温运清利法对非酒精性脂肪性肝病小鼠肝脏自噬水平的调节作用  

Modulation Effect of Wen Yun Qing Li Method on Hepatic Autophagy Level in Mice with Nonalcoholic Fatty Liver Disease

作　　者：袁亚利 谢春娥 毛堂友 张文基 卢心毓 王木源 梁承涛 林政道 李一桐 焦瑶 韩海啸[1] YUAN Yali;XIE Chun′e;MAO Tangyou;ZHANG Wenji;LU Xinyu;WANG Muyuan;LIANG Chengtao;LIN Zhengdao;LI Yitong;JIAO Yao;HAN Haixiao(Dongfang Hospital,Beijing University of Chinese Medicine,Beijing China 100078;Hebei North University,Zhangjiakou Hebei,075000;Shenzhen Bao′an Traditional Chinese Medicine Hospital,Shenzhen Guangdong China 518100)

机构地区：[1]北京中医药大学东方医院,北京100078 [2]河北北方学院,河北张家口075000 [3]深圳市宝安区中医院,广东深圳518100

出　　处：《中医学报》2024年第12期2625-2631,共7页Acta Chinese Medicine

基　　金：国家自然科学基金项目(82074344);中华中医药学会青年人才托举工程项目(CACM-2022-QNRC2-A02);北京中医药大学岐黄英才·优秀青年科技人才培育计划项目(K2023A01)。

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,p6Objective: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 w

关 键 词：非酒精性脂肪性肝病 温运清利法 自噬 苓桂术甘汤 茵陈蒿汤 茵陈苓桂术甘汤 肝脏 小鼠 

分 类 号：R285.5[医药卫生—中药学]