RESUMO
Glycoprotein 2 (GP2) is an autoantigen in Crohn's (CD) and coeliac disease (CeD). We assessed GP2-isoform (GP21-4)-expression in intestinal biopsies of paediatric patients with CD, CeD, ulcerative colitis (UC), and healthy children (HC). Transcription of GP21-4 was elevated in proximal small intestine in CeD and CD patients (only GP22/4) compared to jejunum (CeD/CD) and large bowel (CD). CeD patients demonstrated higher duodenal GP22/4-mRNA levels compared to HC/UC patients whereas CD patients showed higher GP24-mRNA levels compared to UC patients. Duodenal synthesis of only small GP2 isoforms (GP23/4) was demonstrated in epithelial cells in patients/HC and in Brunner glands (also large isoforms) with a more frequent apical location in CD/CeD patients. All four GP2 isoforms interacted with gliadin and phosphopeptidomannan. Gliadin digestion improved binding to GP2 isoforms. GP21-4 binding to CeD/CD-related antigens, elevated duodenal GP21-4-mRNA transcription, and GP2-protein secretion in Brunner glands of CeD/CD patients suggest an autoimmune CeD/CD link.
Assuntos
Glândulas Duodenais , Doença Celíaca , Colite Ulcerativa , Doença de Crohn , Humanos , Criança , Gliadina , Proteínas Ligadas por GPI , Autoanticorpos , Doença de Crohn/genética , Colite Ulcerativa/genética , Isoformas de Proteínas , RNA Mensageiro/genéticaRESUMO
OBJECTIVES: Inborn errors of immunity manifest with susceptibility to infection but may also present with immune dysregulation only. According to the European Society for Immunodeficiencies Registry about 50% of inborn errors of immunity are classified as common variable immunodeficiencies (CVID). In only few CVID patients are monogenic causes identified. IFN regulatory factor-2 binding protein 2 (IRF2BP2) is one of 20 known genes associated with CVID phenotypes and has only been reported in two families so far. We report another IRF2BP2-deficient patient with a novel pathogenic variant and phenotype and characterize impaired B cell function and immune dysregulation. METHODS: We performed trio whole-exome sequencing, determined B cell subpopulations and intracellular calcium mobilization upon B cell receptor crosslinking in B cells. T cell subpopulations, T cell proliferation and a type I IFN signature were measured. Colonoscopy and gastroduodenoscopy including histopathology were performed. RESULTS: The 33-year-old male presented with recurrent respiratory infections since childhood, colitis and RA beginning at age 25 years. We identified a novel de novo nonsense IRF2BP2 variant c.1618C>T; p.(Q540*). IgG deficiency was detected as consequence of a severe B cell differentiation defect. This was confirmed by impaired plasmablast formation upon stimulation with CpG. No serum autoantibodies were detected. Intracellular cytokine production in CD4+ T cells and CTLA4 expression on FOXP3+ Tregs were impaired. Type I IFN signature was elevated. CONCLUSION: The identified loss-of-function variant in IRF2BP2 severely impairs B cell development and T cell homeostasis, and may be associated with colitis and RA. Our results provide further evidence for association of IRF2BP2 with CVID and contribute to the understanding of the underlying pathomechanisms.
