The potential of using blood circular RNA as liquid biopsy biomarker for human diseases

Circular RNA (circRNA) is a novel class of single-stranded RNAs with a closed loop structure. The majority of circRNAs are formed by a back-splicing process in pre-mRNA splicing. Their expression is dynamically regulated and shows spatiotemporal patterns among cell types, tissues and developmental stages. CircRNAs have important biological functions in many physiological processes, and their aberrant expression is implicated in many human diseases. Due to their high stability, circRNAs are becoming promising biomarkers in many human diseases, such as cardiovascular diseases, autoimmune diseases and human cancers. In this review, we focus on the translational potential of using human blood circRNAs as liquid biopsy biomarkers for human diseases. We highlight their abundant expression, essential biological functions and significant correlations to human diseases in various components of peripheral blood, including whole blood, blood cells and extracellular vesicles. In addition, we summarize the current knowledge of blood circRNA biomarkers for disease diagnosis or prognosis.

Comparison between enzyme-linked immunosorbent assay and indirect immunofluorescence for detection of antineutrophil cytoplasmic antibodies / Comparação entre os métodos de ensaio imunoenzimático e imunoflurescência indireta para a pesquisa de anticorpos anticitoplasma de neutrófilos

ABSTRACT Objective To evaluate the performance of enzyme-linked immunosorbent assay and indirect immunofluorescence methods for the detection of antineutrophil cytoplasmic antibodies in a routine clinical laboratory setting. Methods A total of 227 samples were tested by indirect immunofluorescence and enzyme-linked immunosorbent assay with antigen specificity for antiproteinase 3 and antimyeloperoxidase. The proportions of positive samples were compared by McNemar hypotheses and agreement was described by Cohen's Kappa coefficient. Results The agreement of the tests was 96.5%, and the Kappa coefficient obtained was 0.70 (95%CI: 0.50-0.90; p<0.001). Considering indirect immunofluorescence as the gold standard, the sensitivity of the enzyme-linked immunosorbent assay was 0.62 and the specificity was 0.99, with diagnostic accuracy in 96% of cases. Some samples were negative in enzyme-linked immunosorbent assay and positive in indirect immunofluorescence. This situation occurred in all immunofluorescence patterns, but particularly in atypical patterns. Two samples with antiproteinase 3 positivity were considered negative in indirect immunofluorescence. Conclusion The enzyme-linked immunosorbent assay had high specificity but lower sensitivity. The performance of indirect immunofluorescence increases diagnostic sensitivity, while the search for antiproteinase 3 by enzyme-linked immunosorbent assay may also add diagnostic power.

RESUMO Objetivo Avaliar o desempenho das metodologias de ensaio imunoenzimático e imunofluorescência indireta para a detecção de anticorpos anticitoplasma de neutrófilos em um contexto de laboratório clínico de rotina. Métodos Foram testadas 227 amostras pelas metodologias de imunofluorescência indireta e ensaio imunoenzimático com especificidades para anticorpos antiproteinase-3 e antimieloperoxidase. As proporções de amostras positivas foram comparadas por hipóteses de McNemar, e a concordância foi descrita pelo coeficiente Kappa de Cohen. Resultados A concordância dos testes foi 96,5%, e o coeficiente Kappa obtido foi 0,70 (IC95%: 0,50-0,90; p<0,001). Utilizando a imunofluorescência indireta como padrão-ouro, a sensibilidade do ensaio imunoenzimático foi de 0,62 e a especificidade, 0,99, com acurácia diagnóstica em 96% dos casos. Algumas amostras apresentaram resultados negativos por ensaio imunoenzimático e positivos por imunofluorescência. Isso ocorreu em amostras com vários padrões de fluorescência, mas particularmente nos casos com padrões atípicos. Duas amostras com positividade antiproteinase 3 foram consideradas negativas por imunofluorescência. Conclusão Os métodos de ensaio imunoenzimático tiveram alta especificidade, mas sensibilidade inferior. A realização da imunofluorescência indireta aumenta a sensibilidade diagnóstica, ao mesmo tempo que a pesquisa de antiproteinase 3 por ensaio imunoenzimático também pode agregar poder diagnóstico.

Bajos niveles de colesterol de HDL y alto riesgo de enfermedad autoinmune. Dos estudios poblacionales de cohorte con 117.341 individuos incluidos. / Low HDL cholesterol and high risk of autoimmune disease. Two population-based cohort studies including 117, 341 individuals

Antecedentes. HDL es cuantitativamente la lipoproteína más importante en la mayoría de las especies y la evidencia mecanicista sugiere que HDL tendría un papel en la función inmunológica normal. Probamos la hipótesis que sugiere que las concentraciones plasmáticas de HDL están asociadas con el riesgo de enfermedades autoinmunes. Métodos. Se incluyeron 107.954 y 9.387 individuos con mediciones basales de colesterol-HDL provenientes de 2 estudios de la población general: el Estudio de la Población General de Copenhague y el Estudio del Corazón de Copenhague. Los pacientes fueron seguidos mediante el Registro Nacional Danés de pacientes desde el inicio del período 2003-2015 o 1991-1994 hasta 2017, tiempo durante el cual 4.078 y 1.101 individuos desarrollaron enfermedad autoinmune en los 2 estudios respectivamente. Resultados. En el Estudio de la Población General de Copenhague, en comparación a los individuos con colesterol de HDL =2,0 mmol/L (77 mg/dL), los índices de riesgo para cualquier enfermedad autoinmune, ajustados de manera multifactorial fueron 1,06 (IC 95%, 0,94-1,19) para individuos con colesterol-HDL entre 1,5 y 1,99 mmol/L (58 a 77 mg/dL), 1,18 (IC 95%, 1,04-1,35) para individuos con colesterol-HDL entre 1,0 y 1,49 mmol/L (39 a 58 mg/dL) y 1,84 (IC 95%, 1,52- 2,22) para individuos con colesterol-HDL <1,0 mmol/L (39 mg/dL) (p<0,001 para tendencia). Estos resultados fueron similares cuando: se excluyeron los eventos dentro de los 5 años del inicio del estudio, tanto en mujeres como hombres por separado, eventos en el inicio del estudio, independientemente de la inflamación de bajo grado o concentraciones de triglicéridos, para diferentes niveles de apolipoproteína A1 y para definiciones de punto final más restrictivas. Finalmente, el Estudio del Corazón de Copenhague proporcionó una confirmación independiente. Conclusiones: Los bajos niveles de colesterol-HDL se asocian con un alto riesgo de enfermedad autoinmune en individuos de la población general. Nuestros hallazgos observacionales no pueden determinar la causalidad.

