Neutrophil extracellular trap induction by alcohol generates unique low-density neutrophils with defective functions and impaired clearance contributing to liver damage in alcoholic hepatitis

Alcoholic hepatitis (AH) is characterized by severe liver and systemic inflammation and high mortality. Although numerous studies correlated neutrophil counts with poor clinical outcomes, the role of neutrophils in AH is poorly understood. Here, we report that neutrophils contribute to liver damage through increased neutrophils extracellular traps (NET) production in AH patients and mouse models with a concordant significant increase of low-density neutrophils (LDNs) in AH patients. Transcriptome analysis revealed that high-density neutrophils (HDNs) are activated and more prone to release NET, whereas LDNs exhibit exhausted phenotype. We show that alcohol-induced NET release in HDNs induces a unique LDN subset with decreased functionality and reduced clearance. Elimination of both defective HDNs and LDNs through in vivo neutrophil depletion or prevention of NET production with granulocyte colony stimulating factor (G-CSF) treatment ameliorate alcohol-induced liver damage. Our findings uncover alcohol-induced neutrophil phenotypic changes and provide mechanistic insights for therapeutic interventions in AH. Whole transcriptome sequencing of high-density neutrophils (HDNs) and low-density neutrophils (LDNs) isolated from collected blood of acute alcoholic hepatitis patients (n=4) and HDNs from healthy human subjects (n=3).

酒精性肝炎（AH）以严重的肝脏及全身炎症和高死亡率为主要特征。尽管诸多研究显示中性粒细胞计数与不良临床结局存在相关性，但中性粒细胞在AH中的作用机制仍有待明确。本研究发现，在AH患者及匹配的小鼠模型中，中性粒细胞可通过增多中性粒细胞胞外陷阱（Neutrophil Extracellular Traps, NET）的生成参与肝脏损伤，且AH患者体内低密度中性粒细胞（Low-density Neutrophils, LDNs）水平显著升高，该变化在小鼠模型中亦呈现一致性。转录组分析结果显示，高密度中性粒细胞（High-density Neutrophils, HDNs）处于活化状态且更易释放NET，而LDNs则表现出耗竭表型。本研究证实，酒精诱导的HDNs释放NET会催生一类功能受损、清除能力降低的独特LDN亚群。通过体内中性粒细胞耗竭或采用粒细胞集落刺激因子（Granulocyte Colony-stimulating Factor, G-CSF）抑制NET生成，可有效减轻酒精诱导的肝脏损伤。本研究揭示了酒精诱导的中性粒细胞表型改变，并为AH的治疗干预提供了关键的机制见解。本研究对4例急性酒精性肝炎患者外周血中分离得到的HDNs和LDNs，以及3例健康受试者外周血分离得到的HDNs进行了全转录组测序。