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Linfócitos T CD4-Positivos , Fatores de Transcrição , Masculino , Linfócitos B , Mutação , Fenótipo , Humanos , AdultoRESUMO
OBJECTIVES: Anti-neutrophil cytoplasmic antibody (ANCA) directed against proteinase 3 (PR3) is a marker for granulomatosis with polyangiitis, but is also found in patients with inflammatory bowel disease (IBD), mainly ulcerative colitis (UC). The aim of our study was to investigate ANCA and PR3-ANCA in paediatric IBD. METHODS: We tested 326 paediatric IBD patients and 164 controls for anti-Saccharomyces cerevisiae antibodies (ASCA), ANCA (indirect immunofluorescence, IIF) and PR3-ANCA (chemiluminescence immunoassay). We applied the Paris classification for paediatric IBD and documented liver manifestations such as primary sclerosing cholangitis (PSC) and autoimmune hepatitis (AIH). RESULTS: We found PR3-ANCA in 49/121 (40%) of UC, 20/187 (11%) of Crohn disease (CD) and 2/18 (11%) of IBD-unclassified (IBD-U) patients but in none of the controls. 54% UC and 12% CD patients were positive for ANCA (IIF). PR3-ANCA positive UC patients were characterised by more extensive disease (Pâ=â.070). Fourteen of 21 (67%) of UC patients with backwash ileitis were anti-PR3 ANCA-positive (Pâ=â.011). We diagnosed PSC or PSC/AIH in 19 UC and 3 IBD-U patients. Fifteen of 22 (68%) patients with PSC or PSC/AIH were anti-PR3-ANCA positive in contrast to 36 of 117 (32%) patients without PSC (Pâ=â.001). PR3-ANCA positive patients showed higher levels of gamma-glutamyl transferase, alanine transaminase and aspartate transferase (Pâ<â0.001, 0.001, 0.006, respectively). Forty-seven percent of CD and 6% of UC patients were ASCA-IgA positive. PR3-ANCA-positive and -negative patients showed no significant differences concerning age at diagnosis, disease activity, need for drugs, and number of hospitalisations. CONCLUSIONS: Our study provides data for PR3-ANCA as a potential serological marker for paediatric UC and PSC.
Assuntos
Colangite Esclerosante , Colite Ulcerativa , Doença de Crohn , Doenças Inflamatórias Intestinais , Anticorpos Anticitoplasma de Neutrófilos , Biomarcadores , Criança , Colangite Esclerosante/diagnóstico , Colite Ulcerativa/diagnóstico , Doença de Crohn/diagnóstico , Humanos , Mieloblastina , TransferasesRESUMO
The indirect immunofluorescence assay (IIFA) on HEp-2 cells is widely used for detection of antinuclear antibodies (ANA). The dichotomous outcome, negative or positive, is integrated in diagnostic and classification criteria for several systemic autoimmune diseases. However, the HEp-2 IIFA test has much more to offer: besides the titre or fluorescence intensity, it also provides fluorescence pattern(s). The latter include the nucleus and the cytoplasm of interphase cells as well as patterns associated with mitotic cells. The International Consensus on ANA Patterns (ICAP) initiative has previously reached consensus on the nomenclature and definitions of HEp-2 IIFA patterns. In the current paper, the ICAP consensus is presented on the clinical relevance of the 29 distinct HEp-2 IIFA patterns. This clinical relevance is primarily defined within the context of the suspected disease and includes recommendations for follow-up testing. The discussion includes how this information may benefit the clinicians in daily practice and how the knowledge can be used to further improve diagnostic and classification criteria.
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Anticorpos Antinucleares/análise , Doenças Autoimunes/diagnóstico , Técnica Indireta de Fluorescência para Anticorpo/normas , Doenças Autoimunes/imunologia , Biomarcadores/análise , Humanos , Cooperação InternacionalRESUMO
