复方丹参片通过LKB1-AMPK通路干预高脂血症大鼠的作用机制研究

Protective Effect of Compound Danshen Tablets on Hyperlipidemic Rats by Regulating LKB1-AMPK Signal Pathway

目的基于LKB1-AMPK通路探讨复方丹参片对高脂血症大鼠的保护作用及相关机制。方法采用腹腔注射75%蛋黄乳液+连续饲喂高脂饲料4周的方式制备高脂血症大鼠模型。将大鼠随机分为正常组、模型组、辛伐他汀组(0.02 g·kg^(-1))和复方丹参低、中、高剂量组(复方丹参片,1.5、3、6 g·kg^(-1)),每组10只。各组大鼠每日1次给予相应剂量药物灌胃,连续干预4周。采用ELISA法检测血清甘油三酯(TG)、总胆固醇(T-CHO)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)、丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)水平;测定大鼠肝脏系数;采用HE染色法观察肝脏组织病理形态;采用免疫组织化学法检测肝脏组织中AMPK、p-AMPK、LKB1、HMGCR蛋白水平;采用Real-time PCR法检测肝脏组织中AMPK、LKB1、HMGCR mRNA表达水平。结果与正常组比较,模型组大鼠的肝脏系数及血清TG、T-CHO、LDL-C、ALT、AST水平明显升高(P<0.05),HDL-C水平明显降低(P<0.05);肝脏出现脂肪性病变;肝脏组织中AMPK、p-AMPK、LKB1蛋白及mRNA表达水平明显降低(P<0.05),HMGCR蛋白及mRNA表达水平明显升高(P<0.05)。与模型组比较,复方丹参片低、中、高剂量能明显降低高脂血症模型大鼠的肝脏系数及血清TG、T-CHO、LDL-C、ALT、AST水平(P<0.05),升高HDL-C水平(P<0.05);改善肝脏脂肪性病变;升高肝脏组织中AMPK、p-AMPK、LKB1蛋白及mRNA表达水平(P<0.05),降低HMGCR蛋白及mRNA表达水平(P<0.05)。结论复方丹参片可能通过激活LKB1-AMPK通路及抑制HMGCR表达水平来实现对高脂血症大鼠的保护作用。

Objective To study the protective effect of compound Danshen tablets on hyperlipidemic rats and its mechanism based on LKB1-AMPK signal pathway.Methods Hyperlipidemic rat model was induced by intraperitoneal injection of 75% solution of egg yolk and feeding high fat food.Rats were randomly divided into normal group,model group,simvastatin group and compound Danshen group(1.5,3,6 g·kg^(-1)).Animals in each group were given the corresponding dose of drugs daily by gavage,and the intervention was continued for 4 weeks.The serum levels of TG,T-CHO,LDL-C,HDL-C,ALT and AST were measured by ELISA.Liver weight was measured and liver index was calculated.HE staining was applied to observe the pathological morphology of liver tissues.Immunohistochemical method was used to detect the protein expression levels of AMPK,p-AMPK,LKB1 and HMGCR in liver tissues.RT-PCR was used to detect the mRNA levels of AMPK,LKB1 and HMGCR in liver tissues.Results Compared with normal group,there was extensive steatosis in the rats’livers of the model group;the liver index and the levels of TG,T-CHO,LDL-C,ALT,AST were significantly higher(P<0.05)and the level of HDL-C were significantly reduced(P<0.05)in the model group;the levels of AMPK,p-AMPK,LKB1protein and mRNA were significantly reduced(P<0.05) and the levels of HMGCR protein and mRNA were significantly higher(P<0.05)in the model group(P<0.05).In the compound Danshen groups(1.5,3,6 g·kg^(-1)),compound Danshen tablets significantly reduced the liver indexes and the levels of TG,T-CHO,LDL-C,ALT,AST(P<0.05),improved the levels of HDL-C in rats(P<0.05),and improved the pathological changes of liver tissues.It was also found that the levels of AMPK,p-AMPK,LKB1 protein and mRNA were significantly higher(P<0.05),and the levels of HMGCR protein and mRNA were significantly reduced(P<0.05) in liver tissues.Conclusion Compound Danshen tablets had the protective effect on hyperlipidemia rats,and its mechanism may be related to the activation of LKB1-AMPK and down-regulating the expression of HMGCR.