Distinctive features of positive anti-cell antibody tests (indirect immunofluorescence on HEp-2 cells) in patients with non-autoimmune diseases.

OBJECTIVE: The objective of this study was to determine whether characteristics of positive results in the indirect immunofluorescence assay on HEp-2 cells for anti-cell antibodies (HEp-2 IFA) differ between patients with non-autoimmune diseases (NADs) and patients with systemic autoimmune rheumatic diseases (SARDs). METHODS: Cross-sectional observational study comparing HEp-2 IFA test results in three groups: (a) 558 NAD patients comprising four subgroups (cancer ( n = 95), infectious diseases ( n = 148), psychiatric diseases ( n = 163), common non-infectious chronic diseases ( n = 152)); (b) 194 SARD patients; (c) 1217 healthy individuals (HIs). Sera were tested at 1:80 dilution and diluted to the end titer. Slides were analyzed by two independent blinded examiners. RESULTS: A positive HEp-2 IFA test occurred in 102 (18.3%) NAD patients, 170 (87.6%) SARD patients and 150 (12.3%) HIs. The four NAD subgroups did not differ regarding HEp-2 IFA frequency, titer or pattern. HEp-2 IFA titer was higher in NAD patients than in HIs and both had lower titer than SARD patients. Nuclear dense fine speckled pattern was more frequent in NAD patients and HIs than in SARD patients ( p < 0.001). Nuclear homogeneous and nuclear coarse speckled patterns were more frequent in SARD patients than in the other groups ( p < 0.001). The nuclear fine speckled pattern was prevalent in all three groups, but presented a gradient in titer across them; HIs and NAD patients had low and intermediary titers, which were significantly lower than in SARD patients ( p < 0.001). CONCLUSION: Positive HEp-2 IFA frequency, pattern and titer present differential features in NAD and SARD patients, and this attribute adds value to the test in the diagnosis of SARDs.

Improvement of indirect immunofluorescence technique to detect antineutrophil cytoplasmic antibodies and its impact on test positivity rate.

The indirect immunofluorescence (IIF) technique for antineutrophil cytoplasmic antibodies (ANCA) detection is subject to substantial differences across laboratories. This study aimed to assess the impact of improvements in the IIF-ANCA technique on the positivity rate of ANCA tests. A cross-sectional study was performed with serum samples from patients with ANCA-associated vasculitis (AAV), autoimmune hepatitis (AIH), and ulcerative colitis (UC). A paired analysis was performed for IIF-ANCA results using the traditional method and a modified protocol after a series of specific adjustments in the technique based on the protocol of IIF-ANCA test performed at a nation-wide private laboratory in Brazil. ANCA specificity was assessed by ELISA for anti-proteinase 3 (PR3) and anti-myeloperoxidase (MPO) antibodies. Sixty-one patients were evaluated. The positivity rate of IIF-ANCA tests at disease presentation performed at the University reference laboratory was 32.3% in AAV, AIH, and UC patients, whereas the positivity rates of IIF-ANCA and ELISA tests in other laboratories were 75.0 and 72.7%, respectively. After modifications in the IIF-ANCA technique, there was a significant increase in the positivity rate (14.8 vs 34.3%; P=0.0002) and in median titers [1/40 (1/30-1/160) vs 1/80 (1/40-1/80); P=0.0003] in AAV, AIH, and UC patients. UC had the highest increment in positive results from 5.3 to 36.8%. There was poor agreement between MPO- or PR3-ANCA and both IIF-ANCA techniques. In conclusion, modifications in the IIF-ANCA protocol led to a significant improvement in its positivity rate and titers.

Anti-aquaporin-4 antibodies in the context of assorted immune-mediated diseases.

BACKGROUND AND PURPOSES: Anti-aquaporin 4 antibodies are specific markers for Devic's disease. This study aimed to test if this high specificity holds in the context of a large spectrum of systemic autoimmune and non-autoimmune diseases. METHODS: Anti-aquaporin-4 antibodies (NMO-IgG) were determined by indirect immunofluorescence (IIF) on mouse cerebellum in 673 samples, as follows: group I (clinically defined Devic's disease, n=47); group II [inflammatory/demyelinating central nervous system (CNS) diseases, n=41]; group III (systemic and organ-specific autoimmune diseases, n=250); group IV (chronic or acute viral diseases, n=35); and group V (randomly selected samples from a general clinical laboratory, n=300). RESULTS: MNO-IgG was present in 40/47 patients with classic Devic's disease (85.1% sensitivity) and in 13/22 (59.1%) patients with disorders related to Devic's disease. The latter 13 positive samples had diagnosis of longitudinally extensive transverse myelitis (n=10) and isolated idiopathic optic neuritis (n=3). One patient with multiple sclerosis and none of the remaining 602 samples with autoimmune and miscellaneous diseases presented NMO-IgG (99.8% specificity). The autoimmune disease subset included five systemic lupus erythematosus individuals with isolated or combined optic neuritis and myelitis and four primary Sjögren's syndrome (SS) patients with cranial/peripheral neuropathy. CONCLUSIONS: The available data clearly point to the high specificity of anti-aquaporin-4 antibodies for Devic's disease and related syndromes also in the context of miscellaneous non-neurologic autoimmune and non-autoimmune disorders.

