A Novel Multi-organ Male Model of Alcohol-induced Acute-on-chronic Liver Failure Reveals NET-mediated Hepatocellular Death, Which is Prevented by RIPK3 Inhibition

IF 7.1 1区医学 Q1 GASTROENTEROLOGY & HEPATOLOGY Cellular and Molecular Gastroenterology and Hepatology Pub Date : 2025-01-01 DOI:10.1016/j.jcmgh.2024.101446

Martí Ortega-Ribera , Yuan Zhuang , Mrigya Babuta , Veronika Brezani , Radhika S. Joshi , Zsuzsanna Zsengeller , Prashanth Thevkar Nagesh , Yanbo Wang , Roderick Bronson , Gyongyi Szabo

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Abstract

Background & Aims

Alcohol abuse is the most frequent precipitating factor of acute-on-chronic liver failure (ACLF). We aimed at developing an alcohol-induced ACLF model and dissecting its underlying molecular mechanisms.

Methods

ACLF was triggered by a single alcohol binge (5 g/kg) in a bile duct ligation (BDL) liver fibrosis murine model. Liver, kidney, and brain tissues and behavior were assessed in mice. Livers from patients with sclerosing cholangitis with and without ACLF were also evaluated.

Results

In advanced fibrosis induced by BDL, an alcohol binge induced features of ACLF, including significant liver damage, systemic inflammation (increased endotoxin and pro-inflammatory cytokines), and hepatocyte dysfunction compared with BDL alone. ACLF was associated with extrahepatic manifestations, including increased blood urea nitrogen and creatinine, impaired coagulation, and features of encephalopathy. We discovered significantly increased neutrophil count and neutrophil extracellular traps (NETs) in the liver, kidney, and brain in murine ACLF. Livers from ACLF mice showed increased pyroptosis (gasdermin D) and necroptosis (receptor-interacting protein kinase 3 [RIPK3]), when compared with BDL. In vitro, cell-free NETs were induced by alcohol and/or bile acids and triggered pyro-/necroptotic death in hepatocytes. NETosis, pyroptosis, and RIPK3 activation were validated in human livers with ACLF. Moreover, pharmacological inhibition of necroptosis with a RIPK3 inhibitor-ameliorated inflammation, NETs, and liver fibrosis, improving multi-organ ACLF pathophysiology.

Conclusions

Our novel ACLF model triggered by alcohol binge mimics key features of pathophysiology and multi-organ impairment in human ACLF. Our results indicate that neutrophil infiltration and NETs contribute to hepatocyte cell death via pyroptosis and necroptosis in ACLF, identifying RIPK3 as a potential therapeutic target.

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一项新的多器官男性酒精诱导的急性慢性肝衰竭模型显示，net介导的肝细胞死亡可通过RIPK3抑制来预防。

背景和目的：酒精滥用是急性慢性肝衰竭（ACLF）最常见的诱发因素。我们旨在建立酒精诱导的ACLF模型并剖析其潜在的分子机制。方法：在胆管结扎（BDL）肝纤维化小鼠模型中，单次酒精暴饮（5g/Kg）触发ACLF。对小鼠的肝脏、肾脏和脑组织及行为进行了评估。同时对合并/不合并ACLF的硬化性胆管炎患者的肝脏进行了评估。结果：在BDL诱导的晚期纤维化中，与单独BDL相比，酒精狂欢诱导的ACLF特征包括显著的肝损伤、全身性炎症（内毒素和促炎细胞因子增加）和肝细胞功能障碍。ACLF与肝外表现相关，包括BUN和肌酐升高、凝血功能受损和脑病特征。我们发现在小鼠ACLF中，肝、肾和脑的中性粒细胞计数和中性粒细胞胞外陷阱（NET）显著增加。与BDL相比，ACLF小鼠的肝脏显示出更多的焦亡（气真皮蛋白D）和坏死（RIPK3）。在体外，酒精和/或胆汁酸诱导无细胞NETs，并引发肝细胞热/坏死性死亡。在ACLF患者肝脏中证实了NETosis、pyroptosis和RIPK3激活。此外，RIP3抑制剂对坏死下垂的药理学抑制可改善炎症、NETs和肝纤维化，改善多器官ACLF病理生理。结论：我们的新ACLF模型由酗酒引发，模拟了人类ACLF病理生理和多器官损伤的关键特征。我们的研究结果表明，中性粒细胞浸润和NETs通过ACLF中的焦亡和坏死亡参与肝细胞死亡，确定RIPK3是一个潜在的治疗靶点。

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来源期刊

Cellular and Molecular Gastroenterology and Hepatology Medicine-Gastroenterology

CiteScore

13.00

自引率

2.80%

发文量

246

审稿时长

42 days

期刊介绍： "Cell and Molecular Gastroenterology and Hepatology (CMGH)" is a journal dedicated to advancing the understanding of digestive biology through impactful research that spans the spectrum of normal gastrointestinal, hepatic, and pancreatic functions, as well as their pathologies. The journal's mission is to publish high-quality, hypothesis-driven studies that offer mechanistic novelty and are methodologically robust, covering a wide range of themes in gastroenterology, hepatology, and pancreatology. CMGH reports on the latest scientific advances in cell biology, immunology, physiology, microbiology, genetics, and neurobiology related to gastrointestinal, hepatobiliary, and pancreatic health and disease. The research published in CMGH is designed to address significant questions in the field, utilizing a variety of experimental approaches, including in vitro models, patient-derived tissues or cells, and animal models. This multifaceted approach enables the journal to contribute to both fundamental discoveries and their translation into clinical applications, ultimately aiming to improve patient care and treatment outcomes in digestive health.