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Title: Análise do surto respiratório e da atividade fagocítica de neutrófilos humanos em sangue total: aplicação ao estudo de pacientes com fibrose cística
Keywords: Neutrófilos;  Eutrófilos;  Citometria;  Fagocitose;  Surto respiratório;  Fibrose cística;  MEDICINA;  PATOLOGIA INVESTIGATIVA;  Citometria de fluxo
Issue Date: 22-Dec-2010
Abstract: Cystic Fibrosis (CF) is an autosomal recessive disease characterized by the dysfunction of a protein encoded by the gene CFTR (Cystic Fibrosis Transmembrane Conductance Regulator), responsible for regulating the flow of various ions, especially CI-. CF patients present changes in composition and viscosity of exocrine secretions and in the fluid interface lining the airways. These changes interfere with proper elimination of these fluids, thereby increasing the risk of glandular duct obstruction, and favouring subsequent colonization of the lungs with pathogens such Pseudomonas aeruginosa, Staphylococcus aureus and the bacteria of the Burkholderia complex. In these patients, infection and inflammation are interdependent and concomitant factors, however distinct, that contribute to the longterm deterioration of pulmonary function. From an immunological standpoint, the mechanisms of immunity are unable to eradicate these infections. Considering that the mutations in the CFTR gene may directly or indirectly affect the function of the neutrophils by a selective defect of their microbicidal activity and that this defect affects the interaction of neutrophils with pathogens, this study aimed to evaluate, through flow cytometric analyses, the functional activity of human neutrophils, and to further define quantitative differences in the phagocytic activity and in the activation of the respiratory burst between CF patients and healthy individuals. This study included 43 samples from FC patients who were treated in reference center at public institution of the Rio de Janeiro State, as well as 89 samples from healthy individuals. The experiments were conducted using peripheral blood samples collected with anticoagulant. The evaluation of the respiratory burst was made from the oxidation of dihydrorhodamine 123 in rhodamine 123, using phorbol-12-myristate-13-acetate as a cell activator. For the evaluation of phagocytosis, genetically altered bacteria overexpressing green fluorescent protein were used. The most common CFTR mutation found in this study was ΔF508, both in homozygosis (12%) and in association with other CFTR mutations (58%). In the FC patients, both the respiratory burst and phagocytic activity were affected, showing a decreasing trend with increasing age. These results support the view that neutrophils from CF patients have a diminished functional capacity as they grow older, even when neutrophils were not collected from inflammatory sites. These observations suggest that an acquired defect in innate immunity, expressed in circulating neutrophils, develops after several years of disease activity, independently of exposure to the microenvironment of inflammatory
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