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作者中文名	作者英文名	单位中文名	单位英文名	E-Mail
宋紫腾	SONG Zi-teng	天津药物研究院天津市中药质量标志物重点实验室，天津 300301 天津药物研究院释药技术与药代动力学国家重点实验室，天津 300301	Tianjin Key Laboratory of Quality Marker of Traditional Chinese Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China State Key Laboratory of Drug Delivery Technology and Pharmacokinetics, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China
李莹曼	LI Ying-man	天津中医药大学，天津 301617	Tianjin University of Traditional Chinese Medicine, Tianjin 301617, China
徐旭	XU Xu	天津药物研究院天津市中药质量标志物重点实验室，天津 300301 天津药物研究院释药技术与药代动力学国家重点实验室，天津 300301	Tianjin Key Laboratory of Quality Marker of Traditional Chinese Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China State Key Laboratory of Drug Delivery Technology and Pharmacokinetics, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China
韩彦琪	HAN Yan-qi	天津药物研究院天津市中药质量标志物重点实验室，天津 300301 天津药物研究院释药技术与药代动力学国家重点实验室，天津 300301	Tianjin Key Laboratory of Quality Marker of Traditional Chinese Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China State Key Laboratory of Drug Delivery Technology and Pharmacokinetics, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China
张铁军*	ZHANG Tie-jun	天津药物研究院天津市中药质量标志物重点实验室，天津 300301 天津药物研究院释药技术与药代动力学国家重点实验室，天津 300301	Tianjin Key Laboratory of Quality Marker of Traditional Chinese Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China State Key Laboratory of Drug Delivery Technology and Pharmacokinetics, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China
许浚	XU Jun	天津药物研究院天津市中药质量标志物重点实验室，天津 300301 天津药物研究院释药技术与药代动力学国家重点实验室，天津 300301 中国中药协会药食同源物质评价和利用专业委员会，北京 100061	Tianjin Key Laboratory of Quality Marker of Traditional Chinese Medicine, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China State Key Laboratory of Drug Delivery Technology and Pharmacokinetics, Tianjin Institute of Pharmaceutical Research, Tianjin 300301, China Professional Committee on Evaluation and Utilization of Medicinal and Food Homologous Substances, China Association of Traditional Chinese Medicine, Beijing 100061, China

基金项目:国家自然科学基金重点项目（81830111）；天津市科技计划项目中央引导地方科技发展专项（20ZYCGSN00200）

中文摘要:目的基于网络药理学及模式生物秀丽隐杆线虫（以下简称线虫）体内实验探究药食同源物质肉苁蓉延缓衰老的功能因子与作用机制，为其合理开发利用提供实验依据。方法选取肉苁蓉中主要活性成分为研究对象，运用中药系统药理数据库和分析平台（TCMSP）、SwissTargetPrediction、GeneCards、OMIM等数据库预测肉苁蓉发挥功效的潜在靶点，将其输入到STRING网络分析平台，获得活性成分与疾病靶点的蛋白质-蛋白质相互作用（PPI）网络，从而筛选出核心靶点，并借助京都基因与基因组百科全书（KEGG）平台对作用通路进行预测分析，通过Cytoscape软件构建药物-活性成分-靶点-通路网络。将线虫分为对照组及不同质量浓度给药组，采用直接观察法测定线虫寿命、繁殖能力、运动能力；采用试剂盒方法测定线虫体内超氧化物歧化酶（SOD）、过氧化氢酶（CAT）、谷胱甘肽过氧化物酶（GSH-Px）的活性及丙二醛（MDA）、活性氧（ROS）的含量，采用实时荧光定量聚合酶链式反应（qRT-PCR）测定与寿命及应激能力改变相关的基因表达。结果筛选得到的肉苁蓉潜在功能因子可作用于75个衰老相关靶点，经拓扑学分析筛选出热休克蛋白90α（HSP90AA1）、雌激素受体α（ESR1）、GTP酶（HRAS）、凝血酶原（F2）、基质金属蛋白酶2（MMP2）、己糖激酶-2（HK2）、蔗糖异麦芽糖酶（SI）、麦芽糖酶-葡糖淀粉酶（MGAM）8个核心靶点，KEGG分析显示肉苁蓉可干预雌激素信号通路、癌症信号通路、促性腺激素释放素信号通路和胰岛素信号通路等27条通路，发挥延缓衰老作用。肉苁蓉能够明显延长线虫的平均寿命，提高线虫的抗氧化应激能力，增强SOD、CAT和GSH-Px的活性，降低ROS和MDA含量，下调胰岛素受体样基因daf-2及age-1表达水平，上调daf-16及其下游靶基因（sod-3、mtl-1、gst-4、ctl-1和ctl-2）表达水平，同时激活skn-1及热休克相关基因（hsf-1、hsp-16.1和hsp-16.2）的表达。结论基于网络药理学阐释了肉苁蓉延缓衰老可能的作用机制，进一步实验证明了肉苁蓉可以延长线虫的寿命、增强线虫抵抗环境应激能力，其作用机制可能与抑制胰岛素信号通路并激活daf-16下游转录因子及热休克相关基因，从而提高线虫抗氧化应激能力有关。

