基于AMPK/GSK-3β/Nrf2信号通路探讨补骨脂相反相成配伍减毒机制
Detoxification mechanism of psoralea corylifolia reverse compatibility based on the AMPK/GSK-3β/Nrf2 signaling pathway
- 北京中医药大学学报 2023年46卷第5期 页码：665-675
- 作者机构：
1.陕西中医药大学 咸阳 712046
2.南昌大学玛丽女王学院
3.陕西中医药大学第二附属医院
- 作者简介：
王婧，女，在读硕士生
# 卫培峰，男，博士，教授，硕士生导师，主要研究方向：中药新药与中药毒性的临床及实验研究，E-mail：weipeifeng@163.com
- 基金信息：国家自然科学基金项目(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(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(2020XKTD-A04)
DOI：10.3969/j.issn.1006-2157.2023.05.012
中图分类号： R285.5纸质出版日期：2023-05-30，
网络出版日期：2023-03-03，
收稿日期：2022-10-13，
- 稿件说明：
移动端阅览
王婧, 卫文鑫, 李星雨, 等. 基于AMPK/GSK-3β/Nrf2信号通路探讨补骨脂相反相成配伍减毒机制[J]. 北京中医药大学学报, 2023,46(5):665-675.
WANG Jing, WEI Wenxin, LI Xingyu, et al. Detoxification mechanism of psoralea corylifolia reverse compatibility based on the AMPK/GSK-3β/Nrf2 signaling pathway[J]. Journal of Beijing University of Traditional Chinese Medicine, 2023,46(5):665-675.
王婧, 卫文鑫, 李星雨, 等. 基于AMPK/GSK-3β/Nrf2信号通路探讨补骨脂相反相成配伍减毒机制[J]. 北京中医药大学学报, 2023,46(5):665-675. DOI： 10.3969/j.issn.1006-2157.2023.05.012.
WANG Jing, WEI Wenxin, LI Xingyu, et al. Detoxification mechanism of psoralea corylifolia reverse compatibility based on the AMPK/GSK-3β/Nrf2 signaling pathway[J]. Journal of Beijing University of Traditional Chinese Medicine, 2023,46(5):665-675. DOI： 10.3969/j.issn.1006-2157.2023.05.012.
摘要
目的
2
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
2
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
2
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
2
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。
Abstract
Objective
2
To investigate the attenuating effect of reverse compatibility of psoralea corylifolia on rat liver and the underlying mechanism.
Methods
2
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
2
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
2
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.
关键词
补骨脂反成配伍肝毒性AMPK/GSK-3β/Nrf2信号通路大鼠
Keywords
psoralea corylifoliareverse compatibilityhepatotoxicityAMPK/GSK-3β/Nrf2 signaling pathwayrats
references
国家药典委员会. 中华人民共和国药典：2020年版[M]. 一部．北京：中国医药科技出版社，2020: 195.
叶伊琳，唐洁，刘琴，等．复方补骨脂搽剂对豚鼠白癜风模型作用的实验研究[J].北京中医药大学学报，2014，37(4)：252－254.
唐进法，牛璐，王晓艳，等．补骨脂醇提物中8种成分在正常和脂多糖模型大鼠体内的组织分布比较研究[J].中国中药杂志，2022，47(24)：6763－6779.
王娟，周植星，杨莉，等．补骨脂药材UPLC指纹图谱建立及12种主要成分含量测定[J].中草药，2021，52(2)：552－557.
高姝燕，麦丽克姆·麦图如孜，王春，等．基于免疫应激的补骨脂特异质肝毒性作用[J].中国药理学通报，2022，38(12)：1860－1868.
张玥，毕亚男，袁晓美，等．基于靶器官的补骨脂肝毒性成分探讨[J].时珍国医国药，2017，28(8)：1844－1847.
吴晓燕，赫俊杰，阎星旭，等．基于“有故无殒”的补骨脂对不同肾虚证模型大鼠肝脏功能的影响[J].中草药，2023，54(2)：434－441.
贾亚玲，罗俊刚，冯泳．方剂反成配伍纲纪探要[J].中国民族民间医药，2015，24(20)：36－37.
杨军，屈杰，陈丽名，等．《金匮要略》相反相成配伍法及其临床运用初探[J].中国中医基础医学杂志，2022，28(6)：840－843.
孙伟杰. 不同鹿茸加工品的化学成分及生物活性的比较研究[D]. 长春：长春中医药大学，2020.
马超，叶鸿博，林喆，等.4种肾阳虚模型的评价[J].吉林中医药，2019，39(4)：508－511.
张艳春，樊巧玲．方剂相反相成配伍减毒的理论探讨[J].中医药导报，2008，14(9)：13－15.
汪倩，卢鸽，解子婧，等．艾灸对卵巢储备功能减退大鼠Nrf2/HO-1信号通路的影响[J].中国针灸，2021，41(1)：53－58.