BACKGROUND: Based on advances over several decades, a significant number of autoantibodies are aiding the diagnosis, differential diagnosis and even prognostic estimates of patients with connective tissue diseases (CTD). Based on cost and time constraints, multiplex assays are used more and more in routine practice. Here we describe the evaluation results of a novel spot immunoassay (SeraSpot® ANA) for multiplex analysis of the main CTD specific autoantibodies, which is based on autoantigens immobilized on microtitration plates. METHODS: For evaluation of the ANA spot immunoassay, comprising dsDNA, histone, nucleosome, Scl-70, U1-RNP (mixture of U1-RNP-A, C, and 70k), Sm (SmD), RibP (P0), Ro52/TRIM21, Ro60, La/SS-B, CENP-B, Jo-1, PM/ Scl-100, and Ku as target antigens, sera of 489 patients with systemic autoimmune rheumatic disease (SARD), sera of 54 patients with herpes virus infections, and sera of 202 apparently healthy individuals (AHI) were tested. RESULTS: The concordance between the SeraSpot and EIA results of disease specific autoantibodies (AABs) was high (94 - 97% for CENP-B, Jo-1, Scl-70, Sm, La, Ro52 and Ro60 antibodies) to moderate (86.7% for dsDNA antibodies), and no major differences in the frequency of these AABs in patients with CTD were observed. In SLE patients, the SeraSpot results of dsDNA antibodies correlate better with CLIFT results and HEp-2 cell pattern than the EIA results. The diagnostic specificities of SLE, SSc, SjS, and myositis specific AABs are very high (96.5 - 100%) compared to apparently healthy individuals, but lower with regard to serological differentiation of the SARD entities (86.6 - 99.1%). However, very high positive likelihood ratios (10 up to infinite) could be obtained by disease specific AAB profiles. CONCLUSIONS: The SeraSpot® ANA assay allows the detection of 14 AAB specificities used for the diagnosis and dif-ferentiation of CTD in one approach. Although the concordance between the SeraSpot® assay and EIA is moderate for most of the tested AAB specificities, defined AAB profiles are suitable for the correct diagnosis of the CTD entities. If time matters, the authors suggest the parallel employment of immunofluorescence on HEp-2 cells and the SeraSpot® ANA assay for screening and specific AAB determination of patients with suspected CTD.
Assuntos
Autoanticorpos/imunologia , Autoantígenos/imunologia , Doenças do Tecido Conjuntivo/imunologia , Imunoensaio/métodos , Linhagem Celular Tumoral , Doenças do Tecido Conjuntivo/diagnóstico , Humanos , Lúpus Eritematoso Sistêmico/diagnóstico , Lúpus Eritematoso Sistêmico/imunologia , Doenças Reumáticas/diagnóstico , Doenças Reumáticas/imunologia , Sensibilidade e EspecificidadeRESUMO
OBJECTIVES: To analyse the subgroup of early arthritis patients with new onset parvovirus infections for details that may help narrow the population tested. METHODS: From their routine patient charts, patient histories and clinical and serological data were obtained for all 130 patients of the Rheumatology division with parvovirus serology performed. 11 patients had acute parvovirus infections, defined by specific IgM antibodies. 95 patients had a previous infection, 16 were never infected, together forming the n=111 control group, and 8 patients had to be excluded. RESULTS: Most patients with acute parvovirus infection had an acute onset, highly symmetrical polyarthritis of small joints, which was preceded by prodromal symptoms. Positive ANA were frequently found, whereas C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) were only mildly elevated. No frank synovitis was found longer than two weeks after disease onset. Most patients were free of symptoms within three months, and no patient in the parvovirus group developed rheumatoid arthritis or a connective tissue disease. CONCLUSIONS: Parvovirus serology may be helpful in patients with acute polyarthritis of very recent onset, and if they give a history of prodromal symptoms, in particular. In most instances, parvovirus arthritis is an acute disease, which is rapidly self-limiting.
