目的 探讨人参皂苷F₂（GF₂）对α-萘异硫氰酸酯（ANIT）诱导的胆汁淤积肝损伤（CLI）小鼠的作用及机制。方法 将70只雄性昆明小鼠随机分为7组（n＝10）：对照组、单给GF₂（100 mg∙kg^-1）组、模型组、熊去氧胆酸（UDCA，40 mg∙kg^-1）组和GF₂低、中、高剂量（25、50、100 mg∙kg^-1）组。小鼠连续ig给药7 d，于第5天ig给予ANIT（100 mg∙kg^-1）建立胆汁淤积模型。自动生化仪测量血清丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）、碱性磷酸酶（ALP）、总胆汁酸（TBA）、总胆红素（TBIL）、直接胆红素（DBIL）、丙二醛（MDA）、超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）水平，肝组织匀浆上清MDA、SOD、GSH-Px和过氧化氢酶（CAT）水平；ELISA法检测肝组织匀浆上清液中炎症因子肿瘤坏死因子-α（TNF-α）、白细胞介素（IL）-6、IL-1β和脂多糖（LPS）水平；HE染色和Masson染色进行肝组织病理学分析；TUNEL染色观察肝细胞凋亡；免疫组化法观察免疫细胞标志物[中性粒细胞标志物CD11b和Ly6g、巨噬细胞标志物F4/80和T细胞标志物CD3]、肝纤维化标志物[α-平滑肌肌动蛋白（α-SMA）、I型胶原（Collagen I）]、Toll样受体4（TLR4）/髓分化因子88（MyD88）/核因子κB（NF-κB）相关蛋白、转化生长因子-β1（TGF-β1）/Smad相关蛋白、Bcl-2相关X蛋白（Bax）蛋白表达；Western blotting检测TLR4/Myd88/NF-κB、Nrf2/HO-1/NQO1和TGF-β1/Smad、B淋巴细胞瘤-2（Bcl-2）/Bax通路蛋白表达。结果 与模型组比较，GF₂组AST、ALT、ALP、TBA、TBIL和DBIL水平显著降低（P＜0.05、0.01、0.001），GF₂明显改善模型组肝细胞肿胀、空泡化、肝内炎症细胞浸润和坏死，胆管增生和扩张；GF₂显著降低ANIT诱导的炎症因子TNF-α、IL-6、IL-1β、LPS水平（P＜0.05、0.01、0.001），明显降低CD11b、Ly6g、F4/80、CD3表达，显著降低TLR4、Myd88、NF-κB p65和pIκBα的蛋白表达（P＜0.05、0.01）；GF₂组MDA水平降低，SOD、CAT活性和GSH水平恢复，差异显著（P＜0.05、0.01、0.001），且上调Nrf2及其靶基因HO-1和NQO1蛋白的表达（P＜0.05、0.001）；GF₂组肝纤维化明显减轻，α-SMA和Collagen I表达明显下降，且TGF-β1、Smad2、Smad3蛋白表达显著降低（P＜0.05、0.01、0.001）；GF₂减弱CLI小鼠的肝细胞凋亡程度，显著降低促凋亡蛋白Bax的表达、显著增加抗凋亡蛋白Bcl-2的表达（P＜0.05、0.01）。结论 CF₂可抑制TLR4/Myd88/NF-κB通路减轻炎症反应、激活Nrf2通路减轻小鼠氧化损伤、抑制TGF-β1/Smad通路减轻肝纤维化，减轻ANIT诱导的细胞凋亡，进而减轻胆汁淤积小鼠肝损伤。

Objective To investigate the effect of ginsenoside F₂(GF₂) on α-naphthalene isothiocyanate (ANIT) -induced cholestatic liver injury (CLI) in mice and its mechanism. Methods 70 male Kunming mice were randomly divided into seven groups (n= 10): control group, single GF₂ (100 mg∙kg^-1) group, model group, UDCA (40 mg∙kg^-1) group, and low, medium, and high dose GF₂ (25, 50, 100 mg∙kg^-1) groups. The mice were administered orally with the drugs for seven consecutive days, and ANIT (100 mg∙kg^-1) was ig administered on the 5th day to establish a cholestasis model. The automatic biochemistry analyzer was used to measure serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bile acid (TBA), total bilirubin (TBIL), direct bilirubin (DBIL), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) levels, and the levels of MDA, SOD, GSH-Px, and catalase in the liver tissue homogenate supernatant; the enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of inflammatory factors (tumor necrosis factor-α (TNF-α), interleukin (IL)-6, IL-1β, and LPS) in the liver tissue homogenate supernatant; hematoxylin and Masson staining were used for histopathological analysis of the liver tissue; TUNEL staining was used to observe hepatocyte apoptosis; immunohistochemistry was used to detect the expression of immune cell markers (neutrophil markers CD11b and Ly6g, macrophage markers F4/80 and T cell markers CD3), liver fibrosis markers (α-smooth muscle actin (α-SMA), type I collagen (Collagen I)), Toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear factor kappa B (NF-κB) related protein, transforming growth factor-β1 (TGF-β1)/SMAD-associated protein, Bcl-2 associated X protein (Bax) protein expression. Western blotting was used to detect the expression of TLR4/Myd88/NF-κB, Nrf2/HO-1/NQO1, TGF-β1/Smad, B-lymphoblastoma-2 (Bcl-2)/Bax pathway proteins. Results Compared with the model group, the levels of AST, ALT, ALP, TBA, TBIL and DBIL were significantly lower in the GF₂ group (P < 0.05, 0.01, 0.001), and GF₂ significantly improved the liver cell swelling, vacuolization, intrahepatic inflammatory cell infiltration and necrosis, bile duct hyperplasia and dilation in the model group; GF₂ significantly lowered the levels of inflammatory factors TNF-α, IL-6, IL-1β and LPS induced by ANIT (P < 0.05, 0.01, 0.001), and significantly lowered the expression of CD11b, Ly6g, F4/80, CD3, and significantly downregulated the protein expression of TLR4, Myd88, NF-κB p65 and pIκBα (P < 0.05, 0.01); the level of MDA was lowered in the GF₂ group, and the activities of SOD and CAT and the level of GSH were restored, with significant differences (P < 0.05, 0.01, 0.001), and the expression of Nrf2 and its target genes HO-1 and NQO1 was significantly upregulated (P < 0.05, 0.001); the degree of liver fibrosis was significantly alleviated in the GF₂ group, and the expression of α-SMA and Collagen I was significantly downregulated, and the expression of TGF-β1, Smad2 and Smad3 was significantly downregulated (P < 0.05, 0.01, 0.001); GF₂ weakened the degree of liver cell apoptosis in CLI mice, significantly decreased the expression of pro-apoptotic protein Bax and significantly increased the expression of anti-apoptotic protein Bcl-2 (P < 0.05, 0.01). Conclusion CF₂ can inhibit the TLR4/Myd88/NF-κB pathway to alleviate inflammatory responses, activate the Nrf2 pathway to alleviate oxidative damage in mice, inhibit the TGF-β1/Smad pathway to alleviate liver fibrosis, alleviate cell apoptosis induced by ANIT, and thereby alleviate liver damage in cholestasis mice.

2024年度河南省中医药科学研究专项重点项目（2024ZY1030）