RESUMEN
Hirschsprung disease (HSCR, OMIM 142623) involves congenital intestinal obstruction caused by dysfunction of neural crest cells and their progeny during enteric nervous system (ENS) development. HSCR is a multifactorial disorder; pathogenetic variants accounting for disease phenotype are identified only in a minority of cases, and the identification of novel disease-relevant genes remains challenging. In order to identify and to validate a potential disease-causing relevance of novel HSCR candidate genes, we established a complementary study approach, combining whole exome sequencing (WES) with transcriptome analysis of murine embryonic ENS-related tissues, literature and database searches, in silico network analyses, and functional readouts using candidate gene-specific genome-edited cell clones. WES datasets of two patients with HSCR and their non-affected parents were analysed, and four novel HSCR candidate genes could be identified: ATP7A, SREBF1, ABCD1 and PIAS2. Further rare variants in these genes were identified in additional HSCR patients, suggesting disease relevance. Transcriptomics revealed that these genes are expressed in embryonic and fetal gastrointestinal tissues. Knockout of these genes in neuronal cells demonstrated impaired cell differentiation, proliferation and/or survival. Our approach identified and validated candidate HSCR genes and provided further insight into the underlying pathomechanisms of HSCR.
Asunto(s)
Enfermedad de Hirschsprung/genética , Miembro 1 de la Subfamilia D de Transportador de Casetes de Unión al ATP/genética , Animales , Diferenciación Celular/genética , Línea Celular , Proliferación Celular/genética , Supervivencia Celular/genética , Simulación por Computador , ATPasas Transportadoras de Cobre/genética , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica , Técnicas de Inactivación de Genes , Humanos , Lactante , Masculino , Ratones , Proteínas Inhibidoras de STAT Activados/genética , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/genética , Secuenciación del ExomaRESUMEN
INTRODUCTION: A targeted Hirschsprung disease (HD) diagnostic is necessary, as it determines a specific approach primarily based on surgical resection of the affected aganglionic colonic segment. The aim of this study was to evaluate the diagnostic accuracy of a contrast enema (CE) for HD diagnosis and to determine whether it should be performed before or after rectal biopsies (RBs). METHODS: A retrospective observational study of children undergoing RB for HD investigation was performed. In the performed CE, the occurrence and the level of a colonic caliber change (CCC) were recorded and its concordance with the histologically assessed level of aganglionosis by RB and the odds ratio were calculated. RESULTS: A total of 107 cases were included. Sensitivity and specificity for a CCC in CE were 74.1% and 94.6%. A CCC present in CE was associated with a 50-fold increased probability for a histologically proven HD. The overall concordance between a CCC and the histologically assessed level of aganglionosis was high (kappa 0.642, p = 0.003), being correct in 94.4% of cases when the CCC was located in the rectosigmoid, but only in 50% of cases when it was located in more proximal segments. By performing a CE only after HD diagnosis confirmation by RB would avoid 67.5% of CE with no loss of diagnostic accuracy. CONCLUSION: We confirm that CE is a valuable tool for HD diagnosis; however, it should only be performed for subsequent diagnostic and surgical planning following histological confirmation of HD by RB. On the basis of this, an algorithm for an optimized investigation and management of HD is presented.