Assuntos
Artrite Reumatoide/complicações , Infecções por Parvoviridae/complicações , Parvovirus B19 Humano , Adulto , Anticorpos Antivirais/sangue , Feminino , Humanos , Imunoglobulina M/sangue , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: For the serological diagnosis of systemic autoimmune rheumatic diseases, a two-tier approach starting with sensitive antinuclear antibody (ANA) detection by indirect immunofluorescence (IIF) on HEp-2 cells followed by characterization of positive findings with different immunoassays is recommended. To overcome drawbacks of this approach, we developed a novel technique allowing the combination of screening and simultaneous confirmatory testing. For the first time, this creates the basis for second generation ANA testing. METHODS: ANA and autoantibodies (autoAbs) to double-stranded DNA (dsDNA), CENP-B, SS-A/Ro52, SS-A/Ro60, SS-B/La, RNP-Sm, Sm, and Scl-70 were determined by IIF and enzyme-linked immunosorbent assay (ELISA), respectively, and compared to simultaneous analysis thereof by second generation ANA analysis in patients with systemic lupus erythematosus (n=174), systemic sclerosis (n=103), Sjögren's syndrome (n=46), rheumatoid arthritis (n=36), mixed and undetermined connective tissue diseases (n=13), myositis (n=21), infectious disease (n=21), autoimmune liver disease (n=93), inflammatory bowel disease (n=78), paraproteinemia (n=11), and blood donors (n=101). RESULTS: There was very good agreement of second generation ANA testing with classical one by IIF and ELISA regarding testing for ANA and autoAbs to dsDNA, CENP-B, SS-B, RNP-Sm, Scl-70, SS-A/Ro52, and SS-A/Ro60 (Cohen's κ>0.8). The agreement for anti-Sm autoAb was good (κ=0.77). The differences of both approaches were not significant for autoAbs to SS-B/La, RNP-Sm, Scl-70, SS-A/Ro60, and SS-A/Ro52 (McNemar's test, p>0.05, respectively). CONCLUSIONS: Second generation ANA testing can replace the two-tier analysis by combining IIF screening with multiplex confirmative testing. This addresses shortcomings of classical ANA analysis like false-negative ANA findings and lack of laboratory efficiency and standardization.
Assuntos
Anticorpos Antinucleares/sangue , Doenças Autoimunes/diagnóstico , Imunoensaio , Doenças Reumáticas/diagnóstico , Doenças Autoimunes/sangue , Doenças Autoimunes/imunologia , Técnica Indireta de Fluorescência para Anticorpo , Células Hep G2 , Humanos , Doenças Reumáticas/sangue , Doenças Reumáticas/imunologiaRESUMO
Crohn's disease (CrD) and ulcerative colitis (UC) are the main inflammatory bowel diseases (IBD). IBD-specific humoral markers of autoimmunity in the form of autoantibodies have been reported first in the late 1950s by demonstrating the occurrence of autoimmunity in UC, while humoral autoimmunity in CrD can be traced back to the 1970s. Ever since, the pathophysiological role of autoimmune responses in IBDs has remained poorly understood. Notwithstanding, autoreactive responses play a major role in inflammation leading to overt IBD. In CrD, approximately 40% of patients and <20% of patients with UC demonstrate loss of tolerance to antigens of the exocrine pancreas. Glycoprotein 2 (GP2) has been identified as a major autoantigenic target of the so-called pancreatic antibodies. The previously unsolved contradiction of pancreatic autoreactivity and intestinal inflammation in IBD was elucidated by demonstrating the expression of GP2 at the site thereof. Intriguingly, GP2 has been reported to be a receptor on microfold cells of intestinal Peyer's patches, which are believed to represent the origin of CrD inflammation. The development of immunoassays for the detection of antibodies to GP2 has paved the way to investigate the association of such antibodies with the clinical phenotype in CrD. Given the recently discovered immunomodulating role of GP2 in innate and adaptive intestinal immunity, this association can shed further light on the pathophysiology of IBD. In this context, the association of anti-GP2 autoantibodies as novel CrD-specific markers with the clinical phenotype in CrD will be discussed in this review.
Assuntos
Anticorpos/imunologia , Doença de Crohn/imunologia , Proteínas Ligadas por GPI/imunologia , Pâncreas/imunologia , Pâncreas/fisiopatologia , Especificidade de Anticorpos , Autoimunidade , HumanosRESUMO
The International Consensus on ANA Patterns (ICAP) is an ongoing international initiative dedicated to harmonizing technical and interpretation aspects of the HEp-2 IFA test. Comprised of internationally recognized experts in autoimmunity and HEp-2 IFA testing, ICAP has operated for the last 10 years by promoting accurate reading, interpretation, and reporting of HEp-2 IFA images by professionals involved in various areas related to autoimmune diseases, such as clinical diagnostic laboratories, academic research, IVD industry, and patient care. ICAP operates through continuous information exchange with the international community and encourages the participation of younger experts from all over the world. The 7th ICAP workshop has addressed several aspects that originated from this interaction with the international community and has effectively established objective goals and tasks to be delivered over the next two years. Some of these are outlined in this article, including the planning of three audio-visual educational modules to be posted at the www.anapattern.org website, the classification of two novel HEp-2 IFA patterns, the implementation of a project dedicated to continuously updating the information on the clinical and immunologic relevance of the HEp-2 IFA patterns, and the launch of two additional branches of the HEp-2 Clinical and Immunological (HEp-2 CIC) project.
