Influences of chloride and hypochlorite on neutrophil extracellular trap formation.
BACKGROUND
The release by neutrophils of DNA-based extracellular traps (NETs) is a recently recognized innate immune phenomenon that contributes significantly to control of bacterial pathogens at tissue foci of infection. NETs have also been implicated in the pathogenesis of non-infectious diseases such as small vessel vasculitis, lupus and cystic fibrosis lung disease. Reactive oxygen species (ROS) are important mediators of NET generation (NETosis). Neutrophils with reduced ROS production, such as those from patients with chronic granulomatous disease or myeloperoxidase (MPO) deficiency, produce fewer NETs in response to inflammatory stimuli. To better understand the roles of various ROS in NETosis, we explore the role of MPO, its substrates chloride ion (Cl(-)) and hydrogen peroxide (H(2)O(2)), and its product hypochlorite (HOCl) in NETosis.


RESULTS
In human peripheral blood neutrophils, pharmacologic inhibition of MPO decreased NETosis. Absence of extracellular Cl(-), a substrate for MPO, also reduced NETosis. While exogenous addition of H(2)O(2) and HOCl stimulated NETosis, only exogenous HOCl could rescue NETosis in the setting of MPO inhibition. Neither pharmacological inhibition nor genetic deletion of MPO in murine neutrophils blocked NETosis, in contrast to findings in human neutrophils.


CONCLUSIONS
Our results pinpoint HOCl as the key ROS involved in human NETosis. This finding has implications for understanding innate immune function in diseases in which Cl(-) homeostasis is disturbed, such as cystic fibrosis. Our results also reveal an example of significant species-specific differences in NET phenotypes, and the need for caution in extrapolation to humans from studies of murine NETosis.
Department of Pediatrics, University of California San Diego, La Jolla, California, United States of America.
Kathryn
K
Akong-Moore
Ohn A
OA
Chow
Maren
M
von Köckritz-Blickwede
Victor
V
Nizet
D000704:Analysis of Variance; D002712:Chlorides / Q000378:metabolism*; D005110:Extracellular Space / Q000276:immunology* / Q000378:metabolism; D016179:Granulocyte Colony-Stimulating Factor / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D006997:Hypochlorous Acid / Q000378:metabolism*; D007113:Immunity, Innate / Q000276:immunology*; D007377:Interleukin-3 / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009504:Neutrophils / Q000187:drug effects* / Q000276:immunology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013045:Species Specificity
D002712:Chlorides; D007377:Interleukin-3; D017382:Reactive Oxygen Species; D011993:Recombinant Fusion Proteins; C404145:myelopoietin; D016179:Granulocyte Colony-Stimulating Factor; D006997:Hypochlorous Acid; D006861:Hydrogen Peroxide