Intrinsic alterations in peripheral neutrophils from cystic fibrosis newborn piglets

Abstract

BackgroundThe hallmark of the cystic fibrosis (CF) lung disease is a neutrophil dominated lung environment that is associated to chronic lung tissue destruction and ultimately the patient's death. It is unclear whether the exacerbated neutrophil response is primary related to a defective CFTR or rather secondary to chronic bacterial colonization and inflammation. Here, we hypothesized that CF peripheral blood neutrophils present intrinsic alteration at birth before the start of an inflammatory process. MethodsPeripheral blood neutrophils were isolated from newborn CFTR+/+ and CFTR−/− piglets. Neutrophils immunophenotype was evaluated by flow cytometry. Lipidomic and proteomic profile were characterized by liquid chromatography/tandem mass spectrometry (LC-MS/MS), intact cell matrix-assisted laser desorption/ionization mass spectrometry (ICM-MS) followed by top-down high-resolution mass spectrometry (HRMS), respectively. The ability of CF neutrophils to kill pseudomonas aeruginosa was also evaluated. ResultsPolyunsaturated fatty acid metabolites analysis did not show any difference between CFTR+/+ and CFTR−/− neutrophils. On the other hand, a predictive mathematical model based on the ICM-MS proteomic profile was able to discriminate between both genotypes. Top-down proteomic analysis identified 19 m/z differentially abundant masses that corresponded mainly to proteins related to the antimicrobial response and the generation of reactive oxygen species (ROS). However, no alteration in the ability of CFTR−/− neutrophils to kill pseudomonas aeruginosa in vitro was observed. ConclusionsICM-MS demonstrated that CFTR−/− neutrophils present intrinsic alterations already at birth, before the presence of any infection or inflammation.