中文关键词:肉苁蓉延缓衰老功能因子作用机制氧化应激

Anti-aging Functional Factors and Mechanism of Cistanche deserticola

Abstract:Objective To explore the anti-aging functional factors and mechanism of the edible and medicinal substance Cistanche deserticola based on network pharmacology and in vivo experiments of the model organism Caenorhabditis elegans.Methods Potential targets of the major components in C. deserticola were predicted by Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), SwissTargetPrediction, GeneCards, Online Mendelian Inheritance in Man (OMIM), and other databases. The predicted targets were input into the STRING network analysis platform to obtain the protein-protein interaction (PPI) network of the active components and disease targets, so as to screen out the core targets. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was used to predict and analyze acting pathways, and the medicine-active component-target-pathway network was constructed by Cytoscape. C. elegans was divided into a blank group and C. deserticola groups with different concentrations. The life span, reproductive ability, and exercise ability of C. elegans were observed directly. The activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px) and the content of malondialdehyde (MDA) and reactive oxygen species (ROS) in C. elegans were determined by the kit assay. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to determine the expression of genes related to life span and stress ability change in C. elegans.Results The potential functional factors can act on 75 aging-related targets, and 8 core target proteins, including heat shock protein HSP 90-α (HSP90AA1), estrogen receptor (ESR1), GTPase HRas (HRAS), prothrombin (F2), matrix metalloproteinases-2 (MMP2), hexokinase-2 (HK2), sucrase-isomaltase (SI) and maltase-glucoamylase (MGAM), were screened out by topology analysis. KEGG analysis showed that Cistanche deserticola could interfere with 27 pathways including estrogen signaling pathway, cancer signaling pathway, GnRH signaling pathway, and insulin signaling pathway to delay aging. Cistanche deserticola can significantly extend the average survival time and improve the oxidative stress ability of C. elegans, enhance the activities of SOD, CAT and GSH-Px, and reduce the accumulation of ROS and MDA, and the expression of daf-2 and age-1 were decreased, while the expression of daf-16 and its downstream target genes (sod-3, mtl-1, gst-4, ctl-1 and ctl-2) were increased, as well as skn-1 and heat shock related genes (hsf-1, hsp-16.1 and hsp-16.2).Conclusion The network pharmacology is applied to explain the possible mechanism of C. deserticola in anti-aging, and it has proved that C. deserticola can extend the life span of C. elegans and enhance the anti-stress ability. The mechanism may be related to the inhibition of the insulin signaling pathway and activation of the downstream transcription factors of daf-16 and heat shock related-genes, thus enhancing the oxidative stress ability of C. elegans.

keywords:Cistanche deserticola Y.C.Ma anti-aging functional factors mechanism oxidative stress

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