上海交通大学学报（医学版）

上海交通大学学报（医学版） ›› 2025, Vol. 45 ›› Issue (4): 415-425.doi: 10.3969/j.issn.1674-8115.2025.04.003

• 论著 · 基础研究 • 上一篇    下一篇

利拉鲁肽改善非酒精性脂肪性肝病小鼠肝纤维化的作用及其机制

王仁杰1, 祝超瑜2, 方云云2, 肖元元2, 王倩倩2, 宋雯婧2, 魏丽2()   

  1. 1.上海海洋大学水产与生命学院，上海 201306
    2.上海交通大学医学院附属第六人民医院内分泌代谢科，上海 200233
  • 收稿日期:2024-10-30 接受日期:2024-12-20 出版日期:2025-04-28 发布日期:2025-04-28
  • 通讯作者: 魏 丽，主任医师，博士；电子信箱：18930173636@189.cn。
  • 作者简介:王仁杰（1995—），男，硕士；电子信箱：18166332355@189.cn。
  • 基金资助:
    上海市自然科学基金(20ZR1442500)

Effects and mechanisms of liraglutide in ameliorating liver fibrosis in NAFLD mice

WANG Renjie1, ZHU Chaoyu2, FANG Yunyun2, XIAO Yuanyuan2, WANG Qianqian2, SONG Wenjing2, WEI Li2()   

  1. 1.College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China
    2.Department of Endocrinology and Metabolism, Shanghai Sixth People′s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200233, China
  • Received:2024-10-30 Accepted:2024-12-20 Online:2025-04-28 Published:2025-04-28
  • Contact: WEI Li, E-mail: 18930173636@189.cn.
  • Supported by:
    Natural Science Foundation of Shanghai(20ZR1442500)

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摘要：

目的·探讨利拉鲁肽对非酒精性脂肪性肝病（non-alcoholic fatty liver disease，NAFLD）小鼠肝纤维化的作用及其机制。方法·将20只8周龄C57BL/6J小鼠随机分为正常饮食组（Chow组）及甲硫氨酸-胆碱缺乏（methionine-choline deficient，MCD）饮食喂养组（MCD组），每组10只。采用MCD饮食诱导构建NAFLD小鼠。将2组小鼠随机分为4个亚组，分别为Chow＋生理盐水组、Chow＋利拉鲁肽组、MCD＋生理盐水组、MCD＋利拉鲁肽组。每日分别腹腔注射利拉鲁肽（400 μg/kg）或同体积生理盐水4周后，行腹腔注射葡萄糖耐量实验（intraperitoneal glucose tolerance test，IPGTT）。测定小鼠血清天冬氨酸转氨酶（aspartate transaminase，AST）、丙氨酸转氨酶（alanine aminotransferase，ALT）、总胆固醇（total cholesterol，TC）、甘油三酯（triglyceride，TAG）、高密度脂蛋白胆固醇（high density lipoprotein cholesterol，HDL-C）、低密度脂蛋白胆固醇（low-density lipoprotein cholesterol，LDL-C）水平。小鼠处死后取其肝组织测定TAG含量，对肝组织行苏木精-伊红（hematoxylin-eosin，HE）染色、油红O染色及马松（Masson）染色观察肝脏病理结构、脂质沉积和纤维化程度。采用实时荧光定量聚合酶链反应（real-time fluorescent quantitative polymerase chain reaction，qPCR）与Western blotting检测肝组织α平滑肌肌动蛋白（α-smooth muscle actin，α-SMA）、纤连蛋白（fibronectin，FN）、Ⅰ型胶原蛋白α（collagen type Ⅰ α，COL1A）、基质金属蛋白酶9（matrix metallo protein 9，MMP9）、金属蛋白酶组织抑制因子1（tissue inhibitor of metal protease 1，TIMP1）和转化生长因子β（transforming growth factor-β，TGF-β）表达水平，检测Sma和Mad相关蛋白3（Sma- and Mad-related protein 3，SMAD3）及磷酸化SMAD3（SMAD3 phosphorylation，pSMAD3）蛋白表达水平。结果·利拉鲁肽干预后，IPGTT显示小鼠血糖在第15、30、60 min时均有所下降，受试者操作特征曲线（receiver operator characteristic curve，ROC曲线）的曲线下面积（area under the curve，AUC）降低（均P<0.05）。血清生化检测结果显示，利拉鲁肽干预后，MCD组小鼠AST与ALT水平降低（均P<0.001），TC、HDL-C水平升高（均P<0.05），TAG、LDL-C无明显变化。HE染色与油红O染色显示，利拉鲁肽干预后，肝细胞脂滴数量明显减少，气球样变性和炎症浸润程度减轻。Masson染色显示，利拉鲁肽干预后，MCD组小鼠肝脏蓝色胶原纤维数量明显减少。qPCR与Western blotting检测显示，MCD组小鼠肝脏α-SMA、FN、COL1A、TIMP1、TGF-β、pSMAD3/SMAD3表达明显上调，MMP9蛋白表达下降；经利拉鲁肽干预后，MCD组小鼠肝脏α-SMA、FN、COL1A、TIMP1、TGF-β、pSMAD3/SMAD3表达明显降低，MMP9表达上升。结论·利拉鲁肽能缓解NAFLD小鼠肝损伤、肝脏脂质沉积及纤维化程度，并通过调控TGF-β/SMAD3通路和纤维化相关蛋白表达改善小鼠肝功能和肝纤维化。