Background. HDL is quantitatively the most important lipoprotein in most species and mechanistic evidence points toward a role for HDL in normal immune function. We tested the hypothesis that concentrations of HDL cholesterol are associated with risk of autoimmune disease. Methods. From 2 studies of the general population-the Copenhagen General Population Study and the Copenhagen City Heart study-we included 107,954 and 9,387 individuals with baseline measurements of HDL cholesterol. These were followed with the national Danish Patient Registry from baseline in 2003-2015 or 1991-1994 through 2017, during which time 4078 and 1101 individuals developed autoimmune disease in the 2 studies. Results. In the Copenhagen General Population Study, compared to individuals with HDL cholesterol =2.0 mmol/L (77 mg/dL), the multifactorially adjusted hazard ratios for any autoimmune disease were 1.06 (95% CI, 0.94-1.19) for individuals with HDL cholesterol of 1.5-1.99 mmol/L (58-77 mg/dL), 1.18 (95% CI, 1.04-1.35) for individuals with HDL cholesterol of 1.0-1.49 mmol/L (39-58 mg/dL), and 1.84 (95% CI, 1.52-2.22) for individuals with HDL cholesterol <1.0 mmol/L (39 mg/dL) (p for trend <0.001). These results were similar when excluding events within 5 years of baseline, in women and men separately, for events at baseline, irrespective of low-grade inflammation or triglyceride concentrations, for the apolipoprotein A1 part of HDL, and for more restrictive end point definitions. Finally, the Copenhagen City Heart Study provided independent confirmation. Conclusions. Low HDL cholesterol level is associated with high risk of autoimmune disease in individuals from the general population. Our observational findings cannot determine causality.

Prevalence of thyroid autoantibodies in patients with systematic autoimmune rheumatic diseases. Cross-sectional study

ABSTRACT BACKGROUND: Thyroid autoimmunity is more common in patients with rheumatic diseases than in healthy populations. The degree of association seems subject to influence from patients' geographical location. Here, we aimed to ascertain the prevalence of thyroid autoantibodies in a cohort of patients with systemic rheumatic disease and the degree of association between its presence and inflammatory activity. DESIGN AND SETTING: Cross-sectional observational study in a rheumatology unit. METHODS: 301 patients with systemic lupus erythematosus (SLE), 210 with rheumatoid arthritis (RA), 58 with scleroderma (SSc) and 80 with spondyloarthritis (SpA) were studied regarding thyroid function (TSH and T4), anti-thyroglobulin (TgAb) and anti-thyroperoxidase (TPOab) and compared with 141 healthy controls. Disease activity in patients with rheumatic disease was assessed through appropriate indexes. RESULTS: There were more antithyroid antibodies in SLE patients with hypothyroidism (P = 0.01; odds ratio, OR 2.7; 95% confidence interval, CI: 1.20-6.26) and in those without hypothyroidism (P = 0.06; OR 2.4; 95% CI: 1.28-4.55) than in controls. SSc patients also showed: P = 0.03 both with antithyroid antibodies and hypothyroidism (OR 3.4; 95% CI: 1.06-10.80) and without hypothyroidism (OR 3.1; 95% CI: 1.11-0.13). RA and SpA patients had the same prevalence as controls (P not significant). Presence of autoantibodies with and without hypothyroidism was not associated with the activity or functional indexes evaluated. CONCLUSION: SLE and SSc were associated with higher prevalence of thyroid autoantibodies in patients with and without hypothyroidism, unlike SpA and RA. There was no link between thyroid autoantibody presence and disease activity or functional impairment.

Eritrosedimentación extremadamente elevada en una población de adultos en la ciudad de Buenos Aires / Extremely elevated erythrosedimentation in an adult population in the City of Buenos Aires

Resumen Introducción: Una velocidad de eritrosedimentación (VHS) extremadamente elevada, definida como mayor o igual a 100 mm/h, se ha asociado a condiciones graves subyacentes como enfermedades infecciosas, enfermedades del colágeno u oncológicas. Objetivo: Analizar un grupo de pacientes para determinar los diagnósticos de base y las características que se asocian con valores de VHS mayores a 100 mm/h en nuestro medio. Pacientes y Métodos: Estudio tipo observacional de corte transversal, con recolección retrospectiva de datos de pacientes adultos con al menos un valor de VHS mayor o igual a 100 mm/h, registrado en el laboratorio entre enero de 2002 y agosto de 2014 en el Hospital Italiano de Buenos Aires. Resultados: Durante el período evaluado se analizaron 879 pacientes mayores de 18 años. La mediana de los valores de VHS fue 111 mm/h (Rango intercuartil 105-120). La etiología prevalente de VHS elevada fueron las enfermedades infecciosas (41,6%), seguida de malignidad (21,6%) y de autoinmune/inflamatoria (12,9%). El diagnóstico individual más frecuente fue el de neumonía (11,4%), seguido por causa indeterminada (5,9%). Conclusión: En pacientes internados, la causa más frecuente de VHS ≥ 100 mm/h fue las enfermedades infecciosas, mientras que en pacientes ambulatorios la causa más frecuente fue la malignidad.

