【摘要】 目的：探讨温运清利法对非酒精性脂肪性肝病（nonalcoholic fatty liver disease, NAFLD）小鼠肝脏自噬水平的调控作用。方法：将30只小鼠随机分为空白组、模型组、苓桂术甘汤组(5.005 g·kg^-1)、茵陈蒿汤组(5.005 g·kg^-1)、茵陈苓桂术甘汤组(10.01 g·kg^-1)及易善复组(207.48 mg·kg^-1)。除空白组外，其余小鼠喂养高脂饲料10周从而建立NAFLD模型。造模成功后，各组小鼠以相应药物灌胃，每日1次，连续4周。检测小鼠血清谷草转氨酶（aspartate transaminase, AST）、谷丙转氨酶（alanine aminotransferase, ALT）、总胆固醇（total cholesterol, TC）、甘油三酯（triglyceride, TG）、高密度脂蛋白胆固醇（high-density lipoprotein cholesterol, HDL-C）、低密度脂蛋白胆固醇（low-density lipoprotein cholesterol, LDL-C）含量。HE染色观察小鼠肝脏病理形态，RT-qPCR法与Western blot法检测肝组织微管相关蛋白轻链3（microtubule-associated protein light chain 3,LC3）、自噬相关蛋白5（autophagy-related protein 5,ATG5）、p62表达水平，透射电镜检测肝细胞内自噬小体、自噬溶酶体数量。结果：HE染色显示，与空白组比较，模型组小鼠肝细胞结构紊乱，伴见脂肪空泡和炎症细胞浸润。与模型组比较，各个药物组小鼠肝细胞内脂肪空泡数量减少，炎症细胞浸润减轻。与空白组比较，模型组小鼠血清ALT、AST、TC、TG、LDL-C含量升高，HDL-C含量降低（P<0.01）。与模型组比较，各个药物组小鼠血清ALT、AST、TC、TG、LDL-C含量降低（P<0.05）。与模型组比较，温运清利法治疗组小鼠血清HDL-C含量升高（P<0.05）。Western blot显示，与空白组比较，模型组小鼠肝脏LC3B-Ⅱ、ATG5表达水平降低（P<0.01）。与模型组比较，温运清利法治疗组小鼠肝脏LC3B-Ⅱ、ATG5表达水平升高（P<0.05）。RT-qPCR显示，与空白组比较，模型组小鼠肝脏ATG5 mRNA水平降低，p62 mRNA水平升高（P<0.05）。与模型组比较，温运清利法治疗组小鼠肝脏LC3 mRNA、ATG5 mRNA水平升高，p62 mRNA水平降低（P<0.05）。透射电镜显示，与空白组比较，模型组小鼠肝脏自噬小体、自噬溶酶体数量减少，温运清利法治疗后增加。结论：温运清利法可发挥对NAFLD小鼠的肝脏保护作用，其机制可能与调控肝脏自噬水平有关。更多还原

【Abstract】 Objective: To investigate the regulatory effect of Wen Yun Qing Li method on the level of hepatic autophagy in mice with nonalcoholic fatty liver disease（NAFLD）.Methods: Thirty mice were randomly divided into blank group, model group, Linggui Zhugan Decoction group(5.005 g·kg^-1),Yinchenhao Decoction group(5.005 g·kg^-1),Yinchen Linggui Zhugan Decoction group(10.01 g·kg^-1) and Essentiale group(207.48 mg·kg^-1).Except for the blank group, all mice were fed high-fat chow for 10 weeks to establish the NAFLD model.After successful modeling, mice in each group were gavaged with appropriate drugs once daily for 4 weeks.Serum aspartate transaminase（AST）,alanine aminotransferase（ALT）,total cholesterol（TC）,triglyceride（TG）,high-density lipoprotein（HDL）,and cholesterol（HDL） were measured.Total cholesterol（TC）,triglyceride（TG）,high-density lipoprotein cholesterol（HDL-C）,and low-density lipoprotein cholesterol（LDL-C） were detected with HE staining, and the pathological morphology of liver tissue was examined with RT-qPCR and Western blotting.The expression levels of LC3,ATG5 and p62 were detected with RT-qPCR and Western blot, and the numbers of autophagosomes and autophagolysosomes were detected with transmission electron microscopy in hepatocytes.Results: With HE staining it was showed that compared with the blank group, the hepatocytes in the model group were structurally disordered, accompanied by fat vacuoles and inflammatory cell infiltration.Compared with the model group, the number of fat vacuoles in the hepatocytes of mice in each drug group was reduced, and the inflammatory cell infiltration was alleviated.Compared with the blank group, mice in the model group had elevated serum ALT,AST,TC,TG,and LDL-C levels, and reduced HDL-C levels（P<0.01）.Compared with the model group, the serum ALT,AST,TC,TG,LDL-C levels of mice in each drug group were decreased（P<0.05）.Compared with the model group, the serum HDL-C content of mice in the Wen Yun Qing Li method treatment group was increased（P<0.05）.Western blot showed that compared with the blank group, the expression levels of hepatic LC3B-Ⅱ and ATG5 in mice in the model group were decreased（P<0.01）.Compared with that of the model group, the hepatic LC3B-Ⅱ and ATG5 expression levels of mice in the Wen Yun Qing Li method treatment group were elevated（P<0.05）.With RT-qPCR it was showed that the hepatic ATG5 mRNA level of mice in the model group was decreased and the p62 mRNA level of mice in the model group was increased compared with that of the blank group（P<0.05）.Compared with that of the model group, the hepatic LC3 mRNA and ATG5 mRNA levels of mice in the Wen Yun Qing Li method treatment group were increased, and the p62 mRNA level was decreased（P<0.05）.With transmission electron microscopy it was showed that the number of autophagosomes and autolysosomes in the livers of mice in the model group decreased compared with that in the blank group and increased after treatment with Wen Yun Qing Li method.Conclusion: Wen Yun Qing Li method can exert hepatoprotective effects on NAFLD mice, and its mechanism may be related to the regulation of hepatic autophagy level.更多还原