Sapidolide A缓解APAP急性肝损伤的分子机制研究

Acetaminophen (APAP) could cause acute liver injury, which could lead to liver failure and even death. Currently, the only therapeutic drug, N-acetylcysteine (NAC), is limited in clinical application due to the narrow time window. Based on the broad prospects of macrophage polarization in overcoming liver injury, we have previously screened that the extracted monomer of the berries of Baccaurea ramiflora, Sapidolide A (SA) could promote the M2 polarization of macrophages and significantly alleviate acute liver injury induced by APAP in mice. Moreover, SA could synergize with NAC. SA does not directly affect the hepatocyte death induced by APAP in vitro, suggesting that SA may regulate M2 polarization of macrophages to protect APAP-induced acute liver injury, which may be related to DJ1 activation and oxidative stress inhibition. This project will further clarify the effect of SA on APAP-induced acute liver injury, and demonstrate the key role of M2 polarization of macrophage, exploring the molecular mechanism of DJ1-Keap1-Nrf2 signaling pathway in SA-promoted M2 polarization. Our study will not only clarify the function and molecular mechanism of SA in regulating M2 polarization of macrophage protecting APAP-induced acute liver injury, but also provide the potential target and theoretical basis for design and discovery of novel anti-acute liver injury drugs.

对乙酰氨基酚（APAP）可引起急性肝损伤，严重者会导致肝衰竭甚至死亡。目前唯一治疗药物N-乙酰半胱氨酸（NAC）因时间窗窄而临床应用受限。基于巨噬细胞极化在对抗肝损伤中的广阔前景，我们前期筛选发现三丫果提取单体Sapidolide A（SA）能促进巨噬细胞M2型极化，显著缓解APAP引起的小鼠急性肝损伤且与NAC协同增效。SA不直接影响APAP引起的体外肝细胞死亡，提示SA可能调控巨噬细胞M2型极化保护APAP急性肝损伤，此作用可能与DJ1激活抑制氧化应激有关。本课题将进一步明确SA治疗APAP急性肝损伤的效应，证明巨噬细胞M2型极化在其中的关键作用，探索DJ1-Keap1-Nrf2信号通路在SA促进M2型极化中的分子机制。本课题不仅将阐明SA调控巨噬细胞M2型极化保护APAP急性肝损伤的功能和分子机制，还为设计与发现基于巨噬细胞调控APAP急性肝损伤的全新治疗药物提供潜在靶点和理论依据。

研究背景：.对乙酰氨基酚（APAP）可引起急性肝损伤，严重者会导致肝衰竭甚至死亡。肝脏巨噬细胞在APAP急性肝损伤中扮演着重要作用。M1型巨噬细胞产生促炎细胞因子和趋化因子，促进APAP急性肝损伤发展，而M2型巨噬细胞释放抗炎细胞因子，参与死亡细胞的吞噬。近期研究表明巨噬细胞炎症小体在APAP急性肝损伤中发生激活和焦亡，并且依赖于NLRP3炎症小体活性。因此，基于巨噬细胞的靶向策略在治疗APAP急性肝损伤上是极具应用前景的策略。.研究内容：.在前期研究中，我们发现三丫果提取单体Sapidolide A（SA）可以显著缓解APAP急性肝损伤且SA在体外可以显著抑制NLRP3炎性小体的激活。本项目在此基础上，将进一步证明SA在体内对肝脏巨噬细胞和肝实质细胞的作用。同时通过巨噬细胞与L02细胞共培养的体外实验，明确SA 能保护巨噬细胞不受APAP损伤L02肝细胞所致的细胞焦亡的影响。最后，利用APAP急性肝损伤小鼠模型，系统性阐明巨噬细胞NLRP3炎症小体在SA保护APAP急性肝损伤中的关键作用。.研究结果：.（1）首先通过药效学评估，分别用两个剂量SA（25和50 mg/kg, ip）预处理小鼠，再用单剂量APAP (400 mg/kg, ip)，结果表明SA可以治疗APAP急性肝损伤。（2）BMDMs与人肝细胞L02共培养中，SA治疗减少了由LPS/ ATP处理的巨噬细胞的条件培养基(CM)加重的继发性肝细胞损伤。（3）SA以剂量依赖的方式抑制NF-κB的磷酸化活化，从而抑制LPS/ ATP处理的小鼠骨髓源性原发性巨噬细胞(BMDMs)中NOD样受体蛋白3 (NLRP3)炎症小体的激活及细胞焦亡。（4）在AILI小鼠模型上，SA可以抑制巨噬细胞NLRP3炎症小体的激活，从而缓解APAP急性肝损伤的炎症反应。.科学意义：.本研究结果不仅揭示了SA保护APAP急性肝损伤的重要作用，还阐明SA调控巨噬细胞NLRP3炎症小体的激活的分子机制，为设计与发现基于巨噬细胞调控APAP急性肝损伤的全新治疗药物提供潜在策略和理论依据。