Abnormal neutrophil traps and impaired efferocytosis contribute to liver injury and sepsis severity after binge alcohol use

International audienceBACKGROUND & AIMS: Neutrophil extracellular traps (NETs) are an important strategy utilized by neutrophils to immobilize and kill invading microorganisms. Here we studied NET formation and its clearance by macrophages (MØ) (efferocytosis) in acute sepsis following binge drinking.METHODS: Healthy volunteers consumed 2 mL of vodka/kg body weight and blood endotoxin and 16s rDNA was measured. Peripheral neutrophils were isolated and exposed to alcohol followed by phorbol 12-myristate 13-acetate (PMA) stimulation. Mice were treated with three alcohol binges and i.p. LPS to assess the dynamics of NET formation and efferocytosis. In vivo, anti-Ly6G antibody (IA8) was used for neutrophil depletion.RESULTS: Inducers of NETs (endotoxin and bacterial DNA) significantly increased in the circulation after binge alcohol drinking in humans. Ex vivo, alcohol alone increased NET formation but attenuated NET formation upon PMA stimulation. Binge alcohol in mice in vivo resulted in a biphasic response to LPS. Initially, binge alcohol reduced LPS-induced NET formation and resulted in a diffuse distribution of neutrophils in the liver compared to alcohol-naïve mice. Moreover, indicators of NET formation including citrullinated histone H3, neutrophil elastase, and neutrophil myeloperoxidase were decreased at an early time point after LPS challenge in mice with alcohol binge suggesting decreased NET formation. However, in the efferocytosis phase (15 h after LPS) citrullinated histone-H3 was increased in the liver in alcohol binge mice, suggesting decreased clearance of NETs. In vitro alcohol treatment reduced efferocytosis and phagocytosis of NETosing neutrophils and promoted expression of CD206 on MØ. Finally, depletion of neutrophils prior to binge alcohol ameliorated LPS-induced systemic inflammation and liver injury in mice.CONCLUSIONS: Dysfunctional neutrophil NETosis and efferocytosis after binge drinking exacerbates liver injury associated with sepsis.LAY ABSTRACT: Disease severity in alcoholic liver disease (ALD) is associated with significant liver neutrophil presence. It remains unknown how alcohol affects the capacity of neutrophils to control infection, a major hallmark of ALD. We found that binge alcohol drinking impaired important strategies used by neutrophils to contain and resolve infection resulting in increased liver injury during ALD