School of Public Health, Zhejiang Chinese Medical University, Hangzhou 310053, China.
School of Life Science, Zhejiang Chinese Medical University, Hangzhou 310053, China.
Oxid Med Cell Longev. 2022 Mar 20;2022:4557532. doi: 10.1155/2022/4557532. eCollection 2022.
Hepatic metabolic disorder induced by lipotoxicity plays a detrimental role in metabolic fatty liver disease pathogenesis. Cimifugin (Cim), a coumarin derivative extracted from the root of , possesses multiple biological properties against inflammation, allergy, and oxidative stress. However, limited study has addressed the hepatoprotective role of Cim. Here, we investigate the protective effect of Cim against lipotoxicity-induced cytotoxicity and steatosis in hepatocytes and clarify its potential mechanisms.
AML-12, a nontransformed mouse hepatocyte cell line, was employed in this study. The cells were incubated with palmitate or oleate to imitate hepatotoxicity or steatosis model, respectively.
Cim significantly reversed palmitate-induced hepatocellular injury in a dose-dependent manner, accompanied by improvements in oxidative stress and mitochondrial damage. Cim pretreatment reversed palmitate-stimulated TLR4/p38 MAPK activation and SIRT1 reduction without affecting JNK, ERK1/2, and AMPK pathways. The hepatoprotective effects of Cim were abolished either through activating TLR4/p38 by their pharmacological agonists or genetical silencing SIRT1 via special siRNA, indicating a mechanistic involvement. Moreover, Cim treatment improved oleate-induced hepatocellular lipid accumulation, which could be blocked by either TLR4 stimulation or SIRT1 knockdown. We observed that SIRT1 was a potential target of TLR4 in palmitate-treated hepatocytes, since TLR4 agonist LPS aggravated, whereas TLR4 antagonist CLI-095 alleviated palmitate-decreased SIRT1 expression. SIRT1 knockdown did not affect palmitate-induced TLR4. In addition, TLR4 activation by LPS significantly abolished Cim-protected SIRT1 reduction induced by palmitate. These results collaboratively indicated that TLR4-regulated SIRT1 pathways was mechanistically involved in the protective effects of Cim against lipotoxicity.
In brief, we demonstrate the protective effects of Cim against lipotoxicity-induced cell death and steatosis in hepatocytes. TLR4-regulated p38 MAPK and SIRT1 pathways are involved in Cim-protected hepatic lipotoxicity. Cim is a potential candidate for improving hepatic metabolic disorders mediated by lipotoxicity.
脂毒性引起的肝代谢紊乱在代谢性脂肪性肝病发病机制中起有害作用。香豆素（Cim）是从当归根中提取的一种香豆素衍生物，具有抗炎、抗过敏和抗氧化应激等多种生物学特性。然而，关于 Cim 的肝保护作用的研究有限。在这里，我们研究了 Cim 对肝细胞脂毒性诱导的细胞毒性和脂肪变性的保护作用，并阐明了其潜在机制。
采用非转化的小鼠肝细胞系 AML-12 进行本研究。用棕榈酸或油酸孵育细胞，分别模拟肝毒性或脂肪变性模型。
Cim 以剂量依赖的方式显著逆转了棕榈酸诱导的肝细胞损伤，同时改善了氧化应激和线粒体损伤。Cim 预处理逆转了棕榈酸刺激的 TLR4/p38 MAPK 激活和 SIRT1 减少，而不影响 JNK、ERK1/2 和 AMPK 途径。通过其药理学激动剂激活 TLR4/p38 或通过特殊的 siRNA 基因沉默 SIRT1，Cim 的肝保护作用被消除，表明存在一种机制参与。此外，Cim 治疗改善了油酸诱导的肝细胞脂质积累，这种作用可被 TLR4 刺激或 SIRT1 敲低所阻断。我们观察到 SIRT1 是棕榈酸处理的肝细胞中 TLR4 的潜在靶标，因为 TLR4 激动剂 LPS 加重，而 TLR4 拮抗剂 CLI-095 减轻了棕榈酸降低的 SIRT1 表达。SIRT1 敲低不影响棕榈酸诱导的 TLR4。此外，LPS 激活 TLR4 显著消除了 Cim 保护的由棕榈酸引起的 SIRT1 减少。这些结果共同表明，TLR4 调节的 SIRT1 途径参与了 Cim 对脂毒性的保护作用。
总之，我们证明了 Cim 对肝细胞脂毒性诱导的细胞死亡和脂肪变性的保护作用。TLR4 调节的 p38 MAPK 和 SIRT1 途径参与了 Cim 保护的肝脂毒性。Cim 是改善脂毒性介导的肝代谢紊乱的潜在候选药物。
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