Improvement of indirect immunofluorescence technique to detect antineutrophil cytoplasmic antibodies and its impact on test positivity rate

The indirect immunofluorescence (IIF) technique for antineutrophil cytoplasmic antibodies (ANCA) detection is subject to substantial differences across laboratories. This study aimed to assess the impact of improvements in the IIF-ANCA technique on the positivity rate of ANCA tests. A cross-sectional study was performed with serum samples from patients with ANCA-associated vasculitis (AAV), autoimmune hepatitis (AIH), and ulcerative colitis (UC). A paired analysis was performed for IIF-ANCA results using the traditional method and a modified protocol after a series of specific adjustments in the technique based on the protocol of IIF-ANCA test performed at a nation-wide private laboratory in Brazil. ANCA specificity was assessed by ELISA for anti-proteinase 3 (PR3) and anti-myeloperoxidase (MPO) antibodies. Sixty-one patients were evaluated. The positivity rate of IIF-ANCA tests at disease presentation performed at the University reference laboratory was 32.3% in AAV, AIH, and UC patients, whereas the positivity rates of IIF-ANCA and ELISA tests in other laboratories were 75.0 and 72.7%, respectively. After modifications in the IIF-ANCA technique, there was a significant increase in the positivity rate (14.8 vs 34.3%; P=0.0002) and in median titers [1/40 (1/30-1/160) vs 1/80 (1/40-1/80); P=0.0003] in AAV, AIH, and UC patients. UC had the highest increment in positive results from 5.3 to 36.8%. There was poor agreement between MPO- or PR3-ANCA and both IIF-ANCA techniques. In conclusion, modifications in the IIF-ANCA protocol led to a significant improvement in its positivity rate and titers.

Clinical significance of anticardiolipin antibodies in juvenile idiopathic arthritis.

OBJECTIVE: Anticardiolipin antibodies (aCL) have been demonstrated in a large spectrum of autoimmune diseases. However, its occurrence in childhood, in particular in juvenile idiopathic arthritis (JIA), is not well established. The present study addressed the frequency and clinical significance of aCL in a group of JIA patients. METHODS: aCL (IgG and IgM isotypes), antinuclear antibodies (ANA), and rheumatoid factor (RF) were determined in 86 children with JIA (33 systemic, 31 polyarticular and 22 oligoarticular onset type). Thirty-two juvenile systemic erythematosus lupus patients (JSLE) and 52 healthy children formed the control groups. The disease activity and functional status of the JIA patients were scored to study their possible associations with the presence of aCL. RESULTS: Serum aCL levels above the normal range were detected in 28/86 JIA patients (32.5%), 12/32 JSLE patients (37.5%), and 3/52 healthy children (6%). Positive aCL levels were slightly or moderately elevated (usually below 30 GPL and 20 MPL). The presence of aCL was not associated with the presence of ANA or RF. Associations between aCL and clinical parameters, such as disease onset, duration, activity or severity could not be established. No JIA patient had vascular thrombosis, thrombocytopenia or "livedo reticularis". CONCLUSION: aCL occurred in low titers in JIA children, in a similar frequency to that observed in JSLE. No association with JIA clinical parameters or the clinical features classically linked to the antiphospholipid antibody syndrome were observed.

The behavior of the coiled body in cells infected with adenovirus in vitro.

