Investigating the Ameliorative Effect of Gelian Qishen Formula (葛连芪参方) on Non-Alcoholic Fatty Liver Disease in Mice through Inhibiting Hepatic Ferroptosis via the Nrf2/Gpx4 Pathway

Author：GUO Manman1, HENG Wangqin2, GAO Liming1, LU Jielin1, PENG Huiping1, DING Menglei1, WANG Yi1

Unit：1.Traditional Chinese Medicine Hospital of Kunshan, Kunshan Jiangsu 215300, China; 2.The Affiliated Hospital of Nanjing University of Chinese Medicine for Integrated Traditional Chinese and Western Medicine, Nanjing Jiangsu 210028, China

Quote：引用：郭曼曼，衡王琴，高黎明，陆杰霖，彭惠平，丁梦磊，汪怡.基于Nrf2/Gpx4通路抑制肝脏铁死亡探讨葛连芪参方对小鼠非酒精性脂肪性肝病的改善作用[J].中医药导报,2026,32(2):47-52.

DOI：10.13862/j.cn43-1446/r.2026.02.008

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Abstract：

Objective: To investigate the mechanism of Gelian Qishen formula (GQF) in ameliorating non-alcoholic fatty liver disease (NAFLD) in mice. Methods: Mice were randomly divided into a control group, model group, high-dose GQF group (25.0 g/kg), low-dose GQF group (12.5 g/kg), and inhibitor group (12.5 g/kg GQF administered concurrently with intraperitoneal injection of an Nrf2 inhibitor every other day). NAFLD was induced by feeding a high-fat diet (HFD) for 12 weeks to all groups except the control group, which was fed a standard diet, followed by 8 weeks of drug intervention. Hepatic pathology and lipid deposition were observed using HE and Oil Red O staining. Serum and hepatic lipid profiles [total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C)], liver function indices [alanine aminotransferase (ALT), aspartate aminotransferase (AST)], and oxidative stress markers [reduced glutathione (GSH), malondialdehyde (MDA), superoxide dismutase (SOD), total antioxidant capacity (T-AOC)] were measured using commercial assay kits. Serum inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and interleukin-8 (IL-8), were detected by enzyme-linked immunosorbent assay (ELISA). The protein expression levels of Nrf2, Gpx4, HO-1, and xCT in the liver were determined by Western blotting. Results: After HFD feeding, the model group showed significantly increased serum and hepatic lipid contents, liver function indices, and inflammatory cytokine levels (P<0.01), along with elevated hepatic Fe2+ and MDA contents, and significantly decreased levels of GSH, SOD, and T-AOC (P<0.01). Liver tissues exhibited marked steatosis and lipid deposition in model group. After intervention, all treatment groups showed significant reductions in serum and hepatic TC and TG levels, serum ALT and AST levels, and hepatic Fe2+ and MDA contents (P<0.05 or P<0.01), while hepatic SOD and T-AOC levels were significantly increased (P<0.01). Furthermore, the high-dose GQF group and low-dose GQF group exhibited significantly decreased serum and hepatic LDL-C levels and serum TNF-α, IL-6, and IL-8 levels (P<0.01), alongside significantly increased hepatic GSH levels (P<0.01), with markedly alleviated hepatic steatosis and lipid deposition. Western blotting revealed that the protein expression levels of Nrf2, xCT, HO-1, and Gpx4 in the liver were significantly decreased in the model group (P<0.05 or P<0.01). After treatment, the protein expression levels of Nrf2, xCT, HO-1, and Gpx4 significantly increased in the high-dose GQF group and low-dose GQF group (P<0.01). Compared with the low-dose GQF group, the inhibitor group showed significantly lower expression levels of Nrf2, xCT, HO-1, and Gpx4 proteins (P<0.05 or P<0.01). Conclusion: Gelian Qishen formula can effectively ameliorate NAFLD in mice, and the mechanism may be associated with activating the Nrf2/Gpx4 pathway to inhibit hepatic ferroptosis.

Key words：non-alcoholic fatty liver disease; Gelian Qishen formula; Nrf2/Gpx4 pathway; ferroptosis; mice

摘要：目的：探讨葛连芪参方改善小鼠非酒精性脂肪性肝病（NAFLD）的作用机制。方法：将小鼠随机分为空白组、模型组、葛连芪参方高剂量组（25.0 g/kg）、葛连芪参方低剂量组（12.5 g/kg）和抑制剂组（葛连芪参方12.5 g/kg+隔天腹腔注射Nrf2抑制剂）。除空白组喂养普通饲料外，其他组给予高脂饲料喂养12周，制备NAFLD模型；造模成功后，药物干预8周。采用HE和油红O染色观察肝脏病理及脂质沉积情况。试剂盒检测血清和肝脏脂质水平[总胆固醇（TC）、甘油三酯（TG）、高密度脂蛋白胆固醇（HDL-C）、低密度脂蛋白胆固醇（LDL-C）]、肝功能指标[谷丙转氨酶（ALT）、谷草转氨酶（AST）]及氧化应激指标[还原性谷胱甘肽（GSH）、丙二醛（MDA）、超氧化物歧化酶（SOD）、总抗氧化能力（T-AOC）]。酶联免疫吸附试验（ELISA）检测血清炎症因子水平，包括肿瘤坏死因子-α（TNF-α）、白介素-1β（IL-1β）、白介素-6（IL-6）及白介素-8（IL-8）。Western blotting检测肝脏Nrf2、Gpx4、HO-1、xCT等蛋白表达情况。结果：高脂饲料造模后，模型组小鼠血清和肝脏脂质含量、肝功能指标及炎症因子水平显著升高（P<0.01），肝脏亚铁离子、MDA含量上升，GSH、SOD、T-AOC水平显著下降（P<0.01），肝组织呈现明显脂肪变性和脂质沉积。药物干预后，各给药组小鼠血清和肝脏TC、TG含量，血清ALT、AST水平，肝脏亚铁离子含量、MDA含量显著降低（P＜0.05或P＜0.01）；肝脏SOD、T-AOC水平显著升高（P＜0.01）。此外，葛连芪参方高、低剂量组小鼠血清和肝脏LDL-C含量、血清TNF-α、IL-6、IL-8水平显著降低（P＜0.01），肝脏GSH水平显著升高（P＜0.01）；肝脏脂肪变性和脂质沉积明显缓解。Western blotting检测显示，模型组小鼠肝脏组织Nrf2、xCT、HO-1、Gpx4蛋白表达水平显著降低（P<0.05或P<0.01）。治疗后，葛连芪参方高、低剂量组Nrf2、xCT、HO-1、Gpx4蛋白表达量明显升高（P<0.01）。与葛连芪参方低剂量组比较，抑制剂组Nrf2、xCT、HO-1、Gpx4蛋白表达水平显著降低（P<0.05或P<0.01）。结论：葛连芪参方可有效改善小鼠非酒精性脂肪肝病，其作用机制可能与激活Nrf2/Gpx4通路抑制肝脏铁死亡有关。

关键词：非酒精性脂肪性肝病；葛连芪参方；Nrf2/Gpx4通路；铁死亡；小鼠

Release time：2026-03-05

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