RESUMO
Analysis of phosphorylated histone protein H2AX (γH2AX) foci is currently the most sensitive method to detect DNA double-strand breaks (DSB). This protein modification has the potential to become an individual biomarker of cellular stress, especially in the diagnosis and monitoring of neoplastic diseases. To make γH2AX foci analysis available as a routine screening method, different software approaches for automated immunofluorescence pattern evaluation have recently been developed. In this study, we used novel pattern recognition algorithms on the AKLIDES® platform to automatically analyze immunofluorescence images of γH2AX foci and compared the results with visual assessments. Dose- and time-dependent γH2AX foci formation was investigated in human peripheral blood mononuclear cells (PBMCs) treated with the chemotherapeutic drug etoposide (ETP). Moreover, the AKLIDES system was used to analyze the impact of different immunomodulatory reagents on γH2AX foci formation in PBMCs. Apart from γH2AX foci counting the use of novel pattern recognition algorithms allowed the measurement of their fluorescence intensity and size, as well as the analysis of overlapping γH2AX foci. The comparison of automated and manual foci quantification showed overall a good correlation. After ETP exposure, a clear dose-dependent increase of γH2AX foci formation was evident using the AKLIDES as well as Western blot analysis. Kinetic experiments on PBMCs incubated with 5 µM ETP demonstrated a peak in γH2AX foci formation after 4 to 8 h, while a removal of ETP resulted in a strong reduction of γH2AX foci after 1 to 4 h. In summary, this study demonstrated that the AKLIDES system can be used as an efficient automatic screening tool for γH2AX foci analysis by providing new evaluation features and facilitating the identification of drugs which induce or modulate DNA damage.
Assuntos
Técnica Indireta de Fluorescência para Anticorpo , Histonas/isolamento & purificação , Leucócitos Mononucleares/efeitos dos fármacos , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Etoposídeo/farmacologia , Histonas/sangue , Humanos , Leucócitos Mononucleares/imunologiaRESUMO
OBJECTIVES: This paper aims to investigate adherence to, and outcome of, radiographic screening of patients with rheumatoid arthritis (RA) for cervical involvement, given the availability of state of the art disease-modifying anti-rheumatic drug (DMARD) and biological therapies. METHODS: Cervical screening results and clinical information were obtained from the charts of 395 consecutive patients with rheumatoid arthritis who attended an academic rheumatology outpatient clinic in a 3-month interval. This sample was combined with eight patients who underwent C1-C2 fusion at the Department of Orthopaedic Surgery. RESULTS: Reports on cervical spine x-ray films were not found in the charts of 67 patients (17 %), including 21 (8 %) of the 257 patients with a disease duration of ≥5 years. Nevertheless, 17 (7%) of these 257 patients had an increased atlantodental distance. An additional 4 RA patients of the Department of Orthopaedics were added for a total of 21 patients with cervical arthritis, 13 of whom had no cervical symptoms. All 21 patients with cervical arthritis had erosive peripheral arthritis with at least 10 years of disease duration, and were positive for rheumatoid factor. Almost half of these patients were not under adequate DMARD therapy when cervical instability was diagnosed, and none were on biological response modifiers. CONCLUSIONS: Screening for cervical arthritis is still of importance, especially in patients with erosive seropositive disease. In view of the documented incidence, adherence to screening protocols was disappointing.
