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