α-常春藤皂苷对小鼠急性肝损伤自噬相关蛋白表达的影响

Effect of α-hederin on the expression of autophagy related proteins in mice with acute liver injury

目的 研究α-常春藤皂苷(α-hederin)对四氯化碳(CCl4)致小鼠急性肝损伤自噬相关蛋白表达的影响。方法 按体重将ICR小鼠随机分为正常组(等体积2%吐温80溶液),模型组(急性肝损伤模型,等体积2%吐温80溶液),低、高2个剂量实验组(急性肝损伤模型,40,80 mg·kg^(-1)α-常春藤皂苷)和阳性对照组(急性肝损伤模型,150 mg·kg^(-1)联苯双酯)。酶法测定血清谷丙转氨酶(GPT)和谷草转氨酶(GOT)活力,酶联免疫吸附试验(ELISA)法测定小鼠血清中炎症因子肿瘤坏死因子α(TNF-α)和白细胞介素6(IL-6)的水平。苏木精-伊红染色(HE)观察组织病理变化。蛋白质印迹法检测肝组织中细胞自噬相关蛋白微管相关蛋白1轻链3β-Ⅱ(LC3B-Ⅱ)和血管内皮组织酵母自噬基因-1(Beclin-1)蛋白表达水平。结果 在CCl4所致急性肝损伤小鼠实验中,正常组、模型组、低剂量实验组、高剂量实验组和阳性对照组小鼠血清GPT酶活力分别为(10.12±3.86),(97.96±17.07),(48.76±22.23),(34.80±14.38)和(51.57±17.13)U·L^(-1);血清GOT酶活力分别为(14.56±3.13),(39.04±6.21),(27.93±6.66),(24.39±2.60)和(31.44±3.32)U·L^(-1);正常组、模型组、低剂量实验组、高剂量实验组和阳性对照组小鼠血清中TNF-α含量分别为(110.73±24.54),(187.55±28.08),(160.51±14.30),(121.96±20.50)和(114.90±24.65)pg·mL^(-1);这5组小鼠血清中IL-6量分别为(26.38±3.11),(42.01±5.47),(34.13±3.89),(28.62±5.15)和(29.62±4.25)pg·mL^(-1);正常组、模型组和低、高2个剂量实验组LC3BⅡ蛋白表达量分别为2.23±0.36,1.00±0.17,2.47±0.62和3.41±0.49;这4组Beclin-1蛋白表达量分别为1.51±0.14,1.00±0.03,1.23±0.10和1.38±0.12。上述指标,模型组与正常组相比,差异均有统计学意义(P<0.05,P<0.01);低、高2个剂量实验组与模型组相比,差异均有统计学意义(P<0.05,P<0.01)。结论 α-常春藤皂苷对小鼠急性肝损伤具有保护作用,其机制可能与抗炎症反应有关;α-常春藤皂苷可上调急性肝损伤自噬相关蛋白的表达,这可能是其对小鼠急性肝损伤保护作用的机制之一。

Objective The aim is to investigate the effects of α-hederin on the expression of autophagy-related proteins against carbon tetrachloride (CCl4)-induced acute liver injury in mice.Methods According to body weight,ICR mice were randomly divided into five groups:Normal group (equal volume of 2%Tween 80 solution),mode group (acute liver injury model,equal volume of 2%Tween 80 solution),experimental-L,-H groups (acute liver injury model,40,80 mg·kg^(-1)α-hederin) and positive control group (acute liver injury model,150 mg·kg^(-1)bifendate).The activities of glutamic-pyruvic transaminase (GPT) and glutamic-oxaloacetic transaminase (GOT) in serum were determined using commercialized kits.Serum levels of inflammatory cytokines tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) were determined by enzyme linked immunosorbent assay (ELISA) assay.hematoxylin-eosin staining (HE) staining was employed to assess the histopathological changes.The protein expression of microtubule-associated protein light chain 3β-Ⅱ(LC3B-Ⅱ) and Beclin 1 in liver tissue were analyzed by Western blot.Results The activities of serum GPT in normal group,model group,experimental-L group,experimental-H group and the positive control group were (10.12±3.86),(97.96±17.07),(48.76±22.23),(34.80±14.38),(51.57±17.13) U·L^(-1);the activities of serum GOT were(14.56±3.13),(39.04±6.21),(27.93±6.66),(24.39±2.60),(31.44±3.32) U·L^(-1);the content of serum TNF-α were(110.73±24.54),(187.55±28.08),(160.51±14.30),(121.96±20.50),(114.90±24.65) pg·m L^(-1);the content of serum IL-6 were (26.38±3.11),(42.01±5.47),(34.13±3.89),(28.62±5.15),(29.62±4.25) pg·m L^(-1).LC3B-Ⅱprotein levels of liver tissue in normal group,model group,experimental-L group and experimental-H group were 2.23±0.36,1.00±0.17,2.47±0.62,3.41±0.49;Beclin 1 protein levels were 1.51±0.14,1.00±0.03,1.23±0.10,1.38±0.12,respectively.There were statistically significant differences in the above indexes between the model group and the normal group (P<0.05,P<0.01).Compared with model group,there were statistically significant differences in experimental-L and experimental-H groups(P<0.05,P<0.01).Conclusion α-hederin harbors a hepatoprotective effect against acute liver injury induced by CCl4in mice and the mechanism may be related to anti-inflammatory response.α-hederin can up-regulate the expression of autophagy-related proteins in acute liver injury,which may be one of the protective mechanisms of α-hederin against acute liver injury in mice.