Assuntos
Artrite Reumatoide/diagnóstico por imagem , Vértebras Cervicais/diagnóstico por imagem , Programas de Rastreamento/métodos , Padrões de Prática Médica , Fator Reumatoide/sangue , Espondilartrite/diagnóstico por imagem , Centros Médicos Acadêmicos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Assistência Ambulatorial , Antirreumáticos/uso terapêutico , Artrite Reumatoide/sangue , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/epidemiologia , Produtos Biológicos/uso terapêutico , Biomarcadores/sangue , Vértebras Cervicais/efeitos dos fármacos , Distribuição de Qui-Quadrado , Feminino , Alemanha/epidemiologia , Guias como Assunto , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Guias de Prática Clínica como Assunto , Valor Preditivo dos Testes , Prevalência , Prognóstico , Radiografia , Medição de Risco , Fatores de Risco , Espondilartrite/sangue , Espondilartrite/tratamento farmacológico , Espondilartrite/epidemiologia , Fatores de Tempo , Adulto JovemAssuntos
Anticorpos Antinucleares/análise , DNA Topoisomerases Tipo I/imunologia , Técnica Indireta de Fluorescência para Anticorpo/métodos , Algoritmos , Linhagem Celular , Consenso , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Técnica Indireta de Fluorescência para Anticorpo/normas , Humanos , Sociedades CientíficasAssuntos
Anticorpos Antinucleares/análise , Técnica Indireta de Fluorescência para Anticorpo/métodos , Doenças Autoimunes/diagnóstico , Linhagem Celular , Consenso , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Reações Falso-Negativas , Técnica Indireta de Fluorescência para Anticorpo/normas , Humanos , Sociedades CientíficasRESUMO
BACKGROUND: Glycoprotein 2 (GP2) was discovered as the major autoantigen of Crohn's disease (CD)-specific pancreatic autoantibodies (PAB). We investigated anti-GP2 IgA and IgG antibodies as novel serological parameters in CD and assessed their association with distinct disease phenotypes. METHODS: Anti-GP2 and anti-Saccharomyces cerevisiae (ASCA) IgA and IgG were detected by ELISA employing recombinant human GP2 and phosphopeptidomannan, respectively and PAB by indirect immunofluorescence (IIF) in 271 sera, 169 with CD and 102 with ulcerative colitis (UC). As healthy controls 160 adult blood donors and 65 children were included. RESULTS: Anti-GP2 IgG and/or IgA were more prevalent in CD (51/169, 30.2%) than in UC (9/102, 8.9%) patients and in controls (9/225, 4%) (p < 0.001 respectively). ASCA IgG and/or IgA were present in 60/169 (35.5%) in CD and in 7/102 (6.9%) in UC patients (p < 0.001). CD patients with ileocolonic location (L3) showed a significantly higher prevalence of anti-GP2 and ASCA IgA and/or IgG (40/113 and 48/113, respectively; p < 0.05 for both comparisons), whereas CD patients with colonic location (L2) revealed a significantly diminished prevalence for these autoantibody specificities (2/32 and 5/32, respectively, p < 0.05 for both). Anti-GP2 IgG were significantly more prevalent in CD patients with stricturing behaviour (B2) and perianal disease (7/11, p < 0.02) and less prevalent in those with penetrating behaviour (B3) and perianal disease (4/31, p < 0.05). The occurrence of anti-GP2 IgA and/or IgG was significantly more prevalent in CD patients with age at diagnosis of ≤16 years (16/31, p < 0.009). Prevalence of one or more anti-GP2 or ASCA IgA and/or IgG was significantly higher in L3, B2, and A1 and lower in L2 (68/113, 27/41, 23/31, 6/32; p < 0.04, respectively). CONCLUSIONS: Anti-GP2 IgG and IgA, constituting novel CD specific autoantibodies, appear to be associated with distinct disease phenotypes identifying patients at a younger age, with ileocolonic location, and stricturing behaviour with perianal disease.