Background: An extremely elevated erythrosedimentation rate (ESR), defined as equal or higher than 100 mm/h, has been linked to serious underlying conditions, such as infections, connective tissue and oncologic disease. Aim: To analyze a group of patients in order to determine the underlying diagnosis and the characteristics associated with extremely elevated ESR in our environment. Methods: Cross-sectional study of adult patients, who presented with at least one ESR equal or higher than 100 mm/h at Hospital Italiano, in Buenos Aires (Buenos Aires, Argentina) between January 2002 and August 2014. Results: During the previously stated period of time, we analyzed the results of 879 patients. All patients were over 18 years of age. The median for the ESR results was 111 mm/h (interquartile range 105-120). The most prevalent etiology of an elevated ESR was infectious (41.64%), followed by malignancies (21.62%) and autoimmune / inflammatory diseases (12.97%). The most frequent individual diagnosis found was pneumonia (11.49%), followed by undetermined causes (5.92%). Conclusion: When comparing inpatient versus outpatient populations, the most frequent cause was infectious in the former group, while malignancies were the most frequent diagnosis in the latter.

Pancreatitis autoinmune tipo 1: reporte de caso / Autoimmune pancreatitis type 1: a case report

La pancreatitis autoinmune tipo 1 es una enfermedad de baja prevalencia siendo más frecuente en varones, se encuentra incluida dentro de las enfermedades relacionadas a IgG4. Esta patología puede debutar como un síndrome colestásico y el diagnóstico se realiza según los criterios del consenso internacional para pancreatitis autoinmune (ICDC) que incluye una imagen típica, serología, compromiso de otros órganos, histología y respuesta al tratamiento con corticoides. Presentamos el caso de una mujer de 52 años con antecedente de artritis reumatoide sin tratamiento que acude con tiempo de enfermedad de 2 meses caracterizado por dolor abdominal en hipocondrio derecho de moderada intensidad asociado a ictericia, coluria, hipocolia, xeroftalmia, xerostomía y pérdida de peso de 3 kg. Al examen físico se evidencia ictericia, hipertrofia simétrica de glándulas submandibulares, leve dolor en epigastrio. En los exámenes auxiliares existe patrón colestásico con hiperbilirrubinemia a predominio directo. En los estudios de imágenes se evidencia colédoco dilatado, con aumento difuso del volumen del páncreas con captación tardía de contraste. En el estudio inmunológico se evidenció IgG4 en 610 u/L y ANA 1/640. Se inició tratamiento con corticoides con respuesta clínica y de laboratorio favorable. En conclusión, se debe sospechar de pancreatitis autoinmune ante un cuadro de dolor abdominal y colestasis extrahepática con imágenes sugestivas de páncreas inflamatorio, en el que se deben complementar los criterios ICDC para confirmar el diagnóstico

Autoimmune pancreatitis type 1 (AIP) is the pancreatic manifestation of IgG4-related disease. The most frequent presentation of AIP is with obstructive jaundice. For definite diagnosis of type 1 Autoimmune pancreatitis international consensus diagnosis criteria (ICDC) for AIP are used. ICDC criteria include pancreatic parenchymal imaging, ductal imaging, serology, other organ involvement, histology, and response to steroid. We report a 52-years-old woman with rheumatoid arthritis without treatment presented with two months of abdominal pain in up-right quadrant with moderate intensity. She also presented jaundice, acholia, xerophtalmia, xerostomia, and a weight loss of 5 pounds. On examination jaundice, symmetrically enlarged submandibular glands, and epigastric pain was observed. On laboratory, a cholestasis pattern and conjugated bilirubin predominance was found. CT Abdominal, CMR revealed a dilated common bile duct with a diffuse pancreatic enlargement with delayed enhancement. Immunological studies show a IgG4 610 u/l and ANA 1/640. The patient responds to steroid clinically and in the laboratorial values. In conclusion, autoimmune pancreatitis type 1 should be suspected in case of an obstructive jaundice with a pancreatic inflammatory image, and complete ICDC criteria for a definite diagnosis

Severe hypoproteinemia as a harbinger of Ménétrier's disease in autoimmune pancreatitis / Hipoproteinemia grave como indicador de doença de Ménétrier na pancreatite autoimune

Summary Ménétrier's disease is an extremely rare disease of unknown etiology causing gastric mucosal hypertrophy and protein-losing gastropathy. Rare cases of this condition have been reported in patients with autoimmune diseases. However, to the best of our knowledge, Ménétrier's disease associated with autoimmune pancreatitis (AIP) has never been reported. We described a case of severe hypoproteinemia as a harbinger of Ménétrier's disease associated with AIP. The patient was successfully treated with octreotide and high-protein diet, which led to symptomatic remission and significant improvement in serum levels of albumin and recovery of the nutritional status. Thus, in AIP patients presenting with severe and persistent hypoproteinemia without apparent cause, clinicians need to consider Ménétrier's disease in the differential diagnosis. In this setting, endoscopic evaluation with histological examination of gastric biopsy material, including a full-thickness mucosal biopsy of involved mucosa, may be helpful in promptly establishing the diagnosis and allowing appropriate and timely therapy.

