antibodies were commonly found in JIA patients positive for IgG anti-CCP antibodies and IgM RF. IgG anti-CCP antibodies and IgM RF levels were significantly higher in JIA patients with 3 or more anti-citrullinated autoantibody isotypes present. CONCLUSIONS: We have shown that isotypes of anti-citrullinated fibrinogen and α-enolase can be found in the serum of children with JIA of all onset types. Citrullinated autoantibody isotype diversity may indicate a more severe disease course in JIA patients.

Antibodies to histone in the pediatric population: a retrospective chart review.

BACKGROUND: Antibodies to histone have been associated in the adult literature with systemic lupus erythematosus(SLE) and drug induced lupus(DILE). Little data is available regarding the spectrum of pathology that antibodies to histone encompass in the pediatric population. Prior studies suggest an association with SLE, juvenile idiopathic arthritis(JIA), uveitis and linear scleroderma. METHODS: Patient charts were reviewed that contained positive anti-histone antibody testing during a consecutive three year period. Patient diagnosis along with the presence of: anti-histone antibody titer, ANA, and the presence of other autoantibodies to SSA, SSB, Sm, RNP, dsDNA and chromatin were obtained. The frequency of SLE, JIA and DILE was further investigated in specific subsets. RESULTS: 139 individual charts were reviewed containing 41 different diagnoses. The most common diagnosis was hypermobility arthralgia with 22 patients. The most frequent rheumatologic diagnosis was JIA(nonsystemic) with 19. 13 patients in this study were diagnosed with SLE and 2 with DILE. 18 patients had other autoantibody production, of these, 11 had SLE or DILE. Only one of 62 patients with a weak antihistone antibody titer(1.0-1.5) was diagnosed with SLE. When strong titers are present(> 2.5), the antihistone antibody test was associated with a greater than 50% incidence of an underlying rheumatologic disease and ten times higher incidence of SLE than a weak titer. In regards to the frequency of SLE, there was a statistically significant difference between weak and moderate titers and between weak and strong titers. CONCLUSION: The presence of anti-histone antibody was observed in a variety of diagnoses in the pediatric population. Overall, the presence of anti-histone antibodies appears to have poor diagnostic utility for any specific condition. However, diagnostic utility for SLE does appear to improve with higher titers, when combined with other autoantibody positivity. Strength of titer did not appear to be a factor for JIA, but was the most frequently observed rheumatologic disease in this study.

T cell activation by terminal complex of complement and immune complexes.

T cell hyperactivation and complement consumption are prominent features of the immunopathology of systemic lupus erythematosus. Although complement activation is secondary to autoantibodies that form immune complexes (ICs), the trigger for alterations in human peripheral blood T cells is poorly understood. To study the impact (on T cells) of several types of preformed ICs and terminal complement complex, also referred to as C5b-9, we incubated these immune reactants with peripheral blood naive CD4(+) T cells as well as Jurkat cells and analyzed their effects on cellular behavior. We first assembled the C5b-9 in situ on the membrane and observed its assembly primarily on a single site where it promoted aggregation of membrane rafts and recruitment of the CD3 signaling complex. However, C5b-9 alone did not initiate proliferation or commencement of downstream signaling events associated with T cell activation. When T cells were treated with ICs together with nonlytic C5b-9, changes associated with T cell activation by possible antigen engagement then occurred. T cell antigen receptor signaling proteins, including ζ-chain, ZAP-70, Syk, Src, and Lck, were phosphorylated and organized in a synapse-like structure. The cytoskeleton formed F-actin spindles and a distal pole complex, resulting in a bipolar distribution of phosphorylated ezrin-radixin-moesin and F-actin. Furthermore, ICs and nonlytic C5b-9 induced T cell proliferation and IFN-γ production. These results raise the possibility that ICs and the nonlytic C5b-9 modulate T cell-mediated responses in systemic lupus erythematosus and other related chronic inflammatory disorders.

Significance of complement components C1q and C4 bound to circulating immune complexes in juvenile idiopathic arthritis: support for classical complement pathway activation.