Assuntos
Autoanticorpos/imunologia , Doença de Crohn/imunologia , Proteínas Ligadas por GPI/imunologia , Pâncreas/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Autoanticorpos/sangue , Criança , Colite Ulcerativa/sangue , Colite Ulcerativa/diagnóstico , Colite Ulcerativa/epidemiologia , Colite Ulcerativa/imunologia , Doenças do Colo/sangue , Doenças do Colo/diagnóstico , Doenças do Colo/imunologia , Doença de Crohn/sangue , Doença de Crohn/diagnóstico , Doença de Crohn/epidemiologia , Feminino , Humanos , Doenças do Íleo/sangue , Doenças do Íleo/diagnóstico , Doenças do Íleo/imunologia , Masculino , Pessoa de Meia-Idade , Prevalência , Saccharomyces cerevisiae/imunologia , Adulto JovemRESUMO
BACKGROUND: This study investigated whether a dot immunoassay (DIA) can provide simultaneous detection of anti-tissue transglutaminase (tTG), anti-deamidated gliadin (DG) and total IgA antibodies, as required in the work-up of celiac disease (CD) patients. METHODS: Celiac disease patients (n=111) consecutively diagnosed from 2001 to 2011 at the Children's Hospital and Institute of Immunology (Technical University Dresden) were tested for anti-tTG, anti-DG and total IgA by enzyme-linked immunosorbent assay (ELISA) and DIA retrospectively. Blood donors (n=45) and non-CD individuals with low IgA serum levels (n=8) were included as controls. Antibodies to endomysial antigens (EmA) were assessed by indirect immunofluorescence (IIF). RESULTS: Four (3.6%) of 111 CD patients demonstrated an IgA deficiency with total IgA below 50 mg/L by ELISA. Total IgA of the 107 IgA-non-deficient CD patients varied from 70 to 6000 mg/L. All four IgA-deficient CD patients were detected by a reduced reaction control of DIA and demonstrated positive anti-tTG or anti-DG IgG by DIA or ELISA. Detection of anti-tTG and anti-DG by DIA and ELISA showed a very good agreement (IgA: κ=0.972, 0.856, respectively; IgG: 0.921, 0.895, respectively). CONCLUSIONS: Immunodot assay is a reliable and easy-to-use technique for the detection of IgA-deficient CD patients. Simultaneous assessment of anti-tTG and anti-DG IgA antibodies, and IgA deficiency by DIA can improve the efficacy of CD serology.
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Doença Celíaca/diagnóstico , Deficiência de IgA/diagnóstico , Imunoensaio , Adolescente , Adulto , Idoso , Doença Celíaca/imunologia , Criança , Pré-Escolar , Feminino , Humanos , Deficiência de IgA/imunologia , Lactente , Masculino , Pessoa de Meia-IdadeRESUMO
Antibody assessment is an essential part in the serological diagnosis of autoimmune diseases. However, different diagnostic strategies have been proposed for the work up of sera in particular from patients with systemic autoimmune rheumatic disease (SARD). In general, screening for SARD-associated antibodies by indirect immunofluorescence (IIF) is followed by confirmatory testing covering different assay techniques. Due to lacking automation, standardization, modern data management, and human bias in IIF screening, this two-stage approach has recently been challenged by multiplex techniques particularly in laboratories with high workload. However, detection of antinuclear antibodies by IIF is still recommended to be the gold standard method for antibody screening in sera from patients with suspected SARD. To address the limitations of IIF and to meet the demand for cost-efficient autoantibody screening, automated IIF methods employing novel pattern recognition algorithms for image analysis have been introduced recently. In this respect, the AKLIDES technology has been the first commercially available platform for automated interpretation of cell-based IIF testing and provides multiplexing by addressable microbead immunoassays for confirmatory testing. This paper gives an overview of recently published studies demonstrating the advantages of this new technology for SARD serology.