The coiled body is a phylogenetically conserved nuclear organelle whose function is not known. Probes for detection of p80-coilin, an 80 kDa protein enriched in the coiled body, have made possible studies determining the behavior of the coiled body during the cell cycle, in proliferating cells, as well as reports suggesting some relationship of the coiled body to mRNA splicing and to the nucleolus. The objective of this study is to examine the distribution of p80-coilin and nucleolar proteins in cells infected with adenovirus in vitro. HeLa cells grown as monolayers were infected with successive dilutions of type 5 human adenovirus culture and fixed in methanol/acetone at different time points. Single and double indirect immunofluorescence was performed with human autoantibodies to p80-coilin, fibrillarin, NOR-90/hUBF, RNA polymerase I, PM-Scl, and To, as well as rabbit polyclonal serum to p80-coilin (R288) and mouse monoclonal antibody to adenovirus 72-kDa DNA-binding protein. Indirect immunofluorescence (IIF) with anti-p80-coilin antibodies showed that the usual bright dot-like coiled body staining pattern was replaced in infected cells by 1-5 clusters of tiny dots at the periphery of the nucleus. This phenomenon was first detected within 12 h of infection and affected more severely cells with increased length and load of infection. Cells subjected to heat shock presented no such alteration. Double IIF showed cells with abnormal coiled body appearance expressed the viral 72-kDa DNA-binding protein. Nucleolar proteins RNA polymerase I and NOR-90/hUBF became associated with the p80-coilin-enriched clusters and were no longer detected in the nucleolus. Other nucleolar proteins, like PM-Scl and To, remained associated to the nucleolus and were not detected in the newly formed clusters. Fibrillarin had a heterogeneous behavior, being restricted to the nucleolus in some infected cells while in some others it was associated with the p80-coilin-enriched clusters. Thus our results showed that in vitro adenovirus infection induced radical redistribution of nucleolar and coiled body constituents into newly formed structures characterized by clusters of tiny dots in the periphery of the nucleus. The fact that three major proteins involved in rRNA synthesis and processing colocalized with p80-coilin in these clusters may bring additional support to the idea that the coiled body and p80-coilin may be implicated in functions related to the nucleolus.

Antiperinuclear factor and antibodies to the stratum corneum of rat esophagus in juvenile idiopathic arthritis.

We analyzed the frequency and clinical correlates of antiperinuclear factor (APF) and antibodies to the stratum corneum of rat esophagus in 86 children with juvenile idiopathic arthritis (JIA), 32 children with juvenile systemic lupus erythematosus, and 52 healthy children. Forty-two patients with JIA (49%) were positive for APF. No association was observed between APF and current age, sex, JIA subtype, age at disease onset, or disease duration. APF was found in one patient with juvenile systemic lupus erythematosus and in no healthy child. Antibodies to the stratum corneum of rat esophagus were detected in 3 patients with polyarticular JIA. APF may be a valuable tool in the differential diagnosis of JIA.

Aplasia medular prolongada como manifestacao pre-leucemica. / Long-term medullary aplasia as pre-leucemic manifestation

Os autores apresentam um caso de aplasia medular prolongada, em crianca de 7 anos e 8 meses de idade, medicado com androgeno por 9 meses sem melhoria do quadro hematologico, ate que foram constatados 32% de celulas leucemicas ao hemograma o que permitiu, entao, o diagnostico de leucemia mielomonocitica, tendo sido possivel obter remissao completa. Os autores comentam a importancia da identificacao de estados pre-leucemicos

Significado clínico dos anticorpos antinucleares na artrite reumatóide juvenil / Clinical significance of antinuclear antibodies in juvenile rheumatoid arthritis

Os anticorpos antinucleares (AAN) têm sido demonstrados em freqüência extremamente variável na artrite reumatóide juvenil (ARJ), dependendo do substrato empregado e da seleçäo dos pacientes. Além disso, a associaçÝo entre estes anticorpos e a atividade ou a gravidade da doença näo está bem estabelecida. Objetivo: Determinar a freqüência dos AAN e possível associaçöes com diferentes parâmetros clínicos da ARJ, como duraçäo, atividade e gravidade da doença. Material e métodos: 86 pacientes com ARJ foram estudados quanto à presença de AAN (pesquisados pelo método de imunofluorescência indireta em células HEp-2); 32 pacientes com lúpus eritematoso sistêmico juvenil e 52 crianças saudáveis constituíram os grupos-controles. A atividade da doença foi definida segundo uma escala de dois pontos, enquanto a gravidade foi avaliada pelo grau de capacidade funcional. Resultados: AAN foram encontrados em 36 (42 por cento) pacientes com ARJ, sendo mais prevalentes no tipo de início oligoarticular do que no sistêmico (p<0,05). Uma associaçäo com a duraçäo, atividade ou a gravidade da doença näo foi evidenciada (p>0,05), embora uma tendência tenha sido observada em pacientes do tipo de início poliarticular, em atividade de doença (P=0,054). O padräo pontilhado fino foi o mais freqëntemente observado (64 por cento), conquanto outros padröes, como o centrossômico e o do corpo intermediário, também tivessem sido notados. Conclusäo: Anticorpos antinucleares foram encontrados em menor freqüência que aquela descrita na maioria dos trabalhos da literatura para a ARJ, nÝo apresentando associaçäo com o sexo, duraçäo, atividade ou gravidade de doença. Embora o padräo de fluorescência mais freqüente tenha sido o pontilhado, a presença de padröes inusitados de fluorescência nuclear sugere que AAN na ARJ possam apresentar especificidades ainda näo definidas, que poderäo futuramente ser melhor caracterizadas