通讯作者: 叶晓婷, yxt129@163.com

DOI：10.12201/bmr.202409.00013

Esculin improve lipid accumulation in hepatocytes by inhibiting the PERK/eIF2A/ATF4 signaling pathway

• 摘要：目的 初步探讨七叶苷通过抑制PERK/eIF2A/ATF4信号通路改善肝细胞脂肪变性的治疗作用和机制。方法 采用人源正常肝细胞系HL-7702，利用0.5 mmol/L 游离脂肪酸（Free fatty acid，FFA，油酸:棕榈酸=2:1）诱导体外肝细胞脂肪变性模型，经50, 200 μmol/L七叶苷处理24 h。测定细胞内生化指标丙氨酸转氨酶（Alanine Transaminase，ALT）、天冬氨酸转氨酶（Aspartate aminotransferase，AST）、丙二醛（Malonaldehyde，MDA）、还原型谷胱甘肽（Glutathione，GSH）和甘油三酯（Triglyceride，TG）含量。采用尼罗红脂肪荧光染色法检测细胞内脂滴。采用qRT-PCR法检测细胞内脂质代谢过程相关基因的转录水平。采用Western blot（WB）检测细胞内促凋亡因子Caspase-3和Bax的蛋白表达量，以及PERK/eIF2A/ATF4信号通路相关蛋白和磷酸化水平。结果 生化指标检测结果证实，50和200 μmol/L的七叶苷处理显著降低FFA诱导肝细胞内MDA、ALT和AST水平（P < 0.05），并显著增加细胞内GSH水平（P < 0.05）。WB结果显示七叶苷处理能显著降低细胞内促凋亡因子Caspase-3和Bax的蛋白表达量（P < 0.01）。尼罗红染色和TG含量检测结果证实七叶苷处理能显著减少细胞内脂滴堆积和TG含量（P < 0.05）。qRT-PCR结果显示七叶苷处理后肝细胞内PPARγ、FASN、Srebf1、Dgat2、Mvk、Acaca的表达量均显著降低（P < 0.05）。在机制方面，七叶苷处理才能显著降低肝细胞中PERK、eIF2A和ATF4的磷酸化水平（P < 0.05）。结论 七叶苷可通过调控PERK/eIF2A/ATF4信号通路改善肝细胞脂质堆积，对维护肝细胞健康状态起到积极作用。

  关键词： 肝细胞; 脂肪变性; 脂质代谢; PERK/eIF2A/ATF4

   

  Abstract: Objective To explore the effect and mechanism of esculin on hepatocyte steatosis by inhibiting PERK/eIF2A/ATF4 signaling pathway. Methods Human normal liver cell line HL-7702 was used to induce a steatosis model of hepatocytes in vitro with 0.5 mmol/L free fatty acid (FFA, oleic acid: palmitic acid =2:1) and treated with 50, 200 μmol/L esculin for 24 h. After the cell samples were broken by ultrasound, the supernatant was collected and the contents of alanine aminotransferase (ALT), aspartate aminotransferase (AST), malondialdehyde (MDA), reduced glutathione and triglyceride were detected. Intracellular lipid droplets and the transcriptional levels of genes related to intracellular lipid metabolism were detected by Nile red fat fluorescence staining and qRT-PCR, respectively. Western blot was employed to detect the protein expressions of pro-apoptotic factors Caspase-3 and Bax, as well as the protein and phosphorylation levels related to PERK/eIF2A/ATF4 signaling pathway. Results The results confirmed that treatments of 50 and 200 μmol/L of esculin significantly decreased the levels of FFA-induced MDA, ALT and AST in hepatocytes (P < 0.05), and significantly increased the levels of intracellular GSH (P < 0.05). WB results showed that esculin treatment could significantly reduce the protein expression levels of Caspase-3 and Bax (P < 0.01). The results of Nile red staining and TG content detection confirmed that esculin treatment could significantly reduce the accumulation of intracellular lipid droplets and TG content (P < 0.05). The results of qRT-PCR showed that the expression levels of PPARγ, FASN, Srebf1, Dgat2, Mvk and Acaca in hepatocytes were significantly decreased after esculin treatment (P < 0.05). In terms of mechanism, the phosphorylation levels of PERK, eIF2A and ATF4 in hepatocytes were significantly reduced by esculin treatment (P < 0.05). Conclusion Esculin could improve lipid accumulation in hepatocytes by regulating the PERK/eIF2A/ATF4 signalling pathway, which plays a positive role in maintaining the healthy state of hepatocytes.

  Key words: hepatocyte; hepatocyte steatosis; lipid metabolism; PERK/eIF2A/ATF4

  提交时间：2024-09-10

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• 序号	提交日期	编号	操作
  1	2024-08-08	bmr.202409.00013V1	下载

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