The deleterious role of neutrophil extracellular traps (NETs) in pneumococcal pneumonia and therapeutic treatment with adrenomedullin

Abstract

Introduction: Community-acquired pneumonia (CAP) promotes dysregulation of the innate immune system and disruption of the endo-epithelial barrier function, leading to acute lung injury and edema formation. It represents a significant health burden worldwide with high mortality rates despite adequate antibiotic treatment. Streptococcus pneumoniae (S.pn.) is the predominant pathogen causing CAP. Upon S.pn. infection, neutrophils infiltrate and accumulate in the lungs, aiming at bacterial clearance. Neutrophil extracellular traps (NETs) are produced by the infiltrating neutrophils and may contribute to an enhanced inflammation of the tissue. This study investigates if NETs are also released after infection with S.pn. and if these structures contribute to the pneumonia-related tissue damage. It was previously demonstrated that the hormone-peptide adrenomedullin (ADM) protects mice against S.pn.-induced lung epithelial barrier disruption, pulmonary edema formation and extra-pulmonary organ damage. Here, it is further proposed that ADM protects against barrier failure by affecting the process of NET formation. Methods: A murine model for severe pneumococcal pneumonia was assessed to clarify the role of NETs in the disease. For this purpose, we evaluated different strategies aiming at NET degradation or inhibition of the process of NETosis, with DNase or PAD4 inhibition, respectively. Moreover, we targeted histones, the main cytotoxic component of NETs, with polysialic acid. The underlying mechanistic of ADM interference in NET formation were evaluated in vitro. Results: Neutrophils accumulate in the lungs and release NETs after infection with S.pn. NET degradation with DNase attenuated S.pn.-induced lung permeability. Targeting NETs and NETs-related components could reduce the inflammation in the lungs and was beneficial in improving the animal clinical outcome. ADM treatment led to a significant reduction in NET release, suggesting a direct effect of ADM on this process. In vitro, ADM suppressed NET production from activated neutrophils through a mechanism that involves receptor ligation, cAMP production, inhibition of ERK phosphorylation and ROS reduction. Conclusions: The results indicate that NETs have a deleterious role in the pathology of pneumococcal pneumonia and its inhibition may be beneficial for the disease outcome. ADM was shown to have beneficial properties apart from the stabilization of the endo-epithelial barrier, also by inhibiting NET production.