Resumo A doença de Ménétrier é uma condição extremamente rara, de etiologia desconhecida, caracterizada por hipertrofia da mucosa gástrica e gastropatia perdedora de proteína. Casos raros dessa patologia têm sido relatados em pacientes com doenças autoimunes. Até o momento, desconhecemos qualquer relato dessa doença associada à pancreatite autoimune (PAI). Descrevemos um caso de hipoproteinemia grave como indicador de doença de Ménétrier associada à PAI. O paciente foi tratado de forma satisfatória com octreotide e dieta hiperproteica, alcançando remissão sintomática, melhora significativa das concentrações de albumina e recuperação do estado nutricional. Portanto, em pacientes com PAI e hipoproteinemia grave e persistente, deve-se considerar a doença de Ménétrier como um diagnóstico diferencial. Nesses casos, a avaliação endoscópica com biópsia gástrica, incluindo biópsia de toda a espessura da mucosa, pode ser útil no estabelecimento do diagnóstico e do pronto início da terapêutica.

Basophil Markers for Identification and Activation in the Indirect Basophil Activation Test by Flow Cytometry for Diagnosis of Autoimmune Urticaria

BACKGROUND: The indirect basophil activation test using flow cytometry is a promising tool for autoimmune urticaria diagnosis. We aimed to identify better donor basophils (from atopic vs. non-atopic donors and interleukin-3 primed vs. unprimed basophils) and improve basophil identification and activation markers (eotaxin CC chemokine receptor-3 [CCR3] vs. CD123 and CD63 vs. CD203c). METHODS: Donor basophils were obtained from non-atopic and atopic group O donors. Positive control sera were artificially prepared to simulate autoimmune urticaria patients' sera. Patient sera were obtained from nine children with chronic urticaria. Assay sensitivity was compared among each variation by using positive control sera (n=21), applying cutoff values defined from negative control sera (n=20). RESULTS: For basophil identification, a combination of CCR3 and CD123 markers revealed a higher correlation with automated complete blood count (r=0.530) compared with that observed using CD123 (r=0.498) or CCR3 alone (r=0.195). Three activation markers on the atopic donor basophils attained 100% assay sensitivity: CD203c on unprimed basophils, CD63+CD203+ or CD63 alone on primed basophils; however, these markers on the non-atopic donor basophils attained lower assay sensitivity. CONCLUSIONS: For basophil identification markers, a combination of CD123 and CCR3 is recommended, while CD123 alone may be used as an alternative. Donor basophils should be obtained from an atopic donor. For basophil activation markers, either CD203c alone on unprimed basophils or CD203c and CD63 on primed basophils are recommended, while CD63 alone on primed basophils may be used as an alternative.

Síndrome de hipoglicemia autoinmune: Primeros casos en Chile / Insulin autoimmune syndrome: Report of two cases

Insulin autoimmune syndrome (IAS) is characterized by spontaneous hypoglycemia with extremely high insulin levels and the presence of circulating autoantibodies against insulin, in patients who have never been exposed to exogenous insulin. We report two patients with the syndrome. A 36 years old male presenting with hypoglycemia in the emergency room had an oral glucose tolerance test showed basal and 120 min glucose levels of 88 and 185 mg/dl. The basal and 120 min insulin levels were 2,759 and 5,942 μUI/ml. The presence of an insulin secreting tumor was discarded. Anti-insulin antibodies were positive. He was successfully treated with a diet restricted in carbohydrates and frequent meals in small quantities. A 65 years old female presenting with hypoglycemia in the emergency room had the fasting insulin levels of 1,910 µUI/ml. No insulin secreting tumor was detected by images and anti-insulin antibodies were positive. The polyethylene glycol precipitation test showed a basal and after exposition insulin level 1,483 and 114 µUI/ml, respectively. She responded partially to diet and acarbose and required the use of prednisone with a good clinical response.

Doenças autoimunes e autoanticorpos em pacientes pediátricos e seus parentes de primeiro grau com deficiência de imunoglobulina / Autoimmune diseases and autoantibodies in pediatric patients and their first-degree relatives with immunoglobulin A deficiency

Introdução: As manifestações clínicas da deficiência de imunoglobulina A (DIgA) incluem infecções recorrentes, atopia e doenças autoimunes. No entanto, para o nosso conhecimento, as avaliações concomitantes de doenças autoimunes e autoanticorpos em uma coorte de pacientes com DIgA com idade atual > 10 anos e seus parentes não foram feitas. Objetivos: Avaliar doenças autoimunes e presença de autoanticorpos em pacientes com DIgA e seus parentes de primeiro grau. Métodos: Estudo transversal feito em 34 pacientes com DIgA (idade atual > 10 anos) e em seus parentes de primeiro grau. Todos foram acompanhados em um centro terciário brasileiro para imunodeficiência primária: 27 crianças/adolescentes e sete de seus parentes de primeiro grau com diagnóstico tardio de DIgA. Doenças autoimunes e autoanticorpos (anticorpos antinucleares, fator reumatoide e antitireoglobulina, antitiroperoxidase e anticorpos antiendomísio da classe IgA) também foram avaliadas. Resultados: Doenças autoimunes (n = 14) e/ou autoanticorpos (n = 10, quatro deles com autoanticorpos isolados) foram observadas em 18/34 (53%) dos pacientes e seus parentes. As doenças autoimunes mais comuns encontradas foram tireoidite (18%), artrite crônica (12%) e doença celíaca (6%). Os autoanticorpos mais frequentes foram anticorpos antinucleares (2%), antitireoglobulina e/ou antitireoperoxidase (24%). Nenhuma diferença significativa foi observada no sexo feminino, idade no momento do diagnóstico e idade atual em pacientes com DIgA com e sem doenças autoimunes e/ou presença de autoanticorpos (p > 0,05). As frequências de imunodeficiência de primárias na família, autoimunidade em família, atopia e infecções recorrentes foram semelhantes em ambos os grupos (p> 0,05). Conclusão: Doenças autoimunes e autoanticorpos foram observadas em pacientes com DIgA durante o acompanhamento, o que reforça a necessidade de um acompanhamento rigoroso e contínuo durante a adolescência e a idade adulta. .

