中药清肠利肝方对肝细胞自噬的影响及其对小鼠急性肝损伤的保护作用研究

Effect of Qingchang Ligan Formula on hepatocyte autophagy and its protection on acute liver failure in mice

目的探讨中药组方清肠利肝方(QCLG)对D-氨基半乳糖/脂多糖(D-GalN/LPS)所致急性肝衰竭(ALF)小鼠肝细胞自噬的影响。方法将野生型健康C57BL/6雄性小鼠分为正常对照组、ALF模型组、QCLG干预组和QCLG+自噬抑制剂3-MA干预组。ALF模型组:腹腔注射D-GalN/LPS建立ALF模型;QCLG干预组:D-GalN/LPS给药前3天给予QCLG灌胃,1次/日;QCLG+3-MA干预组:D-GalN/LPS给药前3天给予QCLG灌胃,1次/日,D-GalN/LPS给药前24 h尾静脉给予3-MA干预。各组于D-GalN/LPS给药6 h后收集小鼠肝脏组织和血清,检测小鼠血清ALT、AST,免疫印迹、实时荧光定量PCR检测小鼠肝组织自噬水平;抑制自噬后观察各组小鼠死亡率和肝脏损伤。结果与正常对照组比较,ALF模型组ALT、AST水平升高,自噬水平下降;QCLG干预组ALT、AST水平较ALF模型组显著降低,自噬水平升高,差异有统计学意义(P<0.05);抑制自噬后,QCLG的保护性作用被逆转,小鼠死亡率升高,肝脏损伤加重。结论 QCLG可促进D-GalN/LPS所致ALF小鼠肝细胞自噬水平,减轻肝细胞损伤。

Objective To investigate the effect of Qingchang Ligan Formula on hepatocyte autophagy in acute liver failure(ALF) of mice induced by D-galactosamine/lipopolysaccharide.Methods Wild type of healthy C57 BL/6 male mice were randomly divided into normal control group,ALF model group,Qingchang Ligan Formula intervention group,and 3-MA autophagy inhibitor intervention group.ALF model group:ALF model was induced by intraperitoneal injection with D-GalN/LPS;Qingchang Ligan Formula intervention group:3 days before D-GalN/LPS,Qingchang Ligan Formula was given with lavage,once a day;3-MA intervention group:3 days before D-GalN/LPS,Qingchang Ligan Formula was given with lavage,once a day;24 h before D-GalN/LPS,3-MA was given through caudal vein.6 h after D-GalN/LPS administration,the liver tissue and serum of each group were collected.Serum transaminase ALT and AST were assessed.Western blot and real-time fluorescent quantitative PCR were used to detect the expression of autophagy in liver tissue.The mortality and liver damage in mice were observed.Results Compared with the normal control group,the level of transaminase was increased and the level of autophagy decreased in the model group.In Qingchang Ligan Formula intervention group,the level of transaminase was significantly lowered than that of the model group,and the level of autophagy was increased,and the difference was statistically significant(P<0.05).After inhibiting autophagy,the protective effect of Qingchang Ligan Formula was reversed,and the mortality of mice increased and liver damage aggravated.Conclusion Qingchang Ligan Formula can promote the autophagy level of hepatocytes in mice with acute liver failure caused by D-GalN/LPS,and reduce liver cell damage.