Lipocalin-2介导肥胖相关非酒精性脂肪肝炎的作用及机理研究

Nonalcoholic steatohepatitis (NASH) is one of the most common chronic liver diseases closely associated with obesity and metabolic syndrome. Hepatic infiltration of neutrophils and macrophages is the key histopathological feature of NASH. However, the molecular pathways that mediate the hepatic recruitment of these inflammatory cells remain unknown. Lipocalin-2 (LCN2) is a secreted glycoprotein originally isolated from human neutrophils. Our previous studies demonstrated an increased LCN2 level in both blood and liver in obesity-induced NASH animals, and such an elevation of lipocalin-2 is closely associated with augmented hepatic infiltration of neutrophils. Furthermore, we found that LCN-2 deficient mice are resistant to obesity-induced NASH. Therefore, we propose that LCN2 may serve as an important player of obesity-related hepatic inflammation in NASH, possibly by mediating the cross-talk between neutrophils and hepatic macrophages (Kupffer Cells). To test this hypothesis, we will adopt several unique approaches, including generation of LCN2 chimeric mice by bone marrow transplantation, ex vivo co-culture of neutrophils and Kupffer cells, and a series of gain and loss of functional experiments with genetic and pharmacological tools. The findings from this study will not only shed new light on the pathogenesis of obesity-related NASH, but also provide valuable scientific evidence for the future development of effective therapeutic strategies for the treatment of this rapidly growing chronic disease by targeting LCN2 actions in neutrophils.

非酒精性脂肪肝炎（NASH）是一种与肥胖紧密相关，并伴随中性粒细胞等炎性细胞浸润的慢性肝病。但目前对于介导炎性细胞肝组织浸润的分子机制尚未阐明。Lipocalin-2（LCN2）是一种最早从中性粒细胞中分离的糖蛋白。我们在前期研究中发现，LCN2血清及肝内表达水平在NASH动物模型中显著升高；并且，其肝内表达升高与中性粒细胞浸润密切相关。此外，全身性LCN2敲除小鼠中肥胖诱导的肝脏炎症显著减轻。因此，我们假设LCN2作为一种增强肝脏炎症的关键分子，其可能通过介导中性粒细胞与肝脏巨噬细胞（Kupffer细胞）的相互作用而促进NASH的发生与进展。为验证该研究假说，本课题将采用多种特异的研究方案，包括通过骨髓移植形成LCN2髓系细胞嵌合体小鼠、离体中性粒细胞与Kupffer细胞共培养、以及基于药物及基因修饰方法的功能学实验，深入研究LCN2是否可介导NASH炎症反应；并揭示其潜在的分子机制。

项目的背景、主要研究内容、重要结果、关键数据及其科学意义等做简单概述..研究背景：非酒精性脂肪肝炎（NASH）是一种与肥胖紧密相关，并伴随中性粒细胞等炎性细胞浸润的慢性肝病。但目前对于介导炎性细胞肝组织浸润的分子机制尚未阐明。Lipocalin-2（LCN2）是一种最早从中性粒细胞中分离的糖蛋白。我们在前期研究中发现，LCN2 血清及肝内表达水平在NASH 动物模型中显著升高；并且，其肝内表达升高与中性粒细 胞浸润密切相关。此外，全身性LCN2 敲除小鼠中肥胖诱导的肝脏炎症显著减轻。因此，我们假设LCN2 作为一种增强肝脏炎症的关键分子，其可能通过介导中性粒细胞与肝脏巨噬细 胞（Kupffer 细胞）的相互作用而促进NASH 的发生与进展。主要研究内容：①. 通过骨髓移植方案产生LCN2嵌合体小鼠，用以研究LCN-2是否通过调节中性粒细胞功能而启动肝脏炎症；②. 探索LCN-2是否可以通过调节中性粒细胞功能从而介导中性粒细胞与Kupffer细胞相互作用的离体研究研究方案概述；③.通过离体培养小鼠原代中性粒细胞，并通过重组小鼠LCN2蛋白及CXCR2配体蛋白CXCL2刺激原代中性粒细胞，揭示LCN-2介导中性粒细胞功能及干预NASH疾病进展的分子机制研究。重要结果及关键数据：LCN2外周血及肝内表达水平在NASH患者中显着升高；中性粒细胞系统性耗竭可显著改善重组LCN2蛋白诱导的NASH相关肝脏损伤，说明中性粒细胞是LCN2发挥肝脏促炎性效应的核心介导细胞；骨髓细胞中的LCN2嵌合体显著影响HFHC饮食在ApoE 敲除小鼠中诱导NASH；LCN2通过CXCR2增强中性粒细胞迁移；ERK信号传导LCN2对中性粒细胞的调节作用；LCN2的促肝脏炎性反应效应在CXCR2缺失小鼠中显著抑制。研究结论及关键科学意义：本项目研究结果揭示出了中性粒细胞来源的颗粒蛋白LCN-2是介导肥胖相关肝脏代谢性炎症的核心分子，其通过诱导CXCR2表达，从而激活ERK，进而促进中性粒细胞与肝脏巨噬细胞的对话而增强肝脏炎症。研究结果的潜在应用前景为：LCN2-CXCR2-ERK信号通路为临床干预肥胖相关肝脏代谢性炎症，并进行针对NASH进行新型药物研发的重要靶点分子。