Introduction: Clinical manifestations of Immunoglobulin A Deficiency (IgAD) include recur-rent infections, atopy and autoimmune diseases. However, to our knowledge, theconcomitant evaluations of autoimmune diseases and auto antibodies in a cohort of IgADpatients with current age >10 years and their relatives have not been assessed. Objectives: To evaluate autoimmune diseases and the presence of auto antibodies in IgADpatients and their first-degree relatives. Methods: A cross-sectional study was performed in 34 IgAD patients (current age >10years) and their first-degree relatives. All of them were followed at a tertiary Brazilianprimary immunodeficiency center: 27 children/adolescents and 7 of their first-degree rela-tives with a late diagnosis of IgAD. Autoimmune diseases and autoantibodies (antinuclearantibodies, rheumatoid factor, and anti-thyroglobulin, anti-thyroperoxidase and IgA classanti-endomysial antibodies) were also assessed. Results: Autoimmune diseases (n = 14) and/or autoantibodies (n = 10, four of them with iso-lated autoantibodies) were observed in 18/34 (53%) of the patients and their relatives. Themost common autoimmune diseases found were thyroiditis (18%), chronic arthritis (12%)and celiac disease (6%). The most frequent autoantibodies were antinuclear antibodies(2%), anti-thyroglobulin and/or anti-thyroperoxidase (24%). No significant differences wereobserved in the female gender, age at diagnosis and current age in IgAD patients with andwithout autoimmune diseases and/or presence of auto antibodies (p > 0.05). The frequen-cies of primary immunodeficiencies in family, autoimmunity in family, atopy and recurrentinfections were similar in both groups (p > 0.05). Conclusion: Autoimmune diseases and auto antibodies were observed in IgAD patients dur-ing follow-up, reinforcing the necessity of a rigorous and continuous follow-up duringadolescence and adulthood. .

Deteccao de anticorpos antinucleares por imunofluorescencia indireta em celulas HEp-2: definindo a diluicao de triagem adequada para o diagnostico das doencas reumaticas autoimunes / Detection of anti-nuclear antibodies by indirect immunofluorescence on HEp-2 cells: setting the appropriate screening dilution for the diagnosis of autoimmune rheumatic diseases

Objetivo: Definir o título anormal e a diluição de triagem adequada para o teste de FAN (fator antinúcleo) por imunofluorescência indireta em células HEp-2 (FAN HEp-2). Métodos: Realizamos a pesquisa do FAN HEp-2 em amostras de soro de 126 indivíduos saudáveis. As amostras foram triadas na diluição de 1:80, e aquelas positivas diluídas até o título de 1:5120. O título anormal de FAN foi definido como aquele correspondente ao percentil 95 do teste nesta população. A sensibilidade dos diferentes títulos do FAN foi determinada em um grupo de 136 pacientes com diagnóstico de doença reumática autoimune, e a especificidade em um grupo de 118 pacientes com diagnóstico de outras doenças reumáticas. O valor de corte ótimo do teste foi determinado pelo estudo da curva ROC. Resultados: A frequência de FAN positivo em indivíduos saudáveis foi de 13,2%. Não houve diferença na frequência de resultados positivos de acordo com o gênero ou a idade. O título anormal do FAN foi definido como a diluição de 1:160. A diluição dos soros de 1:80 apresentou sensibilidade de 87,7% e especificidade de 67,8%, enquanto a diluição de 1:160 apresentou sensibilidade de 82% e especificidade de 73,7%. Pela análise da curva ROC, a diluição de 1:160 correspondeu ao valor de corte ótimo. Conclusão: O título anormal e o valor de corte ótimo do FAN HEp-2 na população avaliada foram de 1:160. A diluição de 1:160 é, portanto, a diluição de triagem ideal, com melhor especificidade, porém sem comprometimento significativo da sensibilidade diagnóstica do teste. .

Objective: To establish the abnormal title and the appropriate screening dilution for ANA (antinuclear antibodies) test by indirect immunofluorescence on HEp-2 cells (ANA HEp-2). Methods: An analysis of ANA Hep-2 in serum samples from 126 healthy individuals was performed. The samples were screened at a dilution of 1:80, and those positive were diluted to the title of 1:5120. The abnormal title of ANA was defined as that corresponding to the 95th percentile of the test in this population. The sensitivity of the different titles of antinuclear antibodies was determined in a group of 136 patients with a diagnosis of autoimmune rheumatic disease, and the specificity was determined in a group of 118 patients with other rheumatic diseases. The optimal cutoff value of the test was determined by ROC curve analysis. Results: The frequency of ANA positivity in healthy subjects was 13.2%. There was no difference in the frequency of positive results according to gender or age. The abnormal title of ANA was defined as the dilution of 1:160. The 1:80 dilution had sensitivity of 87.7% and specificity of 67.8%, while the 1:160 dilution had sensitivity of 82% and specificity of 73.7%. By ROC curve analysis, a dilution of 1:160 corresponded to the optimal cutoff value. Conclusion: The abnormal title and the optimal cutoff value of ANA HEp-2 in the population was 1:160. Therefore, the dilution of 1:160 is the optimal screening dilution, with better specificity but without significantly compromising the sensitivity of the diagnostic test. .