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Autoanticorpos/química , Autoanticorpos/imunologia , Doenças Autoimunes/diagnóstico , Doenças Autoimunes/imunologia , Técnica Indireta de Fluorescência para Anticorpo/métodos , Testes Sorológicos/métodos , Técnica Indireta de Fluorescência para Anticorpo/instrumentação , Humanos , Testes Sorológicos/instrumentaçãoRESUMO
Why zymogen glycoprotein 2 (GP2), the Crohn's disease (CD)-specific pancreatic autoantigen, is the major target of humoral autoimmunity in inflammatory bowel diseases (IBD) is uknown. Recent evidence demonstrates that GP2 is also present on the apical surface of microfold (M) intestinal cells. As the colon lacks GP2-rich M cells, we assumed that patients with colonic CD are seronegative for anti-GP2. Anti-GP2 antibodies were tested in 225 CDs, including 45 patients with colonic location (L2), 45 with terminal ileum (L1) and 135 with ileocolonic involvement; 225 patients with ulcerative colitis (UC) were also tested. Anti-GP2 reactivity was detected in 59 (26.2%) CDs and 15 (6.7%) UCs (P < 0.001). Only 5 CDs with L2 had anti-GP2 antibodies, compared to 54/180 (30.0%, P = 0.0128) of the CDs with L1 and L3. Anti-GP2 antibody positive CD patients had higher ASCA titres compared to seronegative cases. Amongst the 128 CD patients with previous surgical intervention, 45 (35.0%) were anti-GP2 antibody positive compared to 14/97 (14.0%) without surgical (P < 0.001). Our data support the assumption that ileal inflammation is required for the development of anti-GP2 antibodies in CD, and suggest that the intestine rather than the pancreatic juice is the antigenic source required for the initiation of anti-GP2 antibodies.
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Autoanticorpos/imunologia , Doença de Crohn/imunologia , Proteínas Ligadas por GPI/imunologia , Ileíte/imunologia , Glicoproteínas de Membrana/imunologia , Adulto , Autoantígenos/imunologia , Colite Ulcerativa/imunologia , Feminino , Humanos , Imunoglobulina G/imunologia , Inflamação/imunologia , Doenças Inflamatórias Intestinais/imunologia , Masculino , Pessoa de Meia-Idade , Pâncreas/imunologiaRESUMO
A highly sensitive detection of anti-neutrophil cytoplasmic antibodies to serine proteinase-3 (PR3-ANCAs) aids in the serological diagnosis of autoimmune liver disorders and the prediction of severity in primary sclerosing cholangitis (PSC). Here, we evaluate a novel third-generation ELISA for the detection of PR3-ANCAs. In total, 309 patients with PSC, 51 with primary biliary cholangitis (PBC), and 120 healthy blood donors (BD) were analyzed. For the survival analysis in PSC, the outcome was defined as liver-transplantation-free survival during the follow-up. Positive PR3-ANCA levels were found in 74/309 (24.0%) of patients with PSC. No BDs and one patient with PBC demonstrated PR3-ANCA positivity. PR3-ANCAs were revealed as independent predictors for a poor PSC outcome (study endpoint: liver transplantation/death, log-rank test, p = 0.02). PR3-ANCA positivity, lower albumin levels, and higher bilirubin concentrations were independent risks of a poor survival (Cox proportional-hazards regression analysis, p < 0.05). The Mayo risk score for PSC was associated with PR3-ANCA positivity (p = 0.01) and the disease severity assessed with a model of end-stage liver disease (MELD) and extended MELD-Na (p < 0.05). PR3-ANCAs detected by a third-generation ELISA are diagnostic and prognostic markers for PSC. Their wider use could help to identify patients who are at-risk of a more severe disease.
RESUMO
The relation between infections and autoimmune diseases has been extensively investigated. Multiple studies suggest a causal relation between these two entities with molecular mimicry, hyperstimulation and dysregulation of the immune system as plausible mechanisms. The recent pandemic with a new virus, i.e., SARS-CoV-2, has resulted in numerous studies addressing the potential of this virus to induce autoimmunity and, eventually, autoimmune disease. In addition, it has also revealed that pre-existing auto-immunity (auto-Abs neutralizing type I IFNs) could cause life-threatening disease. Therefore, the topic of the 15th Dresden Symposium on Autoantibodies was focused on autoimmunity in the SARS-CoV-2 era. This report is a collection and distillation of the topics presented at this meeting.