急性肝损伤（ALI）是一种在短时间内由化学物质、酒精和病毒感染等因素影响下肝脏受损的疾病，导致肝细胞损伤。牛膝（Achyranthes bidentata Bl.）具有肝保护作用，引起了广泛关注。本研究首次从牛膝根中分离到一种五环三萜类化合物（牛膝皂苷A, AsA），并研究了其抗ALI活性及其机制。以AsA（10或20 mg/kg, 口服）持续给药1周，然后通过脂多糖（LPS，10 µg/kg）/D-半乳糖胺（D-GalN，700 mg/kg）诱导肝损伤。通过肝脏的HE染色、天冬氨酸氨基转移酶（AST）、丙氨酸氨基转移酶（ALT）和细胞因子的检测来研究ALI相关特征。通过评估肝组织的线粒体形态、线粒体膜电位（MMP）、氧化应激平衡、凋亡、2-DG平均荧光强度、肝组织中的自然杀伤细胞（NK细胞）等指标，评估氧化应激损伤和炎症损伤。使用转录组学和代谢组学分析明确机制。同时，分析了神经酰胺磷酸酯1-磷酸酯（S1P）、神经酰胺磷酸酯激酶1（SPKH1）、神经酰胺磷酸酯1磷酸酸受体1（S1PR1）、神经酰胺磷酸酯1磷酸酸受体3（S1PR3）、TNF受体相关因子2（TRAF-2）、磷酸化 NF-κB p65（p-P65）、NF-κB p65（P65）、原癌基因ras（Ras）、与C3毒素相关的ras样蛋白（Rac）、磷脂酶C（PLC）、白细胞介素6（IL-6）、肿瘤坏死因子α（TNF-α）、白细胞介素1β（IL-1β）、血管细胞黏附分子1（Vcam1）、CC趋化因子配体-2（Ccl2）的mRNA和蛋白质表达水平。通过研究在LPS/D-GalN诱导的AML12细胞中SPHK1转染沉默/过表达对AsA的影响，评估了SPHK1在AsA引起的观察效果中的调节作用。AsA可以改善ALI模型中的肝功能、炎症、线粒体结构和氧化应激。与此同时，AsA导致了与鞘脂信号通路相关的蛋白质表达下调。SPHK1沉默增强了AsA的保护作用，而SPHK1过表达降低了AsA的保护作用，这表明ALI通过AsA的有效分子介导SPHK1调节。通过抑制炎症和氧化应激途径，AsA可以通过SPHK1/S1P/S1PR1途径改善LPS/D-GalN诱导的ALI。这样，为AsA在ALI治疗中的应用提供了分子上的合理性依据。版权所有 © 2023 The Authors. 由Elsevier B.V.出版。保留所有权利。

Acute liver injury (ALI) refers to a disease in which the liver is affected by factors such as chemical substances, alcohol, and virus infection in a short time, resulting in damage to liver cells. Achyranthes bidentata Bl. with the hepatoprotective activity has attracted great attention. In this study, a pentacyclic triterpenoid (Aralia saponin A, AsA) was isolated from roots of Achyranthes bidentata Bl. and its anti-ALI activity, as well as the mechanisms, were investigated for the first time.AsA (10 or 20 mg/kg, i.g.) was administered over a period of 1 weeks, following which liver injury was induced by LPS (10 µg/kg)/D-GalN (700 mg/kg). H&E staining of liver, Aspartate amino transferase (AST), Alanine transaminase (ALT) and cytokines was employed to investigate ALI relevant features. The mitochondrial morphology and levels of mitochondrial membrane potential (MMP), oxidative stress balance, apoptosis, average fluorescence intensity of 2-DG, natural killer (NK) cells in liver tissues were determined to assess the oxidative stress damage and inflammatory injury. Transcriptomics and metabonomics analysis were employed to clarify the mechanisms. Additionally, the mRNA and protein expression levels of Sphingosine 1-phosphate (S1P), Sphingosine kinase-1 (SPKH1), Sphingosine 1 phosphate receptor 1 (S1PR1), Sphingosine 1 phosphate receptor 3 (S1PR3), TNF receptor associated factor 2 (TRAF-2), Phospho-NF- kappaB p65 (p-P65), NF- kappaB p65 (P65), Proto-oncogene ras (Ras), Ras-related C3 botulinum toxin substrate (Rac), Phospholipase C (PLC), Interleukin 6 (IL-6), Tumor necrosis factor α (TNF-α), Interleukin 1β (IL-1β), Vascular cell adhesion molecule 1 (Vcam1), CC chemokine ligand-2 (Ccl2) were analyzed. The mediating role of SPHK1 in the observed effects caused by AsA was assessed by investigatin SPHK1 transfection silencing/overexpression against AsA in AML12 cells induced by LPS/D-GalN.AsA can ameliorate liver function, inflammation, mitochondrial structure and oxidative stress in the ALI model. Meanwhile, AsA led to downregulated expression of proteins associated with sphingolipid signaling pathway. Silencing of SPHK1 led to enhanced protective effects of AsA, while over-expression of SPHK1 led to degraded protective effects of AsA in LPS/D-GalN-induced AML12 cells, suggesting that ALI is regulated by active molecules of AsA by means of SPHK1 mediation.AsA can ameliorate LPS/D-GalN-induced ALI by inhibiting inflammation and oxidative stress via the SPHK1/S1P/S1PR1 pathway. In this way, a molecular justification is provided for AsA application in ALI treatment.Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.