李娜，钟赣生，张晨，等．海藻玉壶汤中海藻不同品种与甘草加减应用对甲状腺肿大大鼠氧化应激及肝脏Nrf2/HO-1通路的影响[J].北京中医药大学学报，2019，42(3)：190－197.
汪倩，卢鸽，李红晓，等．基于Nrf2/HO-1信号通路探讨艾灸对卵巢储备功能减退大鼠的炎性调控机制[J].北京中医药大学学报，2021，44(3)：280－288.
LU MC, JI JA, JIANG ZY, et al. The Keap1－Nrf2－ARE pathway as a potential preventive and therapeutic target: An update [J]. Med Res Rev, 2016, 36(5)：924－963.
BAIRD L, YAMAMOTO M. The molecular mechanisms regulating the KEAP1－NRF2 pathway [J/OL]. Mol Cell Biol, 2020, 40(13): e00099－20 [2022－09－21]. https：//journals.asm.org/doi/10.1128/MCB.00099－20https：//journals.asm.org/doi/10.1128/MCB.00099－20.
BELLEZZA I, GIAMBANCOI I, MINELLI A, et al. Nrf2－Keap1 signaling in oxidative and reductive stress [J]. Biochim Biophys Acta Mol Cell Res, 2018, 1865(5)：721－733.
CARLING D. AMPK signalling in health and disease [J]. Curr Opin Cell Biol, 2017, 45：31－37.
HERZIG S, SHAW RJ. AMPK: guardian of metabolism and mitochondrial homeostasis [J]. Nat Rev Mol Cell Biol, 2018, 19(2)：121－135.
HARDIE DG, SCHAFFER BE, BRUNET A. AMPK: An energy-sensing pathway with multiple inputs and outputs [J]. Trends Cell Biol, 2016, 26(3)：190－201.
CHEN L, SHI X, XIE J，et al. Apelin－13 induces mitophagy in bone marrow mesenchymal stem cells to suppress intracellular oxidative stress and ameliorate osteoporosis by activation of AMPK signaling pathway [J]. Free Radic Biol Med, 2021, 163：356－368.
CHEN L, LIN X, FAN X，et al. Sonchus oleraceus Linn extract enhanced glucose homeostasis through the AMPK/Akt/GSK-3β signaling pathway in diabetic liver and HepG2 cell culture [J/OL]. Food Chem Toxicol, 2020, 136: 111072 [2022－09－21]. https：//linkinghub.elsevier.com/retrieve/pii/S0278691519308622https：//linkinghub.elsevier.com/retrieve/pii/S0278691519308622.
LV H, HONG L, TIAN Y, et al. Corilagin alleviates acetaminophen-induced hepatotoxicity via enhancing the AMPK/GSK3β-Nrf2 signaling pathway [J/OL]. Cell Commun Signal, 2019, 17(1): 2 [2022－09－21]. https：//biosignaling.biomedcentral.com/articles/10.1186/s12964－018－0314－2https：//biosignaling.biomedcentral.com/articles/10.1186/s12964－018－0314－2.
逯明蕾．GSK3β调控非Keap1依赖的Nrf2抗氧化反应参与急性肾损伤到慢性肾脏病转变的研究[D].郑州：郑州大学，2018.
张哲，汪祺，陈建华，等．肉桂多次ig给药在肾阳虚大鼠体内的积累和代谢分析[J].药物评价研究，2022，45(12)：2450－2457.
杜江，李楠，王和鸣．肾虚模型造模方法及相关指标[J].中国组织工程研究与临床康复，2010，14(50)：9433－9436.
肖静，何立群，高建东，等．腺嘌呤与氢化可的松大鼠肾阳虚模型造模方法比较[J].中国比较医学杂志，2008，18(3)：77－80.
李耿，张喆，尹西拳，等．地黄丸类方对肾阳虚大鼠HPA轴的影响[J].中药新药与临床药理，2015，26(3)：320－324.
李苗，曾梦楠，张贝贝，等．麻黄水煎液及拆分组分对肾阳虚水肿大鼠的影响[J].中国实验方剂学杂志，2017，23(23)：91－96.
0
浏览量
26
下载量
0
CSCD
- 网络PDF下载
- BibTeX下载
- Endnote下载
- RefMan 下载
- NoteExpress下载
- NoteFirst下载
关联资源
相关文章
相关作者
相关机构
- 地址：北京市朝阳区北三环东路11号 邮编：100029
- 联系电话：(010)53911857 Email：jbutcm@bucm.edu.cn
- 技术支持由北京北大方正电子有限公司提供 京ICP备09064830号-19
京公网安备11010802024621 - 本系统建议在Chrome、 IE9+ 以上版本浏览器阅读本站内容，360浏览器请切换至极速模式
- Cookies帮助我们提供服务并提供个性化体验。使用本网站，即表示您同意我们使用Cookies