Avaliação do DNA extracelular no escarro de pacientes com fibrose cística e associação com morbidade e gravidade da doença

Abstract

Introduction: High levels of extracellular DNA are present in the sputum of cystic fibrosis (CF) patients, originated from necrotic/apoptotic neutrophils and neutrophil extracellular traps (NETs), a defense mechanism recently described. DNA alters mucus viscosity and contributes to airway obstruction, which predisposes to infections and colonization by bacteria, recurrent exacerbations and lung function decline. However, there are few studies investigating the association between extracellular DNA and NETs release with the disease severity in CF. Objectives: To evaluate the association of the levels of extracellular DNA with lung function, use of antibiotics and hospitalization for pulmonary exacerbation. Methods: This is a cross-sectional study. CF patients older than 5 years old, clinically stable and able to spontaneously expectorate were included. Sputum was collected during the outpatient visit and subsequently processed for quantitation of extracellular DNA. The extracellular DNA traps in sputum were visualized by immunofluorescence microscopy and cellular death profile was determined. Lung function data, airway colonization, number of antibiotic days and hospital stay in the previous 12 months were collected. For the analysis, patients were allocated into two groups according to the median levels of DNA in sputum. Patients or parents signed a consent form and the study was approved by the research ethics committee of the University. Results: We have included 33 patients in the study, mean age 16.3±6.2 years, forced expiratory volume in the first second (FEV1) of 67.0±26.7% and DNA level of 241.9±147.2μg/mL. A significant correlation was observed between DNA and FEV1 (r=-0.60; p<0.001) and forced vital capacity (r=-0.59; p<0.001). However, there was no significant association with age, body mass index, days of antibiotics and hospital stay. There was a significant decrease in FEV1 (52.1±27.8% vs. 81.1±16.2%; p=0.001) and a greater need for hospitalization (68.8% vs. 35.3%; p=0.05) in patients with a higher level of DNA (> 243.0μg/mL). No significant differences were found in the number of exacerbations, days of antibiotics and chronic colonization by Pseudomonas aeruginosa. It was also possible to observe the presence of extracellular DNA traps and a low percentage of necrosis and apoptosis in the sputum. Conclusions: The results of the present study demonstrated that the higher levels of extracellular DNA in the sputum of CF patients is associated with lower lung function and hospitalization, which may be related to the extracellular traps released by neutrophils.