Production of Neutrophil Extracellular Traps Contributes to the Pathogenesis of Francisella tularemia

Abstract

Francisella tularensis(Ft) is a highly virulent, intracellular Gram-negative bacterial pathogen. Acute Ft infection by aerosol route causes pneumonic tularemia, characterized by nodular hemorrhagic lesions, neutrophil-predominant influx, necrotic debris, fibrin deposition, and severe alveolitis. Ft suppresses activity of neutrophils by impairing their respiratory burst and phagocytic activity. However, the fate of the massive numbers of neutrophils recruited to the infection site is unclear. Here, we show that Ft infection resulted in prominent induction of neutrophil extracellular traps (NETs) within damaged lungs of mice infected with the live attenuated vaccine strain of Ft(Ft-LVS), as well as in the lungs of domestic cats and rabbits naturally infected with Ft. Further, Ft-LVS infection increased lung myeloperoxidase (MPO) activity, which mediates histone protein degradation during NETosis and anchors chromatin scaffolds in NETs. In addition, Ft infection also induced expression of peptidylarginine deiminase 4, an enzyme that causes citrullination of histones during formation of NETs. The released NETs were found largely attached to the alveolar epithelium, and disrupted the thin alveolar epithelial barrier. Furthermore, Ft infection induced a concentrationdependent release of NETs from neutrophils in vitro. Pharmacological blocking of MPO reduced Ft-induced NETs release, whereas addition of H 2 O 2 (a substrate of MPO) significantly augmented NETs release, thus indicating a critical role of MPO in Ft-induced NETs. Although immunofluorescence and electron microscopy revealed that NETs could efficiently trap Ft bacteria, NETs failed to exert bactericidal effects. Taken together, these findings suggest that NETs exacerbate tissue damage in pulmonary Ft infection, and that targeting NETosis may offer novel therapeutic interventions in alleviating Ft-induced tissue damage.

Cytokine Signature Induced by SARS-CoV-2 Spike Protein in a Mouse Model

“…We hypothesized that the SARS-CoV-2 mimic would stimulate macrophages to produce inflammatory cytokines including IL-1α, IL-6, and TNF-α, and recruit and activate neutrophils. P38 and extracellular signal-regulated kinase (ERK) are the signaling mediators of NETs formation, while myeloperoxidase (MPO) and elastase, as the peroxidase enzymes of neutrophils, are essential for NETs formation ( 22 , 23 ). The TLR3/dsRNA complex inhibitor can block the interactions between TLR3 and poly(I:C), and the JAK inhibitor blocks the downstream signaling of IL-6.…”

“…Elastase is produced by neutrophils and can mediate lung injury. Inhibitors of MPO and elastase can directly reduce acute lung injury at inflammatory sites ( 23 , 35 , 36 ). The MAPK-p38 signaling pathway plays an essential role in the production of pro-inflammatory cytokines, inhibition of p38 shows promising in COVID-19 therapy ( 37 ).…”

Molecular mechanism of SARS-CoV-2 components caused ARDS in murine model

“…Elastase was produced by neutrophils and can mediate lung injury. Inhibitors of MPO and elastase can directly reduce the acute lung injury at the inflammatory site 21,38,39 . In our study, inhibitors targeting neutrophils include P38 inhibitor, ERK inhibitor, MPO inhibitor and elastase inhibitor can reduce the neutrophil infiltration into lung, but not elastase, but single inhibitor may offer no protection from SARS-Cov-2 viral mimic induced pathogenesis of ARDS, and as such multiple inhibitors may need to be simultaneously utilized.…”

Intestinal infection results in impaired lung innate immunity to secondary respiratory infection

“…Recent studies have revealed that excessive NET formation plays a role in pathogen-associated lung injury, including in models of bacterial pneumonia [16–19]. Our histopathological analysis revealed clusters of pyknotic neutrophils and thrombus formation in mice with prior intestinal infection (Fig 2 and Fig S3).…”