Utilidade da triagem dos anticorpos anti-dsDNA por quimioluminescência, seguida de confirmação por imunofluorescência indireta / Usefulness of anti-dsDNA antibody screening with chemiluminescence followed by confirmation by indirect immunofluorescence

OBJETIVO: Avaliar o desempenho de um imunoensaio quimioluminescente (CLIA) para os anticorpos anti-dsDNA, utilizando como referência o teste de imunofluorescência indireta (IFI) sobre Crithidia luciliae. MÉTODOS: O sistema de automação foi previamente aprovado com 81% de eficiência, 100% de sensibilidade e 82% de especificidade, por processo de validação intrínseca em 179 amostras consecutivas de 169 pacientes no início de 2011. A seguir, esses pacientes foram subdivididos em três grupos de acordo com os resultados da pesquisa dos anticorpos anti-dsDNA nas duas metodologias (reagentes, não reagentes e resultados discrepantes). RESULTADOS: Na análise dos dados: 1) 77% (129/169) dos exames haviam sido solicitados por médicos reumatologistas; 2) 57% (97/169) das amostras eram de pacientes lúpicos; 3) Os resultados de CLIA, reagentes e não reagentes, estavam bem definidos e padronizados; 4) A automação reduziu em 70% as passagens pela técnica manual com segurança e qualidade; 5) A alta prevalência de pacientes lúpicos e com nefrite entre os resultados de CLIA falso-positivos corrobora a hipótese de que o índice real de falsa positividade do CLIA seja menor que o encontrado inicialmente neste estudo.

OBJECTIVE: The purpose of this study was to evaluate the performance of a chemiluminescent immunoassay (CLIA) to detect anti-dsDNA antibodies, using the indirect immunofluorescence test (IIF) on Crithidia luciliae as a reference. METHODS: The automation system demonstrated 81% efficiency, 100% sensitivity and 82% specificity according to the intrinsic validation process performed using 179 consecutive samples from 169 patients in the beginning of 2011. These patients were subsequently divided into 3 groups according to the co-reactivity of anti-dsDNA results using the 2 methods (reactive, non-reactive and discrepant results). RESULTS: Upon data analysis, 77% (129/169) of the tests were requested by rheumatologists, and 57% (97/169) of the samples were from lupus patients. Both the reactive and non-reactive results of the CLIA were well defined and standardised, and automation reduced the manual labor required by 70% in a safe and high-quality manner. Furthermore, the high prevalence of patients with lupus and nephritis among the CLIA false-positive results corroborates the hypothesis that the actual index of CLIA false positivity is lower than that initially found in this study.

A Comparative Study on Serum Immunoglobulin and Tumor Marker Levels in the Patients with Autoimmune Pancreatitis and Pancreatobiliary Malignancies / 대한소화기학회지

BACKGROUND/AIMS: Autoimmune pancreatitis (AIP) often occurs with obstructive jaundice in old age in cases of weight loss, mimicking pancreatobiliary cancer. This study aimed to determine the sensitivity and specificity serum IgG, IgG4 and CEA, CA 19-9 levels for the diagnosis of AIP and their ability to distinguish AIP from pancreatobiliary cancer. METHODS: The level of serums IgG, IgG4 and CEA, CA 19-9 were measured in 413 patients including 125 with AIP, 201 with pancreatic cancer, and 87 with cholangiocarcinoma. RESULTS: Among AIP patients, 43.2% (54/125) showed elevated IgG levels (> or =1,800 mg/dL) and 52% (65/125) showed elevated IgG4 levels (> or =135 mg/dL). Sensitivity and specificity of elevated serum IgG for diagnosis AIP were 43% and 88% respectively, and 52% and 97%, respectively for elevated serum IgG4. When the cut-off value of serum IgG4 was raised to 270 mg/dL (twice the upper limit of normal), the specificity improved to 100%. About 25% of the AIP patients showed an increased level of CA 19-9 at >37 U/mL and about 12.2% of them showed an increased level of CA 19-9 at >100 U/mL. On the contrary, only 1.8% of the AIP patients showed an increased level of CEA at >6.0 ng/mL. CONCLUSIONS: To avoid unnecessary surgeries resulting from a misdiagnosed pancreatobiliary cancer as opposed to AIP, it is necessary to consider both serum immunoglobulin and tumor marker. In particular, because high level of IgG4 (> or =270 mg/dL) and CA19-9 (>100 U/mL) are relatively rare in pancreatobiliary cancer and AIP, respectively, they will be helpful in differential diagnosis.

