The Role of Neutrophils in Alcohol-Induced Liver Damage in Alcoholic Hepatitis

In alcoholic hepatitis (AH), high neutrophil counts correlate with inflammation and poor clinical outcomes. Here, we sought to elucidate the neutrophil-mediated pathogenesis of AH. We revealed that in vivo neutrophil extracellular trap (NET) formation was significantly increased in AH patients and that alcohol alone is sufficient to induce NET formation in neutrophils; thereby, neutrophils increase liver damage through increased NET formation. Moreover, we identify that alcohol-induced NET formation is vital to NETosis and that high-density neutrophils (HDNs) become low-density neutrophils (LDNs) after NET formation in response to alcohol. Through transcriptome profile analysis, we found that genes related to neutrophil activation and immune responses are significantly upregulated in AH HDNs but significantly downregulated in AH LDNs compared to HDNs from healthy subjects. These data suggest that AH HDNs and LDNs have opposing phenotypes: HDNs are activated and more prone to release NETs, while LDNs are functionally incompetent. Consequently, the increase in activated HDNs and defective LDNs are likely associated with an increase in liver damage through NET formation and enhanced susceptibility to infection in AH patients, respectively. Therefore, we evaluated the therapeutic benefits of preventing NET formation in HDNs using peptidyl arginine deiminase 4 (PAD4) inhibition and granulocyte colony-stimulating factor (G-CSF) treatment as well as neutrophil depletion in mice. We observed that in vivo neutrophil depletion and G-CSF treatment prevent NET formation in the liver, thereby significantly reducing liver damage in alcohol-fed mice. Our work identifies the neutrophil/NET-mediated mechanisms of AH pathogenesis and provides insights into therapeutic interventions for AH