当归芍药散联合TLR4抑制剂对大鼠肝纤维化的抑制作用及NF-κB/NLRP3通路的调节作用研究  

Study on Inhibitory Effect of Danggui Shaoyao San combined with TLR4 Inhibitor on Rat Liver Fibrosis and Regulation of NF-κB/NLRP3 Pathway

作　　者：王雯[1] 胡传国[1] 梁伟林 Wang Wen;Hu Chuanguo;Liang Weilin(People's Hospital of Xinjiang Uygur Autonomous Region,Urumqi 830001,China;Jingxi Medical District,PLA General Hospital,Beijing 100144,China)

机构地区：[1]新疆维吾尔自治区人民医院,乌鲁木齐830001 [2]解放军总医院京西医疗区,北京100144

出　　处：《世界科学技术-中医药现代化》2023年第3期1147-1154,共8页Modernization of Traditional Chinese Medicine and Materia Medica-World Science and Technology

基　　金：新疆维吾尔自治区卫生健康委员会自治区卫生健康青年医学科技人才专项科研项目(WJWY-202226):基于TLR4/NF-κB/NLRP3通路探讨当归芍药散对CCl4肝纤维化大鼠的作用机制,负责人:王雯

摘　　要：目的研究当归芍药散联合Toll样受体4(Toll-like receptor 4,TLR4)抑制剂对大鼠肝纤维化的抑制作用及对核因子κB(Nuclear factor kappa-B,NF-κB)/NOD样受体蛋白3(NOD-like receptor protein 3,NLRP3)通路的调节作用。方法大鼠随机分为空白对照组及造模组,造模组采用腹腔注射四氯化碳(CCl_(4))法建立肝纤维化大鼠模型,造模完成后的大鼠随机分为肝纤维化模型组、当归芍药散组、TLR4抑制剂组及联合组,12只/组,当归芍药散组大鼠按照10 g·kg^(-1)(以生药含量计)灌胃给予大鼠当归芍药散,TLR4抑制剂组腹腔注射给予大鼠TAK-242(3 mg·kg^(-1)),联合组同时给予大鼠当归芍药散(10 g·kg^(-1))及TAK-242(3 mg·kg^(-1)),1次/天,连续给药13周,测定大鼠体质量、肝脏重量及肝脏指数,使用全自动生化分析仪测定大鼠血清直接胆红素(Direct bilirubin,DBIL)、总胆汁酸(Total bile acid,TBA)及总胆红素(Total bilirubin,TBIL)水平,使用酶联免疫吸附法(ELISA)测定肝组织细胞间黏附分子-1(Intercellular cell adhesion molecule-1,ICAM-1)、超氧化物歧化酶(Superoxide dismutase,SOD)、丙二醛(Malondialdehyde,MDA)及还原型谷胱甘肽(Glutathione,GSH)水平,Masson染色法检查大鼠肝组织病理学变化,实时定量聚合酶链式反应(Real-time quantitative polymerase chain reaction,Rt-qPCR)测定肝组织NLPR3及NF-κB mRNA水平,免疫印迹法(Western blot)测定肝组织NLPR3及NF-κB蛋白水平,免疫荧光法测定肝组织α-SMA表达。结果与空白对照组比较,肝纤维化模型组大鼠肝脏质量、肝脏指数,血清DBIL、TBA及TBIL水平、肝组织ICAM-1及MDA水平、肝组织NLPR3及NF-κB mRNA及蛋白水平、肝组织α-SMA免疫荧光强度显著升高(P<0.05),体质量、肝组织SOD及GSH水平显著降低(P<0.05),肝组织明显纤维化改变;与肝纤维化模型组比较,当归芍药散组、TLR4抑制剂组及联合组大鼠肝脏质量、肝脏指数,血清DBIL、TBA及TBIL水平、肝组织ICAM-1及MDA水平Objective To study the inhibitory effects of Danggui Shaoyao San combined with TLR4 inhibitors on liver fibrosis in rats and the regulation of NF-κB/NLRP3 pathway.Methods Rats were randomly divided into a blank control group and model group.The model group was intraperitoneally injected with carbon tetrachloride(CCl_(4))to establish a rat model of liver fibrosis.After the model was completed,the rats were randomly divided into liver fibrosis model group,Danggui Shaoyao San group,TLR4 inhibitor group and combination group,12 rats/group,rats in the Danggui Shaoyao San group were given by intragastric administration at the dose of 10 g·kg^(-1)(based on crude drug content)of Danggui Shaoyao San,the TLR4 inhibitor group was intraperitoneally injected with TAK-242(3mg·kg^(-1)),and the combination group was simultaneously given Danggui Shaoyao San(10 g·kg^(-1))and TAK-242(3 mg·kg^(-1)),once a day,for 13 weeks,determination of rat body weight,liver weight and liver index,automatic biochemical analyzer was used to determine levels of serum DBIL,TBA and TBIL,and enzyme-linked immunosorbent assay(ELISA)to determine liver tissue ICAM-1.SOD,MDA and GSH levels,Masson staining method to detect the pathological changes of rat liver tissue,Real-time quantitative polymerase chain reaction(PCR)to detect liver tissue NLPR3 and NF-κB mRNA levels,Western blotting method was used to detect liver tissue NLPR3 and NF-κB protein levels,immunofluorescence method was used to determine the expression level ofα-SMA in liver tissue.Results Compared with the blank control group,the liver mass,liver index,serum DBIL,TBA and TBIL levels,liver tissue ICAM-1 and MDA levels,liver tissue NLPR3 and NF-κB mRNA and protein leves and the level of liver tissueα-SMA immunofluorescence in liver fibrosis model group were increased significantly compared with the blank control group(P<0.05),the body mass,liver tissue SOD and GSH levels were significantly reduced(P<0.05),liver tissue was significantly fibrotic;compared with liver fibrosis model group

关 键 词：当归芍药散 TLR4抑制剂 NF-κB/NLRP3通路 肝纤维化大鼠 

分 类 号：R285.5[医药卫生—中药学]