Calcium dependent mechanisms of neutrophil dysfunction that contribute to cystic fibrosis pathobiology

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 DESCRIPTION (provided by applicant): Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (cftr) gene and characterized by abnormal epithelial ion transport, viscous mucus, chronic bacterial infection, and exaggerated airway inflammation. Opportunistic bacteria that cause lung infection, including Burkholderia cenocepacia (B. cenocepacia), are lethal threat to CF patients. B. cenocepacia is resistant to antibiotics, and host phagocytes fail to clear the infection causing severe inflammation. Neutrophils play essential roles in controlling lung infection and inflammation, however in CF patients; neutrophils cannot eradicate bacteria and promote inflammation. CFTR is a chloride ion channel known to regulate epithelial fluid transport in the lung. The function of CFTR in phagocytes, and how CFTR deficiency affects the inflammatory activities of phagocytic cells is unknown. CFTR mutation has demonstrated to alter directly calcium (Ca2+) homeostasis in airway epithelia, but the impact of this mutation on neutrophil Ca2+-dependent functions remains to be determined. In CF neutrophils, we have found that Ca2+ signaling along with the Ca2+ channel TRPM2 are increased, whereas the production of reactive oxygen species (ROS), and NETosis are deficient. Hence, we hypothesize that CFTR mutation results in increased cytosolic release of Ca2+. Altered Ca2+ homeostasis in turn, positively regulates Ca2+-dependent inflammatory neutrophil functions, but down regulates NADPH activity contributing to neutrophil defective bacterial killing. To test this hypothesis we will: 1). Determine how CFTR mutation impacts calcium signaling pathways in neutrophils 2). Define how the axis TRPM2/NADPH oxidase affects antimicrobial functions of human and mouse CF neutrophils. The successful completion of this proposal will significantly increase our understanding of how neutrophils contribute to CF pathobiology, and will help define novel therapeutic strategies by targeting Ca+2 signaling pathways that modulate CFTR and NADPH oxidase to control infection and inflammation in CF patients.