OBJECTIVES: Immune complexes (ICs) from sera of juvenile idiopathic arthritis (JIA) patients show increased complement opsonisation; however, a definitive role for involvement of the classical or alternative pathway is not entirely clear. To delineate the role of these pathways, we measured activated complement products bound to circulating IC (CICs) in the sera of JIA patients. METHODS: Sera from 100 JIA patients and 22 healthy children were collected. C1q, C4, C3, C3d, and membrane attack complex (MAC) bound to CICs were measured by enzyme-linked immunosorbent assay. Data was compared to IgM rheumatoid factor (RF), IgG anti-cyclic citrullinated peptide (CCP) antibodies, C-reactive protein (CRP), and erythrocyte sedimentation rate (ESR) levels. RESULTS: Mean levels of C1q, C4, and MAC bound to CICs were significantly elevated in JIA patients compared to healthy children. C1q correlated significantly with C4 and MAC bound to CICs and C4 and MAC also demonstrated significant correlation. No significant differences were noted in complement components bound to CICs when evaluating IgM RF, anti-CCP antibody, and CRP positivity. A significant correlation was noted between MAC bound to CICs and ESR. C1q and MAC bound to CICs mean levels were significantly higher in patients with an elevated ESR compared to those with a normal ESR level. CONCLUSIONS: JIA patients have elevated levels of complement components bound to CICs, particularly from the classical pathway. Moreover, classical pathway components were associated with ESR, a marker of disease activity. MAC bound to CICs also correlated significantly with ESR, further supporting the notion of complement-mediated tissue injury that is triggered by IC-mediated classical pathway activation.

Neonatal-onset multisystem inflammatory disease responsive to interleukin-1beta inhibition.

BACKGROUND: 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. METHODS: 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. RESULTS: All 18 patients had a rapid response to anakinra, with disappearance of rash. Diary scores improved (P<0.001) 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 P<0.001), 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. CONCLUSIONS: 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].).

Rheumatoid factors and anticyclic citrullinated peptide antibodies in pediatric rheumatology.

Juvenile idiopathic arthritis (JIA) is a heterogenous childhood disease without reliable biomarkers for monitoring disease progression. Immunoglobulin (Ig) M rheumatoid factor (RF) is used to define a subset of JIA patients, but its significance in JIA is dependent on the method of measurement. In addition to IgM RF, IgA RF has been implicated in determining disease severity in JIA, including functional disability and joint damage. Anticyclic citrullinated peptide (anti-CCP) antibodies have been a valuable diagnostic tool in rheumatoid arthritis, with varied results in their significance in JIA patients. Recent studies have demonstrated the possible usefulness of isotypes of anti-CCP antibodies in monitoring JIA patients to determine disease outcome. Overall, RF isotypes and anti-CCP isotypic antibodies have demonstrated increasing importance in the evaluation of JIA patients to determine which patients may have more aggressive or severe disease and to aid in possible treatment plans to prevent joint damage and disability.

Methylphenidate and dextroamphetamine-induced peripheral vasculopathy.

Methylphenidate and dextroamphetamine are central nervous system stimulants used in the treatment of attention deficit hyperactivity disorders in children. These medications have been associated with cerebral arteritis, renal necrotizing vasculitis, and systemic and pulmonary hypertension. We report 4 patients, 2 on methylphenidate and 2 on dextroamphetamine who presented with acral cyanosis, livedo reticularis, or Raynaud phenomenon. Two patients were found to have a positive ANA at low titers, 1 of whom had histopathologic evidence of stratum malgiphian necrosis with perivascular lymphocytic infiltration on skin biopsy. Two had positive antihistone antibodies. One patient improved after withdrawal of dextroamphetamine; others had worsening of their symptoms on higher doses of medications. These cases indicate the potential for development of acral cyanosis, livedo reticularis, or Raynaud symptoms with these medications and their potential contribution to a vasculopathy.

Scleredema adultorum of Buschke: a case report and review of the literature.

