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

• Published in: Journal of hepatology 2018-11, Vol.69 (5), p.1145-1154
• Main Authors: Bukong, Terence Ndonyi, Cho, Yeonhee, Iracheta-Vellve, Arvin, Saha, Banishree, Lowe, Patrick, Adejumo, Adeyinka, Furi, Istvan, Ambade, Aditya, Gyongyosi, Benedek, Catalano, Donna, Kodys, Karen, Szabo, Gyongyi
• Format: Article
• Language: English
• Subjects: Acetic acid; Alcohol use; Alcoholic hepatitis; Animals; Binge drinking; Binge Drinking - complications; Body weight; Cell death; Citrulline; Deoxyribonucleic acid; DNA; Drinking behavior; Elastase; Extracellular Traps - physiology; Hepatitis, Alcoholic - etiology; Hepatocytes; Histone H3; HMGB1 Protein - physiology; Human health and pathology; Humans; Hépatology and Gastroenterology; Immune clearance; Injuries; Leukocytes (neutrophilic); Life Sciences; Lipopolysaccharides; Liver diseases; Macrophages; Macrophages - physiology; Mice; Mice, Inbred C57BL; Neutrophil depletion; Neutrophil elastase; Peroxidase; Phagocytosis; Phorbol 12-myristate 13-acetate; Sepsis; Sepsis - etiology
• ISSN: 0168-8278; 1600-0641
• DOI: 10.1016/j.jhep.2018.07.005

[Display omitted] •Alcohol treatment induces spontaneous neutrophil extracellular trap (NET) formation.•Alcohol impairs further NET release by the same neutrophils upon subsequent activation.•Alcohol reduces efferocytosis by macrophages, leading to less clearance of NETotic neutrophils.•Depletion of neutrophils in vivo alleviates hepatic injury after binge-drinking. Neutrophil extracellular traps (NETs) are an important strategy utilized by neutrophils to immobilize and kill invading microorganisms. Herein, we studied NET formation and the process of neutrophil cell death (NETosis), as well as the clearance of NETs by macrophages (MΦ) (efferocytosis) in acute sepsis following binge drinking. Healthy volunteers consumed 2 ml of vodka/kg body weight, before blood endotoxin and 16 s rDNA were 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 intraperitoneal lipopolysaccharide (LPS) to assess the dynamics of NET formation and efferocytosis. In vivo, anti-Ly6G antibody (IA8) was used for neutrophil depletion. 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 upon PMA stimulation alcohol attenuated NET formation. Binge alcohol in mice 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 receiving binge alcohol, 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 NETotic neutrophils and promoted expression of CD206 on MΦ. Finally, depletion of neutrophils prior to binge alcohol ameliorated LPS-induced systemic inflammation and liver injury in mice. Dysfunctional NETosis and efferocytosis following binge drinking exacerbate liver injury associated with sepsis. Disease severity in alcoholic liver disease (ALD) is associated with a significant presence of neutr