RESUMO
BACKGROUND AND OBJECTIVE: Familial Mediterranean fever (FMF) is the most frequent hereditary autoinflammatory disease. Its diagnosis relies on a set of clinical criteria and a genetic confirmation on identification of biallelic pathogenic MEFV variants. MEFV encodes pyrin, an inflammasome sensor. Using a kinase inhibitor, UCN-01, we recently identified that dephosphorylation of FMF-associated pyrin mutants leads to inflammasome activation. The aim of this study was to assess whether quantifying UCN-01-mediated inflammasome activation could discriminate FMF patients from healthy donors (HD) and from patients with other inflammatory disorders (OID). METHODS: Real-time pyroptosis and IL-1ß secretion were monitored in response to UCN-01 in monocytes from FMF patients (n=67), HD (n=71) and OID patients (n=40). Sensitivity and specificity of the resulting diagnostic tests were determined by receiver operating characteristic curve analyses. RESULTS: Inflammasome monitoring in response to UCN-01 discriminates FMF patients from other individuals. Pyroptosis assessment leads to a fast FMF diagnosis while combining pyroptosis and IL-1ß dosage renders UCN-01-based assays highly sensitive and specific. UCN-01-triggered monocytes responses were influenced by MEFV gene dosage and MEFV mutations in a similar way as clinical phenotypes are. CONCLUSIONS: UCN-01-based inflammasome assays could be used to rapidly diagnose FMF, with high sensitivity and specificity.
Assuntos
Febre Familiar do Mediterrâneo/diagnóstico , Inflamassomos/efeitos dos fármacos , Interleucina-1beta/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Pirina/efeitos dos fármacos , Piroptose/efeitos dos fármacos , Estaurosporina/análogos & derivados , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Artrite Juvenil/diagnóstico , Síndrome de Behçet/diagnóstico , Estudos de Casos e Controles , Criança , Pré-Escolar , Síndromes Periódicas Associadas à Criopirina/diagnóstico , Febre Familiar do Mediterrâneo/genética , Febre Familiar do Mediterrâneo/imunologia , Feminino , Febre/diagnóstico , Febre de Causa Desconhecida/diagnóstico , Doenças Hereditárias Autoinflamatórias/diagnóstico , Humanos , Testes Imunológicos/métodos , Inflamassomos/imunologia , Inflamassomos/metabolismo , Interleucina-1beta/imunologia , Interleucina-1beta/metabolismo , Lúpus Eritematoso Sistêmico/diagnóstico , Masculino , Deficiência de Mevalonato Quinase/diagnóstico , Pessoa de Meia-Idade , Monócitos/imunologia , Monócitos/metabolismo , Pirina/genética , Pirina/imunologia , Pirina/metabolismo , Sensibilidade e Especificidade , Sepse/diagnóstico , Estaurosporina/farmacologia , Doença de Still de Início Tardio/diagnóstico , Adulto JovemRESUMO
Familial Mediterranean fever (FMF) is the most frequent hereditary systemic autoinflammatory syndrome. FMF is usually caused by biallelic mutations in the MEFV gene, encoding Pyrin. Conclusive genetic evidence lacks for about 30% of patients diagnosed with clinical FMF. Pyrin is an inflammasome sensor maintained inactive by two kinases (PKN1/2). The consequences of MEFV mutations on inflammasome activation are still poorly understood. Here, we demonstrate that PKC superfamily inhibitors trigger inflammasome activation in monocytes from FMF patients while they trigger a delayed apoptosis in monocytes from healthy donors. The expression of the pathogenic p.M694V MEFV allele is necessary and sufficient for PKC inhibitors (or mutations precluding Pyrin phosphorylation) to trigger caspase-1- and gasdermin D-mediated pyroptosis. In line with colchicine efficacy in patients, colchicine fully blocks this response in FMF patients' monocytes. These results indicate that Pyrin inflammasome activation is solely controlled by Pyrin (de)phosphorylation in FMF patients while a second control mechanism restricts its activation in healthy donors/non-FMF patients. This study paves the way toward a functional characterization of MEFV variants and a functional test to diagnose FMF.