摘要
目的探讨补骨脂的反成配伍对大鼠肝脏的减毒作用及作用机制。方法80只SD大鼠采用随机数字表法分为空白组、模型组、补骨脂组、补骨脂+制何首乌1∶1组、补骨脂+制何首乌1∶2组、补骨脂+制何首乌2∶1组、补骨脂+熟地黄1∶1组、补骨脂+五味子1∶1组,每组10只大鼠。除空白组外,其他组皮下注射氢化可的松(25 mg/kg)制备肾阳虚大鼠模型,每日1次,连续注射14 d。大鼠建立肾阳虚模型后,给予相应药物干预,各组给药剂量均为12.6 g/kg,连续灌胃4周。HE染色观察大鼠肝脏组织病理变化。酶联免疫吸附测定检测大鼠血清中谷丙转氨酶(ALT)、谷草转氨酶(AST)、碱性磷酸酶(ALP)、直接胆红素(DBIL)和总胆红素(TBIL)水平,以及肝组织中超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽过氧化物酶(GSH-Px)表达水平。实时荧光PCR检测核因子E2相关因子2(Nrf2)、Kelch样环氧氯丙烷相关蛋白1(Keap1)、血红素加氧酶-1(HO-1)、醌氧化还原酶1(NQO1)mRNA的表达。蛋白质印迹法检测AMP活化蛋白激酶(AMPK)/糖原合成酶激酶-3β(GSK-3β)/Nrf2信号通路中磷酸化AMPK(p-AMPK)、磷酸化GSK-3β(p-GSK-3β)、Nrf2、HO-1的蛋白表达。结果与补骨脂组比较,补骨脂与补阴药制何首乌、熟地黄及五味子配伍后大鼠肝小叶结构改善,炎性浸润细胞减少。与补骨脂组比较,补骨脂+制何首乌1∶2组、补骨脂+熟地黄1∶1组、补骨脂+五味子1∶1组大鼠血清中AST、ALT、ALP、DBIL、TBIL水平均降低(P<0.01);补骨脂+制何首乌1∶2组大鼠肝组织SOD、GSH-Px、MDA的表达改善(P<0.01);补骨脂+制何首乌1∶2组Nrf2、HO-1、NQO1 mRNA水平上调(P<0.01),Keap1 mRNA水平下调(P<0.01);补骨脂+制何首乌1∶2组、补骨脂+制何首乌2∶1组、补骨脂+熟地黄1∶1组、补骨脂+五味子1∶1组p-AMPK、p-GSK-3β、Nrf2、HO-1蛋白表达均升高(P<0.05,P<0.01)。结论补骨脂配伍补阴药制何首乌、熟地黄及五味子可改善补骨脂所致的肝损伤,其作用机制可能是通过激活AMPK/GSK-3β/Nrf2信号通路,抑制氧化应激状态来发挥对肝脏的保护作用,补骨脂与何首乌配伍比例1∶2减毒效果最佳。
Objective To investigate the attenuating effect of reverse compatibility of psoralea corylifolia on rat liver and the underlying mechanism.Methods According to the random number table method,80 SD rats were divided into the blank group,the model group,the psoralea corylifolia group,the psoralea corylifolia+polygonum multiflorum 1∶1 group,the psoralea corylifolia+polygonum multiflorum 1∶2 group,the psoralea corylifolia+polygonum multiflorum 2∶1 group,the psoralea corylifolia+rehmannia glutinosa 1∶1 group,the psoralea corylifolia+schisandra chinensis 1∶1 group,with 10 rats in each group.Rats in all groups except for the blank group were subcutaneously injected with hydrocortisone(25 mg/kg)once a day for 14 days to establish a rat model of kidney yang deficiency.After the model of kidney yang deficiency was established,the rats were given corresponding drugs for 4 weeks;the drug dose was 12.6 g/kg in each group.HE staining was used to observe the pathological changes of rat liver tissue.The serum levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),alkaline phosphatase(ALP),direct bilirubin(DBIL)and total bilirubin(TBIL)and the expression levels of superoxide dismutase(SOD),malondialdehyde(MDA),and glutathione peroxidase(GSH⁃Px)in liver tissue were detected by ELISA.The mRNA expression levels of NF⁃E2⁃related factor 2(Nrf2),Kelch⁃like epichlorohydrin⁃related protein 1(Keap1),heme oxygenase⁃1(HO⁃1),and quinone oxidoreductase NADH1(NQO1)were detected by real⁃time PCR.The protein expression levels of phosphorylated