关键词: 利拉鲁肽, 转化生长因子β, Sma和Mad相关蛋白3, 非酒精性脂肪性肝病, 肝纤维化

Abstract:

Objective ·To investigate the effects of liraglutide on liver fibrosis in mice with non-alcoholic fatty liver disease (NAFLD) and the underlying mechanisms. Methods ·Twenty 8-week-old C57BL/6J mice were randomly divided into a normal chow diet group (Chow group) and a methionine-choline-deficient (MCD) diet group (MCD group), with 10 mice per group. The MCD diet was used to induce NAFLD. Each group was further divided into two subgroups, resulting in four subgroups: Chow+saline, Chow+liraglutide, MCD+saline, and MCD+liraglutide group. After daily intraperitoneal injection of liraglutide (400 μg/kg) or an equivalent volume of saline for 4 weeks, an intraperitoneal glucose tolerance test (IPGTT) was performed. Serum levels of aspartate transaminase (AST), alanine aminotransferase (ALT), total cholesterol (TC), triglyceride (TAG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) were measured. Liver tissues were collected post-euthanasia to assess TAG content. Histopathological changes, lipid deposition, and fibrosis were evaluated via hematoxylin-eosin (HE) staining, Oil Red O staining, and Masson staining. Real-time quantitative PCR (qPCR) and Western blotting were used to analyze the expression of α-smooth muscle actin (α-SMA), fibronectin (FN), collagen type Ⅰ α (COL1A), matrix metalloproteinase 9 (MMP9), tissue inhibitor of metalloproteinase 1 (TIMP1), transforming growth factor β (TGF-β), SMAD3, and phosphorylated SMAD3 (pSMAD3). Results ·The IPGTT revealed that liraglutide intervention reduced blood glucose levels at 15, 30, and 60 min, with a decreased area under the curve (AUC) (both P<0.05). Biochemical analysis showed that liraglutide lowered AST and ALT levels (both P<0.001), increased TC and HDL-C levels (both P<0.05), but had no significant effect on TAG or LDL-C in MCD mice. HE staining and Oil Red O staining revealed reduced lipid droplets, ballooning degeneration, and inflammatory infiltration in hepatocytes after liraglutide treatment. Masson staining indicated decreased collagen fiber deposition in the liver. qPCR and Western blotting analysis demonstrated upregulated expression of α-SMA, FN, COL1A, TIMP1, TGF-β, and pSMAD3/SMAD3, alongside downregulated MMP9 in MCD mice. Liraglutide reversed these changes, lowering α-SMA, FN, COL1A, TIMP1, TGF-β, and pSMAD3/SMAD3 expression while increasing MMP9 expression. Conclusion ·Liraglutide ameliorates liver injury, lipid deposition, and fibrosis in NAFLD mice, through modulation of the TGF-β/SMAD3 pathway and regulating fibrosis-associated protein expression.

Key words: liraglutide, transforming growth factor-β (TGF-β), Sma- and Mad-related protein 3 (SMAD3), non-alcoholic fatty liver disease, liver fibrosis

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