RESUMO
Introducción: La Enfermedad Granulomatosa Crónica (EGC) se presenta como consecuencia de mutaciones en los genes que codifican 5 de las subunidades del sistema NADPH oxidasa humano. Su forma más común es causada por cambios en el gen CYBB que codifica gp91 phox. Objetivo: Identificar el defecto molecular que lleva a la presentación de la EGC. Caso clínico: Paciente de sexo masculino con antecedentes de enfermedad diarreica aguda y abscesos perianales recurrentes desde los 2 meses. A los 6 meses, presentó una inflamación crónica del colon y colitis bacteriana. A los 3 años tenía infecciones en las vías respiratorias inferiores y perianales. Estudio compatible con EGC. El análisis del ADNc identificó expresión anormal del ARNm, lo cual se confirmó al realizar la secuenciación. Específicamente se observó la ausencia del exón 2. Adicionalmente, los datos de la secuenciación del ADNg identificaron una alteración en el sitio aceptor de "splicing" del intrón 1, que incluye una deleción seguida de la inserción de 3 nucleótidos (c.46-14_-11delTTCT insGAA). Conclusiones: Se presenta el primer estudio molecular de un paciente con EGC por defecto de "splicing" reportado en Colombia. La definición de la mutación y su correlación con el fenotipo es importante para proveer una apropiada consejería genética al paciente y su familia.
Chronic granulomatous disease (CGD) is caused by mutations in the genes that encode five of the subunits of the human NADPH oxidase. The most common form is caused by mutations in CYBB, the human gene encoding gp 91 phox. Objective: To identify the molecular defects causing CGD. Case report: A male patient with a history of acute diarrhea and recurrent perianal abscess since two months old. At 6 months, the patient presented a chronic inflammatory disease of the colon and bacterial colitis. After three years, he developed infections in the lower and perianal respiratory tract. The cDNA analysis identified abnormal mRNA expression, which was confirmed by sequencing. Specifically the exclusion of exon 2 was observed. Additionally, gDNA sequencing identified an alteration in the acceptor splice site of intron 1, including a deletion followed by insertion of three nucleotides (c.46-14_-11delTTCT insGAA). Conclusions: The first molecular study of a patient with CGD due to splicing pattern change, reported in Colombia, is presented. The definition of the mutation and its correlation with the phenotype is essential to provide appropriate genetic counseling to patients and their families.
Assuntos
Humanos , Masculino , Lactente , Cromossomos Humanos X , Doença Granulomatosa Crônica/genética , Mutação , NADPH Oxidases/genética , DNA Complementar/genética , RNA Mensageiro/genética , Sequência de Bases , Separação Celular , Éxons , Doença Granulomatosa Crônica/diagnóstico , Citometria de Fluxo , Fenótipo , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , Splicing de RNARESUMO
Background: The cytosolic protein p47-phox (phagocyte oxidase) is one of the essential components of the superoxide generating system in phagocytes and its defect causes approximately 30 percent of the chronic granulomatous disease (CGD) cases. Aim: Two patients were studied, belonging to the same family, without a consanguinous background, in which deficiency or absence of superoxide generation was found together with recurrent and severe infections in one case and benign infections in the second. Methods: The presence of gp91-, p67- and p47-phox in patients and controls was determined by Western Blot analysis of granulocytes. Sequencing of PCR amplified DNA was performed by an enzimatic method. Results: Western Blot analysis showed normal expression of gp91 and p67 and absence of p47-phox. The molecular genetic study demonstrated a homocygotic dinucleotide GT (GT) deletion at the beginning of exon 2 of the p47-phox gene. The same mutation has been found in European, American and Japanese patients. Conclusions: The molecular characterization of this pathology done for the first time in Chile is important for diagnostic classification, patient prognosis, and adequate genetic advice and a possible future therapy