人参皂苷Rg1调节Nrf2/ARE信号通路对大鼠妊娠期肝内胆汁淤积症的影响

The effect of ginsenoside Rg1 on intrahepatic cholestasis of pregnancy in rats by regulating the Nrf2/ARE signaling pathway

目的探讨人参皂苷Rg1(GRg1)调节核因子E2相关因子2(Nrf2)/抗氧化反应原件(ARE)信号通路对大鼠妊娠期肝内胆汁淤积症(ICP)的作用。方法将雌性孕SD大鼠随机分为正常组(Normal组)、ICP模型组(Model组)、低剂量GRg1组(GRg1⁃L组,20 mg/kg GRg1)、高剂量GRg1组(GRg1⁃H组,40 mg/kg GRg1)、高剂量GRg1+Nrf2抑制剂全反式维甲酸(ATRA)组(GRg1⁃H+ATRA组,40 mg/kg GRg1+10 mg/kg ATRA),每组12只。采用苯甲酸雌二醇、黄体酮联合注射的方式建立大鼠ICP模型。给药结束后ELISA法检测大鼠血清总胆红素(TBIL)、丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、碱性磷酸酶(ALP)和总胆汁酸(TBA)、肿瘤坏死因子⁃α(TNF⁃α)、γ⁃干扰素(IFN⁃γ)和白细胞介素⁃1β(IL⁃1β)及肝组织中超氧化物歧化酶(SOD)和丙二醛(MDA)水平。HE染色观察大鼠肝切片的病理学变化。免疫印迹法检测大鼠肝组织Nrf2/ARE信号通路相关蛋白的表达。结果与Normal组相比,Model组大鼠血清ALT、AST、TBIL、ALP、TBA水平均显著升高(P<0.05),Normal组炎症因子TNF⁃α、IFN⁃γ、IL⁃1β和肝组织MDA水平[分别为(248.26±27.64)pg/mL、(153.68±18.47)pg/mL、(189.53±23.21)pg/mL和(2.89±0.36)nmol/mg]均较Model组大鼠[分别为(53.47±8.69)pg/mL]、(24.72±2.94)pg/mL、(46.89±6.82)pg/mL和(1.05±0.14)nmol/mg]显著升高(P<0.05),Normal组肝组织SOD水平和Nrf2、ARE蛋白表达水平[分别为(53.18±8.77)U/mg、0.34±0.03、0.40±0.04]均较Model组大鼠[分别为(128.95±16.34)U/mg、0.87±0.09、0.94±0.09]显著降低(P<0.05);与Model组相比,GRg1⁃L组、GRg1⁃H组大鼠相关指标变化与上述相反(P<0.05)。Nrf2抑制剂减轻了GRg1对ICP大鼠的治疗作用。结论GRg1可能通过激活Nrf2/ARE信号通路对大鼠ICP具有一定的治疗作用。

Objective To investigate the effect of ginsenoside Rg1(GRg1)on intrahepatic cholestasis of pregnancy(ICP)in rats by regulating the nuclear factor erythroid⁃2⁃related factor 2(Nrf2)/antioxidant response ele⁃ment(ARE)signaling pathway.Methods Female pregnant SD rats were randomly separated into a Normal group,an ICP model group(Model group),a low⁃dose GRg1 group(GRg1⁃L group,20 mg/kg GRg1),a high⁃dose GRg1 group(GRg1⁃H group,40 mg/kg GRg1),and a high⁃dose GRg1+Nrf2 inhibitor all trans retinoic acid(ATRA)group(GRg1⁃H+ATRA group,40 mg/kg GRg1+10 mg/kg ATRA),with 12 rats in each group.The combination in⁃jection of estradiol benzoate and progesterone was used to establish a rat ICP model.After administration,the ELISA method was applied to detect the levels of total bilirubin(TBIL),alanine aminotransferase(ALT),aspartate amino⁃transferase(AST),alkaline phosphatase(ALP),total bile acid(TBA),tumor necrosis factor⁃α(TNF⁃α),inter⁃feron⁃γ(IFN⁃γ),interleukin⁃1β(IL⁃1β)in rat serum,and superoxide dismutase(SOD)and malondialdehyde(MDA)in liver tissue.HE staining was applied to observe the pathological changes in rat liver tissue slices.Western Blot was applied to detect the expression of Nrf2/ARE signaling pathway related proteins in rat liver tissue.Results Compared with the normal group,the serum ALT,AST,TBIL,ALP and TBA levels in the model group were obvi⁃ously increased(P<0.05).The inflammatory factors TNF⁃α,IFN⁃γ,IL⁃1βand liver tissue MDA level in the nor⁃mal group[(248.26±27.64)pg/mL,(153.68±18.47)pg/mL,(189.53±23.21)pg/mL and(2.89±0.36)nmol/mg,re⁃spectively]were significantly higher than those in the model group rats[(53.47±8.69)pg/mL,(24.72±2.94)pg/mL,(46.89±6.82)pg/mL and(1.05±0.14)nmol/mg,respectively](P<0.05).The SOD level,Nrf2 and ARE protein ex⁃pression levels in liver tissue of normal group[(53.18±8.77)U/mg,0.34±0.03 and 0.40±0.04,respectively]were significantly lower than those in the model group rats[(128.95±16.34)U/mg,0.87±0.09 and 0.94±0.09,respective⁃ly](P<0.05).Compared with the Model group,the changes in relevant indicators in the GRg1⁃L and GRg1⁃H groups were opposite to the above(P<0.05).Nrf2 inhibitors reduced the therapeutic effect of GRg1 on ICP rats.Conclu⁃sion GRg1 may have a certain therapeutic effect on rat with ICP by activating the Nrf2/ARE signaling pathway.