AMPK(p⁃AMPK),phosphorylated GSK⁃3β(p⁃GSK⁃3β),Nrf2,and HO⁃1,which were involved in the AMPK/GSK⁃3β/Nrf2 signaling pathway,were determined by Western blotting.Results Compared with the psoralea corylifolia group,the structure of hepatic lobules in rats was improved and the inflammatory infiltration cells were reduced after the compatibility of psoralea corylifolia and medicinals for replenishing yin such as polygonum multiflorum,rehmannia glutinosa and schisandra chinensis.Compared with the psoralea corylifolia group,the serum levels of AST,ALT,ALP,DBIL,and TBIL of rats in the psoralea corylifolia+polygonum multiflorum 1∶2 group,the psoralea corylifolia+rehmannia glutinosa 1∶1 group,the psoralea corylifolia+schisandra chinensis 1∶1 group were reduced(P<0.01);the expression levels of SOD,GSH⁃Px and MDA in liver tissue of rats were improved in psoralea corylifolia+polygonum multiflorum 1∶2 group(P<0.01);the mRNA levels of Nrf2,HO⁃1,and NQO1 were up⁃regulated(P<0.01)and the mRNA level of Keap1 was down⁃regulated(P<0.01)in the psoralea corylifolia+polygonum multiflorum 1∶2 group;the protein expression levels of p⁃AMPK,p⁃GSK⁃3β,Nrf2,and HO⁃1 were increased in the psoralea corylifolia+polygonum multiflorum 1∶2 group,the psoralea corylifolia+polygonum multiflorum 2∶1 group,the soralea corylifolia+rehmannia glutinosa 1∶1 group,and the soralea corylifolia+schisandra chinensis 1∶1 group(P<0.05,P<0.01).Conclusion The compatibility of psoralea corylifolia and medicinals for replenishing yin polygonum multiflorum,rehmannia glutinosa and schisandra chinensis can improve the liver injury caused by psoralea corylifolia.The mechanism may be to activate the AMPK/GSK⁃3β/Nrf2 signaling pathway and inhibit the oxidative stress state to play a protective role in the liver.The compatibility ratio of psoralea corylifolia and polygonum multiflorum 1∶2 has the best attenuation effect.
作者
王婧
卫文鑫
李星雨
陈思凯
张萌萌
李瑶
董泰玮
高峰
李敏
奚苗苗
谢允东
卫培峰
WANG Jing;WEI Wenxin;LI Xingyu;CHEN Sikai;ZHANG Mengmeng;LI Yao;DONG Taiwei;GAO Feng;LI Min;XI Miaomiao;XIE Yundong;WEI Peifeng(Shaanxi University of Chinese Medicine,Xianyang 712046,China;Queen Mary School,Nanchang University,Nanchang 330031,Journal of Beijing University of Traditional Chinese Medicine China;Second Affiliated Hospital of Shaanxi University of Chinese Medicine,Xianyang 712021,China)
出处
《北京中医药大学学报》
CAS
CSCD
北大核心
2023年第5期665-675,共11页
Journal of Beijing University of Traditional Chinese Medicine
基金
国家自然科学基金项目(No.82204790)
陕西省“特支计划”人才项目
中医药“双链融合”中青年科研创新团队建设项目(No.2022-SLRH-YQ-008)
陕西中医药大学第二附属医院学科创新团队建设项目(No.2020XKTD-A04)。