OBJECTIVES: To present a case of scleredema adultorum of Buschke associated with hypergammaglobulinemia and review the literature pertaining to this disease. METHODS: Search of MEDLINE (PubMed) was performed using the words "Scleredema Adultorum." Only cases in the English language, with pertinent clinical information for analysis, were included. RESULTS: We present a case of scleredema associated with hypergammaglobulinemia. The skin findings had been progressive for 2 years before he was diagnosed with scleredema. Our patient was found to have a B-cell lymphoma before being diagnosed with scleredema. The progression of skin thickening halted with no apparent correlation to immunosuppressive therapies or chemotherapeutic agents. A total of 165 cases of scleredema adultorum are described in the literature. There are 3 types of scleredema adultorum. Type 1 is usually preceded by a febrile episode and resolves spontaneously. Type 2 is associated with developing paraproteinemias including multiple myeloma. Type 3 is associated with diabetes mellitus. CONCLUSIONS: Scleredema adultorum of Buschke is a rare disorder which is sometimes associated with hypergammaglobulinemia; our patient had a B-cell lymphoma. Physicians should suspect scleredema in any patient with diffuse skin thickening where the hands and feet are spared, particularly if diabetes or a preceding febrile episode were present. Once the diagnosis of scleredema is made, evidence for the presence of hypergammaglobulinemia should be sought.

Lupus-associated pancreatitis.

OBJECTIVES: Involvement of the pancreas in systemic lupus erythematosus is rare. The purpose of this article is to provide a detailed review of lupus-associated pancreatitis. METHODS: We describe 3 patients with lupus-associated pancreatitis and review the English literature of the last 30 years, including the demographic, clinical, therapeutic, and prognostic aspects of this disorder. RESULTS: There were detailed descriptions of 77 patients, 88% were females. Median age was 27 years. In 44% of the patients pancreatitis developed within 1 year of the diagnosis of lupus, and 84% had active lupus at the time of pancreatitis. Abdominal pain was the most frequent pancreatitis-related symptom (88%), followed by nausea or vomiting (67%). In 97% the diagnosis of pancreatitis was based on laboratory evidence of elevated serum amylase or lipase. Abdominal computerized tomography and ultrasonography did not show signs of pancreatic involvement in 24 and 45%, respectively. The mortality rate was 27%; active lupus and some biochemical abnormalities were significantly associated with increased mortality. Treatment with corticosteroids or azathioprine was not associated with increased mortality. On the contrary, mortality was decreased in patients who were treated with these agents after the onset of pancreatitis (20% mortality, compared with 61% among those who were not treated with steroids for their pancreatitis, P = 0.005). CONCLUSIONS: Pancreatitis should be suspected in any SLE patient with abdominal pain. Mortality rate is related to both active lupus and some biochemical markers. In most cases, the onset of pancreatitis appears unrelated to previous treatment with steroids or azathioprine. Moreover, treatment with these medications improves prognosis.

Immune Complexes in Juvenile Idiopathic Arthritis.

Juvenile idiopathic arthritis (JIA) reflects a group of clinically heterogeneous, autoimmune disorders in children characterized by chronic arthritis and hallmarked by elevated levels of circulating immune complexes (CICs) and associated complement activation by-products in their sera. Immune complexes (ICs) have been detected in patients' sera with JIA utilizing a variety of methods, including the anti-human IgM affinity column, C1q solid-phase assay, polyethylene glycol precipitation, Staphylococcal Protein A separation method, anti-C1q/C3 affinity columns, and FcγRIII affinity method. As many as 75% of JIA patients have had IC detected in their sera. The CIC proteome in JIA patients has been examined to elucidate disease-associated proteins that are expressed in active disease. Evaluation of these ICs has shown the presence of multiple peptide fragments by SDS-PAGE and 2-DE. Subsequently, all isotypes of rheumatoid factor (RF), isotypes of anti-cyclic citrullinated peptide (CCP) antibodies, IgG, C1q, C4, C3, and the membrane attack complex (MAC) were detected in these IC. Complement activation and levels of IC correlate with disease activity in JIA, indicating their role in the pathophysiology of the disease. This review will summarize the existing literature and discuss the role of possible protein modification that participates in the generation of the immune response. We will address the possible role of these events in the development of ectopic germinal centers that become the secondary site of plasma cell development in JIA. We will further address possible therapeutic modalities that could be instituted as a result of the information gathered by the presence of ICs in JIA.

Azathioprine therapy for steroid-resistant Henoch-Schönlein purpura: a report of 6 cases.

