清毒汤对重症肝损伤大鼠肠上皮细胞自噬的影响

Effect of Qingdu Decoction on autophagy of intestinal epithelial cells in rats with severe liver injury

目的观察清毒汤对实验性重症肝损伤大鼠血清中内毒素及白细胞介素-17(interleukin-17,IL-17)及白细胞介素-23(interleukin-23,IL-23)和肠组织中自噬相关蛋白LC3、Beclin-1的影响。方法将SPF级Wistar雄性大鼠采用数字表法随机分为正常组、模型组、乳果糖组、中药组,除正常组外,其余各组每日灌胃给予12 mg/kg硫代乙酰胺(thioacetamide,TAA),持续12周,建立TAA致实验性重症肝损伤大鼠模型。第8周末,乳果糖组、中药组每日予以乳果糖1.6 g/kg、清毒汤5.95 g/kg灌胃治疗,连续4周后取材。HE染色观察肝与结肠组织病理变化,透射电镜观察肠上皮细胞中自噬体,酶联免疫吸附法检测大鼠血清中内毒素、IL-17、IL-23的含量,Western blotting法检测肠组织自噬相关蛋白LC3、Beclin-1表达。结果与正常组比较,模型组大鼠肝肠组织大量炎性浸润,自噬体较少,内毒素、IL-17、IL-23浓度显著升高,LC3蛋白及Beclin-1含量均显著下降(P<0.01);经清毒汤干预治疗后,炎性浸润明显减少,自噬体明显增多,内毒素、IL-17、IL-23浓度明显下降,且LC3Ⅱ/LC3I、Beclin-1蛋白含量均显著升高(P<0.01)。结论中药清毒汤可减轻TAA致肝损伤模型大鼠的肝、肠组织损伤,降低内毒素、IL-17及IL-23浓度,并提高自噬体数量及LC3、Beclin-1蛋白含量,说明清毒汤可能通过激活重症肝损伤大鼠的肠上皮细胞自噬,减少炎性因子的释放来降低内毒素的产生,从而减轻肝脏损伤。

Objective To observe the effects of Qingdu Decoction(QDD)on endotoxin,interleukin-17(IL-17),interleukin-23(IL-23)of serum,autophagy-related proteins LC3 and Beclin-1 in experimental rats with severe liver injury.Methods A rat model of severe liver injury induced by thioacetamide(TAA)was established.SPF Wistar male rats were randomly divided into normal group,model group,lactulose(LA)group and QDD group.Except the normal group,the other groups were given 12 mg/kg TAA daily for 12 weeks.At the end of the 8th week,LA group and QDD group were treated with lactulose 1.6 g/kg,Qingdu decoction 5.95 g/kg daily.After 4-week intervention,HE staining was used to observe the pathological changes of liver and intestinal tissues.Transmission electron microscopy was used to observe autophagosomes in intestinal epithelial cells.IL-17,IL-23 and endotoxin levels were measured by enzyme-linked immuno sorbent assay.The expression levels of autophagy-related proteins LC3 and Beclin-1 in intestinal tissues were detected by Western Blot(WB).Results Compared with the normal group,the liver and intestinal tissue of the model group was inflamed,less autophagosomes,and the concentrations of endotoxin,IL-17 and IL-23 were significantly increased,and the contents of LC3 protein and Beclin-1 were significantly decreased(P<0.01).After the intervention of the Qingdu decoction,the inflammatory infiltration was significantly relieved,autophagosomes have increased significantly,the concentrations of endotoxin,IL-17 and IL-23 were significantly decreased,and the contents of LC3Ⅱ/LC3I and Beclin-1 were significantly increased(P<0.01).Conclusion Qingdu Decoction can reduce intestinal damage,reduce the concentration of endotoxin,IL-17 and IL-23,and increase the number of autophagosomes and the content of LC3,Beclin-1 protein in rats with liver injury induced by TAA.It suggests that Qingdu Decoction may alleviate liver damage in rats with severe liver injury by promoting autophagy of intestinal epithelial cells and reducing endotoxin production and the release of inflammatory factors.