Prevalência de anticorpos séricos contra rubéola em doenças autoimunes / Prevalence of rubella serum antibody in autoimmune diseases

INTRODUÇÃO: A associação entre infecções e doenças autoimunes (DAIs) está bem descrita na literatura médica. Vários agentes infecciosos foram implicados como indutores de respostas autoimunes, tais como o parvovírus B19, o vírus Epstein-Barr, o citomegalovírus e os vírus da hepatite. PACIENTES E MÉTODOS: Foram examinamos 1.173 soros de pacientes com 14 doenças autoimunes diferentes e 238 soros de controles saudáveis pareados geograficamente na busca por evidência de infecção rubeólica prévia. Todas as amostras foram testadas para a presença de anticorpos séricos contra rubéola usando-se o sistema Bio-Rad BioPlex 2200. RESULTADOS: Como um grupo, os pacientes com DAIs apresentaram maior prevalência de anticorpos IgM antirrubéola em comparação aos controles saudáveis (11,7% versus 5,4%; P = 0,001). A prevalência de anticorpos IgM antirrubéola foi significativamente maior em 5/14 DAIs, a saber: arterite de células gigantes (33,3%), cirrose biliar primária (24%), síndrome antifosfolipídica (20,6%), polimiosite (16%) e doença intestinal inflamatória (16%). Detectou-se prevalência semelhante de anticorpos IgM antirrubéola nos controles de diferentes países. Detectou-se alta prevalência de anticorpos IgG antirrubéola em pacientes com DAIs (89,9%) e controles. CONCLUSÃO: A prevalência aumentada de anticorpos IgM antirrubéola em DAIs sugere que a rubéola possa desempenhar um papel na etiopatogênese de várias DAIs.

INTRODUCTION: The association between infections and autoimmune diseases (AID) has been well described in the medical literature. Several infectious agents have been implicated as inducers of autoimmune responses, such as Parvovirus B19, Epstein-Barr virus, cytomegalovirus, and hepatitis viruses. PATIENTS AND METHODS: We examined 1,173 sera from patients with 14 different AID and 238 sera from geographically matched healthy controls, for evidence of prior infection with rubella. All samples were tested for the presence of serum antibodies against rubella using the Bio-Rad BioPlex 2200 system. RESULTS: As a group, patients with AID had a higher prevalence of IgM anti-rubella antibodies as compared to healthy controls (11.7% versus 5.4%; P = 0.001). The prevalence of IgM anti-rubella antibodies was significantly higher in 5/14 AID, namely in patients with giant cell arteritis (33.3%), primary biliary cirrhosis (24%), antiphospholipid syndrome (20.6%), polymyositis (16%), and inflammatory bowel disease (16%). A similar prevalence of IgM anti-rubella antibodies was detected among controls from different countries. A high prevalence of IgG anti-rubella antibodies was detected among patients with AID (89.9%) and controls. CONCLUSION: The increased prevalence of IgM anti-rubella antibodies in AID suggests a possible role for rubella in the etiopathogenesis of several AID.

Monoclonal auto-antibodies and sera of autoimmune patients react with Plasmodium falciparum and inhibit its in vitro growth

The relationship between autoimmunity and malaria is not well understood. To determine whether autoimmune responses have a protective role during malaria, we studied the pattern of reactivity to plasmodial antigens of sera from 93 patients with 14 different autoimmune diseases (AID) who were not previously exposed to malaria. Sera from patients with 13 different AID reacted against Plasmodium falciparum by indirect fluorescent antibody test with frequencies varying from 33-100 percent. In addition, sera from 37 AID patients were tested for reactivity against Plasmodium yoelii 17XNL and the asexual blood stage forms of three different P. falciparum strains. In general, the frequency of reactive sera was higher against young trophozoites than schizonts (p < 0.05 for 2 strains), indicating that the antigenic determinants targeted by the tested AID sera might be more highly expressed by the former stage. The ability of monoclonal auto-antibodies (auto-Ab) to inhibit P. falciparum growth in vitro was also tested. Thirteen of the 18 monoclonal auto-Ab tested (72 percent), but none of the control monoclonal antibodies, inhibited parasite growth, in some cases by greater than 40 percent. We conclude that autoimmune responses mediated by auto-Ab may present anti-plasmodial activity.

Steroid Responsive Pancreatic Mass-Forming Type 2 Autoimmune Pancreatitis / 대한소화기학회지

Autoimmune pancreatitis (AIP) has two distinct subsets. Type 1 AIP or lymphoplasmacytic sclerosing pancreatitis is systemic disease with the elevation in serum levels of the IgG4. Type 2 AIP, also called duct-centric pancreatitis, features granulocyte epithelial lesions with duct obstruction in the pancreas without systemic involvement. Here, we report a case of type 2 AIP diagnosed by pathology, which is the first report in Korea. The case is a 56-year-old woman who presented with anorexia and vomiting. Computed tomography revealed mass-like lesion in the pancreatic head and the compression of the distal common bile duct and the head portion of the main pancreatic duct. Serum levels of the IgG4 were normal. Histologic examination revealed a dense neutrophil infiltration in the pancreatic parenchyme associated with extensive fibrosis, thereby confirming the diagnosis of type 2 AIP. The abnormalities in the clinical, laboratory, and radiological findings improved after oral steroid treatment.

Implicações clínicas da persistência de anti-GAD positivo e peptídeo C detectável em pacientes com diabetes melito tipo 1 de longa duração / GADA persistence and detectable C peptide in patients with long standing diabetes mellitus type 1

OBJETIVO: Avaliar se anti-GAD positivo e PC detectável se correlacionam com a presença de outras doenças autoimunes, com controle glicêmico e com risco de retinopatia no diabetes melito tipo 1 (DMT1) > 3 anos de duração. PACIENTES E MÉTODOS: Cinquenta sujeitos com DMT1 foram entrevistados, realizaram fundoscopia e dosaram PC pré e pós-glucagon, HbA1C e anti-GAD. RESULTADOS: Pacientes anti-GAD+ (n = 17) apresentaram maior frequência de doenças autoimunes em relação aos demais (p = 0,02). PC detectável (n = 11) também foi associado ao aumento dessa prevalência (p = 0,03), porém nenhum dos dois parâmetros influenciou na presença de retinopatia diabética. PC detectável não influenciou no controle glicêmico (HbA1C média) (p = 0,28), porém as doses diárias de insulina foram mais baixas (0,62 vs. 0,91 U/kg/dia; p = 0,004) neste grupo. CONCLUSÃO: Apesar de não ser um marcador para outras doenças autoimunes, o anti-GAD+ parece ser não só um sinalizador de autoimunidade pancreática. PC detectável também parece ter papel promissor na detecção dessas comorbidades. Ambos não interferiram na presença de retinopatia, entretanto, o PC detectável se relacionou a menores necessidades de insulina.

