基于非靶向肾脏代谢组学技术研究大黄蛰虫丸改善大鼠肾脏衰老的多靶点作用  

Non-targeted renal metabolomics reveals multi-target effects of Dahuang Zhechong Pills on renal aging in rats

作　　者：刘思逸 刘文杰 涂玥 何伟明[3] 刘欣雨 顾奕旻 温开智 李成娟 韩超 万毅刚[5] 黄煌 LIU Si-yi;LIU Wen-jie;TU Yue;HE Wei-ming;LIU Xin-yu;GU Yi-min;WEN Kai-zhi;LI Cheng-juan;HAN Chao;WAN Yi-gang;HUANG Huang(School of Acupuncture-Moxibustion and Tuina&School of Health Preservation and Rehabilitation,Nanjing University of Chinese Medicine,Nanjing 210023,China;Jiangsu Provincial Traditional Chinese Medicine Technology Engineering Research Center of Health and Health Preservation,Nanjing 210023,China;Department of Nephrology,Jiangsu Province Hospital of Chinese Medicine,Nanjing 210029,China;First Clinical Medical College,Nanjing University of Chinese Medicine,Nanjing 210023,China;Department of Traditional Chinese Medicine,Nanjing Drum Tower Hospital,the Affiliated Hospital of Nanjing University Medical School,Nanjing 210008,China;International Jingfang Insititute,Nanjing University of Chinese Medicine,Nanjing 210023,China)

机构地区：[1]南京中医药大学针灸推拿学院·养生康复学院,江苏南京210023 [2]江苏省中医药健康养生技术工程研究中心,江苏南京210023 [3]江苏省中医院肾内科,江苏南京210029 [4]南京中医药大学第一临床医学院,江苏南京210023 [5]南京大学医学院附属鼓楼医院中医科,江苏南京210008 [6]南京中医药大学国际经方学院,江苏南京210023

出　　处：《中国中药杂志》2024年第11期3002-3011,共10页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金面上项目(82174472,82174294,82374364);江苏省自然科学基金面上项目(BK20211298,BK20231123);江苏省中医药科技发展计划项目(MS2021037);江苏省研究生科研与实践创新计划项目(KYCX232159);江苏高校优势学科建设工程项目(PAPD);南京中医药大学自然科学基金项目(XZR2021087);南京市名中医专家工作室建设项目;南京鼓楼医院中西医协同特色技术发展项目(CZXM2022098)。

摘　　要：观察传统经方“大黄蛰虫丸(DHZCP)”改善肾脏衰老的效果,并探究其潜在的多靶点作用。将大鼠随机分为正常组、模型组、DHZCP组和维生素E(VE)组。首先建立D-半乳糖(D-gal)诱导的大鼠肾脏衰老模型,而后在造模同期,每天分别给予4组大鼠双蒸水、DHZCP混悬液和VE混悬液。在药物干预第60天末,检测大鼠肾脏衰老和损伤的各项指标,包括肾功能,肾脏组织病理学特征,肾脏衰老相关β-半乳糖苷酶(SA-β-gal)染色特征,肾组织Klotho、细胞周期阻滞蛋白和衰老相关分泌表型(SASP)蛋白表达水平。此外,对于大鼠的肾组织分别进行非靶向代谢组学特征分析,筛选潜在生物标志物,富集代谢通路。最后,进行关键作用靶点信号通路的初步验证。结果显示,DHZCP和VE能明显改善衰老模型鼠的肾功能、肾小管/间质组织病理学特征和SA-β-gal染色程度,上调肾组织中Klotho蛋白表达,下调细胞周期阻滞蛋白和SASP蛋白表达。此外,DHZCP和VE能显著调节衰老模型鼠肾脏内源性代谢物;各组共同影响的差异代谢物有21个,其中有5个差异代谢物在衰老模型鼠中显著增加,经DHZCP和VE干预后显著回调,这些代谢物关联脂质代谢和能量代谢等途径;各组曲线下面积为0.88~1;DHZCP主要影响腺嘌呤核糖核苷酸活化蛋白激酶(AMPK)通路、环鸟苷一磷酸-蛋白激酶G(cGMP-PKG)通路、环腺苷酸(cAMP)通路、磷脂酰肌醇3激酶-蛋白激酶B(PI3K-Akt)通路、哺乳动物雷帕霉素靶蛋白(mTOR)通路、自噬通路等;还影响肾素分泌、长寿调节途径、糖尿病心肌损伤以及烟酸和烟酰胺代谢等多条代谢途径。此外,DHZCP和VE能明显上调衰老模型鼠肾组织中AMPK信号通路关键信号分子的蛋白表达。总之,DHZCP和VE在体内能改善衰老模型鼠肾脏衰老和损伤;DHZCP的多靶点作用涉及肾脏内多条信号通路和多条代谢途径;其中,AMPK信号通路可能是其关键作用靶点之一。This study aims to observe the effects of the traditional Chinese medicine prescription Dahuang Zhechong Pills(DHZCP)on renal aging and explore its potential multi-target effects.Rats were assigned into the normal,model,DHZCP,and vitamin E(VE)groups.Firstly,the rat model of D-galactose(D-gal)-induced renal aging was established.During the modeling period,the rats in the 4 groups were administrated with double distilled water,double distilled water,DHZCP suspension,and VE suspension,respectively,by gavage every day.On day 60 of intervention,the indicators of renal aging and injury in rats were measured,including the function,histopathological characteristics,senescence-associatedβ-galactosidase(SA-β-gal)staining,and expression levels of Klotho and proteins associated with cell cycle arrest and senescence-associated secretory phenotype(SASP)in the renal tissue.Moreover,nontargeted metabolomic analysis of the renal tissue was performed for the 4 groups of rats to screen out the potential biomarkers and metabolic pathways.Finally,the signaling pathways of key targets were preliminarily validated.The results showed that DHZCP and VE significantly improved the renal function,histopathological features of renal tubular/interstitial tissue,and degree of SA-β-gal staining,up-regulated the expression level of Klotho,and down-regulated the expression levels of proteins associated with cell cycle arrest and SASP in the renal tissue of the aging rats.In addition,DHZCP and VE regulated the metabolites in the renal tissue of the aging rats.There were 21 common differential metabolites.Among them,5 differential metabolites were significantly increased in the aging rats and recovered after DHZCP or VE treatment,and they were involved in the lipid metabolism and energy metabolism pathways.The areas under the curves of the groups in comparison varied within the range of 0.88-1.DHZCP regulated multiple signaling pathways,such as the adenosine monophosphate-activated protein kinase(AMPK),cyclic guanosine monophosphate-protein kina

关 键 词：大黄蛰虫丸 肾脏衰老 非靶向代谢组学 作用靶点 

分 类 号：R285.5[医药卫生—中药学]