BACKGROUND: A small percentage of children with Henoch-Schönlein purpura (HSP) develop a chronic form of the disease that often requires prolonged corticosteroid therapy. Disease modifying anti-rheumatic agents (DMARDs) or biologics have been successfully used to treat those refractory cases. Azathioprine is a DMARD that has been reported to be effective in HSP nephritis and in adult cutaneous leukocytoclastic vasculitis, a condition with cutaneous histology similar to HSP. CASE PRESENTATION: A description of 6 cases with relapsing HSP without significant renal involvement, treated with azathioprine are reported. All 6 cases met the classification criteria for the diagnosis of HSP, had relapsing symptoms despite corticosteroid use, were successfully treated with azathioprine and were tapered off of corticosteroids. The duration of azathioprine therapy ranged from 7-21 months and no adverse events were reported. CONCLUSIONS: Azathioprine is effective in controlling prolonged relapsing symptoms of HSP, allowing earlier discontinuation of corticosteroids. This report shows that azathioprine can be included in the therapeutic options for relapsing HSP and is the first case series in the literature of azathioprine use in HSP without significant renal involvement.

Antirheumatic drugs in pregnancy and lactation.

OBJECTIVE: To review the toxicity issues of commonly used antirheumatic drugs in pregnancy and lactation. METHODS: A review of the medical literature using Medline database via Ovid was performed to identify the toxicities of antirheumatic drugs in pregnancy and lactation. RESULTS: Many rheumatologic diseases in women often first present during the childbearing years. In most cases, antirheumatic therapy is required for their disease control. Glucocorticoids may be used during pregnancy; however, first-trimester use should be avoided if possible and breastfeeding should occur 4 hours after the last dosing. Nonsteroidal antiinflammatory drugs (NSAIDs) and cyclooxygenase-2 (COX-2) inhibitors should be discontinued 6 to 8 weeks before delivery. NSAIDs are compatible with lactation, although there is potential risk of jaundice and kernicterus. There is insufficient data on COX-2 inhibitors and lactation. Hydroxychloroquine and sulfasalazine may be continued throughout pregnancy and lactation. Methotrexate and leflunomide are contraindicated during pregnancy and lactation. Cyclophosphamide and mycophenolate mofetil should be avoided during pregnancy and lactation. Azathioprine and cyclosporine A could be used with caution during pregnancy if felt there is a need to suppress disease activity. They are not compatible with breastfeeding. There are insufficient data regarding tumor necrosis factor-antagonists, anakinra, and rituximab in relation to pregnancy and lactation. Male patients should be made aware of the effects methotrexate, leflunomide, sulfasalazine, and cyclophosphamide may have on their fertility. CONCLUSIONS: Health care providers should discuss the risks and benefits of antirheumatic therapy during conception, pregnancy, and lactation with their patients. Better maternal and fetal outcomes can be expected if the pregnancy is planned, the rheumatic disease is stable, and if appropriate medication adjustments can be made ahead of time.

Parathyroid hormone bone anabolic action requires Cbfa1/Runx2-dependent signaling.

The Cbfa1/Runx2 (referred to herein as Cbfa1) transcription factor has been shown to be essential for osteoblast differentiation and bone formation during embryogenesis. PTH given intermittently is a proven bone anabolic agent. Here, we investigated whether PTH regulates the expression and/or activity of Cbfa1 in osteoblastic cells and in a rat metatarsal organ culture assay. PTH was found to regulate Cbfa1 mRNA in the rat osteosarcoma cell line UMR106 in a concentration-dependent manner. The effect of PTH was mimicked by forskolin, an activator of adenylate cyclase leading to the protein kinase A pathway. PTH administered intermittently for 5 d in vivo was found to stimulate Cbfa1 protein in the rat proximal tibiae metaphysis. To demonstrate PTH regulation of Cbfa1 activity, a construct containing six tandem Cbfa1 binding elements fused to luciferase was shown to be rapidly stimulated in response to PTH. This stimulation preceded the effects on mRNA regulation and resulted from a protein kinase A-mediated increase in Cbfa1 activity. Finally, using a neonate rat metatarsal organ culture system, we demonstrated dose-dependent anabolic responsiveness to PTH and to Cbfa1 overexpression from an adenoviral construct. We further showed that Cbfa1 antisense oligonucleotides that blocked adenoviral Cbfa1-induced anabolic effects in this organ culture model also abolished the PTH-mediated anabolic increase. These findings suggest a requirement for Cbfa1 in mediating the anabolic effects of PTH. Thus, regulation of Cbfa1 expression or activity is an important mechanism by which PTH controls osteoblast function.