OBJECTIVE: The aim of this study was to evaluate if GADA+ and detectable CP had any influence in other autoimmune diseases, glycemic control, and risks of retinopathy in diabetes mellitus type 1 (T1DM) lasting longer than 3 years of duration. SUBJECTS AND METHODS: Fifty T1DM subjects were interviewed, performed fundoscopic examination, and measured CP before and after glucagon, HbA1C, and GADA. RESULTS: GADA+ (n = 17) had a higher frequency of other autoimmune diseases when compared to GADA (p = 0.02). Detectable CP was also associated with a higher prevalence of these diseases (p = 0.03), although, retinopathy was not influenced by either one. Detectable CP had no influence in the glycemic control (mean HbA1C) (p = 0.28). However, insulin daily doses were lower in this group (0.62 vs. 0.91 U/kg/day; p = 0.004). CONCLUSION: Although not recommend as a marker of other autoimmune diseases, GADA+ seems to be not only a pancreatic autoimmunity signal. Detectable CP may also have some promising influence in detecting these diseases. Neither influenced the presence of retinopathy, but insulin daily requirements were smaller when CP was present.

Markers of autoimmune liver diseases in postmenopausal women with osteoporosis

INTRODUCTION: Osteoporosis is a common complication of chronic liver diseases. However, there is limited information about autoimmune liver diseases as a factor of secondary osteoporosis. Therefore, we aimed to investigate the autoantibodies of autoimmune liver diseases in patients with osteoporosis. METHODS: One hundred fifty female patients with postmenopausal osteoporosis were included. Bone mineral density was measured by dual energy X-ray absorptiometry. We analysized autoantibodies including antinuclear antibodies, liver membrane antibodies, anti-liver/kidney microsomal autoantibodies1, liver-specific protein, antismooth muscle antibodies, and anti-mitochondrial antibodies by indirect immunofluorescence. Serum was assayed for the levels of aminotransferases. RESULTS: The mean age of the patients was 63,13±8,6 years. The mean values of L1-L4 T-scores and femur total T-scores were -3,08±0,58 and -1,53±0,81, respectively. Among the 150 patients with osteoporosis, 14 (9.3 percent) were antinuclear antibodies, four (2.7 percent) were liver membrane antibodies, three (2.0 percent) were anti-liver/kidney microsomal autoantibodies1, and two (1.3 percent) were liver-specific protein positive. None of the patients had anti-mitochondrial antibodies or smooth muscle antibodies positivity. The mean values of levels of aminotransferases were within normal range. CONCLUSIONS: The presence of liver membrane antibodies, liver-specific protein, and anti-liver/kidney microsomal autoantibodies1 has permitted us to see that there may be some suspicious clues of autoimmune liver diseases in patients with osteoporosis as a secondary risk factor. On the other hand, there is a need for comprehensive studies with a larger sample size and studies designed to compare the results with a normal population to understand the clinical importance of our findings.

Helicobacter pylori infection: prevalence in chronic urticaria patients and incidence of autoimmune urticaria (study in Dr. Cipto Mangunkusumo Hospital, Jakarta).

AIM: To determine the prevalence of Hp infection in patients with chronic urticaria (CU) and to evaluate the result of autologous serum skin test (ASST) in CU patients with Hp infections. METHODS: In this cross-sectional study, 16 patients with chronic urticaria and 16 non-urticaria volunteers were investigated (matched for age and sex). All subjects were examined for Hp infection with the 13C-urea breath test. Autologous serum skin test was performed in patients with proven Hp infection. RESULTS: Helicobacter pylori was detected in 12.5% of patients and 0% of the control group. There was no significant difference between the two groups (p = 0.484 using Fisher exact test). Autologous serum skin test was positive in 1 of 2 CU patients with Hp infection. CONCLUSION: In this study, there was no significant difference in the seroprevalence of Hp infection between CU patients and controls. Autologous serum skin test was positive in 1 of 2 CU patients with Hp infection.

Autologous serum skin test in chronic idiopathic urticaria: prevalence, correlation and clinical implications.

Some cases of chronic idiopathic urticaria (CIU) have histamine-releasing IgG autoantibodies in their blood. This disease subgroup is called "autoimmune urticaria". To date, the autologous serum skin test (ASST) is the best in vivo clinical test for the detection of basophil histamine-releasing activity in vitro. This study aimed to find the prevalence of ASST positive cases in Thai patients with CIU, to identify factors related to the positivity of ASST and to find the clinical implications of ASST in CIU. A retrospective study was performed among 85 CIU patients who attended the Urticaria Clinic at the Department of Dermatology, Siriraj Hospital and were willing to perform ASST, from January 2002 to December 2003. Twenty-one (24.7%) patients had a positive ASST. There was no significant difference between patients with positive ASST and negative ASST as to the severity of the disease (wheal numbers, wheal size, itching scores and the extent of body involvement) as well as the duration of the disease.